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  • Meeting abstracts
  • Open Access

10th Congress of International Society of Systemic Auto-Inflammatory Diseases (ISSAID)

Genoa, Italy. 31 March - 3 April 2019
Pediatric Rheumatology201917 (Suppl 1) :18

  • Published:

Oral presentations

Molecular determinants of innate immunity

O01 New MVK mutant mouse avatars of mevalonate kinase deficiency mimic the underlying defect in protein prenylation in patients

Marcia A. Munoz1, Oliver P. Skinner1, Julie Jurczyluk1, Kristen Perry1, Robert Brink2, David Zahra2, Rob J. Arts3, Anna Simon3, Michael J. Rogers1
1Bone Biology; 2Immunology, Garvan Institute of Medical Research, Sydney, Australia; 3Medical Centre, Radboud University, Nijmegen, Netherlands
Correspondence: Marcia A. Munoz

Introduction: Mevalonate Kinase Deficiency (MKD) is a periodic fever syndrome characterised by recurrent bouts of high fever and systemic inflammation. MKD is caused by recessive, hypomorphic mutations in the mevalonate kinase gene (MVK) encoding a key enzyme in the mevalonate pathway. This pathway is responsible for cholesterol synthesis and the production of isoprenoid lipid tags required for post-translational prenylation of proteins. It is believed that inflammation in MKD is triggered by shortage of isoprenoid lipids and defective prenylation of small GTPases. We have found that protein prenylation is indeed compromised in PBMCs from patients and this defect distinguishes MKD from other periodic fever syndromes. However, the link between protein prenylation and inflammation in MKD is still far from understood, largely due to the lack of suitable genetic mouse models.

Objectives: To generate new Mvkmutant mouse models of MKD that mimic the human disease.

Methods: CRISPR/Cas9 gene editing was used to generate different heterozygous mouse lines with hypomorphic mutations in exon 11 of theMvkgene: a V377I substitution (the most frequent mutation in MKD), and 8, 13 or 91 base pair deletions. These lines were then crossed to generate homozygous MvkV377I/V377Ior MvkV377I/deIcompound heterozygous mice. To assess the effect of the mutations on the mevalonate pathway, we measured the accumulation of unprenylated Rab GTPases and Rap1A using an in vitroprenylation assay and western blotting. For quantification of inflammatory cytokines in serum, as well as in culture supernatants from LPS-stimulated PBMCs and bone marrow macrophages, we used ELISA and multiplex cytokine bead arrays.

Results: Homozygous mice carrying complete loss of function deletion mutations were not viable and, as expected, wildtype and heterozygous Mvkmutant mice had normal protein prenylation. However, in a similar pattern to patient PBMCs, MvkV377I/deIimmune cells from blood, spleen and bone marrow had a dramatic accumulation of unprenylated Rab and Rap1A GTPases, with a much milder defect in MvkV377I/V377Icells. This is consistent with reportedly less severe clinical disease associated with the homozygous V377I mutation. Furthermore, MvkV377I/deI mice had slightly elevated levels of inflammatory serum cytokines, including IL-6 and G-CSF, and cultures of PBMCs and bone marrow macrophages responded more robustly to LPS stimulation than cells from control mice. Interestingly, like patient-derived cell lines, the prenylation defect was dramatically enhanced in bone marrow cells from Mvkmutant mice after briefly culturing at higher temperature (39-40oC). Importantly, addition of the missing isoprenoid lipid geranylgeraniol could rescue the prenylation defect in primary cell cultures in vitroand in peritoneal macrophages in vivo.

Conclusion: To our knowledge, we have generated the first genetic mouse avatars of MKD. These mice, like MKD patients, have defective protein prenylation and an exaggerated inflammatory response. Furthermore, as with patient-derived cell lines, Mvkmutant mouse cells are temperature-sensitive, suggesting that elevations in body temperature (e.g. caused by mild infections) could quickly precipitate devastating defects in protein prenylation that lead to systemic inflammatory flares. These mice are exciting new tools to study the pathophysiology of MKD and can be used to develop new therapeutic approaches, such as supplementation with isoprenoid lipids.

Disclosure of Interest

None Declared

O02 Generation and analysis of mice carrying a novel heterozygous missense mutation of a proteasome subunit, PSMB9, in patients with autoinflammation and immunodeficiency

Hiroaki Hemmi1, Nobuo Kanazawa2, Noriko Kinjo3, Satoru Hamada3, Hidenori Ohnishi4, Tsunehiro Mizushima5, Akira Kinoshita6, Koh-Ichiro Yoshiura6, Tsuneyasu Kaisho1
1Department of Immunology, Wakayama Medical University Institute of Advanced Medicine; 2Department of Dermatology, Wakayama Medical University, Wakayama; 3Department of Child Health and Welfare (Pediatrics), University of the Ryukyus Graduate School of Medicine, Nishihara; 4Department of Pediatrics, Gifu University Graduate School of Medicine, Gifu; 5Picobiology Institute, University of Hyogo Graduate School of Life Science, Kamigori; 6Department of Human Genetics, Nagasaki University Atomic Bomb Disease Institute, Nagasaki, Japan
Correspondence: Tsuneyasu Kaisho

Introduction: The proteasome is a large protein complex involved in degradation of unnecessary or useless proteins. Homozygous, compound heterozygous or digenic mutations of proteasome subunits cause autoinflammatory diseases, termed proteasome-associated autoinflammatory syndromes (PRAAS). De novo, heterozygous missense mutation in the proteasome subunit PSMB9 (encodes β1i) gene (hereafter, PSMB9 X mutation), was commonly found in two unrelated patients showing PRAAS-like but distinct manifestations, on which two other posters are presented in this meeting. The PSMB9 X mutation is novel and causes a substitution of an amino acid conserved among multiple species.

Objectives: It is unclear whether and how the PSMB9 X mutation contributes to the manifestations of the patients. In order to clarify this issue, we have generated and analyzed mutant mice carrying the mutation.

Methods: The mice carrying the Psmb9 X mutation were generated with CRISPR/Cas9 technology. Homozygous Psmb9 X mutant mice died within 6 months old. Therefore, the heterozygous Psmb9 X mutant mice were mainly analyzed. Not only biochemical but also immunological analyses including histological and flow cytometry analyses were performed.

Results: Heterozygous Psmb9 X mutant mice appeared healthy at glance. The β1i subunit becomes mature after processing by the proteasome, but this maturation process was impaired in splenocytes of the heterozygous Psmb9 X mutant mice. In the mutant mice, thymus was small and the cortico-medullary junction was unclear. All thymocytes such as CD4+, CD8+, double positive and double negative cells were decreased. In the spleen, B cells as well as CD4+ and CD8+ T cells were decreased. Furthermore, serum levels of immunoglobulins, including IgM, IgGs and IgA, were severely decreased. Dendritic cells (DCs) and all DC subsets were also decreased, although the conventional DC1 subset, defined as CD8α+CD11b- cells, was most severely decreased. Meanwhile, CD11b+ cells consisting mainly of neutrophils and monocytes were increased in the bone marrow. These phenotype of the heterozygous Psmb9 X mutant mice were not identical to, but overlapping significantly with the manifestations of the two patients.

Conclusion: We here generated mutant mice carrying a novel heterozygous missense mutation in the proteasome subunit Psmb9 gene, which was identified in two unrelated PRAAS-like patients. Multiple defects in both innate and adaptive immune cells were observed in the heterozygous Psmb9 X mutant mice and some, although not all, defects were also observed in the two patients. These results indicate that the heterozygous Psmb9 X mutation can be the cause of the PRAAS-like phenotypes in the two patients. The findings that the mutation causes not only autoinflammation but also combined immunodeficiency prompt us to propose a novel category of autoinflammatory diseases distinct from PRAAS as “proteasome-associated autoinflammation and immunodeficiency disease (PRAID)”. The mutant mice are unique and quite useful for clarifying how the proteasome dysfunction leads to various manifestations of PRAID.

Disclosure of Interest

None Declared

Mechanisms of inflammasome activation

O03 Cofilin-1 is an essential redox sensor for NLRP3 inflammasome activation

Wonyong Lee, Yong Hwan Park, Daniel L. Kastner, Jae Jin Chae
NHGRI, Bethesda, United States
Correspondence: Wonyong Lee

Introduction: NLRP3 has a pivotal role in nucleating the inflammasome, a cytoplasmic multiprotein complex that mediates the maturation of the proinflammatory cytokine interleukin-1β (IL-1β) by activating caspase-1. Mutations in the gene encoding NLRP3 cause a spectrum of autoinflammatory diseases, the cryopyrin-associated periodic syndromes (CAPS). The generation of reactive oxygen species (ROS) is one of the major NLRP3 inflammasome activating factors. However, the molecular basis of the relationship between change of cellular redox state and NLRP3 inflammasome activation has not been elucidated.

Methods: We utilized mouse bone marrow-derived macrophage (BMDM) to analyze interaction of cofilin-1 and NLRP3 by co-immunoprecipitation (co-IP). Mouse BMDMs were used to ectopically express wild-type (WT) or mutant cofilin-1 proteins, and to transfect siRNA for knockdown assay. Cofilin-1 knock-in (KI) mice (C39A or C39S) were generated by microinjection of sgRNA and Cas9 ribonucleoprotein (RNP) complex.

Results: To identify an ROS-mediated regulator for NLRP3 inflammasome activation, the immune complexes precipitated by NLRP3 specific antibody from BMDMs of WT or NLRP3-KO mice were analyzed by mass spectrometry. We found cofilin-1, the actin severing protein, as a negative regulator for the NLRP3 inflammasome. Cofilin-1 interacted with the nucleotide-binding domain (NBD) of NLRP3 and dissociated from NLRP3 when the cells were stimulated with known NLRP3 inflammasome activators, such as ATP or nigericin. The NLRP3 inflammasome activators generate ROS that leads to cofilin-1 oxidation, which is intramolecular disulfide bond formation between two cysteine residues at amino acids 39 and 80. This oxidation induces conformational change of cofilin-1 and dissociation from NLRP3, which results in the activation of the NLRP3 inflammasome. Indeed, the assembly of NLRP3 inflammasome components is impaired and the IL-1β release was significantly suppressed in BMDMs ectopically expressing oxidation-resistant mutant cofilin-1 (C39A or C80A). In addition, knockdown of cofilin-1 in LPS-primed BMDMs induced NLRP3 inflammasome activation without activator treatment. We also observed that the interaction of cofilin-1 with the CAPS-associated mutant NLRP3 proteins was substantially diminished relative to WT NLRP3, which resulted in constitutive activation of the NLRP3 inflammasome. To examine the role of cofilin as a redox sensor for NLRP3 inflammasome activation in vivo, we have generated KI mice expressing oxidation-resistant mutant cofilin-1 (C39A or C39S). Unexpectedly, the IL-1β release from the BMDMs of the KI mice was higher than WT BMDMs when the cells were stimulated with ATP after LPS priming. Consistently, IL-1β levels in the serum of KI mice after injection of lipopolysaccharide (LPS) intraperitoneally was significantly higher than WT mice. This inconsistent result may be due to the low level of mutant cofilin-1 in KI mice. Indeed, similarly to the result of cofilin-1 knockdown, LPS-primed KI BMDMs release substantial IL-1β without activators.

Conclusion: Taken together, these findings suggest that cofilin-1 is a key component in regulating the NLRP3 inflammasome in response to ROS. In addition, our data suggest cofilin-1 as a potential therapeutic target for the inflammatory conditions involving the NLRP3 inflammasome, including gout, type 2 diabetes mellitus, atherosclerosis, and Alzheimer’s disease.

Disclosure of Interest

None Declared

O04 Autoinflammatory mutation in NLRC4 reveals an LRR-LRR oligomerization interface

Fiona Moghaddas1,2,3, Ping Zeng4, Yuxia Zhang5, Heike Schuetzle6, Sebastian Brenner6, Sigrun Hofmann6, Reinhard Berner6, Yuanbo Zhao5,7, Bingtai Lu5, Xiaoyun Chen5, Li Zhang5, Suyun Cheng4, Stefan Winkler6, Kai Lehmberg8, Scott W. Canna9, Peter E. Czabotar10,11, Ian P. Wicks2,11,12, Dominic De Nardo2,11, Christian Hendrich6,13,14, Huasong Zeng4, Seth L. Masters2,11
1Clinical Immunology and Allergy, The Royal Melbourne Hospital, Melbourne; 2Inflammation Division, The Walter and Eliza Hall Institute of Medical Research; 3Department of Medical Biology, The University of Melbourne, Parkville, Australia; 4Department of Rheumatology; 5Immunology Laboratory, Guangzhou Women and Children’s Medical Centre, Guangzhou, China; 6Department of Pediatrics, University Hospital and Faculty of Medicine Carl Gustav Carus, Dresden, Germany; 7Department of Chemical Biology, Guizhou Medical University, Guiyang, China; 8Division of Pediatric Stem Cell Transplantation and Immunology, University Medical Center Hamburg Eppendorf, Hamburg, Germany; 9Pediatric Rheumatology/RK Mellon Institute, Children’s Hospital of Pittsburgh ofUPMC, Pittsburgh, United States; 10Structural Biology Division, The Walter and Eliza Hall Institute of Medical Research, Parkville; 11Department of Medical Biology, The University of Melbourne, Melbourne; 12Rheumatology Department, The Royal Melbourne Hospital, Parkville, Australia; 13Department of Women’s & Children’s Health, nstitute of Translational Medicine, University of Liverpool; 14Department of Pediatrics Rheumatology, Alder Hey Children's NHS Foundation Trust Hospital, Liverpool, United Kingdom
Correspondence: Fiona Moghaddas

Introduction: Monogenic autoinflammatory disorders are characterised by dysregulation of the innate immune system. A significant number of this broadening group of disorders are caused by gain-of-function mutations in inflammasome forming proteins, such as NLRC4. A number of mutations in NLRC4 have been described, leading to a spectrum of NLRC4-associated autoinflammatory disorders (NLRC4-AID).

Objectives: We studied two patients with early onset macrophage activation syndrome caused by the same de novo mutation in NLRC4 (c.G1965C, p.W655C). Unlike other mutations in NLRC4 described to date, p.W655 is located within the leucine rich repeat (LRR) domain. For this reason, we investigated mechanisms by which this mutation contributes to the pathogenesis of autoinflammatory disease.

Methods: Next generation and Sanger sequencing techniques were used for genetic analysis. ELISA was performed to quantify serum cytokine levels. In vitro, inflammasome complex formation was quantified using flow cytometric analysis of Apoptosis-associated Speck-like protein containing a Caspase recruitment domain (ASC) specks.Monocyte-like cell lines were generated by genetic deletion of NLRC4 from THP-1 cells using CRISPR/Cas9 techniques followed by lentiviral transduction of wild type (WT) or mutant NLRC4 cDNA. Cell death and release of IL-1b/IL-18 were quantified using flow cytometry and ELISA respectively.

Results: Both reported patients succumbed to macrophage activation syndrome early in life, associated with increased IL-18 serum levels. The NLRC4 mutation identified, c.G1965C/p.W655C, caused increased ASC speck formation in vitro. In THP-1 cells, introduction of c.G1965C/p.W655C NLRC4 resulted in increased cell death, IL-1b and IL-18 production. The enhanced response was independent of NLRP3 and caspase-8. ASC contributed to p.W655C NLRC4 mediated cytokine release, but not cell death. p.W655 is located at the interface between adjacent LRR domains in the oligomeric inflammasome structure. Mutation of p.W655 activates the NLRC4 inflammasome complex by engaging with two interfaces on the opposing LRR domain. One key set of residues (p.D1010, p.D1011, p.L1012 and p.I1015) participates in LRR-LRR oligomerization when it is triggered by NLRC4-AID mutations or T3SS effector (PrgI) stimulation of the NLRC4 inflammasome complex.

Conclusion: This is the first report of a mutation in the LRR domain of NLRC4 causing NLRC4-AID. c.G1965C/p.W655C NLRC4 increases inflammasome activation, leading to constitutive IL-18 production and increased IL-1b release upon priming, where ASC contributes to the cytokine response, but not to cell death. Data generated from various NLRC4 mutations suggests that the tryptophan at p.W655 does not tolerate substitution, and provides evidence that the LRR-LRR interface has an important, previously unrecognized role in oligomerization of the NLRC4 inflammasome complex.

Disclosure of Interest

None Declared

Multifactorial vs monogenic autoinflammatory diseases

O05 Canakinumab, on a reduced dose or a prolonged dose interval without concomitant corticosteroids and methotrexate, maintains efficacy in systemic juvenile idiopathic arthritis patients in clinical remission

Pierre Quartier1, Ekaterina Alexeeva2, Carine Wouters3, Inmaculada Calvo4, Tilmann Kallinich5, Bo Magnusson6, Nico Wulffraat7, Xiaoling Wei8, Alan Slade9, Ken Abrams9, Alberto Martini10
1AP-HP, Institut des Maladies Génétiques (IMAGINE), and Université Paris-Descartes, Necker-Enfants Malades Hospital, Paris, France; 2National Medical Research Center of Children's Health and Sechenov First Moscow State Medical University of the Ministry of Health of the Russian Federation, Moscow, Russian Federation; 3Gasthuisberg University Hospital,Leuven, Belgium; 4Hospital Universitario La Fe, Valencia, Spain; 5Charité Berlin Campus Virchow, Berlin, Germany; 6Karolinska University Hospital, Stockholm, Sweden; 7University Medical Center Utrecht, Utrecht, Netherlands; 8China Novartis Institutes for Biomedical Research Co., Ltd, Beijing, China; 9Novartis Pharmaceuticals Corporation, East Hanover, United States; 10Universita di Genova Pediatria II, Genova, Italy
Correspondence: Ekaterina Alexeeva

Introduction: Treatment with canakinumab (CAN), a selective, human anti-IL-1β monoclonal antibody, has shown sustained therapeutic effect along with corticosteroid dose reduction/discontinuation in patients with systemic juvenile idiopathic arthritis (SJIA), in a long-term extension study (NCT00891046).1

Objectives: To evaluate the efficacy and safety results from a study evaluating 2 different canakinumab tapering regimens in SJIA patients who were in clinical remission (NCT02296424).

Methods: This Phase 3b/4 study had two parts. In Part I, 182 patients with inactive disease from the extension study1 (cohort 1) and CAN-naïve patients (cohort 2) with active disease were administered subcutaneous CAN 4 mg/kg q4w. Per protocol titration off corticosteroids and/or methotrexate was attempted during Part I. Eligible patients (inactive disease for 24 consecutive weeks and being corticosteroid- and methotrexate-free for at least 4 weeks) advanced to Part II. Patients were randomised to either a 3-step CAN dose reduction regimen (2mg/kg/q4w, followed by tapering to 1 mg/kg/q4w and then discontinuation) or dose interval prolongation regimen (4mg/kg q8w, followed by tapering to 4 mg/kg/q12w and then discontinuation); patients advanced to the next tapering step if inactive disease was maintained for 24 weeks. The primary objective was to evaluate if at least 40% of patients were able to maintain inactive disease status for at least 24 consecutive weeks on either 2mg/kg q4w or 4mg/kg q8w.

Results: In Part II, a total of 75 patients were randomised to a dose reduction (n=38) or dose interval prolongation (n=37) in CAN tapering regimen. The proportion of patients who maintained inactive disease for 24 consecutive weeks exceeded the predefined threshold of 40% for Step 1 of: the reduced CAN dose (71%; 2 mg/kg q4w) and prolonged dose interval (84%; 4 mg/kg q8w) treatment arms. A total of 68% (26/38) and 79% (30/37) of the dose reduction and interval prolongation arms, respectively were successful in Step 2, while only 33% (25/75) of patients successfully discontinued CAN and maintained inactive disease for 24 consecutive weeks. Adverse events (AEs) and serious AEs observed within the 2 treatment cohorts and across Parts I and II were similar without any specific pattern or relationship to patients’ disease status at baseline or treatment regimen. The most frequent AEs were common infections such as nasopharyngitis, upper respiratory tract infection, and pharyngitis followed by SJIA-related events such as rash, pyrexia and arthralgia. Clinical laboratory abnormalities were consistent with expected findings in patients with active SJIA and the known safety profile of CAN.

Conclusion: SJIA patients who are able to maintain inactive disease status on CAN monotherapy can successfully taper CAN by either reducing the dose or prolonging the dosing interval. However, only a minority of patients successfully discontinued CAN treatment for 24 weeks. The safety profile for both CAN titration regimens was similar and consistent with other CAN SJIA studies. No new safety signals were identified.


1. Brunner et al. Arthritis Rheumatol.2016; 68 (S10).

Disclosure of Interest

P. Quartier Consultant for: Abbvie, Lilly, Novimmune, Novartis and SOBI,Speaker Bureau of: AbbVie, Lilly, Novartis and SOBI, E. Alexeeva Grant / Research Support from: Roche, Abbott, Pfizer, Bristol-Myers Squibb, Centocor, Novartis, C. Wouters Consultant for: GSK, Roche, Pfizer , I. Calvo: None Declared, T. Kallinich Speaker Bureau of: Novartis, B. Magnusson: None Declared, N. Wulffraat Consultant for: Novartis, X. Wei Employee of: Novartis, A. Slade Conflict with: Novartis Pharmaceuticals CorporationShareholder of: Novartis Pharmaceuticals Corporation,Employee of: Novartis Pharmaceuticals Corporation, K. Abrams Conflict with: Novartis Pharmaceuticals CorporationShareholder of: Novartis Pharmaceuticals Corporation,Employee of: Novartis Pharmaceuticals Corporation, A. Martini: None Declared

O06 IL-18:CXCL9 ratio as a predictor of treatment response in patients with systemic juvenile idiopathic arthritis treated with canakinumab

Tanja Hinze1, Christoph Kessel1, Claas Hinze1, Julia Seibert2, Hermann Gram2, Dirk Foell1
1Department of Pediatric Rheumatology and Immunology, Muenster University Hospital, Muenster, Germany; 2Novartis Pharma, Basel, Switzerland
Correspondence: Tanja Hinze

Introduction: Canakinumab, a monoclonal anti-interleukin (IL)-1β antibody, is highly effective for treating patients with systemic juvenile idiopathic arthritis (SJIA) but biomarkers predicting treatment response are desirable.

Objectives: The objective of this study was to analyze the association of various serum biomarkers with treatment outcomes.

Methods: Serum samples from 54 patients treated with canakinumab in an open-label long-term extension study were studied by Luminex at different time points during the study, including days 1 (baseline), 3, 15 and weeks 4, 8, 24, 48. Treatment outcomes included modified pediatric American College of Rheumatology (pACR) 30/50/70/90/100 responses within 15 days of treatment, clinically inactive disease (CID) according to the Wallace criteria within 15 days of treatment and sustained complete response, defined as pACR100or CID within 15 days of treatment plus no disease flare or macrophage activation syndrome (MAS) during the study. Data were analysed using non-parametric testing and receiver operating characteristic (ROC) analysis.

Results: Within 15 days of treatment with canakinumab, 79%/77%/68%/49%/34% of the patients reached a modified pACR 30/50/70/90/100 response and 34% CID. Within a median follow-up of 23 months, 12 of 54 (22%) patients had a sustained complete response and 5 (9%) had developed MAS. Biomarkers did not correlate significantly with age, duration of disease or active joint count. Most biomarkers were elevated when compared to healthy controls at baseline and some rapidly decreased within 15 days of therapy (IL-1RA, IL-6, IL-18 and S100A12). A pattern was apparent when comparing responders and non-responders; responders had higher IL-18 and IFN-γ levels and lower CXCL9 levels at baseline, most emphasized by the IL-18:CXCL9 and the IFN-γ:CXCL9 ratios (Table 1). As determined via ROC analyses, these ratios had good accuracy in predicting treatment responses relating to pACR30/50/70/90/100/CID responses at day 15 and sustained complete response (area under the curve [AUC] for IL-18:CXCL9 ratio 0.79/0.75/0.67/0.72/0.77/0.71 and 0.80, respectively; and for the IFN-γ:CXCL9 ratio 0.79/0.76/0.71/0.75/0.77/0.76 and 0.79). Higher baseline CXCL9 levels predicted future MAS during the course of the study (ROC analysis AUC = 0.77).

Conclusion: Several serum biomarkers were markedly elevated in patients with SJIA at baseline. A dysregulation of the IL-18-IFN-γ-CXCL9 axis is present in patients with SJIA, confirming findings from other investigators. However, our findings indicate that patients with a lower response to IL-18 and IFN-γ, as measured by CXCL9, an IFN-γ-induced chemokine, may have a better clinical response to canakinumab treatment. These findings will have to be confirmed in other cohorts.

Disclosure of Interest

T. Hinze Grant / Research Support from: Study was funded by Novartis Pharma , C. Kessel Grant / Research Support from: Study was funded by Novartis Pharma, C. Hinze Grant / Research Support from: Study was funded by Novartis Pharma, J. Seibert Employee of: Novartis Pharma, H. Gram Employee of: Novartis Pharma, D. Foell Grant / Research Support from: Study was funded by Novartis Pharma

Table 1 (abstract O06).

Median (range) IFN-gamma: CXCL9 and IL-18:CXCL9 ratios at baseline


IL-18:CXCL9 ratio at baseline

IFN-γ:CXCL9 ratio at baseline



P value*



P value*

pACR30 at day 15

2.54 (0.03-557.31)

0.34 (0.01-1.62)


5.27 (0.19-68.32)

0.82 (0.24-4.85)


pACR50 at day 15

2.37 (0.03-557.31)

0.36 (0.01-3.76)


5.23 (0.19-68.32)

1.35 (0.24-5.38)


pACR70 at day 15

2.54 (0.03-557.31)

0.57 (0.01-42.27)


5.27 (0.19-68.32)

1.56 (0.24-20.03)


pACR90 at day 15

3.69 (0.03-557.31)

0.57 (0.01-42.27)


7.51 (0.19-68.32)

1.88 (0.19-20.03)


pACR100 at day 15

3.99 (0.05-557.31)

0.57 (0.01-42.27)


8.35 (0.19-68.32)

1.97 (0.19-43.46)


CID at day 15

3.84 (0.03-557.31)

0.66 (0.01-42.81)


8.34 (0.62-68.32)

1.97 (0.19-41.52)


Sustained complete response

5.52 (0.03-557.31)

0.65 (0.01-277.69)


8.35 (0.62-68.32)

2.01 (0.19-43.46)


*Mann-Whitney U test

Update of autoinflammatory diseases

O07 The NIH cohort study of DADA2 patients: novel insights into pathophysiology and treatment with TNF inhibitors

Qing Zhou1,2, Natalie Deuitch1, Dan Yang3, Natalia Sampaio Moura1, Xiaomin Yu4, Oskar Schnappauf1, Patrycja Hoffmann1, Deborah Stone1, Amanda Ombrello1, Manfred Boehm3, Daniel Kastner1, Ivona Aksentijevich1
1NHGRI/NIH, Bethesda, United States; 2Zhejiang University, Hang Zhou, China; 3NHLBI/NIH; 4NIAID/NIH, Bethesda, United States
Correspondence: Qing Zhou

Introduction: Deficiency of Adenosine Deaminase 2 (DADA2) is a recessively inherited disorder caused by a loss of functional ADA2 protein. A broad spectrum of features, including systemic inflammation, cutaneous, neurologic, musculoskeletal, and immunological manifestations have now been associated with DADA2. Given the highly polymorphic nature of the ADA2 and the complex clinical presentations of DADA2, large cohort studies are critical to advancing our knowledge of ADA2’s role in disease manifestation and pathogenesis.

Objectives: To expand on the genetics of patients with DADA2 and explore the pathophysiology and the underlying mechanisms of TNF inhibitor response in these patients.

Methods: We performed Sanger sequencing of the ADA2 gene (previously known as CECR1) and measured ADA2 enzyme activity in serum samples of patients and family members. We used flow cytometry, intracellular cytokine staining, transcriptome analysis, immunohistochemistry and cell differentiation experiments to define an inflammatory signature in DADA2 patients and studied their response to TNF inhibitor treatment.

Results: We have identified 60 patients with DADA2 and detected 10 pathogenic variants, which had not previously been associated with DADA2 – p.R9W, p.R34W, p.W204C, p.E244A, p.D329N, p.L351Q, p.A357T, p.P425A, p.P435A, p.W501*.Patients tested for ADA2 enzymatic activity had significantly decreased protein activity. Symptoms were highly variable among patients, even in cases with identical genotypes.

We identified distinct inflammatory signatures. Patients had significantly higher subsets of CD14+ inflammatory monocytes than healthy controls. Phosphorylation of STAT1 was upregulated in patients CD4+ cells and monocytes following stimulation. Additionally, we observed increased gene expression of IP-10 in patients’ monocytes and macrophages. Furthermore, we observed strong NF-κB signaling in patients. Intracellular cytokine staining and qPCR for IL-1β, IL-6, and TNF cytokines were elevated in patients’ monocytes.

After TNF inhibitor treatment, IFN and NF-κB inflammatory signatures were normalized. Analyses of post-treatment skin, lung and brain biopsies showed minimal perivascular TNF and resolution of inflammatory myeloid cell infiltrates. Immunostaining of skin biopsies revealed intact blood vessels with normal endothelial layers after treatment. Together, the data provides evidence that TNF inhibition both reduces systemic inflammation and improves endothelial integrity in the small vessels.

ADA2 deficiency affects the differentiation of monocytes to anti-inflammatory M2 macrophages. Treatment with TNF-inhibitors rescued the impairment of M2 differentiation as demonstrated by improved cell morphology and a higher number of M2 macrophages.

Conclusion: We report 10 novel pathogenic variants in ADA2, which is valuable for future DADA2 molecular diagnostic. Most important, we showed the cellular mechanism underlying effective treatment with anti-TNF therapies. DADA2 vasculitis is strongly related to the presence of activated myeloid cells and is reversible.

Disclosure of Interest

None Declared

O08 Recommendation on colchicine dosing and definition of colchicine resistance/intolerance in the management of FMF

Seza Ozen1, Erdal Sag1, Eldad Ben-Chetrit2, Marco Gattorno3, Ahmet Gul4, Philip Hashkes5, Isabelle Kone-Paut6, Helen Lachmann7, Elena Tsitsamis8, Marinka Twilt9, Fabrizio de Benedetti10, Jasmin B. Kuemmerle-Deschner11
1Pediatric Rheumatology, Hacettepe University, Ankara, Turkey; 2Rheumatology Unit, Hadassah-Hebrew University Hospital, Jerusalem, Israel; 3Clincal Pediatrics and Rheumatology, Gaslini Institute, Genova, Italy; 4Department of Internal Medicine, Division of Rheumatology, Istanbul University, Istanbul, Turkey; 5Pediatric Rheumatology Unit, Department of Pediatrics, Shaare Zedek Medical Center, Jerusalem, Israel; 6Service de rhumatologie pédiatrique, CHU de Bicêtre, Le Kremlin-Bicêtre, France; 7Royal Free Campus, National Amyloidosis Centre, London, United Kingdom; 8First Department of Pediatrics, Aghia Sophia Childrens Hospital, University of Athens Medical School, Athens, Greece; 9Rheumatology, Department of Pediatrics, Alberta Children’s Hospital, University of Calgary, Calgary, Canada; 10Division of Rheumatology, Ospedale Pediatrico Bambino Gesù, Rome, Italy; 11Department of Pediatrics, Division of Pediatric Rheumatology, University Hospital Tuebingen, Tuebingen, Germany
Correspondence: Jasmin B. Kuemmerle-Deschner

Introduction: FMF is the most common monogenic autoinflammatory disease and colchicine is the drug of choice for the treatment. However, about 5-10% of FMF patients do not respond to colchicine even when they are fully compliant. Anti-IL1 treatments are used for the patients who are resistant to colchicine. Treatment with IL-1 inhibitors have been shown to be effective in clinical trials and in several case series.

Objectives: The objective of this report is to produce evidence-based recommendations to define “colchicine resistance“, as well as compliance and intolerance, to guide rheumatologists and other health professionals in the treatment and follow-up of patients with colchicine-resistant FMF.

Methods: A consensus meeting with 12 experts followed a systemic literature review and Delphi questionnaire. The expert committee consisted of adult and pediatric rheumatologists with expertise in FMF. Parameters for colchicine resistance/intolerance/compliance derived from the literature were evaluated by a pre-meeting online questionnaire. All parameters were discussed with a nominal group technique during the meeting. Recommendations were accepted if more than 80% agreement was reached. If agreement was below 80% a second round of discussion was held.

Results: The systematic literature review yielded 264 articles. Of these, 38 were selected for expert review. After the literature review, Delphi survey, and round table discussion, recommendations that reached consensus levels were:

Colchicine is the drug of choice for the treatment of FMF and compliance is a critical issue. For the following statements, it is assumed that the patient is compliant with colchicine.

When utilizing colchicine to treat FMF, it is recommended to adjust the dose based on disease activity with the maximal dose in children depending on age (and weight).

The maximum recommended colchicine dose for the treatment of FMF is between 1-3 mg per day depending on age, limited by signs of toxicity and tolerability.

For a patient receiving the maximum tolerated dose of colchicine, resistance to colchicine is defined as ongoing disease activity (as reflected by either recurrent clinical attacks (average one or more attacks per month over a three-month period), or persistently elevated CRP or SAA in between attacks (depending on which is available locally)), in the absence of any other plausible explanation.

AA amyloidosis develops as a consequence of persistent inflammation, which may be a manifestation of colchicine resistance.

Colchicine intolerance, which generally manifests as GI symptoms (such as diarrhea and nausea), is common and can limit the ability to achieve or maintain the effective dose. Dose-limiting toxicity is rare and may include elevated LFT, leukopenia, azoospermia etc.

Active disease and intolerance to colchicine affect quality of life.

Various patient reported outcomes to be used to guide FMF disease management were outlined.

Conclusion: The suggested recommendations are intended to improve patient care in FMF, to make a personalized treatment plan.

Disclosure of Interest

S. Ozen Speaker Bureau of: Novartis, SOBI, Pfizer, E. Sag: None Declared, E. Ben-Chetrit: None Declared, M. Gattorno: None Declared, A. Gul: None Declared, P. Hashkes Grant / Research Support from: Novartis,Consultant for: Novartis, I. Kone-Paut: None Declared, H. Lachmann: None Declared, E. Tsitsamis: None Declared, M. Twilt: None Declared, F. de Benedetti: None Declared, J. Kuemmerle-Deschner Grant / Research Support from: Novartis, SOBI,Consultant for: Novartis, SOBI

Oral communications – clinical

O09 The clinical spectrum of the deficiency of adenosine deaminase 2 (DADA2) continues to expand

Karyl Barron1, Amanda Ombrello2, Debra Stone2, Patrycja Hoffmann2, Tina Romeo2, Anne Jones2, Natalia Sampaio Moura2, Oskar Schnappauf2, Ivona Aksentijevich2, Jenna Bergerson1, Ariane Soldatos3, Camilo Toro2, Dan Kastner2
1NIAID; 2NHGRI; 3NINDS, NIH, Bethesda, United States
Correspondence: Karyl Barron

Introduction: The original reports of the deficiency of adenosine deaminase 2 (DADA2) in 2014 emphasized early-onset lacunar strokes, livedoid rash, intermittent fevers and early-onset polyarteritis nodosa. Since then, there have been reports of antibody deficiency, pure red cell aplasia, and cytopenias observed in DADA2 patients.

Objectives: To document the range of clinical manifestations of DADA2.

Methods: 46 patients were enrolled in an IRB approved study at the NIH. Sequencing of ADA2, the gene encoding ADA2 (adenosine deaminase 2), was performed on all patients. All underwent extensive clinical, laboratory & radiologic evaluation.

Results: We evaluated 46 patients with DADA2 (24 F/22 M) including 6 sibling pairs & 2 families with 3 affected individuals. All patients had biallelic germline mutations in ADA2. Serum ADA2 enzyme activity levels were obtained in 32 patients and revealed absent to low levels compared to age-matched controls.

The 46thpatient, seen 5 years after bone marrow transplantation for presumed GATA2 deficiency, was not included in our summary calculations.

32 patients (71%) reported a history of recurrent fevers. 6 patients (13%) had diffuse lymphadenopathy.

Skin involvement was seen in 38 patients (84%) including livedo in 34 (76%), cutaneous polyarteritis nodosa in 27 (60%), and Raynaud’s in 9 (20%).

22 patients (49%) had a history of at least one stroke.Brain MRI showed evidence of ischemic infarcts in l6/22 (73%), 5 had both ischemic & hemorrhagic strokes (23%) and 1 had a hemorrhagic stroke (5%). There were 55 strokes in the 22 patients, the majority occurring in the brain stem and cerebellum (38%) and deep brain nuclei (36%). The average age at the time of the first stroke was 5.6 years (range 5 months-20 years). Stroke patients had an average of 3 strokes (range of 1-11). 3 patients manifest severe sequelae of hemorrhagic strokes.

Abdominal ultrasound revealed hepatomegaly in 20 patients (44%) & splenomegaly in 26 (58%). Portal hypertension was observed in 7 patients (16%). Liver biopsies revealed hepatoportal sclerosis in 5 patients and focal nodular regenerative hyperplasia in 2. Abdominal MRA was abnormal in 7/13 patients, revealing arteritis and aneurysms.

Significant peripheral vasculopathy was seen in 4 patients, one requiring multiple amputations of gangrenous digits. Systemic hypertension was observed in 11 patients (24%).

Laboratory evaluation revealed hypogammaglobulinemia in 26 patients (58%). Immunoglobulin replacement was required in 10 patients. Lymphocyte phenotyping revealed arrested B cell class switching in 24/34 patients (71%) and decreased memory T cells in 11/34 (32%). Severe hematologic abnormalities, including anemia, leukopenia, lymphopenia and/or thrombocytopenia in 18 patients (40%), with 6 developing pancytopenia and 3 pure red cell aplasia. Immune mediated neutropenia was observed in 12 patients. ESR and C-reactive protein were elevated in 73% & 86%, respectively.

6 patients underwent bone marrow transplantation with 4 patients successfully engrafted (2 requiring a second transplant), and 2 recently transplanted.

Conclusion: The spectrum of DADA2 continues to expand to include ischemic and hemorrhagic strokes, cutaneous findings, portal and systemic hypertension, hematologic abnormalities, vascular pathology, immune deficiency and bone marrow failure. As the phenotypic presentation is likely to continue to expand, it is important to investigate any new complaints.

Disclosure of Interest

None Declared

O10 Serum S100A8/A9 (calprotectin) in familial mediterranean fever and carriers of MEFV mutations does not correlate with disease activity

Ruth Pepper1, Mathew Hutchinson2, Scott R. Henderson3, Sarah K. Todd3, Alan D. Salama3, Philip N. Hawkins4, Dorota Rowczenio4, Helen J. Lachmann4
1Centre for Nephrology, UCL; 2Rheumatology, University College Hospital; 3Centre for Nephrology; 4National Amyloidosis Centre, UCL Division of Medicine and Royal Free Hospital NHS Foundation Trust, London, United Kingdom
Correspondence: Ruth Pepper

Introduction: Familial Mediterranean Fever (FMF) is caused by mutations in MEFV. The protein product pyrin is expressed in monocytes, neutrophils and eosinophils. Acute inflammatory attacks are accompanied by a dramatic hepatic acute phase response. S100A8/A9 is damage associated molecular pattern and a TLR4 ligand expressed in neutrophils, monocytes and early infiltrating macrophages.

Objectives: We aimed to investigate S100A8/A9 in 39 patients with FMF, 45 healthy carriers and 16 wild type controls.

Methods: All patients were genotyped. Patients and healthy controls (HC) serum S100A8/A9 levels, cell surface expression on monocytes and neutrophils as well as intracellular peripheral blood mononuclear cells (PBMC) expression were measured by flow cytometry (FACS). CD14 cells were isolated and following overnight incubation with or without LPS, S100A8/A9 was measured in the supernatants by ELISA. Patient and HC monocyte apoptosis was compared.

Results: Serum levels were measures in 84 samples from 31 patients with homozygous or compound mutations (median 9061ng/ml [range 500-38470], 79 samples from 39 symptomatic patients who were MEFV heterozygotes (median 9394ng/ml [range 1744-38119], 80 samples from 45 individuals with MEFV variants but without clinical features of FMF (median 10939ng/ml [range 2447->40000]. There was no difference in calprotectin concentrations between the different mutations and no correlation with levels of the hepatic acute phase response, CRP or SAA. All the groups described had significantly higher levels than healthy controls (n=16 median 2836ng/ml [range 1058-6175])(p<0.001). Minimal monocyte and neutrophil cell surface expression was detectable. Following LPS stimulation there was significantly more S100A8/A9 detected in the supernatants in patients than healthy control CD14. There was also a trend to an increased intracellular monocyte S100A8/A9 expression.

Conclusion: Patients with pyrin mutations both with and without clinical disease have greatly elevated serum S100A8/A9 levels without detectable cell surface expression in well-controlled disease with a trend to an increased monocyte intracellular expression. Upon monocyte stimulation with LPS, increasedS100A8/A9 is secreted. The exact mechanism by which these patients, especially those with mutations but no clinical disease, demonstrate sustained elevated serum S100A8/A9 remains to be elucidated but does not appear to result in a significant clinical sequelae.

Disclosure of Interest

None Declared

O11 PAPA syndrome: novelties from the Eurofever registry

Roberta Caorsi, Daniela Marotto, Antonella Insalaco, Angelo Marzano, Joost Frenkel, Graciela Espada, Immaculada Calvo Penades, Marijia Jelusic, Maria Cristina Maggio, Joost Swart, Esther Hoppenreijs, Ozgur Kasapcopur, Fabrizio De Benedetti, Marco Gattorno, The Pediatric Rheumatology International Trial Organization (PRINTO) and the Eurofever Project
Center for Autoinflammatory Diseases and Immunodeficiency, Istituto G. Gaslini, Genova, Italy
Correspondence: Roberta Caorsi

Introduction: PAPA syndrome is a very rare autoinflammatory condition. Few data are nowadays available about the clinical characteristic, the response to treatment and the outcome of this disease.

Objectives:To analyse the data of the PAPA patients enrolled to the Eurofever registry.

Methods: the data analysed in the study were extracted from the Eurofever registry, which is hosted in the PRINTO website ( The patients were included in the study in the presence of mutations in the PSTPIP1 gene or, in genetically negative patients, in the presence of at least two of the following clinical manifestation: recurrent pyogenic arthritis, pyoderma gangrenousm or skin abscess with negative cultural tests. Demographic data, clinical manifestations and response to treatment were analysed.

Results: In may 2018 baseline and clinical information were available of near 4000 patients in the Eurofever registry. Of the 36 patients classified as PAPA syndrome, 2 were excluded from the study. 34 PAPA patients, from 11 different centers, were analysed: the genotype was confirmatory in 29 patients, while in 5 was not available. 10 patients were of the same family, in 4 cases one parent was affected (2 included in the registry), while in other 8 patients the family history was negative. At the time of enrolment, 15 patients were in the paediatric age, while 19 were adults. The mutations detected in the PSTPIP1 gene were E250Q (13 pts), E250K (5 pts), A230T (3 pts), G258A (3 pts), E277D (2 pts), E257G (1 pt), G940A (1pts) and R365W (1 pts).

The disease course was recurrent in 24 patients, while the other 10 presented a chronic disease course with periodic recrudesces. Joint and skin involvement were present at disease onset in 24 and 9 patients respectively. In other 12 patients skin involvement appeared over time. 20 out of the 34 patients presented clinical manifestations not typical of PAPA syndrome (psoriasis, uveitis, osteolytic bone lesions, chronic renal failure, muscular abscesses, gastrointestinal symptoms anaemia and hepatosplenomegaly).

10 patients were treated with NSAID with partial and poor response in 6 and 4 patients respectively, while steroids caused a complete or partial control of disease manifestations in 6 and 10 patients respectively. Five patients were treated with methotrexate with partial response. Etanercept was used in 6 patient with complete response in 2 and partial in 4, adalimumab in 4 patients (1 partial and 1 complete responders, 2 failure) and anakinra in 9 patients (3 partial and 6 complete responders). 2 patients were treated with Canakinumab with complete response.

Conclusion: This study enlightens the phenotypic variability of PAPA syndrome. The unusual clinical manifestations and the lack of the clinical triad of the disease may be responsible for the under recognition of this disease. Between biologic drugs, IL-1 inhibitors were more effective in the analysed cohort of patients.

Disclosure of Interest

None Declared

O12 New classification criteria for recurrent autoinflammatory diseases applied to an independent cohort: experience from the JIR cohort database

Glory Dingulu1, Sophie Georgin-Lavialle2, Isabelle Koné-Paut3, Pascal Pillet4, Anne Pagnier5, Etienne Merlin5, Daniela Kaiser6, Alexandre Belot7, Michael Hofer8, Véronique Hentgen9
1Centre Hospitalier Versailles, Le Chesnay; 2Service de Médecine Interne-CEREMAIA, Centre Hospitalier Universitaire Tenon, Paris; 3Service de Rhumatologie Pédiatrique-CEREMAIA, Centre Hospitalier Universitaire Le Kremlin Bicêtre, Le Kremlin Bicêtre; 4Service d’Accueil des Urgences Pédiatriques, Centre Hospitalier Universitaire Pellegrin, Bordeaux; 5Service de Pédiatrie Générale, Centre Hospitalier Universitaire Clermont Ferrand, Clermont Ferrand, France; 6Service de Pédiatrie Générale, Centre Hospitalier Cantonal Luzern, Luzern, Switzerland; 7Service de Néphrologie-Rhumatologie Pédiatrique, Centre Hospitalier Universitaire Mère-Enfant, Bron, France; 8Service de Rhumatologie Pédiatrique, Centre Hospitalier Universitaire Vaudois, Lausanne, Switzerland; 9Service de Pédiatrie Générale-CEREMAIA, Centre Hospitalier Versailles, Le Chesnay, France
Correspondence: Glory Dingulu

Introduction: New classification criteria for the inherited periodic fever syndromes Cryopyrin Associated Periodic Syndrom (CAPS), Tumour Necrosis Factor Receptor Associated Periodic Syndrom (TRAPS), Familial Mediterranean Fever (FMF), Mevalonate Kinase Deficiency (MKD), have recently been developed during a Consensus Conference held in Genoa in March 2017.

Objectives: The aim of our study was to compare these new classification criteria for monogenic recurrent fever syndromes with the diagnoses of clinicians in a real-life setting.For this purpose we used the JIRcohort database, an international platform gathering data of patients with pediatric inflammatory disease.

Methods: Patients with recurrent fever syndrome and complete clinical and genetic data were enrolled to the study from the auto-inflammatory module of the JIRcohort database. Patients genotype were characterized with HRF pathogenicity classification. A score from 0 to 2, 0 (no mutation), 1 (non-confirmatory genotype) and 2 (confirmatory genotype) was attributed to each gene screened in one patient. The new Genoa classification criteria were applied to all the patients and then compared to the diagnosis of the treating physician. The treating physician diagnosis was considered as standard reference. If the treating physician hesitated between two or more diagnoses, patients were redefined as Syndrome of Unexplained Recurrent Fever (SURF). SURF and PFAPA patients were pooled together. Finally, for each criteria sensitivity and specificity were determined before an analytical study, describing true positive, false positive and false positive patients.

Results: 455 patients were included:10 CAPS, 11 MKD, 27 TRAPS, 122 FMF and 285 SURF/PFAPA patients.

CAPS classification criteria showed a 60 % sensitivity and 98% specificity. 6 patients were true positive patients with confirmatory and non-confirmatory NLRP3 genotype. 4 were false negative patients with non-confirmatory genotype or no mutation in NLRP3. 8 were false positive with no mutation in NLRP3.

TRAPS classification criteria showed a 100 % sensitivity and 98% specificity. 22 were true positive patients with confirmatory and non-confirmatory TNFRSF1A genotype. 5 patients were false positive with non-confirmatory genotype or no mutation in TNFRSF1A.

FMF classification criteria showed a 96 % sensitivity and 88% specificity. 117 patients were true positive with confirmatory, non-confirmatory and non mutated MEFV genotype. 5 were false negative with non-confirmatory and non mutated genotype. 37 werefalse positive patients with patients with non-confirmatory genotype or non mutated genotype or no MEFV screening.

MKD classification criteria showed a 64 % sensitivity and 66% specificity. 7 patients were true positive patients with confirmatory MVK genotype.4 were false negative patientswith non-confirmatory genotype or non mutated MVK genotype. 148 were false positive patients with non mutated MVK genotype.

Conclusion: This work is the first to study Genoa criteria, in real-life setting, in a cohort of patients seen with recurrent fever. Genoa criteria showed tremendous performance for patients with confirmatory genotype and helped classifying patients with non-confirmatory genotype.

Genoa classification criteria were less effective when patients did not display at least one genetic variant. Implementation of biological criteria in MKD might improve MKD criteria performance.

The major limit of our study is the lack of a proper gold standard when genotype is not confirmatory. Nevertheless our study shows that the new classification criteria are of a high risk of misclassification in patients displaying a recurrent fever syndrome without genetic test.

Disclosure of Interest

None Declared

O13 Impaired platelet functions in patients treated with colchicine

Özlem Çimen1, Selcan Demir2, Erdal Sağ2, Armağan Keskin1, Yelda Bilginer2, Şule Ünal Cangül3, Seza Özen2
1Department of Pediatrics; 2Department of Pediatric Rheumatology; 3Department of Pediatric Hematology, Hacettepe University Medical Faculty, Ankara, Turkey
Correspondence: Selcan Demir

Introduction: Colchicine has been used in the treatment of Familial Mediterranean Fever (FMF) since 1972. Apart from the inhibiting mitosis in all cells, colchicine has an anti-inflammatory effect by inhibiting activation and migration of neutrophils. Colchicine is a safe drug at recommended doses, but it can cause rare side effects including hematological findings such as lymphopenia, thrombocytopenia and neutropenia.

Objectives: In this study we aimed to define the adverse effect of colchicine on platelet function and its clinical relevance.

Methods: A total of 220 FMF patients between June 2016-2017, followed at Hacettepe University Pediatric Rheumatology Department and were on colchicine treatment for at least one year, were included to the study.

Results: Among the selected 220 FMF patients, 100 of them (54% female) described hematological symptoms when questioned in detail. The mean age of these patients was 11.74 ± 4.86 years. The mean cumulative colchicine exposure was 5.7±3.8 years.

The most common referral symptom was frequent epistaxis (79%) followed by easy bruising (69%), and menstrual disorder including prolonged or heavy menstrual bleeding (21.8% among female patients). Among these 100 patients, 36 of them had prolonged bleeding time and impaired platelet aggregation test. Patients who had abnormal platelet function tests (the group with abnormal bleeding time) were receiving higher colchicine doses (median 0.05 vs 0.03 mg/kg/day; p:0.001) compared to the patients who had normal platelet function tests (bleeding time normal group) However there were no significant difference in terms of cumulative colchicine exposure (median 6.5 vs 4.5 years; p:0.07) and total platelet counts (median 288500 vs 279000/mm3; p:0.61). Patients with abnormal platelet function tests (bleeding time abnormal group) also had more epistaxis (47% vs 7%; p<0.001), bruising (51% vs 3%; p<0.001) and dysmenorrhea (among female patients 100% vs 26%; p<0.001) (Figure 1 and 2). Colchicine was not reduced in these patients and no life-threatening event was observed.

Conclusion: In our study, we have shown prolonged bleeding time for the first time in the literature. Colchicine may cause microtubule inhibition in platelets as well as in other cells and impair platelet function. Further prospective studies are needed to clarify the significance of this side effect.

Disclosure of Interest

None Declared

O14 Autoinflammatory disorders in patients with myelodysplastic syndrome: the role of distinctive karyotypes and somatic mutations

Mark Kacar1, Abdulla Watad2, Nicola Bragazzi3, Qiao Zhou2, Catherine Cargo4, Dennis McGonagle2, Sinisa Savic2
1Department of Clinical Immunology and Allergy, St James University Hospital; 2LIRMM, University of Leeds, Leeds, United Kingdom; 3Department of Health Sciences, University of Genoa, Genoa, Italy; 4Department of Haematology, St James University Hospital, Leeds, United Kingdom
Correspondence: Mark Kacar

Introduction: A higher prevalence of autoimmune and autoinflammatory complications has been reported in patients with myelodysplastic syndrome (MDS). The exact cause of this remains to be elucidated.

Objectives: To determine the correlation between patients’ demographic, clinical and molecular (karyotype, somatic genetic mutation status) features of MDS with specific autoinflammatory complications and long-term outcome.

Methods: This was a retrospective study of 140 MDS patients referred to the Haematological Malignancy Diagnostic Service (HMDS) in Leeds, UK, between 2012-2018. As part of their diagnostic workup, all patients had karyotype assessment and targeted genetic sequencing performed. Patients’ medical records were examined to collect demographic and clinical information, and to identify patients with autoinflammatory complications. Patients were classified as having ‘non-specific autoinflammatory features’ if CRP was found to be elevated (>10.0 mg/L) on 5 or more separate occasions and this elevation could not be explained by infection, malignancy or autoimmunity. Chi-squared test, Student t-test, analysis of variance (ANOVA), univariate and multivariate logistic regression analyses were performed.

Results: The average age was 77.08±11 years (median 79 years), with a male (n=91, 65.0%) preponderance. The 72 patients who had non-specific autoinflammatory features (51%) tended to be younger (75.15±11.23 years versus 79.15±11.92, p=0.0395), and more frequently had arthritis (n=25, 34.7%, versus n=12, 17.6%, p=0.0225), arthralgia (n=32, 44.4%, versus n=18, 26.5%, p=0.0271), skin rash (n=22, 30.6%, versus n=10, 14.7%, p=0.0261), and pleuritis (16, 22.2%, versus n=3, 4.4%, p=0.0022). 26.4% of MDS patients had a well-defined diagnosis of autoinflammatory disorder, with neutrophilic dermatosis and polymyalgia rheumatic occurring most commonly. Mutations affecting the transcription factor pathway (NPM1, RUNX1, BCOR, WTI, TP53, MYD88) (OR 3.15 [95%CI 1.04-9.56], p=0.0426) and deletion of chromosome 5 (OR 3.37 [95%CI 1.01-11.22], p=0.0479) were associated with autoinflammatory complications in general. Stratifying the patients into a “well-defined” and “non-specific” autoinflammatory disease group showed that deletion of chromosome 7 was associated with well-defined conditions, whilst deletion of chromosome 5 was linked with a non-specific autoinflammatory status. Furthermore, a higher rate of acute leukaemia transformation was reported in MDS patients with autoinflammatory status (n=25, 34.7%, versus n=8, 11.8%, p=0.0002).

Conclusion: Autoinflammatory conditions were found to be more prevalent than expected in patients with MDS and were linked to a worse prognosis. Transcription factor pathway gene mutations and an abnormal karyotype were also associated with autoinflammation. Autoinflammatory features were associated with malignant transformation, hinting at the possibility that treatment of the autoinflammation might play a role in preventing disease progression. Further studies are required to replicate our findings and study the effect of anti-inflammatory therapy on disease progression.

Disclosure of Interest

None Declared

Table 1 (abstract O14).

See text for description



OR [95%CI]

Statistical significance

Overall autoinflammation

Transcription factor pathway

3.15 [95%CI 1.04 to 9.56]


Deletion of chromosome 5

3.37 [95%CI 1.01 to 11.22]


Well-defined autoinflammatory disease

Transcription factor pathway

4.50 [95%CI 1.04 to 19.47]


Deletion of chromosome 7

6.13 [95%CI 1.16 to 32.33]


Number of mutations

3.39 [95%CI 1.08 to 10.66]


Unspecified inflammatory state

Deletionof chromosome 5

3.57 [95%CI 1.02 to 12.48]


O15 A novel MEFV mutation associated with an autosomal dominant FMF complicated by AA amyloidosis in a large British kindred

Dorota Rowczenio, Taryn Youngstein, Hadija Trojer, Charalampia Papadopoulou, Tamer Rezk, Philip Hawkins, Helen Lachmann
National Amyloidosis Centre, UCL, London, United Kingdom
Correspondence: Dorota Rowczenio

Introduction: Hereditary systemic autoinflammatory diseases (SAIDs) are rare genetic disorders characterised by recurrent, spontaneously resolving episodes of fever and systemic inflammation that predominantly involves serosal surfaces. AA amyloidosis is the most serious complication of SAIDs and is associated with proteinuric renal dysfunction that progresses to end stage renal failure and premature death.

Objectives: To find a genetic cause in a large British family with a dominantly inherited autoinflammatory syndrome complicated by AA amyloidosis.

Methods: Initially the Next Generation Sequencing (NGS) targeting 20 autoinflammatory genes was performed in the index patient and his sister, both of whom have been diagnosed with AA amyloidosis. There was a clear autosomal dominant inheritance affecting three generations. Upon finding a genetic cause the DNA obtained from other affected family members were analysed by Sanger sequencing.

Results: The index case was diagnosed with sJIA aged 7 and developed end stage renal failure in his early twenties. He had haemodialysis for four years and underwent renal transplantation at the age of 32. He was referred to the National Amyloidosis Centre (NAC) with a suspicion of AA amyloid deposits in the transplanted kidneys. He reported suffering with fever accompanied by severe abdominal and chest pain, arthritis, erythema and night sweats from early life. Interestingly having been started on colchicine for post-transplant gout he felt significantly better. Subsequently his sister developed nephrotic syndrome due to AA amyloidosis. She describes similar symptoms throughout most of her life. Their parents were of white British origin from a non-consanguineous kindred. Their father had died aged 52 years from complications immediately following a cadaveric renal transplantation. The post-mortem examination of his renal biopsy revealed deposition of AA amyloid fibrils. The index case grandmother also suffered with cyclical episodic abdominal cramping and had a history of osteoarthritis. The index case’s two paternal cousins and two of their children described similar symptoms.

NGS revealed a single MEFV allele to be affected in both the index case and his sister, resulting in the p.P373L variant in exon 3. Subsequently this variant was confirmed by Sanger sequencing in all living affected members. The mutant allele was not identified in the unaffected cases. All symptomatic individuals were treated with colchicine which suppressed their FMF related inflammation. We sequenced SAA1 gene, as homozygosity for the SAA1.1 allele is a known susceptibility factor for AA amyloidosis, but all cases were heterozygous.

Conclusion: In the Northern Caucasian population FMF is extremely rare in comparison to the Mediterranean region. Typically FMF is an autosomal recessive disorder, nonetheless very rare cases of dominantly inherited disease have previously been reported, namely caused by the deletion of methionine residue at position 694 identified in British patients and three substitutions affecting threonine 577: p.T577N, p.T577S and p.T577A found in British, Turkish and Dutch patients respectively.

Here we report a novel MEFV variant p.P373L causing dominant FMF in three generations of a large British family and in three cases this was complicated by AA amyloidosis indicating a severe pathogenicity of this variant.

Consent for publication has been obtained from patient


Disclosure of Interest

None Declared

Oral communications – Immunology

O16 PYRIN inflammasome dysregulation in FMF patients: implication for a fast diagnostic test

Thomas Henry, Flora Magnotti, Alexandre Belot, Yvan Jamilloux
Inserm U1111, CNRS UMR5308, Univ. Lyon, ENS Lyon, CIRI, Lyon, France
Correspondence: Thomas Henry

Introduction: Familial Mediterranean Fever (FMF) is usually associated with bi-allelic mutations in the MEFV gene. MEFV encodes Pyrin, an inflammasome sensor leading to IL-1β release and a fast cell death termed pyroptosis. The relationship between MEFV mutations and Pyrin inflammasome regulations is still poorly understood. Furthermore, due to the large number of MEFV variants and a substantial proportions of FMF patients presenting mutations in a single MEFV allele, the genetic confirmation of the FMF diagnosis is often unconclusive.

Objectives: The objective of the study were:
  1. 1.

    to understand at the molecular levels the Pyrin inflammasome dysregulation in monocytes from FMF patients

  2. 2.

    to assess whether monitoring Pyrin inflammasome activation could lead to a novel functional diagnostic test for FMF


Methods: Monocytes from healthy donors (HD) or FMF patients were isolated and treated with a kinase inhibitor targeting the PKC superfamily, which includes PKN1/2 known to regulate Pyrin inflammasome activation. IL-1β release andcell death kinetics were monitored.

In parallel, a human monocyte cell line was engineered to modelize FMF and HD monocytes and assess the role of Pyrin phosphorylation and inflammasome components in cell death and cytokine release.

Results: Dephosphorylation of Pyrin was sufficient to trigger Pyrin inflammasome activation in monocytes from FMF patients while no inflammasome activation was observed in monocytes from healthy controls.

Using a human monocyte cell line, we demonstrated that dephosphorylation of Pyrin was similar in cells expressing wild-type or mutated MEFV but that a mutated MEFV was necessary and sufficient to progress to active inflammasome upon PKC superfamily inhibition.

Finally, thanks to a cohort of FMF patients, we demonstrate that FMF patients can be efficiently and specifically diagnosed based on the response of their monocytes to PKC superfamily inhibitors.

Conclusion: Our results demonstrate that Pyrin dephosphorylation is sufficient to trigger inflammasome activation in monocytes from FMF patients but not from healthy donors. This indicates that in healthy donors, the progression to an active Pyrin inflammasome requires two independently-controlled steps and that the second mechanism of control is deficient in monocytes from FMF patients.

Our study also demonstrates that monitoring inflammasome activation upon PKC superfamily inhibition in monocytes can discriminate FMF patients from patients with other inflammatory conditions opening the way to a fast functional test to diagnose FMF.

Disclosure of Interest

None Declared

O17 Unraveling the molecular pathogenesis of proteasome-associated autoinflammatory syndromes

Frédéric Ebstein1, Anja Brehm2, Sébastien Küry3, Thomas Besnard3, Bertrand Isidor3, Stéphane Bézieau3, Pawel Stankiewicz4, Elke Krüger1
1Institut für Medizinische Biochemie und Molekularbiologie (IMBM), Universitätsmedizin Greifswald, Greifswald; 2Institut für Biochemie, Charité Universitätsmedizin Berlin, Berlin, Germany; 3Service de Génétique Médicale, CHU de Nantes, Nantes, France; 4Dept of Molecular and Human Genetics, Baylor College of Medicine, Houston, United States
Correspondence: Frédéric Ebstein

Introduction: In most eukaryotic cells, the degradation of abnormal and/or regulatory proteins is ensured by large multi-subunit ATP-dependent proteases known as proteasomes which consist in two distinct sub-complexes, a 20S core particle and a 19S regulatory particle. Since the early 2010s, an increasing number of loss-of-function mutations have been identified in genes encoding proteasome subunits including PSMB8, PSMB9, PSMB4, PSMA3 and PSMD12 and/or the proteasome maturation protein POMP. Fascinatingly, depending on the subunit affected, such genomic alterations result in the development of two seemingly distinct phenotypes, namely: (i) systemic autoinflammation or (ii) cognitive impairment. Herein, mutations of the 20S core particle subunits (i.e. PSMB8, PSMB9, PSMB4 and PSMA3) and/or POMP are typically associated with a group of autoinflammatory syndromes sharing the same constellation of clinical signs and frequently referred to as chronic atypical neutrophilic dermatosis with lipodystrophy and elevated temperature (CANDLE) and/or proteasome-associated autoinflammatory syndromes (PRAAS). By contrast, genetic disruption of the subunits of the 19S regulatory particle (i.e. PSMD12) leads to a syndromic form of intellectual disability (ID).

Objectives: Our aim in this study is to determine whether syndromic ID disorders caused by variants in genes encoding 19S proteasome subunits share similarities with CANDLE/PRAAS in their etiology and/or pathogenesis particularly regarding inflammation.

Methods: Peripheral blood mononuclear cells (PBMC) from subjects carrying 19S proteasome loss-of-function mutations diagnosed with syndromic ID as well as related control were collected and subjected to RNA isolation and protein extraction prior to quantitative PCR and western-blot analysis, respectively.

Results: Like CANDLE/PRAAS, syndromic ID disorders caused by genomic alterations in 19S proteasome genes were associated with perturbed proteasome function, as evidenced by increased levels of intracellular ubiquitin-protein conjugates. Most importantly, our data show that PBMC from patients with syndromic ID exhibit high transcription levels of various IFN-induced genes (ISG) including CXCL10, CXCL9, IFI44 and IFI44L. Interestingly, the strength of the IFN signatures in these patients correlated with the magnitude of the unfolded protein responses (UPR) initiated by proteasome dysfunction, suggesting a cause-and-effect relationship between endoplasmic reticulum (ER) stress and inflammation.

Conclusion: In this work, we provide evidence on the association of 19S proteasome dysfunction with the generation of autoinflammation in subjects diagnosed with syndromic ID disorders. From these data, we expect to convey a more integrated picture of the pathophysiology of syndromic ID and identify new therapeutic targets for the treatment of cognitive impairment.

Disclosure of Interest

None Declared

O18 A novel knock-in mouse model of CAPS that develops amyloidosis: therapeutic efficacy of proton pump inhibitors

Arinna Bertoni1, Sonia Carta2, Chiara Baldovini3, Federica Penco1, Enrica Balza2, Silvia Borghini4, Marco Di Duca4, Emanuela Ognio5, Paolo Nozza3, Francesca Schena1, Patrizia Castellani2, Claudia Pastorino1, Carola Perrone1, Laura Obici6, Alberto Martini7, Isabella Ceccherini4, Marco Gattorno1, Anna Rubartelli2, Sabrina Chiesa1
1Centro Malattie Autoinfiammatorie ed Immunodeficienze, IRCCS Istituto G. Gaslini; 2Unità di Biologia Cellulare, IRCCS Ospedale Policlinico San Martino; 3Anatomia Patologica; 4Genetica Medica, IRCCS Istituto G. Gaslini; 5S.S Animal Facility, IRCCS Ospedale Policlinico San Martino, Genova; 6Centro per lo Studio e la Cura delle Amiloidosi Sistemiche, Fondazione IRCCS Policlinico San Matteo, Pavia; 7Direzione Scientifica, IRCCS Istituto G. Gaslini, Genova, Italy
Correspondence: Arinna Bertoni

Introduction: Cryopyrin associated periodic syndromes (CAPS) are a group of autoinflammatory diseases linked to gain-of-function mutations in the NLRP3 gene that cause uncontrolled IL-1β secretion. CINCA syndrome is the most severe CAPS disease characterized by central nervous system disabilities with a long-term risk of secondary amyloidosis.

Proton pump inhibitors (PPIs), commonly used as inhibitors of gastric acidproduction, also display anti-inflammatory properties, making them promising drugs in sepsis and in inflammatory disorders.

Objectives: To develop a novel NLRP3 knock-in (KI) mouse model of CAPSto evaluate amyloid deposition and to test alternative therapeutic approaches.

Methods: We generated KI mice by engineering N475K mutation associated with CAPS phenotype into mouse Nlrp3 gene. KI and Wild Type (WT) mice received PPIs or PBS intraperitoneally and were analyzed for survival, inflammation, cytokine secretion, and amyloidosis development. Cytokines secretion from bone marrow derived dendritic cells (BMDCs) and peritoneal macrophages (PMs) was evaluated by ELISA. Hystological analysis of all organs was evaluated by hematoxylin and eosin staining. Amyloid deposition was quantified through Congo Red staining.

Results: Mutant NLRP3 KI mice displayed features that recapitulates the immunological and clinical phenotype of CAPS. These mice had systemic inflammation, with high levels of serum pro-inflammatory cytokines compared to WT controls. Hystological analysis revealed the presence of acute and chronic inflammatory cell infiltrates and amyloid deposits in spleen, liver and kidneys. As in CAPS monocytes , BMDCs and PM from KI mice showed a strong increase in IL-1β, IL-18, and IL-1α secretion and decreased levels in interleukin-1 receptor antagonist (IL-1Ra), the naturally occurring IL-1b inhibitor.

PPIs treatment of KI mice showed a clear clinical impact with improvement of inflammatory conditions and regression of amyloid deposits. Remarkably, BMDCs and PMs from PPI-treated mice presentedreduced secretion of pro-inflammatory cytokines and re-established the levels of IL-1RA.

Conclusion: NLRP3 KI mice display a CAPS phenotype with many characteristics of autoinflammation, including amyloidosis. The therapeutic effectiveness associated with lack oftoxicity indicates that PPIs could represent relevant adjuvants to the anti-IL-1 drugs in IL-1 driven diseases.

Disclosure of Interest

None Declared

O19 T cell defects in patients with ARPC1B germline mutations account for combined immunodeficiency

Immacolata Brigida1, Matteo Zoccolillo1,4, Maria Pia Cicalese1,2,3, Laurène Pfajfer5-9, Federica Barzaghi2,4, Serena Scala1, Carmen Oleaga-Quintas10,11, Jesus A. Alvarez12, Lucia Sereni1, Stefania Giannelli1, Claudia Sartirana1, Francesca Dionisio1, Luca Pavesi13, Marta Benavides-Nieto14,15, Luca Basso-Ricci1, Paola Capasso1, Benedetta Mazzi16, Jeremie Rosain10,11,28, Nufar Marcus17, Yu Nee Lee18, Raz Somech18, Massimo Degano19, Giuseppe Raiola20, Roberta Caorsi21, Paolo Picco21, Marcela Moncada Velez12, Joelle Khourieh11,12, Andrés Augusto Arias12,29, Aziz Bousfiha22, Thomas Issekutz23, Andrew Issekutz23, Bertrand Boisson11,12,24, Kerry Dobbs25, Anna Villa1,26, Angelo Lombardo1,3, Benedicte Neven14, Despina Moshous14,15, Jean-Laurent Casanova11,12,14,24,27, José Luis Franco12, Luigi D Notarangelo25, Cristina Scielzo13, Stefano Volpi21,30, Loïc Dupré5-9, Jacinta Bustamante11,12,24,28, Marco Gattorno21,31‡, and Alessandro Aiuti1,2,3‡
1San Raffaele Telethon Institute for Gene Therapy, SR-TIGET; 2Pediatric Immunohematology, San Raffaele Scientific Institute, Milan; 3Vita-Salute San Raffaele University, Milan, Italy; 4Department of Systems Medicine, Tor Vergata University, Rome, Italy; 5INSERM, UMR1043, Centre de Physiopathologie de Toulouse Purpan, Toulouse, France; 6CNRS, UMR5282, Toulouse, France; 7Université Toulouse III Paul-Sabatier, Toulouse, France; 8Ludwig Boltzmann Institute for Rare and Undiagnosed Diseases, Vienna, Austria; 9CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria; 10Laboratory of Human Genetics of Infectious Diseases, Necker Branch, INSERM U1163, 75015 Paris, France, EU; 11Paris Descartes University, Imagine Institute, 75015 Paris, France, EU; 12Group of Primary Immunodeficiencies, Department of Microbiology & Parasitology, School of Medicine, University of Antioquia UdeA, Medellin, Colombia; 13Division of Experimental Oncology, Unit of B-cell Neoplasia, San Raffaele Scientific Institute, Milan; 14Pediatric Hematology-Immunology Unit, Necker Hospital for Sick Children, AP-HP, 75015 Paris, France, EU; 15Genome Dynamics in the Immune System, Université Paris Descartes – Sorbonne Paris; 16Immunogenetics Laboratory, HLA & Chimerism, Dept. of Immunohematology & Blood Transfusion, IRCCS Ospedale San Raffaele, Milano, Italy; 17Kipper Institute for Allergy and Immunology, Schneider Children’s Medical Center of Israel, Petach Tikva, Israel, affiliated with Sackler School of Medicine, Tel Aviv University, Tel Aviv, Israel; 18Pediatric Department A and the Immunology Services, “Edmond and Lily Safra” Children's Hospital, Jeffrey Modell Foundation Center, Sheba Medical Center, Tel Hashomer affiliated with Sackler School of Medicine, Tel Aviv University, Tel Aviv, Israel; 19Division of Immunology, Transplantation, and Infectious Diseases, Biocrystallography Unit. San Raffaele Scientific Institute, Milan; 20Unità Operativa di Pediatria, Azienda Ospedaliera “Pugliese-Ciaccio” di Catanzaro; 21U.O. Clinica Pediatrica e Reumatologia, Istituto Giannina Gaslini, Genova, Italy; 22Clinical Immunology Unit, Department of Pediatrics, King Hassan II University, Ibn-Rochd Hospital, Casablanca, Morocco; 23Department of Pediatrics & Department of Microbiology-Immunology, Dalhousie University, Halifax, Nova Scotia, Canada; 24St. Giles Laboratory of Human Genetics of Infectious Diseases, Rockefeller Branch, The Rockefeller University, New York, NY 10065, USA; 25Laboratory of Clinical Immunology and Microbiology, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, USA; 26Istituto di Ricerca Genetica e Biomedica (IRGB), Consiglio Nazionale delle Ricerche (CNR), Milan Unit, Milan, Italy; 27Howard Hughes Medical Institute, NY, USA; 28Center for the Study of Primary Immunodeficiencies, Assistance Publique-Hôpitaux de Paris AP-HP, Necker Hospital for Sick Children, Paris, France, EU; 29School of Microbiology, University of Antioquia UdeA, Medellin, Colombia; 30Università degli Studi di Genova, Genova, Italy; 31UOSD Centro Malattie e Autoinfiammatorie e Immunodeficienze, Istituto Giannina Gaslini, Genova, Italy
Correspondence: Alessandro Aiuti

Introduction: ARPC1B is a key factor for the assembly and maintenance of the ARP2/3 complex, involved in actin branching from an existing filament. Germline mutations in ARPC1B have been recently described in six unrelated patients, with clinical features of combined immunodeficiency, whose neutrophils and platelets but not T lymphocytes were studied.

Objectives: We hypothesized that ARPC1B-deficiency may also lead to cytoskeleton and functional defects in T cells

Methods: Next-generation sequencing in 6 patients; flow cytometry, proliferation and migration, confocal and electron microscopy) to characterize defects in T cells; lentiviral-mediated gene transfer for genetic correction of ARPC1B.

Results: We identified bi-allelic mutations in 6 unrelated patients with severe infections, autoimmune manifestations and platelet defects showing altered protein structure, and absent/low expression of the ARPC1B protein. Confocal microscopy showed altered expression of ARPC1B with actin. T cells displayed impaired TCR-mediated proliferation and SDF1-α directed migration. Gene transfer of ARPC1B in patient’s T cells using a lentiviral vector restored ARPC1B expression, leading to improved T-cell proliferation in vitro. In 2 patients normal ARPC1B levels in a fraction of lymphocytes were associated with in vivo somatic reversion and improved T cell migration in vitro. In one of the patient somatic revertant was present only in memory CD8+ T-cells, which showed improved in vitro T-cell migration.

Conclusion: Inherited ARPC1B deficiency alters T cell cytoskeletal dynamics and functions, contributing to the clinical features of combined immunodeficiency.

Disclosure of Interest

None Declared

O20 B cell defect in ADA2 deficiency patients

Francesca Schena1, Federica Penco1, Stefano Volpi1,2, Claudia Pastorino1, Roberta Caorsi1, Arinna Bertoni1, Francesca Kalli3, Daniela Fenoglio3,4,5, Annalisa Salis6, Ignazia Prigione1, Paola Bocca1, Francesca Antonini7, Antonella Insalaco8, Alice Grossi9, Gianluca Damonte6, Isabella Ceccherini9, Gilberto Filaci3,4,5, Alberto Martini1,2, Elisabetta Traggiai10, Marco Gattorno1
1UOSD Centro Malattie Autoinfiammatorie e Immunodeficienze and Clinica Pediatrica e Reumatologia, IRCCS Istituto Giannina Gaslini; 2Università degli studi di Genova; 3Center of Excellence for Biomedical Research; 4Department of Internal Medicine, Clinical Immunology Unit, Università di Genova; 5Ospedale Policlinico San Martino; 6Department of Experimental Medicine and Center of Excellence for Biomedical Research, Università di Genova; 7Core Facilities Flow-Cytometry and Cell imaging Lab, Istituto Giannina Gaslini, Genova; 8Division of Rheumatology, Department of Pediatric Medicine, IRCCS, Bambino Gesù Children’s Hospital, Roma; 9Medical Genetics, IRCCS Istituto Giannina Gaslini, Genova, Italy; 10Novartis Institutes for Biomedical Research, Basel, Switzerland
Correspondence: Francesca Schena

Introduction: Adenosine Deaminase 2 Deficiency (DADA2) is an autoinflammatory disease characterized by systemic vasculopathy, strokes and mild immunodeficiency, mainly affecting B cell compartment. The defect is due to a loss of function mutation of ADA2 gene, coding for Adenosine Deaminase 2, a protein which regulates the catabolism of extracellular adenosine.

Objectives: Hypogammaglobulinemia and recurrent infections are associated to DADA2. We therefore investigated phenotype and in vitro B and T cell responses in DADA2 patients to address if ADA2 mutation affects B and T cell function and in particular we focused on B cell-T cell interaction.

Methods: 14 patients carrying loss of function mutations in ADA2 were examined. They showed clinical history with livedo reticularis, fever, vasculitis and neurological symptoms. We analyzed peripheral B and T cell phenotype by flow cytometry, in vitro B-cell proliferation and differentiation to Immunoglobulin secreting cells in response to TLR9 agonist and T cell help. Moreover B cells isolated from DADA2 patients or HD have been cultured in co-culture with CD4+ T cells and in vitro B cell proliferation has been evaluated by CFSE dilution, whereas B cell differentiation and immunoglobulin secretion in response to TLR9 agonist and T cell help have been evaluated by ELISA and ELISPOT assay. Simultaneously cytokine production from Tfh cells has been analyzed.

Results: Flow-cytometric analysis of DADA2 peripheral blood showed a significant reduction of switch memory B cells and an increased frequency of CD21low B cells. Regarding T cell compartment memory CD4+ and CD8+ T cells are significantly reduced; interestingly we identified an expansion in frequency of circulating Tfh cells.

Then we investigated a role of ADA2 in B cells: we found that ADA2 is expressed in all B cell subsets, secreted but its enzymatic activity is strongly impaired in DADA2. We show that Naïve B cells from DADA2 patients are impaired in their proliferation, suggesting an intrinsic defect due to the mutation.

We also addressed the interaction between B and helper T cells; DADA2 CD4+ T cells showed an impairment in the IL21 production and a downregulation of CD40L, suggesting a functional defect of Tfh cells; then we found that proliferation and differentiation of patients’ B cells were not sustained from patients’ CD4+ T cells.

Conclusion: Our findings suggest that ADA2 mutations could lead to an intrinsic defect in B cell function and to a reduced T cell dependent B cell response.

Disclosure of Interest

None Declared

O21 Periodic Fever, Aphthous Stomatitis, Pharyngitis and Adenitis (PFAPA) syndrome and obstructive sleep apnea are distinct inflammatory disorders of oropharyngeal lymphoid tissue

Kalpana Manthiram1, Silvia Preite2, Fatma Dedeoglu3, Maranda Lawton3, Pamela Mudd4, Hemalatha Srinivasalu4, Greg Licameli3, Kathryn Edwards5, Pamela Schwartzberg2, Daniel Kastner1
1National Human Genome Research Institute; 2National Institute of Allergy and Infectious Diseases, NIH, Bethesda; 3Boston Children’s Hospital, Harvard Medical School, Boston; 4Children’s National Medical Center, George Washington University School of Medicine, Washington; 5Vanderbilt University School of Medicine, Nashville, United States
Correspondence: Kalpana Manthiram

Introduction: Tonsillectomy is one of the most common surgical procedures in children and is performed typically for recurrent tonsillitis or obstructive sleep apnea (OSA), and less frequently for periodic fever, aphthous stomatitis pharyngitis, cervical adenitis (PFAPA) syndrome. The pathogenesis of OSA and PFAPA is unknown.

Objectives: In order to understand the pathogenesis of these two disorders, we studied the immunologic profile of tonsils removed from children with PFAPA and OSA in comparision with control tonsils.

Methods: After getting informed consent, we obtained tonsils from children undergoing tonsillectomy for (1) PFAPA (N=12) and (2) OSA (N=12), and from (3) children undergoing tonsillectomy during oropharyngeal anatomic correction surgery like pharyngeal flap preparation (“controls”, N=9).Mononuclear cells from the tonsils were separated by standard methods and cells were analyzed by flow cytometry and gene expression with Nanostring. Cytokine production by isolated tonsillar mononuclear cells was measured by flow cytometry following phorbol 12-myristate 13-acetate (PMA) and ionomycin stimulation for 4 hours ex vivo.

Results: Tonsils from patients with PFAPA, which were removed during asymptomatic intervals, had evidence of germinal center suppression with significantly fewer T follicular helper cells, more T follicular regulatory cells, fewer plasma cells and reduced IgG class switching compared to tonsils from either OSA patients or controls. Gene expression analysis revealed downregulation of pro-inflammatory genes in tonsillar myeloid cells from patients with PFAPA. However, upon stimulation with PMA and ionomycin, CD4+ T cells from tonsils of patients with PFAPA produced more IFNγ and IL-17 than that of controls, but less than those of patients with OSA.

On the other hand, tonsils from patients with OSA, had significantly larger germinal center areas by histology compared to those from patients with PFAPA and controls. Moreover, in tonsils from OSA patients, we found higher IFNγ production by CD4+ T, CD8+ T and NK cells, and greater IL-17 production by CD4+ T cells and NK cells upon stimulation with PMA and ionomycin when compared with tonsils from PFAPA patients and controls.

Conclusion: We show that tonsils from both patients with PFAPA and OSA exhibit evidence of immune dysregulation. During asymptomatic periods, PFAPA patients display myeloid cell and germinal center suppression in the tonsils, suggesting a compensatory response to inflammatory flares. However, upon stimulation, CD4+ T cells produce high levels of pro-inflammatory cytokines. We hypothesize that alternating periods of heightened immune activation and suppression lead to the periodicity of PFAPA. In comparison, tonsils from patients with OSA display relatively constant germinal center hypertrophy likely due to unchecked chronic T cell activation. Further studies are necessary to understand the genetic and environmental risk factors for T cell activation in both of these disorders and why PFAPA patients may have cycling of inflammatory periods in the tonsils while OSA patients have chronic tonsillar inflammation.

Disclosure of Interest

None Declared

O22 PFAPA syndrome: NK cells infiltrating tonsils support the crucial role of innate immunity in the pathogenesis

Sabrina Chiesa1, Roberta Caorsi2, Francesca Bellora3, Mariella Della Chiesa3, Ilaria Ingrosso1, Federica Penco1, ArinnaBertoni1, Claudia Pastorino1, Ignazia Prigione1, Alessia Omenetti2, Martina Finetti2, Silvia Borghini4, Angela Sementa5, Roberto D’Agostino6, LuciaSemino6, Alberto Martini7, Cristina Bottino3, Marco Gattorno1
1Centro Malattie Autoinfiammatorie e Immunodeficienze; 2Clinica Pediatrica e Reumatologica, IIRCCS G. Gaslini; 3Medicina Sperimentale, University of Genoa; 4Laboratorio di Genetica Molecolare; 5Anatomia Patologica; 6UO Otorinolaringoiatria, IIRCCS G. Gaslini; 7Clinica Pediatrica e Reumatologica, University of Genoa, Genova, Italy
Correspondence: Sabrina Chiesa

Introduction: Periodic fever, aphthous stomatitis, pharyngitis, and cervical adenitis (PFAPA) syndrome is a more frequent cause of recurrent fever in children. The exact etiology of this pediatric disorder is still unknown. Elevated serum levels of IL-1β, IL-18, IL-6, and IFNγ suggest innate immunity dysregulation as the key pathomechanism of PFAPA attacks. Palatine tonsils are sites where innate immunity leads to the onset of adaptive immunity, mediated by B and T lymphocytes. Natural killer (NK) cells, the most important effectors of the innate lymphoid cells (ILCs), play a fundamental role in innate immune responses.

Objectives: Tonsillectomy is one of the therapeutic options for PFAPA patients. We tested whether specific infiltrating inflammatory cells in pediatric tonsils contribute to PFAPA pathogenesis.

Methods: Tonsils were collected from 2 groups of pediatric patients undergoing tonsillectomy: PFAPA patients (n=32) and children with bacterial tonsillitis (control group, CG) (n=25). Phenotypic analysis of subpopulations of tonsil cell suspensions and tissues was performedby flow cytometry and immunohistochemistry.

Results: During the asymptomatic phase of disease the number of monocytes did not differ between the PFAPA and control tonsils. We observed a considerable recruitment of NK cells in tonsils of PFAPA patients respect to CG. In particular, we detected a significant expansion of both CD56bright CD16- and CD56dim CD16+ NK cell subsets in PFAPA samples compared to CG. A fine characterization of activating and inhibitory NK receptors suggested a crucial role of CD56dim CD16+ cell subset in PFAPA disease. Specially, activating receptors, as natural cytotoxicity receptors (NCRs) and NKG2D, were higher in NK cells of PFAPA patients than in NK cells from CG. Remarkably, CD56+ NK cells from PFAPA tonsils werecytotoxic and contained much more perforine and granzyme than those from CG tonsils. NK cells from PFAPA tonsils exhibited increased IFN-g production, compared to NK from CG. Accordingly, plasmacytoid dendritic cells, the main source of type I interferon cytokines, were significantly increased in PFAPA tonsils. We detected a higher number of naïve and a significantly lower percentage of effector memory CD4+ and CD8+ T cells in PFAPA tonsils compared to CG. Additionally, PFAPA tonsils presented a significant decrease of both functional follicular helper T cells (CXCR5+ICOS+) and T regulatory cells (CD39+Foxp3+). Finally, CD19+CD38+ B cellswere reduced in this cohort of PFAPA tonsils.

Conclusion: These results suggest aninvolvement of NK cells in the pathogenesis of PFAPA and support the crucial role ofinnate immunity in the disease. Nonetheless, the abundant and activated CD56+NK cells might shape adaptive immunity that is impaired in the tonsils of PFAPA patients.

Disclosure of Interest

None Declared

Novel AID pathways and genes

O23 Identification of rare coding variants in IL-1-related pathways in patients with adult-onset still’s disease

Giulio Cavalli1,2, Rosanne Van Deuren2, Peer Arts2, Marloes Steerhower2, Paolo Sfriso3, Paola Galozzi3, Serena Colafrancesco4, Roberta Priori4, Luca Cantarini5, Stefano Rodolfi1, Elena Baldissera1, Frank van der Veerdonk2, Lorenzo Dagna1, Alexander Hoischen2, Charles A. Dinarello2,6
1Unit of Immunology, Rheumatology, Allergy and Rare Diseases (UniRAR), Vita-Salute San Raffaele University, Milan, Italy; 2Department of Medicine, Radboud University Medical Centre, Nijmegen, Netherlands; 3Rheumatology, University of Padua, Padua; 4Rheumatology, Sapienza University, Rome; 5Rheumatology, University of Siena, Siena; 6Department of Medicine, University of Colorado Denver, Aurora, CO, Italy
Correspondence: Giulio Cavalli

Introduction: Adult-onset Still's disease (AOSD) is a rare autoinflammatory disease characterized by fever, arthritis, and multi-organ involvement. Inflammation in AOSD is mediated by interleukin (IL)-1β, as confirmed by the dramatic clinical efficacy of selective blockers of this cytokine. The genetic predisposition to this rampant IL-1-driven inflammation remains nevertheless elusive. Previous studies failed to identify associations between polymorphisms in the genes encoding IL-1 and AOSD, thus pointing at more complex genetic mechanisms. This ‘missing heritability’ cannot be adequately investigated with traditional techniques for genetic partitioning, such as GWAS, which only assess common variants and polymorphisms. Studies focusing on highly penetrant rare variants or different types of mutations (i.e. small copy-number variations; insertions/deletions) are warranted.

Objectives: We hypothesized that genetically determined changes in IL-1-related pathways resulting in excessive IL-1β activity lead to the development of autoinflammation in AOSD. Scope of this study was to unravel the combined mutational variation of a network of IL-1-related receptors, pathways, counter-regulators, and cellular processes possibly involved in the pathogenesis of AOSD and IL-1-mediated inflammation in general.

Methods: We collected clinical, demographic, and genetic data from a large cohort of 76 AOSD patients and developed an innovative platform based on molecular inversion probes (MIP) technology for performing highly multiplexed targeted-resequencing. This allows efficient sequencing of the coding sequence of 48 genes related to the IL-1-pathway, and allows studying rare and common variants in one assay. We have also screened 500 healthy controls, and 1000s of samples with other disorders using the same assay.

Results: We identified rare and unique (i.e. private variants) in the IL1 pathway in several individuals with AOSD. Whether any these are involved in a strong predisposition to AOSD is currently followed-up. Rare genetic variants have been identified in six IL-1-pathway ‘clusters’:
  1. 1.

    Deregulated activation of the inflammasome and release of IL‑1β and IL-18.

  2. 2.

    IL-1 family receptors and intracellular signaling mediators.

  3. 3.

    Other pro-inflammatory cytokines and receptors.

  4. 4.

    Regulatory molecules, including IL-1Ra or IL-37.

  5. 5.

    Cellular processes regulating production of IL-1 and IL-18 (i.e. autophagy).

  6. 6.

    Production of ROS, which function as markers of cellular damage and trigger inflammation.


Conclusion: Unraveling the genetic bases of inflammation in AOSD deepens our understanding of the human innate immunome. Of note, this study platform may serve for the genetic analysis of other IL-1-mediated conditions, including gout and other autoinflammatory diseases, whose genetic predisposition remains elusive. Equally important, the identification of pathways amenable to targeting with small molecules or biologics may translate into remarkable clinical implications.

Disclosure of Interest

None Declared

O24 Trisomy 8-associated autoinflammatory disease (TRIAD) is characterized by increased monocyte activation

Kalpana Manthiram1, Alina Dulau-Florea2, Deborah Bruns3, Amanda Ombrello1, Karyl Barron4, Tina Romeo1, Anne Jones1, Karin Weiss1, Sandro Perazzio5, Isabelle Kone-Paut6, Nora Al-Mutiari7, Troy Torgerson5, Daniel Kastner1
1National Human Genome Research Institute; 2Department of Laboratory Medicine, NIH, Bethesda; 3Department of Counseling, Quantitative Methods, and Special Education, Southern Illinois University Carbondale, Carbondale; 4National Institute of Allergy and Infectious Diseases, NIH, Bethesda; 5Department of Pediatrics, University of Washington School of Medicine, Seattle, United States; 6Bicêtre University Hospital, APHP, CEREMAIA, University of Paris Sud, Paris, France; 7Department of Pediatrics, Sabah Hospital, Kuwait City, Kuwait
Correspondence: Kalpana Manthiram

Introduction: Many adults with acquired trisomy 8 and myelodysplasia have been reported to have Behçet’s-like inflammatory disease. A few patients with constitutional trisomy 8 mosaicism and ulcerative disease have also been reported. The spectrum of phenotypic abnormalities, inflammatory profiles, and treatment responses of patients with constitutional trisomy 8 are not well-characterized.

Objectives: We analyzed the clinical and immunologic features of a cohort of patients with trisomy 8 mosaicism in order to understand the pathogenesis of the disorder.

Methods: Whole blood gene expression was analyzed with the Nanostring Human Immunology panel. Copy number of two genes on the p and q arms of chromosome 8 were determined by digital droplet PCR in sorted CD3+ T cells, CD19+ B cells, and CD14+ monocytes from peripheral blood to determine the percentage of mosaicism in different cell populations. Platelet electron microscopy was performed in a clinical laboratory.

Results: Eleven patients with trisomy 8 mosaicism ranging in age from 5 to 37 years were recruited. Eight-two percent had recurrent fever, 82% had oral ulcerations, and 72% had severe oral ulcerations larger than 1 cm or lasting more than one week. Five patients reported genital, esophageal, or colonic ulcers. Nine patients had cognitive or motor delay. Two patients had symptoms of a bleeding diathesis with subdural hematoma, petechiae, and/or menorrhagia with normal platelet counts; the platelets of these patients had fewer dense granules by electron microscopy. Patients had elevated peripheral absolute monocyte count (average 970 cells/uL), but none had chronic myelomonocytic leukemia, acute myeloid leukemia or myelodysplasia. Whole blood gene expression revealed upregulation of IL-1-, TLR- and NF-ĸB-related genes. CD14+ monocytes had a significantly higher percentage of cells with trisomy 8 compared to CD3+ T cells and CD19+ B cells (chromosome 8 copy number was 2.8 in CD14+ cells vs. 2.5 in CD19+ [p=0.01] and 2.3 in CD3+ cells [p =0.001]). Four out of 5 patients had partial improvement on daily colchicine, 3 out of 6 reported improvement with intermittent or daily anakinra, 2 out of 3 had improvement on TNFα inhibitors (etanercept or infliximab), and one patient improved following hematopoietic stem cell transplant.

Conclusion: We report the spectrum of clinical and immunologic manifestations in patients with trisomy 8 mosaicism, a disease we name trisomy 8-associated autoinflammatory disease (TRIAD). Patients with TRIAD present with (1) recurrent fever and/or severe mucosal ulcerations, (2) bleeding diatheses, and (3) developmental delay. Patient monocytes are elevated in number and have higher percentage of trisomy 8, indicating that an extra copy of chromosome 8 may confer a survival advantage preferentially to monocytes. In addition, activation pathways in monocytes are upregulated, and patients have symptom improvement in response to blockade of TNFα and IL-1, prominent cytokines associated with myeloid cell activation. Like monocytes, megakaryocytes and platelets develop from a common myeloid progenitor suggesting that cell development in the myeloid lineage may be affected. Further studies are underway to better characterize the inflammatory and survival pathways in myeloid cells with trisomy 8.

Disclosure of Interest

None Declared

Deficiencies in key regulatory signals

O25 A combined immunodeficiency with severe infections, inflammation and allergy caused by ARPC1B deficiency

Stefano Volpi1,2, Maria Pia Cicalese3, Paul Tuijnenburg4,5, Anton T. J. Tool6, Eloy Cuadrado7, Hamid Ahanchian8, Raed Alzyoud9, Zeynep C. Akdemir10, Federica Barzaghi11, Alexander Blank12, Bertrand Boisson13, Cristina Bottino14, Roberta Caorsi15, PaoloPicco15, Jean-Laurent Casanova13,16, Sabrina Chiesa17, Ivan Kingyue Chinn18, Gregor Dückers19, Anselm Enders20, Hans Christian Erichsen21, LisaR. Forbes18, Tomasz Gambin22,23, Marco Gattorno24, Ehsan G. Karimiani25, Silvia Giliani26, Michael S. Gold27, Marwan Abu-Halaweh28, Eva-Maria Jacobsen12, Machiel H. Jansen29,30, Jovanka R. King27, Ronald M. Laxer31, James R. Lupski22,32, Emily Mace18, Stefania Marcenaro33, Reza Maroofian34, Alexander B. Meijer35, Tim Niehues19, Luigi D. Notarangelo36, Jordan Orange18, Ulrich Pannicke37, Chris Pearson38, Patrick J. Quinn27, Ansgar Schulz12, Filiz Seeborg18, Asbjørg Stray-Pedersen39, Hasan Tawamie40, Ester M. M. van Leeuwen30, Alessandro Aiuti11, Rae Yeung31,41, Klaus Schwarz37,42, Taco W. Kuijpers29,43
1Clinica Pediatrica e Reumatologia, Centro per le malattie Autoinfiammatorie e Immunodeficienze, Istituto Giannina Gaslini; 2DINOGMI, Università degli Studi di Genova, Genova; 3Pediatric Immunohematology, San Raffaele Hospital and San Raffaele Telethon Institute for Gene Therapy (SR-TIGET), Milan, Italy; 4Department of Pediatric Immunology, Rheumatology and Infectious diseases, Emma Children’s Hospital, Amsterdam UMC, University of Amsterdam; 5Department of Experimental Immunology, Amsterdam Infection & Immunity Institute; 6Department of Blood Cell Research; 7Department of Immunopathology, Sanquin Research and Landsteiner Laboratory AMC, University of Amsterdam, Amsterdam, Netherlands; 8Department of Allergy and immunology, School of medicine, Mashhad university of Medical Sciences, Mashhad, Iran, Islamic Republic Of; 9Immunology, Allergy and Rheumatology section- Bone Marrow Transplantation for Primary Immunodeficiency Disorders, Queen Rania Children's Hospital, Amman, Jordan; 10Baylor-Hopkins Center for Mendelian Genomics of the Department of Molecular and Human Genetics, Baylor College of Medicine, Huston, United States; 11Pediatric Immunohematology, San Raffaele Hospital and San Raffaele Telethon Institute for Gene Therapy (SR-TIGET), Milan, Italy; 12Department of Pediatrics, University Medical Center Ulm, Ulm, Germany; 13St Giles Laboratory of Human Genetics of Infectious Diseases, Rockefeller Branch, The Rockefeller University, New York, United States; 14Department of Experimental Medicine (DIMES), University of Genoa; 15Centro per le Malattie Infiammatorie e Immunodeficienze, Clinica Pediatrica e Reumatologia, Istituto Giannina Gaslini, Genova, Italy; 16Pediatric Hematology-Immunology and Rheumatology Unit, Necker Hospital for Sick Children, APHP, Paris, France; 17Clinica Pediatrica e Reumatologia, Centro per le malattie Autoinfiammatorie e Immunodeficienze Istituto Giannina Gaslini, Genova, Italy; 18Department of Pediatrics, Section of Allergy, Immunology, and Rheumatology & Center for Human Immunobiology, Texas Children's Hospital, Houston, United States; 19Center for Child and Adolescent Medicine, Helios-Clinic, Krefeld, Germany; 20Department of Immunology and Infectious Disease, John Curtin School of Medical Research and Centre for Personalised Immunology, Australian National University, Canberra, Australia; 21Section of Paediatric Medicine and Transplantation, Division of Paediatric and Adolescent Medicine, Oslo University Hospital, oslo, Norway; 22Baylor-Hopkins Center for Mendelian Genomics of the Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, United States; 23Institute of computer science, Warsaw University of Technology, Warsaw, Poland; 24. Clinica Pediatrica e Reumatologia, Centro per le malattie Autoinfiammatorie e Immunodeficienze Istituto Giannina Gaslini, Genova, Italy; 25Molecular and Clinical Sciences Institute, St. George’s, University of London, Cranmer Terrace, London, United Kingdom; 26Medical Genetics Unit and “A. Nocivelli” Institute for Molecular Medicine, Spedali Civili Hospital, Department of Molecular and Translational Medicine, University of Brescia, Brescia, Italy; 27Discipline of Pediatrics, School of Medicine, University of Adelaide and Department of Allergy and Clinical Immunology, Women's and Children's Health Network, Adelaide, Australia; 28Department of Biotechnology and Genetics Engineering in Philadelphia University, Jordan, United States; 29Emma Children’s Hospital, Amsterdam UMC, University of Amsterdam, Department of Pediatric Immunology, Rheumatology and Infectious diseases; 30Amsterdam UMC, University of Amsterdam, Department of Experimental Immunology, Amsterdam Infection & Immunity Institute, Amsterdam, Netherlands; 31Division of Rheumatology, Department of Paediatrics and Department of Medicine, University of Toronto, The Hospital for Sick Children, Toronto, Canada; 32Department of Pediatrics, Baylor College of Medicine, Houston, United States; 33Istituto Giannina Gaslini, Genova, Italy; 34Medical Research, RILD Welcome Wolfson Centre, Exeter Medical School, Royal Devon and Exeter NHS Foundation Trust, Exeter and Genetics and Molecular Cell Sciences Research Centre, St George's University of London, London, United Kingdom; 35Department of Plasma proteins, Sanquin Research and Landsteiner Laboratory AMC, University of Amsterdam, Amsterdam, Netherlands; 36Laboratory of Clinical Immunology and Microbiology, National Institute of Allergy and Infectious Diseases, Bethesda, United States; 37Institute for Transfusion Medicine, University Ulm, Ulm, Germany; 38Department of General Medicine, Women's and Children's Health Network, Adelaide, Australia; 39Norwegian National Unit for Newborn Screening, Division of Pediatric and Adolescent Medicine, Oslo University Hospital, Oslo, Norway; 40The Institute of Human genetics of Leipzig, Leipzig, Germany; 41Departments of Paediatrics, Immunology, Institute of Medical Science, University of Toronto, Cell Biology Program, The Hospital for Sick Children, Toronto, Canada; 42Institute for Clinical Transfusion Medicine and Immunogenetics Ulm, German Red Cross Blood Service Baden-Wuerttemberg – Hessen, Ulm, Germany; 43Department of Blood Cell Research, Sanquin Research and Landsteiner Laboratory AMC, University of Amsterdam, Amsterdam, Netherlands
Correspondence: Stefano Volpi

Introduction: Genetic defects in regulatory proteins of the cytoskeleton are known to cause different syndromes, mostly dominated by hematologic and immune phenotypes. Recently has been reported a novel syndrome of combined immunodeficiency, allergy and autoinflammation caused by mutations in the ARPC1B gene, one of the seven subunits of the ARP2/3 complex, which regulates actin polymerization.

Methods: We report the natural history, clinical manifestations, genetics, and immunohematological findings in 14 patients from 12 families with ARPC1B deficiency.

Results: Although consanguinity was not revealed by clinical history in five families, all cases carried homozygous mutations in ARPC1B. The mutations resulted in undetectable or reduced protein expression. Early-onset gastrointestinal bleeding and skin rash were common findings (7/14 and 9/14, respectively), whereas bacterial (12/14) and viral (10/14) infections (including warts, molluscum and CMV infections), vasculitis (9/14) and growth failure (10/14) became prominent features later in life. The majority of children developed severe allergy in the presence of extensive eczema (8/14) and a universal increase in IgA and IgE levels. Moderate thrombocytopenia was present in the majority of patients while overt bleeding tendency was absent after infancy. Immunophenotyping showed a B-cell lymphocytosis and abnormalities in T- and NK-lymphocyte subsets. In vitro T- and B-lymphocyte proliferation and in vivo response to vaccination were normal. Most noticeable was the strongly reduced regulatory T-cell function in those patients tested.

Conclusion: In conclusion, our cohort delineates the spectrum of clinical, hematological and immunological manifestations of subjects with ARPC1B deficiency. The disease appears progressive in most cases and challenging to manage clinically with prophylactic measures and immunosuppression alone.

Disclosure of Interest

None Declared

O26 ARPC1B-deficiency causes defective cell migration, loss of actin polymerization and hyperresponsiveness in zebrafish and patient-derived cells

Gabriella Leung1,2, Aleixo M. Muise2,3
1Cell Biology; 2Gastroenterology, Hepatology and Nutrition; 3Inflammatory Bowel Disease Centre, Hospital for Sick Children, Toronto, ON, Canada
Correspondence: Gabriella Leung

Introduction: ARPC1B-deficiency is a rare paediatric genetic disorder identified in 2017 with autoinflammatory components. Primary symptoms include cutaneous vasculitis, increased susceptibility to infection, microthrombocytopenia, and colitis. ARPC1B is a component of the Arp2/3 complex which regulates branched actin polymerization. Its expression is limited to the haematopoietic compartment, however its function in macrophages and B cells has not been characterised.

Objectives: To determine the effect of ARPC1B-deficiency on macrophage migration and B cell activation.

Methods: Mutant Arpc1b-deficient zebrafish were generated by targeting CRISPR-Cas9 to exon 4 of arpc1b in AB zebrafish embryos, and crossing F1 mutants to mpeg1-GFP+ fish. Four days post-fertilization, tails were cut transversely just distal to the notochord, and GFP+ cells imaged over an 8 hr period. Cells were tracked individually and migration behaviour quantified using Volocity. EBV-transformed lymphoblasts (LCLs) were derived from three previously characterised ARPC1B patients. Patient 1 has a null mutation (c.387_388insCT, L90fs). Patients 2 and 3 are brothers with two SNPs (c.434C>T, A105V; c832G>A, A238T). LCLs were also derived from the parents of Patient 2 and 3, heterozygous for both mutations. Cell phenotype was assessed by immunoblotting, immunofluorescence, and flow cytometry. Calcium flux was analysed by flow cytometry using ratiometric dye Fura Red-AM and stimulating LCLs with anti-IgG. The VCA domain of WASP binds and activates the Arp2/3 complex and was used to stimulate pyrene-actin polymerization activity. Protein interaction between ARPC1B vs. ARPC1A to VCA and WASP were also tested by immunoprecipitation.

Results: ARPC1B-deficient zebrafish were significantly smaller, and monocytes migrated slower compared to WT fish in response to injury. In the patient LCLs, loss of ARPC1B protein, compensatory upregulation of isoform ARPC1A, and loss of total F-actin were confirmed. Patient 1 LCLs were unusually adherent in the absence of stimulation, suggesting constitutive activation. To measure activation, LCLs were stimulated with anti-IgG to measure calcium flux; Patient 1 had a statistically significant higher calcium flux peak compared to control. To address whether Arp2/3 function was compromised, LCL lysates were stimulated with (activating domain) VCA and actin assembly was measured. Although both ARPC1B and ARPC1A isoforms were found to interact with VCA and full-length WASP in immunoprecipitation experiments, VCA-induced actin polymerization in cell lysates was completely abolished in Patients 1-3. Patient 1 was treated with an allogeneic HSCT in January 2018.

Conclusion: Loss of ARPC1B leads to impaired cell migration, loss of Arp2/3 function, and hyperactivation in B cells, which together likely contribute to the dysfunctional immune phenotype. HSCT may represent a curative option for patients with severe ARPC1B-deficiency.

Disclosure of Interest

None Declared

Nucleic acid sensing and interferon

O27 Heterozygous mutations in COPA are associated with enhanced type I interferon signalling

Marie-Louise Frémond1, Alice Lepelley1, Carolina Uggenti2, Maria José Martin-Niclos1, Marine Depp2, Vincent Bondet3, Darragh Duffy3, Gillian I. Rice4, Mary Brennan5, Caroline Thumerelle6, Siham Boulisfane6, Marie Legendre7, Serge Amselem7, Thierry Molina8, Nadia Nathan9, Yanick J. Crow2
1Laboratory of Neurogenetics and Neuroinflammation, Imagine Institute, Paris, France; 2Centre for Genomic and Experimental Medicine, Institute of Genetics and Molecular Medicine, Edinburgh, United Kingdom; 3Immunobiology of Dendritic Cells, Institut Pasteur, Paris, France; 4Division of Evolution and Genomic Sciences, Manchester Academic Health Science Centre, Manchester; 5Department of Paediatric Rheumatology, Royal Hospital for Sick Children, Edinburgh, United Kingdom; 6Pediatrics Department, CHRU de Lille, Lille; 7Genetic Department and Inserm UMR S933, Trousseau Hospital-APHP and Sorbonne Université; 8Pathology Department, Necker Hospital-APHP; 9Inserm UMR S933 and Pediatric Pulmonology department and Reference Centre for Rare Lung Diseases, RespiRare, Trousseau Hospital-APHP and Sorbonne Université, Paris, France
Correspondence: Marie-Louise Frémond

Introduction: Heterozygous mutations in COPA, encoding coatomer protein subunit alpha, cause an autosomal dominant inflammatory syndrome associating lung, joint and renal disease, showing some overlap with STING-associated vasculopathy with onset in infancy (SAVI). Mutations were originally described to cause endoplasmic reticulum (ER) stress and priming of a T helper 17 response. More recently, increased transcription of interferon (IFN)-stimulated genes (ISGs) was reported in blood circulating cells of affected individuals. However, the precise pathophysiology of this disease remains unclear.

Objectives: To better decipher the mechanism of COPA syndrome.

Methods: We studied 8 patients from 3 unrelated families, each segregating a heterozygous mutation in COPA. We assessed type I IFN status by IFNa ultra-sensitive digital quantification in plasma, STAT1 phosphorylation and RNA expression of ISGs in whole blood from patients. In vitro assays also were performed in HEK293T and THP-1 cells to study IFN signalling in the context of COPA mutations.

Results: We observed commonalities in the lung pathology between COPA and SAVI, as well as an IFN signature, raised levels of IFNa protein in the serum and phosphorylation of STAT1 in patient T cells. In a cellular model of HEK293T, phosphorylation of IRF3 and increased ISG expression were observed in cells co-transfected with wild type STING and mutant COPA plasmids. In THP-1 cells, short hairpin RNA knockdown of COPA induced IFN signalling that was abrogated in the absence of STING.

Conclusion: Our data suggest that mutations in COPA lead to constitutive activation of type I IFN signalling through STING. Based on these results, one patient has been treated with the JAK1/2 inhibitor ruxolitinib for the last 12 months. How COPA interacts with ER-resident STING remains to be investigated.


Watkin et al, Nat Genet 2015;47:654-60.

Volpi et al, Clin Immunol 2018;187:33-36.

Disclosure of Interest

None Declared

O28 Sting-associated vasculopathy in mice requires adaptive immunity but not cGAS or type I interferon

Hella Luksch1, Angela Rösen-Wolff1, Alexander Gerbaulet2, W. Alexander Stinson3, Brock G. Bennion3, Gowri Kalugotla4, Wei Qian3, Catherine A. Miner4, Jonathan Miner5
1Department of Pediatrics, University Clinic Carl Gustav Carus, TU Dresden; 2Department of Immunology, Faculty of Medicine,TU Dresden, Dresden, Germany; 3Department of Pathology and Immunology; 4Department of Medicine, Washington University School of Medicine; 5Departments of Pathology and Immunology, Medicine and Molecular Microbiology, Washington University School of Medicine, Saint Louis, St. Louis, United States
Correspondence: Angela Rösen-Wolff

Introduction: It is assumed that monogenic interferonopathies are mediated by type I interferon (IFN) inducing autoinflammation. For instance, it has been shown that a gain-of-function mutation in STING (STING N153S) up-regulates type I IFN-stimulated genes (ISGs) and results inperivascular inflammatory lung disease in mice. The corresponding mutation in humans also causes lung disease. It is thought that signaling via the cGAS-STING pathway and subsequent activation of type I IFN, IFN regulatory factors (IRF) 3/7, and ISGs are involved.

Objectives: We decided to characterize the role of cGAS, IRF3/7, the type I IFN receptor (IFNAR1), and adaptive immunity in the spontaneous inflammatory lung disease in STING N153S mice.

Methods: Hence, we crossed STING N153S mice to animals lacking cGAS, IRF3, IRF7, IFNAR1, adaptive immunity, alph/beta T cells, and mature B cells. As read out we evaluated the mice for development of spontaneous inflammatory lung disease.In addition we generated bone marrow chimeric mice and examined severity of the lung disease and survival of the transplanted animals for 322 days.

Results: We found that spontaneous inflammatory lung disease in STING N153S mice developed independently of cGAS, IRF3, IRF7, and type I IFN signaling. Bone marrow transplantation experiments revealed that certain aspects of STING N153S-associated disease are intrinsic to the hematopoietic system. In additon, we discovered that Rag1-/- STING N153S mice have histologically normal lungs without perivascular infiltrates.Tcr beta-/- STING N153S animals developed a mild lung phenotype.

Conclusion: Spontaneous inflammatory lung disease in STING N153S mice develops independently of cGAS and type I IFN signaling.STING N153S depends on adaptive immunity to induce inflammatory lung disease in mice.

Disclosure of Interest

None Declared

New frontiers in the treatment of SAID

O29 Preclinical studies of gene therapy for deficiency of adenosine deaminase type 2 (DADA2)

Ying Hong, Marina S. Casimir, Barbara Jensen, Benjamin Houghton, Adrian Thrasher, Paul Brogan, Despina Eleftheriou
Infection, immunity and inflammation, UCL Great Ormond Street Institute of Child Health, London, United Kingdom
Correspondence: Ying Hong

Introduction: Deficiency of adenosine deaminase type 2 (DADA2) is an autosomal recessive autoinflammatory disease and vasculitis, caused by loss-of-function mutations in ADA2. Treatment with anti-TNF-α is effective for the autoinflammation and vasculitis of DADA2, but may add to the burden of immunosuppression, is expensive, and is required for life. This treatment may also not effectively treat bone marrow failure or the associated immunodeficiency. Since haematopoietic stem cell transplantation (HSCT) may be curative, but is toxic, we are currently exploring gene therapy for DADA2.

Objectives: (i) develop specific self-inactivating (SIN) lentivirus vectors encoding ADA2 cDNA; (ii) explore the efficacy of gene transfer using this vector in monocyte derived macrophages (MDM) from DADA2 patients; and in an ADA2 knockout (KO) monocyte cell-line (THP-1) model generated using CRISPR/CAS9.

Methods: We generated an ADA2-SIN lentivirus under the control of the elongation-1a-factor (EFS) promoter, that we then used to transduce ADA2 KO THP-1/ wild type (WT) control THP-1 cells; or patient and healthy control-derived MDM, at different multiplicity of infection, and examined the following:
  1. (i)

    ADA2 protein expression.

  2. (ii)

    ADA2 enzyme activity

  3. (iii)

    M1/M2 macrophage immunophenotype. MDM and THP-1 differentiated macrophages were incubated with IL-4, IL-10 and IL-13 to obtain M2 polarized macrophages or with IFN-γ/LPS to induce polarization to M1. Immunophenotyping was assessed in transduced or non-transduced M1 and M2 polarised macrophages using qPCR, flow cytometry, and ELISA-cytokine quantification in culture supernatants.

  4. (iv)

    macrophage induced endothelial activation. Transduced or non-transduced MDM from DADA2 patients and healthy controls were also cultured with endothelial cells, and CD62E expression (marker of endothelial activation) was examined with flow cytometry

Results: In the THP-1 KO cell line we observed:
  1. (i)

    full restoration of ADA2 protein expression and ADA2 enzyme activity in transduced ADA2 KO THP-1 cells compared to non-transduced cells;

  2. (ii)

    amelioration of M1 proinflammatory molecule gene expression: polarized M1 cells derived from transduced ADA2 KO THP-1 cells exhibited a similar gene expression profile for proinflammatory cytokines (TNF-α, CXCL-10, STAT-1, and IL-1β) to that observed in M1 derived from WT control THP-cells; in comparison, non-transduced ADA2 KO THP-1 cells exhibited significant up-regulation of gene expression for these proinflammatory cytokines (p<0.0001).

Using DADA2 patient derived cells (n=3) we also established:
  1. (iii)

    full restoration of ADA2 protein expression and enzyme activity: transduction of DADA2 MDM led to full restoration of ADA2 protein expression and enzyme activity to levels observed in healthy controls (p=0.0001);

  2. (iv)

    amelioration of M1 proinflammatory gene expression in patient cells: transduced DADA2 MDM were restored to levels seen in healthy control MDM cells for proinflammatory cytokine gene expression (p=0.0001); cytokine production at protein level (p=0.01); and reduced M2 apoptosis (p=0.0001).

  3. (v)

    prevention of endothelial activation: transduction of DADA2 MDM significantly prevented endothelial activation in co-culture experiments, compared with non-transduced DADA2 MDM, that continued to induce significant endothelial activation (p=0.0002).


Conclusion: We have used a gene therapeutic approach to successfully demonstrate rescue of the immunophenotype of DADA2 using a THP-1 ADA2 KO cell line model; and primary cells from DADA2 patients, thus providing proof of principle that gene therapy might work in patients with DADA2. Next steps now include: 1. gene correction in CD34 cells from DADA2 patients; and 2. safety studies in animal models.

Disclosure of Interest

None Declared

O30 Experience with and management of HLH-like toxicities following chimeric antigen receptor T-cell therapy for treatment of relapsed/refractory pre-B ALL

Amanda K. Ombrello1, Bonnie Yates2, Haneen Shalabi2, Terry J. Fry3, Nirali N. Shah2
1NHGRI; 2NCI, NIH, Bethesda; 3Children's Hospital of Colorado, Denver, United States
Correspondence: Amanda K. Ombrello

Introduction: Chimeric antigen receptor T-cell (CAR-T) therapy is a highly effective form of adoptive cell immunotherapy combining antigen specific targeting capabilities with T-cell based cytotoxicity. Particularly effective against B-cell antigens (CD19/CD22), CAR-T cell activation leads to a systemic inflammatory response called cytokine release syndrome (CRS) that can further evolve into hemophagocytic histiocytosis (HLH) symptomatology.

Objectives: To evaluate the presentation, incidence and management of HLH toxicities in children/young adults with relapsed/refractory pre-B acute lymphoblastic leukemia (pre-B ALL) treated on a phase I study of anti-CD22 CAR-T cell therapy ( NCT02315612).

Methods: Using modified diagnostic criteria to define CAR-T cell related HLH, it was established in those with ferritin of > 100,000 ng/mL and at least 1 of the following: > grade 3* AST/ALT elevation or hyperbilirubinemia; > grade 3* oliguria or increase in serum creatinine; > grade 3* pulmonary edema; or hemophagocytosis in the bone marrow. Serial inflammatory markers (ferritin, CRP) and serum cytokines were prospectively monitored from CAR-T cell infusion through day +28 (+/-4) and retrospectively analyzed comparing peak values in those who did/did not develop HLH. Treatments included supportive care, glucocorticoids +/- anakinra.

*Grading as per Common Terminology Criteria for Adverse Events, v4.03

Results: In 52 subjects, 46 experienced CRS, of whom 37 (80.4%) achieved complete remission and 18 (39.1%) developed HLH. Median ferritin in those with/without HLH was 206740 vs. 22758 (ng/mL) (Table 1). Clinical manifestations included: > grade 3* creatinine (n=2); > grade 3* AST/ALT elevation or hyperbilirubinemia (n=23, including 6 without HLH), > grade 3* pulmonary edema (n=5, including 2 without HLH); and hemophagocytosis in the bone marrow (n=9). Cytokine profiling demonstrated significantly higher levels of IFN-y, IL-1B, IL-6, IL-10, TNFa and MIP-1a (Table 1). Limited paired samples of sIL-2R showed statistically significant increase from baseline (median level 1254) to HLH presentation (median 9310), with 6/9 having substantial increase (peak 123700). All had resolution of HLH symptoms, except 1 who died from gram-negative rod sepsis complications prior to resolution. Three had asymptomatic lab abnormalities that self-resolved (median 14 days) without intervention. Anakinra monotherapy (median 5 mg/kg/day) was used in 3 and the remainder (n=3) received anakinra + steroids. Both regimens resolved HLH. There was no statistically significant difference in underlying leukemia burden or efficacy following CAR-T cell therapy in those who did/did not develop HLH. Notably, use of anakinra +/- steroids did not diminish therapeutic efficacy of CAR-T cells.

Conclusion: Ferritin and cytokine profiling revealed HLH patients had a different inflammatory response independent of disease burden. Anakinra +/- steroids was effective and did not impede CAR-T cell expansion. Further analysis to identify HLH-predictive parameters and optimizing interventions to treat/prevent these complications are ongoing.

Disclosure of Interest

None Declared

Table 1 (abstract O30).

See text for description

Inflammatory Marker

No HLH, median (25-75% IQR)

HLH, median (25-75% IQR)

p value (1- tailed)


22758 (3554-52686)

206740 (171968-420273)


Cytokines* (pg/mL)


352.2 (196.7-1041)

2800 (1838-2900)



0.77 (0.45-2.09)

3.51 (1.02-48.95)



41.58 (18.83-214.5)

904.5 (264.1-1480)



55.94 (22.02-154)

338.7 (128.1-567.4)



12.77 (9.17-23.62)

27.1 (16.2-43.91)




223.8 (157-422.2)


*For ferritin, n=19 (No HLH) and n=18 (HLH) due to initial lack of ferritin monitoring. All other cytokines, n=27 (No HLH) and n=18 (HLH)

Novel targets and therapies in autoinflammation

O31 Novel NLRP3 targeted therapy in caps

Laela M. Booshehri1, Matthew McGeough1, Ben Keer1, Milos Lazic2, Christopher McBride2, Davide Povero2, James Veal2, Gretchen Bain2, Hal M. Hoffman1
1Pediatrics, University of California San Diego, La Jolla; 2Jecure Therapeutics, San Diego, United States
Correspondence: Hal M. Hoffman

Introduction: Cryopyrin-associated periodic syndrome (CAPS) is an autoinflammatory disease characterized by a hyperactive inflammasome leading to the overproduction of interleukin-1b (IL-1b). Assembly of the NLRP3 inflammasome is central to the CAPS disease process resulting in subsequent activation and prolific release of inflammatory cytokines, which further propagate inflammation and disease. Current therapies for CAPS patients directly target IL-1b or IL-1 receptor to mitigate excess inflammation caused by NLRP3 activation. While there are a number of new compounds in pre-clinical development that target NLRP3 directly, there is little data on the efficacy of these compounds in CAPS versus healthy controls.

Objectives: To study the ex vivo efficacy of a novel NLRP3 selective small-molecule inhibitor compound 1 (C1) in monocytes from CAPS patients with multiple Nlrp3 mutations as compared to healthy controls and in bone marrow derived dendritic cells from murine Nlrp3 mutant CAPS models as compared to wild type mice, with the goal of identifying an effective NLRP3 specific inhibitor for CAPS patients.

Methods: Peripheral blood mononuclear cells from 7 CAPS patients with 6 different NLRP3 mutations and 4 healthy donor controls were isolated by gradient centrifugation, and monocytes were allowed to adhere for 4 hours prior to treatment and stimulation. Samples were obtained under an approved Institutional Review Board protocol for human subjects. Bone marrow was isolated from MWS Nlrp3A350V/+ CreT, FCAS Nlrp3L351P/+ CreT, and NOMID Nlrp3D301N/+ CreT conditional knock-in mice and cells were cultured with GMCSF for 1 week and treated with tamoxifen 1 day prior to drug treatment to induce expression of the mutation. Cells were treated with C1 at varying concentrations prior to stimulation with LPS (mutants) or LPS and ATP (controls), and supernatants were collected after overnight incubation for analysis of IL-1b by ELISA. C1 was also orally administered to MWS Nlrp3A350V/+ CreT mice prior to in vivo tamoxifen administration to determine in vivo efficacy and pharmacodynamics. Whole body and spleen weights were measured and blood was obtained for complete blood counts in treated and untreated mice. Animal studies were performed in accordance with the University of California San Diego and IACUC policies and procedures.

Results: Robust inhibition of IL-1b release from C1 treated cells was shown with comparable activity across multiple human NLRP3 mutations and murine Nlrp3 mutant models. Compound efficacy and pharmacodynamics were also shown to be similar between CAPS mutant cells and cells from respective healthy donors or wild-type controls. Daily oral administration of C1 was well tolerated and demonstrated efficacy in preventing weight loss, splenomegaly, and neutrophilia in MWS Nlrp3A350V/+ CreT mice.

Conclusion: The novel NLRP3 inhibitor C1 was shown to significantly reduce LPS induced IL-1b release in cells from CAPS patients and murine Nlrp3 mutant models indicating direct and effective inhibition of mutant NLRP3. C1 and other emerging NLRP3 inhibitors present an additional avenue for future CAPS patient therapy by directly targeting the inflammasome. Similar efficacy of C1 on both normal and mutant NLRP3 likewise indicate potential applications in other inflammatory diseases beyond CAPS.

Disclosure of Interest

None Declared

O32 Preclinical efficacy of NLRP3 small molecule inflammasome inhibitors: implications for future treatment of autoinflammatory syndromes

Angela Abad-Perez1, Stefan Frischbutter1,2, Niklas A. Mahnke1, Jens v. Kries3, Marc Nazaré4, Marcus Maurer1, Jörg Scheffel1,2, Karoline Krause1,2
1Department of Dermatology, Venereology and Allergology; 2Autoinflammation Reference Center Charité (ARC2), Charité Universitätsmedizin Berlin; 3Screening Unit Cell Biology and High Content Screen; 4Medicinal Chemistry, Leibniz-Forschungsinstitut für Molekulare Pharmakologie (FMP), Berlin, Germany
Correspondence: Angela Abad-Perez

Introduction: Systemic autoinflammatory diseases (SAIDs) are characterized by abnormally increased inflammation affecting different organs. They are mediated predominantly by the cells and molecules of the innate immune system. Inflammasome activation represents the critical pathogenic mechanism shared by most SAIDs. Current treatment strategies are limited to downstream cytokine blockade. Specific inflammasome inhibitors are not available so far.

Objectives: To address this unmet medical need, we performed a high content screening of more than 60.000 small molecules from the compound collection of the …Leibniz-Forschungsinstitut für Molekulare Pharmakologie (FMP)”, which includes the ChemBioNet and LOPAC®1280 libraries as well as donations of academic chemists, FDA approved drugs (Selleck library) and a natural product collection from AnalytiCon Discovery.

Methods: For the primary screen, we used a fluorescent murine inflammasome reporter cell line to detect ASC speck formation, a marker of inflammasome activation. Compounds were selected based on their inhibitory capacity on ASC speck formation, as observed by automated fluorescence microscopy, and IL-1ß release after activation with canonical NLPR3-inflammasome inducers ATP and nigericin. The 10 most potent and druggable hit compounds were tested in peripheral blood mononuclear cells (PBMCs) obtained from venous blood of patients with Schnitzler syndrome (N=8), familial Mediterranean fever, FMF (N=8) and from unmatched healthy donors (N=18). In vitro effect on cellular IL-1ß release was measured by ELISA.

Results: Selected compounds proved to efficiently inhibit the secretion of IL-1ß in PBMCs from both patients and healthy controls in a dose dependent manner, validating their inhibitory capacities in human and murine cellular assays. Among these compounds were known anti-inflammatory drugs such as auranofin and a VEGFR2 tyrosine kinase inhibitor. The median inhibitory capacity upon stimulation with lipopolysaccharide (LPS) and ATP at 10 μM ranged between 50% to 80% with IC50s in the low μM region. A similar inhibitory profile could be observed for the previously reported inflammasome inhibitor MCC950, which was included in our assays as a reference substance. Moreover, compounds had similar efficacy in inflammasome inhibition PBMCs obtained from patients and healthy controls.

Conclusion: Based on our results in murine and human cells in vitro, small molecule inflammasome inhibitors my complement current treatment options for SAIDs in the future.

Disclosure of Interest

None Declared

Oral communications – new diseases

O33 Biallelic loss of function mutations in sharpin cause autoinflammation

Hirotsugu Oda1, David Beck1, Kalpana Manthiram1, Hye Sun Kuehn1, Natalia Sampaio Moura1, Rao Anand2, Mariana Kaplan1, Douglas Kuhns3, Wanxia Li Tsai1, Hiroyuki Yoshitomi4, Junya Toguchida4, Gustavo Gutierrez-Cruz1, Jeremy Davis1, Massimo Gadina1, Jennifer Stoddard1, Kazuhiro Iwai4, Sergio Rosenzweig1, Luigi Notarangelo1, Daniel L. Kastner1, Ivona Aksentijevich1
1NIH, Bethesda, United States; 2Manipal Hospital, Bangalore, India; 3NIH, Frederick, United States; 4Kyoto University, Kyoto, Japan
Correspondence: Hirotsugu Oda

Introduction: The linear ubiquitination chain assembly complex (LUBAC) consists of HOIP, HOIL1 and SHARPIN and mediates linear ubiquitination. LUBAC is essential for NF-κB signaling and thus proper innate and adaptive immunity. Patients with HOIP and HOIL1 deficiencies have been reported to have immunodeficiency, autoinflammation and amylopectinosis. Although mice deficient in Sharpin have severe TNF-dependent skin inflammation due to enhanced apoptosis and necroptosis of the keratinocytes, the role of SHARPIN in human diseases is unknown.

Objectives: We aimed to investigate a 14 year-old boy from a consanguineous family in India with polyarthritis, parotitis, hepatosplenomegaly and colitis associated with anorectal fistula, but without skin manifestations or any history of severe infections. The patient’s symptoms dramatically improved on anti-TNF therapy.

Methods: We performed whole exome sequencing to identify the genetic cause of the patient's phenotypes.

Results: We identified a homozygous frameshift mutation in SHARPIN in our proband (c.220dupC). Patient derived dermal fibroblasts have no detectable SHARPIN protein with markedly reduced HOIP, and HOIL1 suggesting destabilization of the LUBAC complex. These cells also displayed impaired canonical NF-κB activity, as exemplified by induction of IκBα phosphorylation and nuclear translocation of p65. However, in contrast to HOIP and HOIL1 deficiencies, the patient’s monocytes did not show hyperresponsiveness to IL-1β stimulation.SHARPIN deficient patient fibroblasts demonstrated enhanced apoptosis induced by FAS stimulation as compared to control cells, which parallels the enhanced apoptosis observed in the Sharpin deficient mice. We knocked out SHARPIN in a human immortalized osteoblast cell line (hFOB1.19) and interestingly, despite attenuated NF-κB activity, these cells secrete higher amounts of IL-6 more rapidly after IL-1β stimulation than control cells.

Conclusion: We identified the first case of human SHARPIN deficiency in a patient with autoinflammation. Molecular consequences of the SHARPIN deficiency are currently being investigated in comparison to other LUBAC deficiencies.

Consent for publication has been obtained from patient


Disclosure of Interest

None Declared

O34 A loss-of-function mutation in USP43, a deubiquitinase gene, is linked to an interferon-mediated autoinflammatory disorder with proteasome defects

Hongying Wang1, Qing Zhou1, Anna Kozlova2, Vasili Burlakov2, Daniel Kastner1, Ivona Aksentijevich1, Anna Shcherbina2
1Inflammatory Disease Section, National Human Genome Research Institute (NHGRI) / NIH, Bethesda, United States; 2National Research and Practical Center of Pediatric Hematology, Oncology and Immunology, Moscow, Russian Federation
Correspondence: Hongying Wang

Introduction: Deubiquitinase enzymes (DUBs) function in the removal of poly-ubiquitin chains from substrate proteins to regulate their activity and degradation by the ubiquitin-proteasome system (UPS). Deficiency of DUB activity may lead to excessive immune signaling as has been observed in patients with haploinsufficiency of A20 (HA20) and OTULIN deficiency. Through whole exome sequencing analysis (WES), we identified a novel homozygous missense mutation (c. 2509G>A; p. E837K) in a deubiquitinase encoding gene USP43, in a Russian patient of Tatar ancestry. The patient presented with early-onset recurrent fevers, rash, subcutaneous skin nodules, lipodystrophy, and prominent arthritis, and this phenotype was suggestive of the CANDLE syndrome.

Objectives: USP43 is a poorly characterized ubiquitin specific protease (USP).We aimed to study the disease-causing mechanism underlying this novel mutation, E837K, as well as the biological function and targets of USP43.

Methods: We performed WES in the patient’s family and RNA sequencing in whole blood samples. We generated a fibroblast cell line and EBV-transformed B cells from the patient’s primary cells. We used USP43 knockdown and CRISPR/Cas9 knockout in 293T cells to study the effects of the USP43 depletion. A series of USP43 truncated mutants were generated to study the effect of protein domains on its DUB activity. Immunoprecipitation and immunoblot, luciferase assays, serum and plasma cytokine profiling, immunofluorescence, real-time PCR, and flow cytometry were used to investigate abnormalities in patient-derived cells.

Results: We found that the novel USP43 mutation leads to an upregulation in interferon signaling and causes an impairment in the proteasome-mediated protein degradation pathway. The patient’s EBV-transformed B cells had increased phospho-STAT levels in response to interferon stimulation and spontaneously produced a significantly higher level of IL-6. This cellular phenotype was rescued by transfection with wild-type USP43, which suggests that this mutation is loss-of-function.The patient’s primary cells showed decreased proteasome activity and excessive accumulation of K48-ubiquinated proteins following stimulation with a proteasome inhibitor MG132. Similarly, transient knockout of USP43 in 293T cells led to increased levels of ubiquitinated proteins. Reintroducing wildtype USP43 to EBV-transformed patient B cells markedly decreased the expression of ubiquitinated proteins. These data suggest that the mutant USP43/E837K protein loses the ability to remove K48-ubiquitin chains from target proteins. Overexpression of a series of truncated USP43 mutants with K-48 Ub chains in 293T cells confirmed that the C-terminal domain is required for the DUB function of USP43. In addition, USP43 mutants possessing the E837K mutation lost the ability to clear accumulated K48-Ub chains. Interferon-stimulated patient’s EBV cells and fibroblasts showed decreased protein levels of PSMB8.PSMB8 encodes the catalytic subunit of the immunoproteasome. Co-transfection of USP43/E837K mutant with PSMB8 in 293T cells reduced the expression of PSMB8 precursor protein compared to cells transfected with USP43 wild type.

Conclusion: Our data suggest that the loss-of-function mutation in USP43 decreases immunoproteasome activity and causes an upregulation in type I interferon signaling, similar to what is observed in patients with CANDLE. Treatment with a JAK inhibitor has been very effective in controlling the disease activity in this patient. To our knowledge, this is the first report of a human disease caused by mutation in USP43.

Disclosure of Interest

None Declared

O35 A novel autoinflammatory disease characterized by neonatal-onset cytopenia with autoinflammation, rash, and hemophagocytosis (NOCARH) due to aberrant CDC42 function

Michael T. Lam1,2,3, Simona Coppola4, Oliver H. Krumbach5, Giusi Prencipe6, Antonella Insalaco6, Cristina Cifaldi7,8, Immacolata Brigida9, Serena Scala9, Marcello Niceta10, Andrea Ciolfi10, Alexandre F. Carisey1,2, Mohammad Akbarzadeh5, Andrea Finocchi7,8, Franco Locatelli11, Caterina Cancrini7,8, Alessandro Aiuti9,12,13, Mohammad R. Ahmadian5, Jordan S. Orange2, Fabrizio De Benedetti6, Marco Tartaglia10
1Department of Pediatrics, Baylor College of Medicine, Houston; 2Department of Pediatrics, Columbia University, Irving Medical Center, New York; 3Translational Biology and Molecular Medicine Graduate Program and Medical Scientist Training Program, Baylor College of Medicine, Houston, United States; 4National Center for Rare Diseases, Istituto Superiore di Sanità, Rome, Italy; 5Institute of Biochemistry and Molecular Biology II, Medical Faculty of the Heinrich-Heine University, Düsseldorf, Germany; 6Division of Rheumatology; 7Department of Pediatrics, Ospedale Pediatrico Bambino Gesù, IRCCS; 8Department of Systems Medicine, University of Rome Tor Vergata, Rome; 9San Raffaele Telethon Institute for Gene Therapy (SR-TIGET), IRCCS San Raffaele Scientific Institute, Milan; 10Genetics and Rare Diseases Research Division; 11Department of Pediatric Hematology and Oncology, Ospedale Pediatrico Bambino Gesù, IRCCS, Rome; 12Pediatric Immunohematology, San Raffaele Scientific Institute; 13Vita Salute, San Raffaele University, Milan, Italy
Correspondence: Antonella Insalaco

Introduction: Despite continuous advances in the identification of novel causative genes, several patients with a clinical autoinflammatory phenotype remain unclassifiable.

Objectives: to describe a novel hematological and autoinflammatory disorder in three unrelated patients caused by a de novo missense mutation of CDC42

Methods: Whole exome sequencing was used to identify the novel variant. The functional impact of altered CDC42 function on hematopoiesis and inflammation was assessed through patient peripheral blood and bone marrow analyses, protein behavior and immune and non-immune cell functioning through in vitro biochemical and functional assays and in vivo C. elegans modeling.

Results: Patients shared the same de novo missense mutation of CDC42 (NM_001791, Chr1:22417990, c.556C>T, p.R186C).Disease features included neonatal-onset cytopenia with dyshematopoiesis, autoinflammation, rash, and hemophagocytosis (collectively termed NOCARH syndrome) (Table). An altered hematopoietic compartment (prevalence of early differentiation elements and substantially decreased clonogenic progenitors) was demonstrated. Complementary assays documented the unique consequences of this mutation on CDC42 localization and function, and its disruptive effect on cell behavior and developmental processes, possibly linked to actin dysregulation. Increased secretion of IL-1β, and particularly of IL-18, was observed via ex vivo spontaneous release from unstimulated bone marrow mononuclear cells and by high levels in bone marrow supernatants and plasma. IFNγ was alsoincreased and correlated to CXCL9 levels which were strictly related to ferritin levels. Treatment with anakinra and emapalumab, a monoclonal antibody to IFNγ, was identified as critical in the survival of one patient, who underwent successful hematopoietic stem cell transplantation.

Conclusion: The p.R186C amino acid substitution in CDC42 underlies a novel, unique syndrome where CDC42 functional dysregulation has pleiotropic effects, causing hematopoietic disturbance, hyperinflammation, and immune impairment. Early recognition and control of HLH, through neutralization of IFNγ, followed by hematopoietic stem cell transplantion, appear to be crucial to survival.

Disclosure of Interest

None Declared

Table 1 (abstract O35).

See text for description

Outcome and status

Patient 1

Patient 2

Patient 3

Alive, 6 yrs

Dead, 6 mos

Dead, 1.5yrs





Skin rash








Hemophagocytic lymphohistiocytosis




Gastrointestinal symptoms








Acute phase response




Bone marrow dysplasia




O36 PSMB10, The last immunoproteasome gene missing for PRAAS (Proteasome-Associated Autoinflammatory Syndrome)

Guillaume Sarrabay1,2, Déborah Méchin1,2, Aicha Salhi3, Guilaine Boursier1, Cécile Rittore1, Yanick Crow4, Gillian Rice5, Tu-Ahn Tran2,6,7, Renaud Cezar7, Darragh Duffy8, Vincent Bondet8, Lakhtar Boudehane9, Sylvie Grandemange1,2, Florence Apparailly2, Isabelle Touitou1,2
1Department of Medical Genetics, Rare diseases and Personalized medicine, Rare and Autoinflammatory diseases unit; 2IRMB, INSERM, CHU Montpellier, Univ Montpellier, Montpellier, France; 3Dermatology department, Alger medicine University, Alger, Algeria; 4Laboratory of Neurogenetics and Neuroinflammation, Institut Imagine, Paris Descartes University, Paris, France; 5Division of Evolution and Genomic Sciences, School of Biological Sciences, Faculty of Biology, Medicine and Health, University of Manchester, Manchester Academic Health Science Centre, Manchester, United Kingdom; 6Paediatrics department, University Hospital Nimes; 7Immunology department, CHU Nîmes, Univ Montpellier, Nîmes; 8ICD Unit, Inserm U1223, Institut Pasteur, Paris, France; 9Paediatrician office, Liberal, Sétif, Algeria
Correspondence: Guillaume Sarrabay

Introduction: PRAAS defines a clinical spectrum encompassing JMP (joint contractures, muscle atrophy, microcytic anemia and panniculitis-induced childhood-onset lipodystrophy syndrome), NNS (Nakajo-Nishimura syndrome) and CANDLE (chronic atypical neutrophilic dermatosis with lipodystrophy and elevated temperature). PRAAS is caused by autosomal recessive, autosomal dominant, or digenic mutations in several genes encoding for either constitutive proteasome subunits (PSMB4, PSMA3), immunoproteasome subunits (PSMB8, PSMB9) or chaperone protein (POMP). We describe here a 3-year-old female patient with clinical features evocative of PRAAS in whom no mutations have been found using our 62 gene panel sequencing that includes known PRAAS genes.

Objectives: The aim of the study was to identify the molecular cause responsible for the PRAAS phenotype in this patient.

Methods: We performed a trio-based whole exome sequencing (WES) in the patient and her parents. Functional assays were conducted to confirm the pathogenic effect of the mutated candidate gene. They included: enzymatic protease activity in the patient’s peripheral blood monocyte cells (PBMC) and in transfected HEK293T cells, interferon (IFN) signature and IFNα dosage, multiplexed cytokines measurement from patient’s serum, and protein maturation assays by WB (western-blot) analyses in transfected wild-type and mutant HEK293T cells.

Results: The patient is a 3-year-old female patient of Algerian descent, born to related parents. She developed a cutaneous rash on the 7th day of life and became febrile at the age of one year. The rash was polymorphic, annular shaped and predominantly periorbital She failed to thrive and has long-lasting hepatosplenomegaly. She has an emaciated face, a distinctive nose, and long and gracile fingers. She had elevated acute phase reactants, microcytic anemia and hypertriglyceridemia. She exhibited partial response to steroid and methotrexate treatment and relapsed when the doses were lowered. WES revealed a homozygous missense mutation in the candidate gene PSMB10, located in the N-terminal part of protein which is cleaved in the mature form. This variant is absent from the GnomAD cohort, and predicted to be pathogenic according to in silico bioinformatic tools. The patient had a positive interferon signature and elevated IFNα protein in the serum on the one occasion tested. Cytokines multiplexed measurement showed raised IL-6, TNFα, MIG and MCP-3 in the patient’s serum whereas IL-1β level was similar to healthy pediatric controls. WB assays in HEK293T cells showed defective cleavage of the mutant protein upon IFNγ induction compared to the wild-type protein. Patient’s PBMC and mutant HEK293T cells showed an alteration in trypsin-like proteasome activity.

Conclusion: We report here a patient with clinical and biological criteria consistent with PRAAS, with a homozygous PSMB10 mutation. This is the third and last immunoproteasome subunit involved in this disease, and this new gene responsible for PRAAS expands the number of genes involved in this spectrum.

Consent for publication has been obtained from patient


Disclosure of Interest

None Declared

O37 WNT6 mutation causes an early onset granulomatosus intestinal disease with recurrent hemophagocytic lymphohistiocytosis (HLH)

Claudia Bracaglia1, Daniela Knafelz2, Fiammetta Bracci2, Antonella Insalaco1, Giulia Marucci1, Manuela Pardeo1, Giusi Prencipe1, Ivan Caiello1, Antonia Pascarella1, Marcello Niceta3, Francesca Pantaleoni3, Andrea Ciolfi3, Bronislava Papadatou2, Marco Tartaglia3, Giuliano Torre2, Fabrizio De Benedetti1
1Division of Rheumatology; 2Hepatology, Gastroenterology and Nutrition Unit; 3Genetics and Rare Diseases Research Division, IRCCS Ospedale Pediatrico Bambino Gesù, Rome, Italy
Correspondence: Claudia Bracaglia

Introduction: Use of NGS in patients with unclassifiable disease lies a possible approach to the identification of novel disease causing genes.

Objectives: We report a patient with an early onset inflammatory bowel disease with granulomatous lesions and recurrent HLH episodes carrying a missense mutation in the WNT6 gene.

Methods: A trio based Whole Exome Sequencing (WES) approach was used. Cytokine levels were measured by multiplex assay and by specific ELISAs.

Results: Ten years old Caucasian boy affected by early onset pan-colitis from 9 months of age. Since the disease onset the patient is on glucocorticoid treatment with amino acidic enteral nutrition and oligo antigenic diet. Because of recurrent disease relapses at any attempt of glucocorticoid withdrawal, azathioprine and cyclosporine treatments were also added. At 2 years of age he received total colectomy with ileostomy. Because of insufficient disease control, treatment with a TNF-inhibitor (infliximab) was started with apparent improvement of intestinal symptoms.However, persistent granulomatous inflammatory disease of the distal portion of the ileus-rectal anastomosis persisted. Moreover, the patient presented recurrent HLH episodes that required high dose of glucocorticoid and cyclosporine-A treatment. Except one HLH episode related to a varicella zoster infection, the other HLH events were most likely triggered by his underlying inflammatory condition. During the HLH episodes levels of IL-18 were moderately elevated (10.880 pg/ml) the IFN-gamma induced chemokine CXCL9 was markedly high (21.871 pg/mL) and remained markedly elevated also during clinical and laboratory HLH remission (3.121 pg/ml and 9.929 pg/ml respectively). Considering the early disease onset, primary immunodeficiency and early intestinal bowel disease onset were genetically ruled out as well as chronic granulomatosis diseases through extensive NGS panels. WES revealed carriage of a private (MAF: 1/125568, TOPMED), predicted pathogenic (CADD: 31), homozygous variant of WNT6 (c.793G>C; p.(Asp265His); NM_006522.3). The patient is now partially controlled on low dose of oral glucocorticoid (0.1 mg/kg), cyclosporine-A (5mg/kg) and antimicrobic treatment.

Conclusion: WNT signalling has been primarily described as a regulatory pathway in ontogeny and homeostatic processes. Schaale at al. demonstrated that WNT6 is expressed in granulomatous lesions in the lung of Mycobacterium tuberculosis–infected mice. Moreover, they found that the transcription factor c-Myc is significantly induced in murine macrophages by WNT6. This identifies WNT6 as a novel factor driving macrophage polarization toward an M2-like phenotype, suggesting a role for WNT6 in macrophage differentiation. Our case suggests defective function of WNT6 might be involved in the development of a granulomatous disease. WNT6 role in macrophage differentiation and polarization might also be important in the activation of the IFN-gamma pathway and in recurrent HLH episodes.


K. Schaale et al. Wnt6 Is Expressed in Granulomatous Lesions of Mycobacterium tuberculosis–Infected Mice and Is Involved in Macrophage Differentiation and Proliferation. J Immunol 2013; 191:5182-5195.

Consent for publication has been obtained from patient


Disclosure of Interest

C. Bracaglia: None Declared, D. Knafelz: None Declared, F. Bracci: None Declared, A. Insalaco: None Declared, G. Marucci: None Declared, M. Pardeo: None Declared, G. Prencipe: None Declared, I. Caiello: None Declared, A. Pascarella: None Declared, M. Niceta: None Declared, F. Pantaleoni: None Declared, A. Ciolfi: None Declared, B. Papadatou: None Declared, M. Tartaglia: None Declared, G. Torre: None Declared, F. De Benedetti Grant / Research Support from: Novartis, Novimmune, Hoffmann- La Roche, SOBI, AbbVie, Pfizer

O38 NFIL3 mutations alter immune homeostasis and sensitize for arthritis pathology

Stephanie Humblet-Baron1, Susan Schlenner2, Emanuela Pasciuto2, Vasiliki Lagou1, Oliver Burton2, Teresa Prezzemolo1, Steffie Junius1, Carlos Roca1, Cyril Seillet3, Cynthia Louis3, James Dooley1, Kylie Luong3, Erika Van Nieuwenhove1, Ian P Wicks3, Gabrielle Belz3, Adrian Liston1, Carine Wouters4
1Immunology and Microbiology, Center for Brain and disease research, KU Leuven-VIB; 2Immunology and Microbiology, Center for Brain and disease research, KU Leuven - VIB, LEUVEN, Belgium; 3Walter and Eliza Hall Institute of Medical Research, Melbourne, Australia; 4KU Leuven and University Hospitals Leuven, LEUVEN, Belgium
Correspondence: Stephanie Humblet-Baron

Introduction: Juvenile idiopathic arthritis (JIA) is the most common of the childhood rheumatic diseases. JIA is characterized as juvenile-onset persistent arthritis with no defined cause. A high degree of clinical heterogeneity is observed within the JIA group of diseases, thought to reflect a diversity in genetic and environmental factors and mechanistic drivers. JIA shows similarities to adult autoimmune diseases,but also has similarities to autoinflammatory diseases, such as genetic associations to innate inflammatory pathways and response to IL-1β blockade. The recent success in identifying monogenic causes of autoinflammatory diseases suggests that monogenic causes may also underlie a subset of JIA patients.

Objectives: NFIL3 is a key immunological transcription factor, with knockout mice studies identifying functional roles in multiple immune cell types. Despite the importance of NFIL3, little is known about its function in humans.

Methods: Here we characterized a kindred of two monozygotic twin girls with juvenile idiopathic arthritis at the genetic and immunological level, using whole exome sequencing, single cell sequencing and flow cytometry. Parallel studies were performed in a mouse model.

Results: The patients inherited a novel p.M170I in NFIL3 from each of the parents. The mutant form of NFIL3 demonstrated reduced stability in vitro. The potential contribution of this mutation to arthritis susceptibility was demonstrated through a pre-clinical model, where Nfil3-deficient mice upregulated IL-1β production, with more severe arthritis symptoms upon disease induction. Single cell sequencing of patient blood quantified the transcriptional dysfunctions present across the peripheral immune system, converging on IL-1β as a pivotal cytokine.

Conclusion: NFIL3 mutation can sensitize for arthritis development, in mice and humans, and rewires the innate immune system for IL-1β over-production.

Disclosure of Interest

None Declared

O39 EROS/CYBC1 mutations: a novel cause of chronic granulomatous disease and more

David C. Thomas1, Louis M. Charbonnier2, Andrea Schejtman3, Hasan Aldhekri4, Eve Coomber5, Elizabeth Dufficy6, Anne Beenken1, James Lee1, Simon Clare7, Anneliese Speak7, Adrian Thrasher8, Giorgia Santilli8, Hamoud Al-Mousa9, Fowzan Alkuraya10, Talal Chatila11, Kenneth Smith1
1Department of Medicine, University of Cambridge, Cambridge, United Kingdom; 2Paediatrics, Harvard, Boston, United States; 3University College London, London, United Kingdom; 4Department of Paediatrics,King Faisal Specialist Hospital and Research Center , Riyadh, Saudi Arabia; 5Wellcome Trust Sanger Institute, Cambridge; 6Medicine, University of Cambridge, Camridge; 7WTSI, Cambridge; 8Institute of Child Health, UCL, London, United Kingdom; 9Paediatrics; 10Genetics, KFSH, Riyadh, Saudi Arabia; 11Paediatrics, Harvard University, Boston, United States
Correspondence: David C. Thomas

Introduction: The multi-subunit phagocyte nicotinamide adenine dinucle- otide phosphate oxidase generates reactive oxygen species and is crucial for host defence. Deficiencies in individual subunits (gp91phox, p22phox, p47phox, p67phox, and p40phox) cause chronic granulomatous disease (CGD), but some patients with CGD do not have mutations in these genes. We recently found that Eros, a hitherto undescribed protein, is essential for the generation of reactive oxygen species because it is necessary for protein (but not mRNA) expression of the gp91phox-p22phox heterodimer, which is almost absent in Eros-deficient mice. Eros-/- animals succumb quickly following infection with Salmonella typhimurium or Listeria monocytogenes. Eros is highly conserved and has a human orthologue CYBC1 (alias C17ORF62), hereafter referred to asCYBC1 gene and essential for reactive oxygen species (EROS) protein.

Objectives: We asked:
  1. 1.

    Whether the gene fulfilled the same function in humans.

  2. 2.

    Whether mutaions in juman EROS/CYBC1/C17ORF62 could cause a human disease


Methods: We performed CRISPR-mediated deletion of CYBC1/EROS in PLB-985 cells and identified 2 clones with 8 bp and 1 bp deletions, respectively. Neither clone expressed detectable EROS protein. We also identified a patient with a homozygous EROS/CYBC1/C17ORF62 who was subsequently diagnosed with chronic granulomatous disease secondary to this mutation.

Results: We show that the function of CYBC1/EROS is conserved in human cells. Knockout of EROS in cell lines or primary human ips derived macrophages results in abswnt gp91phox-p22phox expression and aboloishges the phagocyte respiratory burst.We also describe a case of CGD secondary to a homozygous CYBC1/EROSmutation that abolishes EROS protein expression. This work demonstrates the fundamental importance of CYBC1/EROSin human immunity and describes a novel, 6thcause of CGD.

However, EROS also regulates the expression of other proteins. Eros-/- macrophages also express very low levels of P2X7, a ligand gated ion channel that functions as a danger receptor by binding extracellular ATP released from damaged or dying cells. and driving activation of the NLRP3 inflammasome. We show that Eros co-immunoprecipiates with P2X7 and that P2X7 driven calcium flux and inflammasome activation are markedly abnormal in Eros-/- cells. Eros also affects T cell biology, underlining key roles beyond NADPH oxidase activation.

Conclusion: This work demonstrates the fundamental importance of CYBC1/EROSin human immunity and describes a novel, 6thcause of CGD as well as highlighting role of EROS that are independent of the geneartion of reactive oxygen species.

Disclosure of Interest

None Declared

O40 Cold-induced urticarial autoinflammatory syndrome related to factor XII activation

Jörg Scheffel1, Niklas Mahnke1, Zonne Hofman2, Steven de Maat2, Jim Wu1, Hanna Bonnekoh1, Reuben Pengelly3, Sarah Ennis3, John Holloway3, Martin Church1, Marcus Maurer1, Coen Maas2, Karoline Krause1
1Charite - Universitaetsmedizin Berlin, Berlin, Germany; 2University Medical Center Utrecht, Utrecht, Netherlands; 3University of Southampton, Southampton, United Kingdom
Correspondence: Karoline Krause

Introduction: Early onset cold-induced urticarial rash with systemic inflammatory symptoms are hallmarks of hereditary autoinflammatory diseases caused by gene mutations of the innate immune pathway, e.g. nucleotide receptor protein 3 (NLRP3). However, in many cases genetic tests are negative, suggesting the existence of unrecognized genetic variants.

Methods: We studied eight members of a four-generation family, four of whom were affected. Genetic analysis involved exome sequencing followed by targeted Sanger sequencing. Functional analyses included immunoblotting, mononuclear cell stimulation and immunohistochemistry. We generated recombinant protein variants and assessed cytokines and proteins in plasma and skin.

Results: Affected patients had cold-induced urticarial rash, arthralgia, chills, headache and malaise associated with an autosomal-dominant inheritance. Genetic studies identified a novel deleterious variant in gene F12 (T859A, resulting in p.W268R) which encodes coagulation factor XII (FXII). Occurrence of the mutation segregated with disease status. Immunoblotting for FXII exhibited a distinct 50kDa band that was also present in recombinant W268R-mutated proteins suggesting unusual fragmentation and spontaneous activation of FXII. Furthermore, we observed contact system activation with reduced plasma prekallikrein and profound cleavage of high molecular weight kininogen, representing bradykinin production. Skin and blood neutrophils were found to be a prominent source of FXII. Interleukin-1ß (IL-1ß) was upregulated in lesional skin and in mononuclear cells of healthy donors exposed to recombinant proteins. In accordance with these findings, treatment with icatibant (bradykinin-B2-antagonist) or anakinra (interleukin-1-antagonist) reduced disease activity in patients.

Conclusion: We identified a novel autoinflammatory syndrome characterized by a substitution in the F12 gene resulting in activation of the contact system and cytokine-mediated inflammation.

Disclosure of Interest

None Declared

Poster presentations – Monday 1 April

Guided poster tour 1A

PT1A01 Multi-omics analysis of ADA2 deficiency in Japanese cohort

Hiroshi Nihira1, Kazushi Izawa1, Takahiro Yasumi1, Moeko Ito2, Sachiko Iwaki-Egawa2, Yoji Sasahara3, Hirokazu Kanegane4, Tadateru Yasu5, Tomohiro Kubota6, Syuji Takei6, Dai Keino7, Etsuro Nanishi8, Hidetoshi Takada9, Shoichi Ohga8, Syunsuke Kajikawa10, Makio Takahashi11, Naoko Nakano12, Osamu Ohara13, Toshio Heike14, Junko Takita1, Ryuta Nishikomori1
1Pediatrics, Kyoto University, Kyoto; 2Life Sciences, Hokkaido University of Science, Sapporo; 3Pediatrics, Tohoku University, Sendai; 4Pediatrics, Tokyo Medical and Dental University, Tokyo; 5Pediatrics, Nagasaki Medical Center, Omura; 6Pediatrics, Kagoshima University, Kagoshima; 7Pediatrics, St. Marianna University, Kawasaki; 8Pediatrics, Kyusyu University, Fukuoka; 9Pediatrics, Tsukuba University, Tsukuba; 10Neurology, Kyoto University, Kyoto; 11Neurology, Osaka Red Cross Hospital, Osaka; 12Pediatrics, Ehime University, Toon; 13Applied Genomics, Kazusa DNA Research Institute, Kisarazu; 14Pediatrics, Hyogo Prefectural Amagasaki General Medical Center, Amagasaki, Japan
Correspondence: Hiroshi Nihira

Introduction: Adenosine deaminase type 2 deficiency (DADA2) is caused by recessive loss-of-function variants in ADA2. Most of DADA2 patients reveal systemic vasculopathy consistent with polyarteritis nodosa and large phenotypic variability has been reported [1, 2, 3]. However, pathogenesis of DADA2 remains unclear.

Objectives: The objective of this study is to reveal clinical and genetic characteristics of Japanese DADA2 patients, and to gain insight into the pathogenesis of DADA2 by multi-omics analysis.

Methods: We performed the genetic analysis of the ADA2 gene and measured ADA2 activity of the patients from 2016 to 2018 in Japan. Multi-omics analysis had been done in 4 out of 8 DADA2 patients and 4 healthy donors using their peripheral blood mononuclear cells (PBMCs). The samples were taken before and after introduction of anti-TNFα agents (meaning acute and remission phase) in the patients.

Results: We found 8 DADA2 patients. In this cohort, central neurological manifestations were present in 5 (63%), including asymptomatic small lacunar infarction. Seven subjects (88%) had livedo racemose, but there was no digital ulcer or necrosis. Low levels of IgG and IgM were revealed in 3 (37.5%) and 5 (62.5%) patients respectively, but there was no recurrent infectious episode in this case series. There were two (25.0%) who revealed pure red cell aplasia (PRCA); one revealed only anemia without any inflammation, the other revealed anemia transiently and recovered from it spontaneously but he revealed chronic inflammation afterwards. All 8 patients received anti-TNFα agent and all except one with CsA-dependent PRCA were well controlled.

We identified 6 previously described and 4 novel variants in ADA2, which included two that we reported before [3]. Overexpression of ADA2 variant constructs in HEK 293 cells showed that some variants had comparable protein expression levels to wild-type in cell lysate but most of them were not secreted and all the variants had low or absent ADA2 enzyme activities.

In multi-omics analysis, differentially expressed (DE) genes were analyzed at the mRNA and protein levels. We found 64 and 58 genes that were differentially expressed in acute phase vs control and remission phase vs control in common at the transcriptome and proteome levels respectively. Gene ontology analysis of these datasets revealed constitutive up-regulation of type 1 and type 2 interferon pathway. Some genes were common to both datasets.

Conclusion: We have found 8 DADA2 patients in Japan and identified some novel disease-causing variants. Using multi-omics analysis, we also have found differentially expressed genes in DADA2 patients. Some genes were consistently up-regulated even in the remission period. This may provide further insights into the pathogenesis of DADA2.


[1] Zhou Q., et al. N Engl J Med, 2014.

[2] Navon Elkan P., et al. N Engl J Med, 2014.

[3] Meyts I., Aksentijevich I. J Clin Immunol, 2018.

[4] Nihira H., et al. Scand J Rheumatol, 2017.

Disclosure of Interest

None Declared

PT1A02 The clinical and immunological profiles of haploinsufficiency of A20 in Japan

Hidenori Ohnishi1, Tomonori Kadowaki1, Norio Kawamoto1, Tomohiro Hori1, Kenichi Nishimura2, Chie Kobayashi3, Tomonari Shigemura4, Shohei Ogata5, Yuzaburo Inoue6, Tomoki Kawai7, Eitaro Hiejima7, Kazushi Izawa7, Tadashi Matsubayashi8, Kazuaki Matsumoto9, Masatoshi Takagi9, Kohsuke Imai9, Ryuta Nishikomori7, Shuichi Ito2, Toshio Heike7, Osamu Ohara10, Tomohiro Morio11, Hirokazu Kanegane12, Toshiyuki Fukao1
1Pediatrics, Gifu University Graduate School of Medicine, Gifu; 2Pediatrics, Yokohama City University, Kanagawa; 3Child Health, Faculty of Medicine, University of Tsukuba, Ibaraki; 4Pediatrics, Shinshu University School of Medicine, Matsumoto; 5Pediatrics, Kitasato University Hospital, Kanagawa; 6Allergy and Rheumatology, Chiba Children’s Hospital, Chiba; 7Pediatrics, Kyoto University Hospital, Kyoto; 8Pediatrics, Seirei Hamamatsu General Hospital, Shizuoka; 9Community Pediatrics, Perinatal and Maternal Medicine, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo; 10Applied Genomics, Kazusa DNA Research Institute, Chiba; 11Pediatrics and Developmental Biology; 12Child Health and Development, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan
Correspondence: Hidenori Ohnishi

Introduction: A20, encoded by the TNFAIP3gene, is a negative regulator of the tumor necrosis factor (TNF)-nuclear factor (NF)-κB signaling pathway. Recently, the haploinsufficiency of A20 (HA20) caused by heterozygous mutations in the TNFAIP3gene was identified to cause early onset autoinflammatory disease resembling Behçet’s disease.

Objectives: In this study, we performed a multicenter survey investigating HA20 patients found in Japan and characterized immunological profile.

Methods: We summarized the detailed clinical manifestations, genetic analyses and several immunological parameters of Japanese patients with HA20. Serum cytokine levels and the production levels of IL-1β and TNF-α from the peripheral blood mononuclear cells (PBMCs) were measured using ELISA. Multicolor flowcytometry analysis was performed. To detect A20 protein expression, immunoblot analysis was performed for PHA blast derived from the patients and A20 transfected HEK293T cells. The NF-κB reporter gene activity was analyzed using the dual-luciferase reporter assay system.

Results: A total 32 patients from 10 independent families were enrolled in this study. Age of onset was 0 to 20 years. Three mutations in the TNFAIP3gene were previously reported; however, seven were novel. All these mutations were evaluated to be functionally pathogenic by several in vitroassays. The production levels of proinflammatory cytokines such as TNF-α and IL-1β from PBMCs were increased. In the detailed analysis of lymphocyte subsets including T, B and NK cells, regulatory T cells (Treg) were increased in all analyzed patients. The increase of double-negative T (DNT) and T helper 17 cell (Th17) cells were observed in 7 and 3 out of 18 analyzed patients, respectively. In addition, follicular helper T cells (Tfh) were significantly increased especially in younger patients. Memory B cells were decreased in most of analyzed patients. Intriguingly, in the complications of HA20 patients, 9 out of 32 of them had not only autoinflammatory phenotypes but also several autoimmune disorders including psoriatic arthritis, Hashimoto’s thyroiditisand autoimmune lymphoproliferative syndromewere observed. The immune dysregulation derived from the defect of A20 may cause the increase of the autoimmune related T cell subsets and the secondary Treg expansion as well as the phonotype of the previously reported A20 knockout mice.

Conclusion: Our study revealed unexpected variation in phenotypes of HA20 including autoimmunity. In the analysis of lymphocyte subsets for HA20 patients, the characteristic findings such as the increase of Treg and the decrease of memory B cells were observed. The increase of DNT, Th17 and Tfh cells may be involved in onset of several autoimmune disorders.

Disclosure of Interest

None Declared

PT1A03 Use of SIGLEC1/CD169 as a biomarker for monogenic interferonopathies

Banu Orak1,2, Axel Panzer3, Manuela Theophil3, Elke Krüger4, Frédéric Ebstein4, Barbara Zieba4, Nadine Unterwalder5, Christian Meisel5, Tilmann Kallinich2
1Center for chronically sick children, Charité University Medicine Berlin; 2Department of Pediatrics, Division of Pneumology, Immunology with intensive Medicine, Charité University Medicine Berlin; 3Pediatric Neurology, DRK Klinikum Berlin-Westend, Berlin; 4Institute of Medical Biochemistry and Molecular Biology, University Medicine Greifswald, Greifswald; 5Department of Immunology, Labor Berlin GmbH, Berlin, Germany
Correspondence: Banu Orak

Introduction: Monogenic Interferonopathies represent a rare group of inflammatory diseases with difficulties in early diagnosis. Expression of SIGLEC1, also known as CD169, on monocytes is the second highest interferon stimulated gene (ISG) in systemic lupus erythematodes (SLE).A correlation of SIGLEC1 expression with ISG in SLE is well established. Furthermore, SIGLEC1 seems to estimate disease activity more accurately than anti-dsDNA antibodies.

Objectives: To show the relevance of SIGLEC1 as a diagnostic marker for detection of Interferonopathies.

Methods: Eight patients with genetically confirmed monogenic Interferonopathies were included. Clinical data, classical inflammatory markers and blood count were obtained by patients file. SIGLEC1 expression was measured by flow cytometry with a highly standardized quantitative assaywith a reference range in healthy controls less than 2500 SIGLEC1 molecules/monocyte.In order to quantify the antigen expression by every single cell QuantiBRITE™ PE tubes were applied. Additionally, transcriptional level of SIGLEC1, IFI44L, IFI27, ISG15 and RSAD2 as type I Interferon stimulated genes were assessed by real-time PCR.

Results: All patients showed homozygous mutations. Three patients displayed TREX-1, two patients IFIH-1, two patients SAMDH1 and one patient RNASE2HB mutations. Mean age of patients was 12 years (min. 6 months, max. 49 years, SD+/- 17 years). Six of eight patients showed neurological symptoms consistent with Aicardi-Goutières-Syndrome like neurological development retardation and microcephaly. Five patients showed abnormalities on brain MRI, like periventricular calcifications or corpus callosum thinning. Two patients (homozygous for IFIH-1 mutation) were diagnosed with Singleton-Merten-Syndrome presenting abnormal ossification of extremities and dental anomalies.One patient with homozygous TREX1 mutation presented with postnatal glaucoma, microcephaly, developed sensorimotor polyneuropathia and suffered from recurrent fever with persistent chilblain lesions.

All eight patients (100%) showed elevated results for SIGLEC1 expression (mean molcules/monocyte +/- SD: 10272 +/- 3746) without having high levels of standard inflammatory markers. In six patients elevated SIGLEC1 expression showed dysregulation of the type 1 interferon pathway prior to genetic testing. In three patients with unclear disease phenotype measuring expression of SIGLEC1 contributed to establish the right diagnosis. On transcriptional level SIGLEC1 and the other ISGs were also elevated in comparison to healthy controls.

Conclusion: In all patients with monogenic Interferonopathies like Aicardi-Goutières-Syndrome high expression of SIGLEC1 was observed, either before diagnosis was established or during disease course.

Therefore, SIGLEC1 qualifies as an easy accessible and cheap diagnostic marker with short turnaround time to screen patients with suspected Interferonopathy.

Disclosure of Interest

None Declared

PT1A04 Screening of patients with idiopathic polyarteritis nodosa, granulomatosis with polyangiitis, and microscopic polyangiitis for deficiency of adenosine deaminase 2

Oskar Schnappauf1, Monique Stoffels2, Ivona Aksentijevich1, Amanda Ombrello1, Natalia Sampaio Moura1, Karyl Barron1, Daniel Kastner1, Peter Grayson3, Peter Merkel4, on behalf of Vasculitis Clinical Research Consortium
1National Human Genome Research Institute (NHGRI), National Institutes of Health (NIH); 2National Human Genome Research Institute (NHGRI), National Institutes of Health; 3National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS), National Institutes of Health (NIH), Bethesda; 4Division of Rheumatology and the Department of Biostatistics, Epidemiology, and Informatics (DBEI), University of Pennsylvania, Philadelphia, United States
Correspondence: Oskar Schnappauf

Introduction: Deficiency of adenosine deaminase 2 (DADA2) is the first described monogenic vasculitis. Patients usually present in childhood, but age of onset, disease severity, and organ involvement of DADA2-associated vasculitis is highly variable. Clinical manifestations of DADA2 overlap with the typical features of necrotizing vasculopathies such as polyarteritis nodosa (PAN), granulomatosis with polyangiitis (GPA), and microscopic polyangiitis (MPA).

Objectives: This study aimed to test the prevalence of DADA2 in patients with presumed idiopathic PAN, GPA or MPA.

Methods: Patients (n=117) with idiopathic PAN, all of whom tested negative for hepatitis B virus infection, and patients (n=1107) with GPA or MPA were screened for mutations in ADA2. To further assess the pathogenicity of identified variants on a functional level, ADA2 activity was determined on available serum samples of patients with PAN.

Results: Nine of 118 patients with PAN (7.6%) were identified as having rare missense variants in ADA2 with a minor allele frequency of < 0.005. Four patients (3.4%) were homozygous or compound heterozygous for variants in ADA2. Of the seven distinct variants present in these four patients, G47A, G47W, R169Q, E328K, F355L, and G383S had previously been reported as causative for DADA2. The remaining variant, P106S, is a rare variant predicted to be damaging to protein function by in silico algorithms. Five additional patients were carriers for the monoallelic variants R34W, T65M, M309I, V349I, and Y453C. R34W and Y453C were reported in DADA2 before, while the three remaining variants are of unknown clinical significance. None of the patients with GPA or MPA were biallelic for rare missense variants in the ADA2 gene but 32 individuals (2.9%) were carriers for monoallelic rare missense variants.

Serum samples of patients with PAN were available on the individuals with the G383S/G383S and E328K/F355L genotypes and showed markedly reduced ADA2 enzyme activity, comparable to levels seen in patients with DADA2. ADA2 activity of three of the four available serum samples on monoallelic carriers was not reduced, confirming the non-pathogenicity of T65M, M309I, and V349I. The serum sample on the individual carrying the pathogenic variant Y453C showed ADA2 activity in the range of carriers. ADA2 enzyme activity testing of the remaining serum samples revealed one additional individual with strongly reduced ADA2 activity levels as well as five individuals with enzymatic activity in the range of carriers. Sanger sequencing of the ADA2 gene in these individuals did not identify any pathogenic variants and indicates the presence of cryptic mutations undetectable by conventional sequencing. In summary, for five out of 118 patients with PAN the diagnosis of DADA2 can be applied.

Conclusion: This is the first study to report biallelic pathogenic variants in ADA2 in patients with adult-onset, idiopathic PAN, and demonstrates that DADA2 specifically accounts for a subset of patients with idiopathic PAN but not GPA or MPA. Given the potential efficacy of TNF-inhibitors in DADA2, that anti-TNF treatment is not the conventional therapy in PAN, and the consequences for other family members, these findings suggest that ADA2 testing and/or ADA2 activity testing should be considered in patients with HBV-negative idiopathic PAN, especially in patients with an early onset of this potentially life-threatening disease.

Disclosure of Interest

None Declared

PT1A05 Diagnosis and long term management of type I interferonopathies in a pediatric rheumatology center

Stefano Volpi1,2, Elettra Santori3, Margherita Ricci1, Paolo Picco1, Alessandra Tesser4, Gillian I. Rice5, Roberta Caorsi1, Alice Grossi6, Isabella Ceccherini6, Alberto Martini2, Yanick J. Crow7,8, Alberto Magnasco9, Alberto Tommasini4, Fabio Candotti3, Marco Gattorno1
1Centro per le Malattie Infiammatorie e Immunodeficienze, Clinica Pediatrica e Reumatologia, Istituto Giannina Gaslini; 2DINOGMI, Università degli Studi di Genova, Genova, Italy; 3Allergy and Immunology, Lausanne University Hospital, Lausanne, Switzerland; 4IRCCS Buro Garofalo, Trieste, Italy; 5Genetic Medicine, Manchester Academic Health Science Centre, University of Manchester, Manchester, United Kingdom; 6UOC Genetica Medica e UOSD Genetica e Genomica delle Malattie Rare, Istituto Giannina Gaslini, Genova, Italy; 7Centre for Genomic and Experimental Medicine, MRC Institute of Genetics and Molecular Medicine, University of Edinburgh, Edinburgh, United Kingdom; 8Institute Imagine, University Paris Decartes, Paris, France; 9Nephrology, Dialysis, Transplantation Unit, Istituto Giannina Gaslini, Genova, Italy
Correspondence: Stefano Volpi

Introduction: in recent years several genetic diseases linked to pathologic activation of type 1 interferon (IFN) pathway have been described

Objectives: To identify type I interferonopathies in a cohort of children with early-onset rheumatic disease and to report the long-term efficacy and side effect of pathway-specific treatment with a Janus Kinase (JAK) inhibitor

Methods: Patients were selected based on the presence of any of the following: i) early onset (infancy or before puberty) inflammatory SLE-like symptoms; ii) vasculopathy (skin ulcers, chilblains, strokes) iii) panniculitis with or without lipodystrophy iv) persistent or recurrent systemic inflammation with or without lung involvement v) polyarthritis with lung involvement. Type 1 IFN activation was assessed by quantitative PCR, measuring the expression of 6 type 1 interferon-related genes (IFI27, IFI44L, IFIT1, ISG15, RSAD2, SIGLEC1) in peripheral blood. In selected patients, molecular analysis was performed using Sanger sequencing, a NGS panel of 41 inflammatory-related genes or whole exome sequencing. Off label therapy with the JAK inhibitor Ruxolitinib was considered in patients with a definitive genetic diagnosis and incomplete disease control by standard treatment

Results: We screened 324 patients evaluated in the rheumatology unit of our Institute. 132 out of 324 patients had a positive type 1 IFN signature. Based on the clinical presentation and the result of the IFN signature we further analyzed a subset of patients for an underline genetic defect. In 6 patients we identified pathogenic mutations affecting TMEM173 (p.V155M in one patient and p.R281Q in a second patient), DNASE2 (p.D121V), DNASE1L3 (c.289_290delAC in 2 patients) and COPA (p.R233H) genes were identified. Therapy with Ruxolitinib was started in 4 patients and allowed to partially control disease manifestations and reduce or stop steroids, however, we observed the following limitation: the patient with DNASE1L3 mutations and a severe kidney involvement at therapy onset, despite the control of his systemic symptoms progressed to kidney failure; one patient with SAVI and a severe lung involvement experienced severe recurrent viral lung infections requiring intensive care and extracorporeal membrane oxygenation (ECMO) in one episode; the patient with DNASE2 mutationsexperienced a Herpes Zoster infection controlled with antiviral therapy and decreasing Ruxolitinib dosage.

Conclusion: Patients with type 1 interferonopathies might present with symptoms overlapping pediatric rheumatic diseases. The combination of type 1 interferon pathway activation assessment and NGS is an effective strategy for the diagnosis of this heterogeneous class of diseases and can guide the choice for targeted therapies. JAK inhibitors represent a therapeutic resource in controlling these difficult-to-treat patients, however further studies are needed to assess their efficacy and long-term safety

Disclosure of Interest

None Declared

PT1A06 Results of international Delphi survey for the diagnosis, investigation and management of deficiency of ADA2

Taryn A.-B. Youngstein1, Eugene P. Chambers2, on behalf of DADA2 Foundation, Helen J. Lachmann1, DADA2 Delphi Study Participants
1National Amyloidosis Centre, UCL Division of Medicine, London, United Kingdom; 2Vanderbilt University Medical Centre, Vanderbilt University, Nashville, United States
Correspondence: Taryn A.-B. Youngstein

Introduction: DADA2 is phenotypically heterogenous in its presentation, even within families with the same mutation, and the absence of a murine ortholog has placed emphasis on the study of known cases of the disease.

Objectives: We conducted an E-Delphi study to explore consensus on the diagnosis, investigations and management of those with suspected disease.

Methods: A Delphi method over two rounds of consensus building was used. Invitation email to participate in the study was sent to all published authors on DADA2 and physicians and scientists known to the DADA2 Foundation, a patient advocacy group supporting research into DADA2.Consensus was defined as > 80% agreement.

Results: 69 respondents contributed to Round 1 and 53 respondents in Round 2, 75% of respondents were paediatricians, the rest adult physicians, a nurse specialist and basic scientists.

Consensus was reached in 1. Suggestive diagnostic features; Livedoid Rash, Stroke, Fever, Digital Gangrene, 2. The combined use of ADA2 gene sequencing and ADA2 enzyme activity levels to confirm diagnosis. 3. Use of anti-TNF therapy in acute and chronic presentations, 4. Indefinite use of anti-TNF until more information is available, and 5. The avoidance of anticoagulation.

A wide variety in practice was identified in the use of baseline and follow up investigations including imaging.

Conclusion: There is consensus that rash, stroke, fever, digital gangrene are highly suggestive of DADA2 but there may be a separate haematological phenotype.Gene sequencing and ADA2 activity levels in combination are the diagnostic gold standard but not generally available.There is not yet consensus on the baseline screening investigations, such as neuroimaging, for these cases. Follow-up investigations are currently guided by clinical course and practice remains highly variable.

92% of treating physicians believe that anti-TNF therapy is the correct treatment approach currently, and its use should be indefinite until more information is available.

The authors recommend the creation of a detailed international case registry and a standardised series of baseline and follow-up investigations during this accelerated learning phase about this recently described disease.

Disclosure of Interest

None Declared

PT1A07 Somatic mutations in the NLRP3-inflammasome gene in late adulthood-onset chronic urticaria

Eman Assrawi1, Camille Louvrier1, Fawwaz Awad1, Clemence Lepelletier2, JD Bouaziz2, William Piterboth1, Florence Moinet3, Philippe Moguelet4, Claire Jumeau1, Laetitia Cobret1, Elma El-Khouri1, Philippe Duquesnoy1, Marie Legendre1, Sophie Georgin-Lavialle5, Gilles Grateau5, Sonia Athina Karabina1, Serge Amselem1, Irina Giurgea1
1Sorbonne Université, inserm UMRS 993; 2Hôpital Saint-Louis , Service de Dermatologie, Paris; 3Centre Hospitalier Universitaire de Martinique, Service de médecine interne, Martinique; 4Hôpital Tenon , Anatomie et cytologie pathologiques; 5Hôpital Tenon, Service de médecine interne, Paris, France
Correspondence: Eman Assrawi

Introduction: Chronic urticaria is a common dermatological disorder and one of the most prominent symptoms of Cryopyrin-Associated Periodic Syndrome (CAPS), a systemic autoinflammatory condition. Besides urticaria, CAPS cardinal symptoms are fever, arthralgia and deafness; however, absence of pathognomonic symptoms makes this diagnosis challenging. NLRP3, the disease-causing gene, encodes the cryopyrin, which upon activation initiates NLRP3 inflammasome assembly and proinflammatory cytokine secretion. Familial cases of CAPS are due to heterozygous germ-line NLRP3 mutations; however, sporadic cases, more often identified in children, are related to de novo or somatic NLRP3 mutations.

Objectives: We aimed to establish the etiological diagnosis of two elderly unrelated patients presenting with idiopathic chronic urticaria for two decades.

Methods: Molecular study of patients’ DNA was performed using a NGS panel targeting genes involved in autoinflammatory disorders. Functional analyses of the identified NLRP3 variants were performed using two cellular models, HEK293T cells (stably expressing ASC-GFP and pro-caspase1-FLAG) to study ASC speck formation, and THP1 cells to assess IL1β secretion.


In two sporadic unrelated patients, we identified two mosaic NLRP3 mutations: a novel in-frame deletion (c.926_934del, p.Gly309_Phe311del) and a recurrent CAPS mutation (c.1705G>A, p.Glu569Lys). In whole blood DNA, the mosaicism level was of 17.2% in the first patient and of 11% in the second one. The patients, who are about 70 years old, presented for two decades with late onset chronic idiopathic urticaria, occasionally associated with fever, arthralgia and myalgia. Deafness was diagnosed at the age of 50 years in the first patient, and of 70 years, after the identification of a NLRP3 mutation, in the second one.

To assess the pathogenicity of the identified variants, we studied the activation of NLRP3 inflammasome after transient expression of NLRP3 wild-type (WT) or of NLRP3 carrying either the p.Gly309_Phe311del or the p.Glu569Lys mutation. Both mutations were found to significantly increase ASC speck formation andIL1β secretion as compared to NLRP3-WT.

The diagnosis of CAPS was therefore established on the bases of these molecular and functional data. Accordingly, complete remission was achieved with anti-interleukin 1 receptor antagonists in both patients.

Finally, we studied the mosaicism level in several cell types from both patients and showed a wide distribution profile of the mutant alleles, suggesting that, in both cases, the mutational event occurred early during embryogenesis.

Conclusion: In late adulthood-onset chronic urticaria, the search for autoinflammatory markers and for somatic NLRP3 mutations may have important diagnostic and therapeutic issues. Importantly, despite the onset of the disease after 50 years old, NLRP3 mutations are not restricted to myelomonocytic cells.

Disclosure of Interest

None Declared

PT1A08 Idiopathic recurrent pericarditis: clinical findings and treatment approach

Camilla Celani1, Silvia Federici1, Anna Tulone1, Brigitte Bader Meunier2, Virginia Messia1, Manuela Pardeo1, Claudia Bracaglia1, Pierre Quartier Dit Maire2, Fabrizio De Benedetti1, Antonella Insalaco1
1Division of Rheumatology, IRCCS, Ospedale Pediatrico Bambino Gesù, Rome, Italy; 2Unité d’Immunologie-Hématologie et Rhumatologie pédiatrique, Hôpital Necker-Enfants, Paris, France
Correspondence: Camilla Celani

Introduction: Recurrent pericarditis affects 15-30% of patients with acute pericarditis. The etiology is poorly understood, with about 80% being idiopathic. Several treatment options are available for recurrences, including NSAIDs, colchicine, glucocorticoides and IL-1 inhibitors (i.e. Anakinra). Standardized guidelines for the management of these patients are still lacking

Objectives: To analyze clinical findings and treatment approach in a cohort of pediatric patients with recurrent pericarditis

Methods: Patients with at least two episodes of idiopathic pericarditis, followed at two Pediatric Rheumatology centers between 2006 and 2018, were included

Results: A total of 42 patients (18 males ) were included. Mean age at disease onset was 11.8 years (range 4-17). Chest pain and fever were the presenting symptoms in all patients. In 47% pleural effusion was detected. Laboratory tests showed increased white blood cell count (mean 14.509/mm3), C-reactive protein (mean 18.01 mg/dl) and erythrocyte sedimentation rate (mean 39 mm/h) in all patients. The first episode was variably treated: 18/42 (43%) received NSAIDs alone, 5/42 (11.9%), colchicine alone or associated to NSAIDs and 3/42 patients (7%) received antibiotics alone. 16/42 (38%), not responsive to NSAIDs or colchicine, received glucocorticoides. Patients who received glucocorticoids at the first episode relapsed earlier (median time of 2.1 months range 10 days-5 months), than patients treated with NSAIDs ( 6.6 months range 10 days -24 months) or with colchicine (5 months range 10 days-5 months) (p<0.05). In our study, initial treatment of the first episode did not affect the number of subsequent flares. To evaluate treatment strategy at relapses, we divided our study population in two groups: Group 1 (20 pts) in which recurrence was treated with NSAIDs, colchicine or glucocorticoid (alone or combined); group 2 (22 patients) in which anakinra was started. Among patients belonging group 2, 9 received anakinra at first relapse, 7 at the second, 2 at the third and 2 at the fourth. Anakinra treatment was followed by a prompt resolution of symptoms and inflammatory signs within 2 days. During daily treatment with full dose anakinra, no relapses were reported over a median of 13.3 months (range 5-24 months). In 13 out of 22 patients, anakinra was gradually tapered reducing the days of administration during the week. Four of these patients relapsed. The mean time from the start of anakinra to tapering was 17±4 months (range 14-23 months) in the 4 patients who experienced a relapse versus 14±4 months (range 7-21 months) in patients who did not flare, with no statistical difference. Among the 22 patients belonging to group 2 anakinra was finally discontinued in 11 after a mean time of 23.4 months (range 12-36). Among these, 8 relapsed after anakinra withdrawal (including 2 of the 4 patients already relapsed during tapering). Only 3 patients didn’t present any relapse (up to 20.3 months of follow-up). All patients who relapsed responded quickly to the reintroduction of anakinra

Conclusion: Our study confirms the lack of a standardized treatment approach in patients with recurrent pericarditis. Patients treated with glucocorticoid at first episode relapse before than those treated with other drugs. Anakinra is an effective treatment; however, tapering/discontinuation of the drug lead to relapses in several cases. Further experience on larger population is needed to define the best treatment duration and approach to withdrawal of IL-1 inhibitor

Disclosure of Interest

None Declared

Guided poster tour 1B

PT1B01 Monocytes proteomic profile of patients with different autoinflammatory diseases: a new approach to characterize these diseases

Federica Penco1, Andrea Petretto2, Chiara Lavarello2, Ilaria Gueli3, Arinna Bertoni1, Alessia Omenetti3, Claudia Pastorino1, Marco Gattorno1
1Centro Malattie Autoinfiammatorie ed Immunodeficienze; 2Laboratorio Core Facilities - Proteomica e Metabolomica Clinica; 3Clinica Pediatrica e Reumatologica, Istituto Giannina Gaslini, Genova, Italy
Correspondence: Federica Penco

Introduction: Autoinflammatory diseases are a group of inherited diseases characterized by early onset and systemic inflammation, often manifesting with unexplained fevers.These pathologies are usually caused by mutations in genes involved in the regulation of innate immune response with a consequent inflammatory phenotype. The most common genetically defined periodic fevers are Familial Mediterranean Fever (FMF), Cryopyrin-associated periodic syndromes (CAPS), TNF receptor-associated periodic syndrome (TRAPS) and mevalonate kinase deficiency/hyperimmunoglobulin D syndrome (MKD/HIDS). Some patients show clinical features similar to autoinflammatory diseases but no genetic mutation has been found.

Objectives: Our aim is to evaluate the differences in the expression of proteins or pathway in monocytes, and plasma metabolites in patients with autoinflammatory diseases compared with healthy subjects to clusterize and better understand the mechanisms underlying different genetically defined disorders and try to characterize the genetically undefined pathologies.

Methods: Monocytes, purified from peripheral blood and incubated for 4 hours with or without LPS, were collected from 5 patients for each pathology (FMF, CAPS, TRAPS and MKD) and healthy donors. The samples have been processed by iST protocol. Each digested sample was analyzed by high-resolution liquid chromatography and tandem mass spectrometry (LC-MS/MS) based on Orbitrap technology. The quantification strategy is a label-free approach (LFQ) available in MaxQuant suite.

Results: Here we identified a median of about 5000 proteins from the monocyte samples of each 4000 is quantified by LFQ approach. PCA analysis and Person’s correlation show good reproducibility of data and a good separation between the different groups. The data were then submitted to an appropriate statistic. The T-Tests highlighted the differentially expressed proteins and through the use of Cytoscape with the ClueGo app we obtained the differently regulated pathways in the different conditions. It has also been constructed, starting from significative proteins, a network, related to disease using the information of String Disease db. This was done to highlight the proteins associated with the disease as well as to reveal new possible biomarkers. We observed that the expression of proteins is differently enriched according to the different conditions. For each autoinflammatory disease, a list of significantly modulated proteins was obtained: some of which are already known to be related to the disorders, while others have not yet been described. In FMF, MEFV, RhoA and some related proteins were significantly up-regulated together with genes linked to the interferon pathway. In TRAPS relevant proteins turn up related to the maintenance of Golgi and cellular trafficking. The bioinformatics analysis allows us to better understand the functional interaction between these monocytes proteins and map which are involved in the diseases. Proteins thus analyzed were then contextualized in dominant pathways for each pathology through Cytoescape network analysis.

Conclusion: Here, we addressed how a high-resolution proteomics approach could be used to better understand the biology of autoinflammatory diseases. The characterization of a broad spectrum of proteins and their interaction network will allow us to identify new biomarkers for the different pathologies and better comprehend and recognize the genetically undefined disorders.

Disclosure of Interest

None Declared

PT1B02 Prulipotent stem cell derived myeloid cell lines for dissecting the mechanism of autoinflammation

Megumu Saito
Center for iPS Cell Research and Application, Kyoto University, Kyoto, Japan

Introduction: Autoinflammatory disorders (AIDs) is defined as an disorders associated with dysregulation of innate immune systems. In typical case, patients with AIDs show various inflammatory symptoms, such as periodic fever, skin rash and sterile serositis. Human induced pluripotent stem cell (iPSC) models of AIDs are supposed to be feasible and useful, because 1) most of typical AIDs are monogenic, 2) responsible cells are usually innate immune cells which can be robustly differentiated from PSCs, and 3) improvement of therapeutic or diagnostic approach is still needed in many AIDs. We therefore focused on establishing PSC models of AIDs, and applied these models to identification of genetic and biological background of patients’ pathophysiology.

Results: We established iPSCs from patients with AIDs such as CINCA/NOMID, Nakajo-Nishimura syndrome (NNS), and Blau syndrome (BS). We also corrected the disease-causing mutations by genome editing technology in selected clones. iPSCs were then differentiated into hematopoietic cells, especially into monocytic lineage cells. To stably obtain functional monocytic cells, monocytic progenitor cells derived from iPSCs were immortalized. The immortalized monocytic cell lines (MLs) were further differentiated into mature macrophages and then used for functional assays. The monocytic cells from AID-iPSCs showed increased secretion of proinflammatory cytokines such as IL-6, TNF and IL-1β. In case of CINCA/NOMID, iPSCs established from a NLRP3-mutation negative patient were useful for the identification of somatic NLRC4 mutation of the patient. The AID-iPSCs were also used for dissecting underlying disease mechanism, and suitable for high-throughput phenotypic screening.

Conclusion: We applied iPSC technology for studying autonflammatory disorders. When combined with other novel technologies such as next generation sequencing and genome editing, iPSC-based phenotypic dissection was useful for diagnosis of the patients and understanding the disease mechanism.

Disclosure of Interest

None Declared

PT1B03 Loss of protein prenylation in human monocytes promotes the formation of an NLRP3-dependent inflammasome in a model of mevalonate kinase deficiency

Oliver Skinner1, Julie Jurczyluk1,Paul Baker2, Seth Masters2, Avril Robertson3, Kate Schroder4, Sam Mehr5, Marcia Munoz1, Michael Rogers1
1Bone Biology Division, Garvan Institute of Medical Research, Darlinghurst, Sydney; 2Inflammation Division, Walter and Eliza Hall Institute of Medical Research, Melbourne; 3School of Chemistry and Molecular Biosciences; 4Institute of Molecular Bioscience, University of Queensland, Brisbane; 5Dept of Allergy/Immunology, Royal Children's Hospital, Melbourne, Australia
Correspondence: Oliver Skinner

Introduction: Mevalonate kinase deficiency (MKD) is an autoinflammatory disease caused by mutations in an enzyme of the mevalonate pathway, leading to loss of isoprenoid lipids necessary for protein prenylation. Defective prenylation in MKD is thought to trigger inflammasome activation, IL-1β release and recurrent episodes of systemic inflammation. However, how loss of prenylation causes inflammasome activation remains controversial. Recent studies have suggested that lack of Rho or K-Ras prenylation leads to assembly of the Pyrin inflammasome in macrophages, whereas others have shown that disruption of the mevalonate pathway in monocytes triggers IL-1β release via NLRP3 inflammasome activation.

Objectives: To determine which type of inflammasome is activated in human monocytes upon loss of protein prenylation in MKD.

Methods: THP-1 human monocytic cells were treated for 24 hours with 5μM simvastatin (SIM) to pharmacologically block the mevalonate pathway, followed by stimulation with LPS or Pam3CSK4 (TLR4 and TLR2 agonists, respectively) to induce an inflammatory response. IL-1β and IL-18 release was quantified by ELISA whilst caspase-1 enzyme activity, ASC immunostaining and permeability to propidium iodide were used as measures of inflammasome formation and pyroptosis. To assess the contribution of Pyrin or NLRP3, we used inducible CRISPR/Cas9 knockout THP-1 cells, as well as the small molecule inhibitor of NLRP3, MCC950. Finally, inflammasome activation was examined in peripheral blood mononuclear cells (PBMCs) from an MKD patient (bearing V377I/H20N mutations in MVK) and heterozygous parents.

Results: SIM treatment of THP-1 cells caused a clear accumulation of unprenylated Rab and Rap1A proteins, without affecting cell viability. Furthermore, LPS or Pam3CSK4 stimulation of SIM-treated cells resulted in a 4-fold increase in IL-1β and IL-18 release and significantly higher ASC-containing speck formation, caspase-1 activity and pyroptosis. Importantly, all these effects were reversed to normal levels by restoring protein prenylation using geranylgeraniol (the missing lipid metabolite necessary for protein prenylation). In support of a predominant role for NLRP3, MCC950 completely inhibited the stimulatory effect of SIM on LPS-induced ASC speck formation, caspase-1 activation, IL-1β release and pyroptosis. All of these were also abolished by knocking out NLRP3, whereas deletion of Pyrin had no effect. Similarly, LPS stimulation of MKD patient-derived PBMCs resulted in much higher IL-1β release compared to heterozygous parents, which was completely blocked by NLRP3 inhibition with MCC950.

Conclusion: We clearly demonstrate that lack of prenylation in human monocytic cells, using statin treatment (a pharmacologic model of MKD) or using authentic cells from an MKD patient, leads to enhanced formation of an NLRP3-dependent, Pyrin-independent, inflammasome upon TLR2/4 stimulation. In contrast to reports that lack of prenylation activates the Pyrin inflammasome in macrophages, these findings indicate a prominent additional role for NLRP3 in the pathogenesis of MKD and demonstrate that targeting the Pyrin inflammasome in isolation may not be sufficient to resolve all the pathology associated with MKD. Rather, approaches to overcome the metabolic defect in the mevalonate pathway and restore normal protein prenylation could be more effective at preventing broader inflammasome activation.

Disclosure of Interest

None Declared

PT1B04 Performance of targeted NGS for routine diagnosis of autoinflammatory diseases

Guilaine Boursier1, Cécile Rittore1, Déborah Méchin2, Muriel Gutierrez1, Florian Milhavet1, Guillaume Sarrabay2, Isabelle Touitou2
1Department of Medical Genetics, Rare diseases and personalized medicine, Rare and autoinflammatory diseases unit, CHU Montpellier, Univ Montpellier; 2IRMB, INSERM, Univ Montpellier, Department of Medical Genetics, Rare diseases and personalized medicine, Rare and autoinflammatory diseases unit, CHU Montpellier, Montpellier, France
Correspondence: Guilaine Boursier

Introduction: Monogenic systemic autoinflammatory diseases (SAIDs) are characterized by mutations in genes coding for proteins involved in innate immunity. Since the discovery of the first gene MEFV (OMIM 608107) responsible for familial Mediterranean fever, more than 30 new conditions have been identified, notably through high-throughput sequencing approaches. Currently, next generation sequencing (NGS) allows the simultaneous investigation of multiple genes at a manageable cost. We present here our 4 years of experience of targeted NGS as a routine diagnostic for SAIDs.

Objectives: To evaluate the performance of a panel of 49 genes targeting well-defined autoinflammatory diseases and clinical concordance after retro-phenotyping.

Methods: DNAs from 577 patients clinically suspected for SAIDs (age 3 months to 79 years) were sequenced by NGS between September 2014 and December 2018. The libraries were prepared using Nextera (Illumina) or SureSelect (Agilent) Target Enrichment Capture custom kits. Sequencing reactions were performed on MiSeq or NextSeq500 equipment (Illumina). The mutations were classified according to a 5-class scale provided by the Infevers database or according to the American College of Medical Genetics and Genomics (ACMG) guidelines. Epidemiological data, clinical symptoms and biological markers were collected on a form provided with all genetic diagnosis requests.

Results: Almost a half of patients (261/577) had at least one mutation. Mutations that were probably pathogenic (class 4) or clearly pathogenic (class 5) accounted for one third of the mutations and were detected in only 30/49 genes. We identified 87% (100/115) missense variants and 11% (13/115) truncating mutations including whole gene deletions. The genes in which we identified most of the pathogenic mutations were MEFV, MVK, PSMB8, ADA2, NLRP3 and RNASEH2B then NOD2, PSTPIP1 and SLC29A3. The most recurrent pathogenic variants were MVK:p.(Val377Ile), MEFV:p.(Met694Val) and RNASEH2B:p.(Ala177Thr). The yield of conclusive genetic diagnosis (one class 4 or 5 mutation including mosaicism in a dominant condition or two non-allelic mutations in a recessive condition) using this NGS strategy was 8% over this 4-year period. We observed a rather good, though incomplete clinical concordance in patients with a genetic diagnosis.

Conclusion: The simultaneous investigation of multiple genes using targeted NGS is a successful routine diagnostic for SAIDs and is of particular interest for the detection of low-level mosaic mutations and copy number variations. However, our targeted NGS approach has resulted in genetic confirmation in a relatively small proportion of patients. One of the reasons may be related to the stricter definition we have used compared to previous reports. On the other hand, SAID genes or molecular mechanisms that are still unknown are likely to be found. Finally, a better clinical filter for ordering genetic tests through a consultation with experts could be encouraged.

Disclosure of Interest

None Declared

PT1B05 Late-onset TRAPS with low-grade mosaicism in TNFRSF1A

Barend P. Kant1, Marco J. Koudijs1, Ruben van‘t Slot2, Joyce van Kuik3, Lisanne M. Sikkema1, Joost Frenkel4, Anna Simon5, Mariëlle E. van Gijn1
1Department of Genetics; 2Center for Molecular Medicine; 3Department of Pathology; 4Department of Pediatrics, University Medical Center Utrecht, Utrecht; 5Department of Internal Medicine, Radboudumc Expertisecenter for Immunodeficiency and Autoinflammation, Radboud University Medical Center, Nijmegen, Netherlands
Correspondence: Barend P. Kant

Introduction: The diagnosis of patients with systemic autoinflammatory diseases (SAID) is difficult which can result in delayed treatment and irreversible organ damage. In several patients, low grade mosaicism of an autosomal dominant form of SAID has been detected. In NLRP3, even mosaic mutations with allele frequency <5% in whole blood can result in severe disease. In recent years, high-grade mosaic mutations have been detected in the TNFRSF1A gene in patients with late-onset TRAPS. With a new screening assay, we investigate whether low-grade mosaic mutations in TNFRSF1A might contribute to autoinflammatory disease.

Objectives: To examine the presence of low-grade mosaicism in a patient with clinically suspected late-onset TRAPS.

Methods: DNA was extracted from whole blood, FACS sorted hematopoietic cells and non hematopoietic cells. Mosaic mutation screening was performed using a single molecule molecular inversion probe (smMIP) assay. Positive results were confirmed with droplet digital (dd)PCR technology.

Results: Our patient is a 70 year old male with a history of episodes of fever with migratory erythematous rash and myalgia since his 30s. Also pleuritis, cervical lymphadenopathy and eye involvement were recorded and there was a single episode with abdominal pain and vomiting. On average, the episodes lasted for 2 weeks and occurred every 5 weeks. Family history was negative. Complete remission of the symptoms was achieved after starting treatment with anakinra.

We previously screened the TNFRSF1A gene by Sanger sequencing and next generation sequencing (NGS) with negative results. With our smMIP assay, we detected a c.269C>A p.(Thr90Asn) likely pathogenic missense mutation with 1,3% allele frequency in whole blood. This result was confirmed by ddPCR. The mutation was also present in sorted granulocytes, monocytes, T and B cells, but not in non-hematopoietic cells. This distribution is different from late-onset CAPS patients who were found to have myeloid restricted mosaic mutations in NLRP3.

Conclusion: We report the first low-grade mosaic variant in TNFRSF1A. Based on clinical symptoms, favorable response to anakinra and genetic findings, we diagnosed our patient with mosaic TRAPS. Our data suggest that even mutations present in a very small number of cells can cause systemic autoinflammatory disease.

Consent for publication has been obtained from patient


Disclosure of Interest

None Declared

PT1B06 Heterozygous TNFAIP3 mutation as the cause of an interferon-mediated neuroinflammatory disorder

Ciara M. Mulhern1, Ying Hong1, Ebun Omoyinmi1, Dara McCreary1, Marina Casimir1, Cheryl Hemingway2, Felice D’Arco2, Paul Brogan1, Despina Eleftheriou1
1Infection, Inflammation and Rheumatology, Great Ormond Street Institute of Child Health; 2Neurology Department, Great Ormond Street Hospital for Children, NHS Foundation Trust, London, United Kingdom
Correspondence: Ciara M. Mulhern

Introduction: Heterozygous loss-of-function mutations in TNFAIP3 lead to haploinsufficiency of A20 (HA20) commonly manifesting with severe orogenital ulceration and ocular inflammation. Central nervous system (CNS) inflammation has been observed in patients with haploinsufficiency of HA20 but CNS involvement as the sole clinical manifestation of heterozygous TNFAIP3 mutation in humans has never been described however. Herein, we report on the case of an 8 year old female of non-consanguineous Pakistani-Indian descent who presented with left sided focal seizures and hemiparesis, acute uveitis and congitive decline. Magnetic resonance imaging (MRI) of her brain revealed contrast-enhancing T2 hypointense intracranial mass lesions, in the grey matter of the paracentral lobule and the thalamus on the left with surrounding oedema. Brain biopsy revealed necrotising granulomatous inflammation. Brain CT revealed intracerebral calcification. Extensive screening of infectious or malignant causes was negative.She failed to respond to multiple anti-inflammatory treatments but had an excellent radiological and clinical response to an oral JAK-STAT inhibitor suggesting this was likely an interferon (IFN) mediated inflammatory disease.

Objectives: To use next generation sequencing to identify the genetic cause of the progressive neuroinflammatory disorder in this case and characterise the mechanisms underpinning neuroinflammation

Methods: Whole exome sequencing (WES; illumina MiSeq) was carried out in all family members. Expression of phosphorylated- p65, IRF3, STAT1 and STAT3 in both patient and healthy control derived peripheral blood mononuclear cells was assessed by flow cytometry. Meso Scale Discovery (MSD) assays was used to quantify cytokines in patient serum. qPCR assays measured the expression levels of IFN stimulated gene expression. Co-immunoprecipitation analysis assessed the binding capacity of the mutated protein to Tank-binding kinase 1 (TBK1)

Results: WES revealed a heterozygous missense p.T647P mutation in TNFAIP3 as the cause of the progressive neuroinflammatory disorder in this case. Patient-derived cells exhibited enhanced phosphorylation of the p65 transcription factor, and increased expression of NF-ƙB mediated proinflammatory cytokines. The mutated p.T647P A20 protein failed to control interferon-regulatory-factor-3 (IRF3) activation and interferon (IFN)-dependent gene transcription, in comparison to healthy control cells. We also show that the mutant A20 protein cannot efficiently bind to TBK1, in order to turn off IRF3 activation leading to an enhanced IFN signature in the patient. We believe that failure to regulate IFN-mediated immune responses, was the driver of the CNS inflammation observed. Furthermore, treatment with an oral JAK 1/2 inhibitor resulted in marked clinical improvement, complete radiological resolution of neuroinflammation, and normalisation of IFN-stimulated gene expression in whole blood. IFN mediated immune responses were also impaired in an additional disease control case of HA20 in a 4 year old heterozygote for the p.N98Tfs25 TNFAIP3 variant.

Conclusion: We describe for the first time heterozygous p.T647P missense mutation in TNFAIP3 as the cause ofprogressive neuroinflammation. The p.T647P mutated A20 proteinfailed to control IRF3 activation and IFN dependent transcription. Treatment with an oral JAK 1/2 inhibitor was highly effective. Our report now adds TNFAIP3 mediated neuroinflammation to the ever-expanding group of monogenic interferonopathies with propensity to CNS involvement.

Consent for publication has been obtained from patient


Disclosure of Interest

None Declared

PT1B07 Systemic evaluation of genetic and biochemical testing for deficiency of ADA2 (DADA2) from the NIH patient cohort

Natalia Sampaio Moura, Oskar Schnappauf, Natalie Deuitch, Qing Zhou, Daniel Kastner, Ivona Aksentijevich
Inflammatory Disease Section, National Institutes of Health, Bethesda, United States
Correspondence: Natalia Sampaio Moura

Introduction: Deficiency of adenosine deaminase 2 (DADA2) is an autosomal recessive autoinflammatory disorder caused by loss-of-function mutations in the ADA2 gene, which encodes the adenosine deaminase 2 (ADA2) enzyme. Patients with DADA2 can present with many different manifestations including early-onset lacunar stroke, recurrent fever, hepatosplenomegaly, livedo reticularis, immune dysregulation, and hematopoietic abnormalities. They also exhibit absent or significantly reduced ADA2 enzyme activity. Despite these conserved features, some individuals display discordance between clinical characteristics and genetic or enzyme activity testing.

Objectives: The goal of our retrospective study is to evaluate sensitivity of diagnostic testing for DADA2 in our CLIA-certified laboratory at the National Institutes of Health (NIH). We compared the diagnostic efficacy of the different types of assays, such as Sanger sequencing, ADA2 enzymatic analysis and multiplex ligation-dependent probe amplification (MLPA).

Methods: We reviewed all ADA2 Sanger sequencing results of individuals tested between 2014 and 2018, including confirmatory testing in referral patients, and determined the diagnostic yield of this method in patients we suspected to have DADA2 based on clinical phenotype. We then analyzed if subsequent ADA2 enzyme assay and/or MLPA further increased our diagnostic yield.

Results: Out of 190 individuals with clinical characteristics suggestive of DADA2, 56 patients tested positive for biallelic mutations (29%). Eleven tested as carriers for a monoallelic pathogenic mutation (6%), and 123 (65%) were mutation-negative based on conventional Sanger sequencing. ADA2 enzyme activity assay was performed on 45 patients who had available serum samples. The assay corroborated the DADA2 diagnosis on 30 patients and identified 7 individuals with monoallelic mutations (detected via Sanger) from three different families as patients due to low enzyme activity. MLPA successfully identified a second pathogenic copy number variation (CNV) in all patients who displayed low ADA2 activity. Addition of ADA2 protein assay and MLPA increased our DADA2 diagnostic rate to 33%. Thus, we observed a 13.8% (4/29) relative increase in our diagnostic yield due to the incorporation of these testing modalities.

Conclusion: Our results suggest that the serum ADA2 enzyme assay should be considered first tier testing for patients with suspected DADA2, given that pathogenic mutations in ADA2 are highly heterogeneous and not always detectable by Sanger sequencing alone. Sanger sequencing in combination with MLPA remain indispensable assays to support and confirm enzymatic testing when indeterminate or unclear, and to identify causal variants. Detection of mutations at the DNA level is important for genetic counseling. Targeted next-generation testing is another method able to identify a diverse scope of pathogenic mutations and has the potential to increase the diagnostic yield of genetic testing.

Disclosure of Interest

None Declared

Monogenic autoinflammatory diseases (clinical)

P1001 Is there any difference between M694V heterozygote and non-exon 10 mutations on symptoms onset and response to colchicine treatment?

Hatice Adiguzel Dundar1, Serkan Turkucar1, Ceyhun Acari1, Ozge Altug Gucenmez2, Balahan Makay2, Erbil Unsal1
1Department of Pediatrics, Pediatric Rheumatology Unit, Dokuz Eylul University Faculty of Medicine; 2Pediatric Rheumatology Unit, Dr. Behcet Uz Childrens’ Hospital, Izmir, Turkey
Correspondence: Hatice Adiguzel Dundar

Introduction: Familial Mediterranean fever (FMF) is the most common inherited autoinflammatory syndrome throughout the world. It is caused by mutations of the MEFV gene encoding a protein called pyrin. The most frequent genotype-phenotype correlation is in a certain part of exon 10, especially M694V mutation. There are also a group of patients with non-exon 10 mutations, who have a similar clinical spectrum of the disease.

Objectives: We aim to investigate the genotype-phenotype differences between M694V heterozygote mutations and non-exon 10 mutations.

Methods: Data charts of children (n=431) with FMF from Dokuz Eylul University childrens’ hospital and Dr.B.Uz childrens’ hospital were reviewed. Patients were divided into two groups with regard to having M694V heterozygote or non-exon 10 mutations. Genotype-phenotype features and response to treatment were compared.

Results: There were M694V heterozygote mutations in 128 (29.7%) patients and non-exon 10 mutations in 303 (70.3%) patients. The follow-up period was 54.5 (33-105) months. There was no difference between the age of symptoms onset, the age of diagnosis, and the diagnosis delay time. The family history in patients with M694V heterozygote mutation was statistically positive compared to non-exon 10 mutation group (p:0.000). The symptoms of joint involvement as arthritis were significantly higher in the M694V heterozygotegroup (p:0.026). Additionally, biological agent need due to colchicine unresponsiveness was statistically higher in M694V heterozygote group than group with non-exon 10 mutation (p:0.004) (Table 1).

Conclusion: There is a significant difference between children with M694V and non-exon 10 mutations, even when the M694V mutation is present in one allele only. Family history with FMF, musculoskeletal symptoms, and unresponsiveness to colchicine are main parameters.

Disclosure of Interest

None Declared

P1002 A case report of aicardi goutieres syndrome type 5 in two siblings mimicking juvenile idiopathic arthritis

Buthaina Al Adba1, Hajar Dauleh2
1Paediatric Rheumatology; 2Paediatric, Sidra Medicine, Doha, Qatar
Correspondence: Buthaina Al Adba

Introduction: Aicardi-Goutières syndrome (AGS) is a genetically determined encephalopathy characterized by calcification of the basal ganglia and white matter, demyelination, and raised levels of lymphocytes and IFNα in the cerebrospinal fluid [1]. Neurological dysfunction becomes clinically apparent in infancy and manifests as progressive microcephaly, spasticity, dystonia, and psychomotor retardation. Expression of interferon-regulated genes (IGS) in peripheral blood is also upregulated, which is sustained over time [2]. The genes mutated in AGS have been defined to encode proteins implicated in the metabolism of nucleic acids TREX1, the RNASEH2 complex and SAMHD1,which may all function as cellular nucleases [3].

Objectives: Arthritis and progressive arthropathy with distal joint contractureshave been reported in SAMHD1 mutation along with neurological symptoms [4]. We are describing two siblings with SAMHD1 mutation who only presented with early onset polyarthritis and no systemic or CNS manifestations.

Methods: Chart review of clinical data includes IFN signature and molecular analysis.

Results: Two brothers of consanguineous parents with no significant family history of auto inflammatory or autoimmune disease, were born healthy, at term with normal growth parameters.Patient one is a six yearold boy who presented at age of four with one-year history of joint pain and morning stiffness. He had active arthritis in his both wrists, knees and elbows. Lab testing revealed normal blood work ESR, CRP, and negative ANA and RF. Interestingly HLAB27 was present.A diagnosis of JIA was made and he was started on naproxen, and had intra-articular in jectionsarticular injections twice. His arthritisHis arthritis was difficult to control and Etanercept andsubcutaneous Methotrexate were started. His arthritis improved significantly,with residual mild disease atboth wrist joints.His brother, patient two is a 3 year old boy who presented similarly at age of 18 months with morning stiffness and abnormal gait for one-month duration. He had active arthritis in both knees, hips and wrists. He had similar lab findings and positive HLAB27.He was treated with naproxen, a short course of oral steroid and Methotrexate. He responded faster than his brother, but continue to have mild arthritis atboth wrists. Due to difficult to treat, early arthritis in two consanguineous brothers, wesent genetic testing with whole exome sequencing (WES) to look for other causes of their arthritis.Genetic testing for both siblings showed Homozygous mutation at the SAMHD1 gene for the P.Arg 290His (CGT>CAT):c.869G >A.Both parents were heterozygous for the same mutation. The R290H variant in the SAMHD1 gene has been reported previously in association with AGS in an affected individual with multiple clinical features including CNS symptoms and arthritis [5].Peripheral blood analysis of type I interferon-related biomarkers demonstrated that both siblings have an interferon signature characteristic of Aicardi-Goutieres syndrome (Fig 1). Both brothers continued to have normal development with no CNS symptoms or rash and their eye exam showed no glaucoma, which was previously reported in AGS with the SAMHD1 mutation. Despite being asymptomatic from CNS disease we are considering brain MRI to look for inflammatory cerebral vasculopathy that has been reported previously [6].

Conclusion: We are reporting a spectrum of AGS with a SAMHD1 mutation that mimicks difficult to treat early onset polyarticular arthritis without any CNS or skin manifestations. This illustrates the need to consider mutation analysis of SAMHD1 in similar presentations as CNS and skin disease may evolve later. Finally, even though there has been partial response to treatments,we may consider changing therapy to Janus Kinase inhibitors to achieve complete remission of arthritis.

Consent for publication has been obtained from patient


Disclosure of Interest

None Declared

P1003 Phenotypic and genotypic characteristics and damage accrual of monogenic autoinflammatory diseases other than familial Mediterranean fever from the pediatric rheumatology Arab group (PRAG)

Sulaiman Al-Mayouf1, Abdulaziz Almutairi1, Safia Albrawi2, Abdulatif AlEnazi3, Basil Fatallah4, Abdulallh Alsonbul1, Mohammed Abu-shukair5, Raed Alzyoud5, Adel Alwahadneh5, Mabruka Zlenti6, Ebtisam Kawaja6, Khloud Khawaja7, Zakia Almusawi8, Wafa Madan8, Muna AlMutairi9, Nora Almuatiri10 and Pediatric Rheumatology Arab Group (PRAG)
1King Faisal Specialist Hospital and Research Center, Riyadh, Saudi Arabia; 2Royal Hospital, Muscat, Oman; 3King Fahad Medical City, Riyadh, Saudi Arabia; 4AlJalial Children Hospital, Dubai, United Arab Emirates; 5Queen Rania Children Hospital, Amman, Jordan; 6Tripoli Children Hospital, Tripoli, Libya; 7Mafraq Hospital, Abu Dhabi, United Arab Emirates; 8Salmaniya Hospital, Bahrain, Bahrain; 9Al Adan Hospital; 10AlSabah Hospital, Kuwait, Kuwait
Correspondence: Sulaiman Al-Mayouf

Introduction: Monogenic autoinflammatory diseases (AIDs) are a group of rare hereditary recurrent multisystem inflammatory diseases.The available published data from Arab countries about monogenic AIDs other tahn Familial Mediterranean fever (FMF) is very limited.

Objectives: To report the phenotype, genotype and response to treatment in Arab children with monogenic AIDs other than FMF, with focus on accrual damage.

Methods: We retrospectively reviewed patients with clinical and/ or genetically proven monogenic AIDs other than FMF seen between 1990 and 2018 at 10 rheumatology clinics from seven Arab countries. Data were collected at the last follow-up visit comprising history of consanguinity, age at onset and diagnosis, follow-up duration, clinical and laboratory findings, as well as the damage accrual and death related to monogenic AIDs.

Results: Seventy (46 female) patients with monogenic AIDs were analyzed. Consanguinity rate among the enrolled patients was 74.6% and a family history of AIDs was present in 60%. The mean age at disease onset was 3.2±2 years and mean duration of follow-up was 7.5±4 years. The initial diagnosis was inaccurate in 47% and the mean diagnosis delay was 4±3 years. The most frequent monogenic AIDs were LACC1 associated monogenic disorders (monogenic JIA and monogenic Crohn’s) (23) followed by cryopyrin-associated periodic syndrome (CAPS) (12), tumor necrosis factor receptor associated periodic syndrome (TRAPS) (12), hyperimmunoglobulinemia D syndrome (HIDS) (9), Majeed’s syndrome (6) and eight patients with other monogenic AIDs. Musculoskeletal involvement was the main feature in LACC1 associated monogenic disorders while fever and mucocutaneous and gastrointestinal involvement were the most prevalent features in the other monogenic AIDs. Genetic testing was performed in 67 patients, 69% had genetically confirmed disease. Patients with mutation c.T850C p.C284R in exon 4 of LACC1 had severe arthritic changes. Three CAPS patients with NLRP3 mutation had cognitive impairment and one with significant hearing and ocular damage. Two HIDS patients had homozygous p.V3771 mutation and other two patients with p.V3771/compound heterozygous MEFV: p.E148Q/p.P369S/p.R408G. Three different LPIN2 mutations were recorded for Majeed’s syndrome. Overall, growth failure was the most frequent (36%), followed by cognitive impairment (13%). There were three deaths due to infection.

Conclusion: The number of genetically confirmed patients with monogenic AIDs other than FMF are not uncommon among Arab children probably due to a high consanguinity rate. Diagnostic delay and high damage accrual emphasize the need for more awareness and early referral to specialized centers.

Disclosure of Interest

None Declared

P1004 Auto-inflammatory diseases: a retrospective single center study in Saudi Arabia

Abdulrahman A. Alrasheed1, Ashwag Al Harthi2, Banan Al Rewaithi2, Wafa Suwairi2, Jubran Al Qanatish2, Fayhan Al Roqi2
1King Abdullah Specialized Children Hospital; 2Pediatric, King Abdullah Specialized Children Hospital, Riyadh, Saudi Arabia
Correspondence: Abdulrahman A. Alrasheed

Introduction: The auto-inflammatory conditions represent an emerging and over-expanding series of diseases unified by a dysregulation of innate immunity. The current increase in the prevalence of chronic inflammatory diseases makes this subject of interest. Early recognition and treatment with immunomodulator agents have the potential to improve the outcomes and the quality of life.

Objectives: The purpose of this review is to describe the clinical features, investigations, therapeutic modalities and outcome of auto-inflammatory syndromes seen at King Abdullah Specialized Children’s Hospital, Riyadh, Saudi Arabia.

Methods: We carried out a descriptive retrospective analysis of pediatric patients with auto-inflammatory diseases who were seen and managed at a single tertiary center, in Riyadh, Saudi Arabia, between 2004 and 2018. Multiple measures were investigated including time of disease onset, clinical features, investigations, treatment, and outcome.

Results: 67 patients (30 females and 37 males) with auto-inflammatory diseases were identified and formed the basis of the study (Table 1).

Autoinflammatory diseases include Systemic Juvenile Idiopathic Arthritis (sJIA) 46%, Chronic Recurrent Multifocal Osteomyelitis (CRMO) 15%, Familial Mediterranean fever (FMF) 7%, Hyperimmunoglobulin D syndrome (HIDS) 6%, Periodic fever, aphthous stomatitis, pharyngitis, cervical adenitis (PFAPA) 6%, Neonatal onset multisystemic inflammatory disease (NOMID) 3%, STING-associated vasculopathy with onset in infancy (SAVI) 3%, ), Aicardi-Goutières syndrome (AGS) 1%, Deficiency of Adenosine Deaminase 2 (DADA2) 5%, Deficiency of interleukin thirty-six receptor antagonist (DITRA) 1%, Blau syndrome 1% and Uncharacterized Auto-inflammatory Disease (UAD) 3%.

The mean age at initial presentation was 58.2 months. The median Erythrocyte Sedimentation Rate (ESR) and C - Reactive protein (CRP) were 72 mm/hr and 86 mg/dl, respectively. Treatment modalities that have been used include Disease-modifying antirheumatic drugs (DMARDs) and/ or biologic therapy including Anti TNF alpha, Anti IL-1 or Anti IL-6 and others (Table1).

Conclusion: We believe this cohort provides the common features of auto-inflammatory diseases in Saudi Children and the outcome after frequently used target therapies. Early recognition and treatment of auto-inflammatory diseases with immunomodulator agents have the great potential to alleviate the debilitating symptoms and to improve the quality of life.

Disclosure of Interest

None Declared

Table 1 (abstract P1004).

See text for description


No. of patients


gene study

treatment and outcome




Negative HLH gene done for patients with MAS

good response to Anakinra or Tocilizumab




Negative geneexcept 1 patient with CRMO and Familial hyperphosphatemic tumoral calcinosis who has homozygous mutation in GALNT3 gene

3 patients with mild CRMO responded to NSAIDs only.

2 patients required either pamidronate or Methotrexate.

4 patients required combination of Infliximab and Methotrexate.

1 patient who had unidentified auto- inflammatory disease responded partially to Abatacept but not to Anakinra, Adalimumab or Tocilizumab.




Heterozygous mutation in MEFV gene (Met694Val)

Good response to Colchicine




Negative periodic fever gene panel

4 patients received no treatments with less frequent attacks during follow up and one free of attacks after tonsillectomy




Mutation in MVK gene

3 patients treated with Anakinra showed good response

1 patient had infrequent attacks not on treatment




Negative gene

Good response to anti IL-1 therapy (Anakinra and Canakinumab)





Mutation in TMEM-173 gene (STING gene) for SAVI and RNASEH2A for AGS

1 SAVI patient treated with Ruxolitinib with significant improvement while the other died

AGS treated with Ruxolitinib with good response




mutation in CECR1 gene

2 patients treated with adalimumab and GCSF with good response, and the 3rd patient recently diagnosed




mutation inIL36RN gene

Response was lost on etanercept, Anakinra and then Ustekinamab

Currently on Adalimumab with no relapse so far

Blau syndrome + crohn’s disease



mutations in NOD2/CARD15 gene

Treated with Azathioprine and steroid with partial response

P1005 Clinical and genetic characteristics of myalgia in Armenian children with familial Mediterranean fever

Gayane G. Amaryan1, Tamara F. Sarkisian2, Nune G. Mkrtchyan3, Marina M. Papazyan4, Artashes E. Tadevosyan5
1National Pediatric Centre for Familial Mediterranean Fever of “Arabkir” Medical Centre - Institute of Child and Adolescent Health, Department of Pediatrics Yerevan State Medical University; 2Centre of Medical Genetics and Primary Health Care, Depatment of Medical Genetics Yerevan State Medical University; 3National Pediatric Centre for Familial Mediterranean Fever of “Arabkir” Medical Centre - Institute of Child and Adolescent Health, Department of Pediatrics Yerevan State Medical University; 4NationalPediatric Centre for Familial Mediterranean Fever, “Arabkir” Medical Centre - Institute of Child and Adolescent Health; 5Department of Public Health and Health Care Organization , Yerevan State Medical University, Yerevan, Armenia
Correspondence: Gayane G. Amaryan

Introduction: Familial Mediterranean Fever (FMF) as an ethnic disease is wide-spread in Armenia. In most cases FMF manifests in childhood. Myalgia is an essential feature of musculoskeletal symptoms (MSM) of FMF, which can be presented as acute muscle attacks - spontaneous myalgia (SM) and exercise-induced myalgia (EIM), as well as prolonged protracted febrile myalgia (PFM). Awareness of these symptoms are important for early diagnosis and differential diagnosis of FMF.

Objectives: to study the frequency of different types of myalgia in Armenian children with FMF and their correlation with MEFV genotype and disease severity.

Methods: A group of 715 children with FMF was observed at the National Pediatric Centre for FMF (438 boys, 277 girls, mean age 8.64±0.17). The diagnosis of FMF was based on the Tel-Hashomer criteria and detection of 12 MEFV mutations common for Armenians using Viennalab Diagnostics molecular-genetic assay. For statistical analysis standard statistical Epi-Info 2000 Program was performed. For comparison of two nominal variables in table “two by two” Yaet’s corrected for continuity chi-square test was used, significance level p<0.05

Results: Myalgia was observed in 37.5% (268 out of 715) of FMF patients and manifested mainly as SM and /or EIM in 34.7% (248 patients), as well as PFM in 2.7% (20 patients).

SM and EIM manifested as transient pains of the legs without fever, disappearing after resting or taking NSAIDs. The frequency of development of SM and EIM varied from 43.6% to 31.2% depending on the genotype and was relatively common in M694V and V726A heterozygotes compared to homozygotes(χ2= 3.41; p> 0.05). Risk of development of SM and EIM was associated with severity of FMF (χ2= 23.20, p <0.0001) and was higher in severe and moderate course of FMF compared to mild 5.6 times (χ2= 18.28; P <0.0001) and 1.9 times (χ2= 3.77; P <0.05) respectively.

We observed prolonged PFM in 2.7% of FMF patients with more severe phenotypes, which were characterized by 4 times higher risk of early onset (χ2= 5.94; p = 0.015) (mean age 3 years), as well as late diagnosis of FMF (9.42 ± 0.72) and the delayed start of colchicine therapy . This cohort of patients had also high frequency of severe FMF attacks with generalized febrile muscle pain during 1-3 weeks, prevalence of acute recurrent arthritis, erysipelas-like erythema. PFM manifestation usually was after 5-6 year of FMF onset. Because of PFM is considered also as vasculitis, steroid therapy was started along with colchicine. Development of PFM was associated with severe M694V-homozygous genotype (χ2 = 8.27; p <0.02) and was observed at 4.6% of M694V-homozygous patients.

Conclusion: The frequency of different types of myalgia in Armenian children with FMF is high (37.5%), especially acute muscle attacks - SM and/or EIM (34.7%). PFM is diagnosed in 2.7% patients.

The development of SM and EIM is associated with the severity of FMF and does not depend on the MEFV genotype. The risk of development of PFM, as a part of prolonged MSM, is associated with both: the MEFV genotype (mostly M694V- homozygous) and the severity of the disease.

This allows to consider PFM as a marker of severe course of FMF and early disease onset and the M694V homozygous genotype as a risk factor for the development of PFM.

MEFV mutation genetic screening is recommended for Armenian paediatric patients with different types of myalgia, especially with PFM, for early diagnosis of FMF, treatment and prevention of complications.

Disclosure of Interest

None Declared

P1006 The impact of aging on familial Mediterrinean fever patients

Okan Aydin1, Serdal Ugurlu1, Bugra Egeli2, Ece Soykut2, Deniz Demir2, Huri Ozdogan1
1Division of Rheumatology, Department of Internal Medicine, Cerrahpasa Medical Faculty; 2University of Istanbul - Cerrahpasa, Istanbul, Turkey
Correspondence: Okan Aydin

Introduction: Familial Mediterranean Fever (FMF) is a monogenic autoinflammatory disorder with innate immune activation with an onset before age 20 in approximately 90% of the patients. There is scarce data on the effect of aging on FMF patients over 40 years of age

Objectives: This study aims to collect data on FMF patients who have survived over 40 years of age. Here we report our preliminary data on disease course and treatment status and comorbidities of our patients with FMF.

Methods: Among the FMF patients who have been followed in our FMF outpatient clinic with a pool of approximately 5000 patients, those who have aged 40 and over are being included to the study. As by today 180 patients are considered for evaluation. The files of patients were reviewed and a standard questionnaire was used to interview the patients. Here we report the results of 100 of these patients (56%) who were contacted for this purpose. These patients were questioned on their demographic characteristics, comorbid conditions, colchicine treatment details, and attack information. In order to see the trend of the change in the parameters assessed , the patients were divided into two groups based on their present age(Group 1: 40-50 years, Group 2: ≥50 years).

Results: A total of 100 (78 F, 22M) patients were evaluated. There were 61(46F, 15M) patients aged between 40-50 years and 39 (32F, 7M) over 50. The demographic characteristics and clinical features of these patients are given in Table 1.Besides 3, all patients were still on colchicine regularly. Ninety-six percent of the patients declared overall benefit from colchicine therapy, however 38% experienced a side effect related to this treatment.Over 88% of the patients reported decrease in severity and frequency of FMF attacks.The mean daily colchicine dose was lower in the age 50 and over group (1.7±0,77 mg versus 1,35 ±0,38 mg). There were no patients with AA amyloidosis in neither age group. The mean duration from the last attack increased from 15.3 ±19.7 months to 35.6± 52 months in the older patients. One or more additional disease was present in 75% of this patient group.Among the comorbidities hypertension was the most frequent, diagnosed in 25% of the patients, followed by hypothyroidism (16%), diabetes mellitus (10%) and cardiac disease (5% ). Sixty-five of the patients were recieving other medications in addition to colchicine.

Conclusion: According to our preliminary data the majority of the patients continue to take colchicine after age of 40. However the frequency of FMF attacks as well as daily colchicine dose decrease as the patients get older.With well designed trials stopping colchicine treatment may be considered in a subgroup of patients after 50 years of age. Approximately ¾ of the FMF population over 40 years of age has a comorbidity that neccecitates additional medications which underlines the need for special attention.

Disclosure of Interest

None Declared

Table 1 (abstract P1006).

Clinical course and co-morbidities in two age groups over 40 years


Group 1*


Group 2**



Sex (F:M); current age (mean±SD) (yr)

(46 :15) ;45.5 ± 2.29

(32:7); 57.05 ± 6.81


Mean duration since the last episode, (mean±SD, mo)

15.3 ± 19.7 (1-60)

35.67 ± 52.05 (1-276)


Number of patients on colchicine therapy, n (%)

59 (96.7)

38 (97.4)


Mean colchicine dose, mg/day (current)




Number of patientswith decrease in attack severity, n (%)


35 (89,7)


Number of patients with decrease in attackfrequency, n (%)


37 (94,8)


Co-morbidities, n (%)

 Hypertension, n (%)


13 (33.33)


 Hypotyroidism, n (%)




 Type 2 Diabetes Mellitus, n (%)




 Rheumatological diseases, n (%)




 Cardiac disease, n (%)




 Malignancies, n (%)




 Additionalmedications (number of patients, %)

36 (61)

29 (74.3)


P1007 Comorbidities in familial Mediterranean fever

Ummusen Kaya Akca1, Banu Balci Peynircioglu2, Zehra S. Arici1, Edibe Avci2, Zulfiye Y. Akkaya Ulum2, Engin Yilmaz2, Yelda Bilginer1, Seza Ozen1
1Pediatric Rheumatology; 2Medical Biology, Hacettepe University, Ankara, Turkey
Correspondence: Banu Balci Peynircioglu

Introduction: Familial Mediterranean Fever (FMF) is a periodic fever syndrome, characterized by recurrent episodes of fever and serosal inflammation accompanied with high acute phase reactants.The analysis of possible comorbidities is important to understand the impact of these conditions on clinical care and whether they share a common etiological pathway.

Objectives: We aimed to evaluate the comorbidities associated with FMF patients in a large genetically diagnosed cohort.

Methods: We retrospectively evaluated the medical records of FMF patients who were followed up at Department of Pediatric Rheumatology in Hacettepe University between 2000 and 2015. This study was approved by the Research Ethics Committee and was conducted in accordance with the Declaration of Helsinki. The diagnosis of FMF was made according to Tel Hashomer diagnosis criteria for patients who applied prior to April 2009 and to the Turkish FMF pediatric diagnosis criteria after April 2009. The FMF patients who had homozygous or compound heterozygous mutations were included in the study. Comorbidities associated with FMF were divided into three groups; associated with increased inflammation, associated with FMF and incidental.

Results: A total of 1999 patients were enrolled in the study. Of all 1999 FMF patients, 636 were children (31.8 %), 1029 were males (51.4%), with a mean age of 31.60±16.01 years. The mean follow up time was 4.50±3.99 years (median:3.84 range from 0.21-29.4 years). 880 of 1999 (44%) FMF patients had homozygous MEFV gene mutation, the most common mutation was M694V homozygous. The remaining were compound heterozygous. 656 patients (32.8%) had one or more than one comorbidity associated with FMF. Ankylosing spondylitis was the most common comorbidity associated with increased inflammation while the most common comorbidity in FMF related comorbidities was renal amyloidosis. The frequency of ankylosing spondylitis, henoch schonlein purpura, juvenile idiopathic arthritis, polyarteritis nodosa (PAN), multiple sclerosis (MS) and Behçet’s disease were increased in patients with FMF when compared to those in the literature. Systemic lupus erythematosus was observed less frequently in the patients with FMF than in the population. While the increase in the frequency of MS was 3.3 times, the frequency of PAN was increased 110 times.

Conclusion: This study shows that FMF is a hereditary disease associated with significant comorbidity. We also confirm that inflamatory and rheumatic diseases are more common in FMF.

Disclosure of Interest

None Declared

P1008 Bone metabolism in systemic autoinflammatory diseases (SAIDS): a case- control study from Padova cohort

Sara Bindoli1, Giulio Franceschet2, Paola Galozzi1, Martina Zaninotto3, Valentina Camozzi2, Paolo Sfriso1
1Rheumatology Unit, Department of Medicine; 2Endocrinology Unit, Department of Medicine; 3Department of Laboratory Medicine, University of Padova, Padova, Italy
Correspondence: Sara Bindoli

Introduction: Systemic autoinflammatory diseases (SAIDs) represent a group of disorders characterized by recurrent fever attacks, polyserositis, skin, musculo-skeletal and articular manifestations. A dysregulation of the innate immune response in a genetic predisposed host leads to the development of SAIDs. In our study we included patients affected by Familial Mediterranean Fever (FMF), TNF Receptor Associated Periodic Syndrome (TRAPS) and Hyper-IgD Syndrome (HIDS). It is assessed that chronic inflammation, perpetuated by pro-inflammatory cytokines, may exert a role on bone metabolism leading, in the long run, at the onset of osteoporosis (OP). However, how OP may occur in the context of autoinflammatory diseases remains partially unknown.

Objectives: The aims of our study are focused on the assessment of bone metabolism in patients affected by SAIDs compared to healthy subjects and on the relationship between bone turnover markers (Receptor activator of nuclear factor kappa-Βligand, RANKL and osteoprotegerin, OPG) and serum inflammation markers (serum amyloid A, SAA).

Methods: 40 adults patients referring to the Rheumatology Unit of Padova University Hospital affected by FMF, TRAPS and HIDS and 40 healthy subjects were recruited between March and June 2018. Fasting blood samples were collected in order to determinate calcium (Ca), phosphorus (P), magnesium (Mg), 24-h urine calcium, 24-h urine phosphorus, albumin, parathyroid hormone (PTH), Vitamin D, creatinine, serum amyloid A (SAA), c-terminal telopeptide of type I collagen (CTX), bone alkaline phosphatase (b-ALP). Moreover, serum OPG and RANK-L were determined by a commercially available ELISA kit (Pantec, Turin, Italy). Femur and lumbar dual-energy X-ray absorptiometry (DXA) was performed with the QDR Bone Densitometer Discovery (Hologic Inc.,Waltham, MA). Trabecular Bone Score (TBS) was calculated on DXA lumbar images, using iNsight Software (version; Medimaps, Merignac, France). The statistical analysis was performed using Mann-Whitney U Test.

Results: We did not observe a statistically significant difference between bone mineral density (BMD) and TBS of patients compared to controls (p=0.5037 and p=0.8031). Also, the values of phospho-calcic metabolism samples were not statistically different between patients and controls. As expected, SAA levels were significantly higher in patients (p= 0.0144), and interestingly also OPG levels were significantly higher if compared to the healthy subjects (p= 0.0018). For b-ALP, CTX and RANK-L no differences were observed between the two groups (p=0.1466, p=0.8861, and p= 0.7890 respectively).

Conclusion: Patients of our cohort affected by FMF, TRAPS and HIDS do not present an increased risk of OP compared to the healthy controls. Indeed, TBS and BMD are similar between the two groups highlighting a preserved bone quality in our patients. Interestingly, OPG levels are higher compared to controls and this could suggest a protective role and a bone re-balancing activity in a context of inflammation. Finally, a regulatory effect of serum amyloid A on bone homeostasis should be taken into account.

Disclosure of Interest

None Declared

P1009 Two cases of hyperzincaemia/hypercalprotectinaemia show novel phenotypic properties

Anikó Szabó1, Péter Blazsó1, Viktória Sümegi2, Tibor Kalmár1, Zoltán Maróti1, Csaba Bereczki1
1Department of Paediatrics; 2Department of Rheumatology and Immunology, University of Szeged, Szeged, Hungary
Correspondence: Péter Blazsó

Introduction: Hyperzincaemia/hypercalprotectinaemia (Hz/Hc) is a recently discovered autoinflammatory disorder showing monogenic, autosomal dominant inheritance pattern. Until now the p.E250K and p.E257K missense mutations of PSTPIP1 gene have been found in the background. These result in the extreme overproduction of the pro-inflammatory cytokine IL-1β leading to the striking elevation of serum calprotectin and zinc through positive feedback loops. Uncontrolled inflammation elicited by the continuous activation of these danger signals end up in the classical constellation of systemic and cutaneous inflammation, hepatosplenomegaly, arthritis, pancytopenia, and failure to thrive.

Objectives: We aimed to characterize the clinical phenotypes accompanied the mutations of PSTPIP1 gene found in two unrelated Hungarian male patients. These data were compared to the clinical presentations of the already published cases and novel symptoms were demonstrated.

Methods: Clinical cases of both boys were investigated thoroughly. In order to clarify the diagnosis targeted exome sequencing panel (Illumina TruSight One) was applied in each case to pre-screen for possible mutations. Nucleotide variants in the probands and in their parents were confirmed by Sanger sequencing.

Results: A p.E250K mutation was detected in one patient and a p.E257K in the other. Both variants were de novo and heterozygous. p.E250K in the 2,5-year-old boy was associated with additional recurrent non-infectious diarrhea, macrocephaly and pericardial fluid on the top of the classical signs. p.E257K in the 7-year-old boy was joined with haemangiomatosis, mental retardation, autism and overweight besides the already known manifestations of Hz/Hc.

Conclusion: Atypical and potentially misleading clinical findings in Hz/Hc underline the importance of genetic testing in order to get to the proper diagnosis. The presented cases might extend our knowledge of the phenotypic traits observable in Hz/Hc.

Consent for publication has been obtained from patient


Disclosure of Interest

None Declared

P1010 Identification of novel NLRC4 and IL2RA variants in a family cohort with juvenile-onset arthritis and rash

Jessica L. Bloom1, Megan L. Curran1, Scott Canna2, Harold Hoffman3, Elena Hsieh4
1Department of Pediatrics, University of Colorado, Aurora; 2Departments of Pediatrics and Immunology, University of Pittsburgh, Pittsburgh; 3Departments of Pediatrics and Medicine, University of California San Diego, San Diego; 4Departments of Pediatrics and Immunology and Microbiology, University of Colorado, Aurora, United States
Correspondence: Jessica L. Bloom

Introduction: Identification of genetic etiologies of autoinflammatory syndromes can inform targeted therapy to improve outcomes.

Objectives: We aimed to identify a genetic etiology for an underlying autoinflammatory syndrome in a 3-year-old boy presenting with failure to thrive, rash, and polyarthritis since infancy without significant fevers. The patient’s mother and maternal aunt also had similar symptoms since infancy.

Methods: We obtained a history, exam, routine laboratory evaluation, chromosomal microarray, immune phenotyping and functional assays, and genetic sequencing of familial autoinflammatory syndromes via INVITAE.

Results: The patient’s first examination showed small and large joint polyarthritis and maculopapular rash. Laboratory results included anemia, positive ANA, negative RF and anti-CCP, mildly raised ferritin, and very elevated platelet level, LDH, ESR, CRP and IgG. He had recurrent diarrhea and tested positive for Campylobacter. INVITAE’s autoinflammatory panel showed two heterozygous variants of unknown significance (VUS): NLRC4, exon 4, c.741_742insAlu (p.Leu247fs) and IL2RA, exon 2, c.76G>C (p.Asp26His).

Genetic testing revealed the same two variants in the patient’s mother and aunt, while unaffected relatives had one or the other (see Table 1). The patient and mother are HLA-B27+ and have the same 2.8 Mb duplication from 3q28 to 3q29 on chromosomal microarray including IL1RAP. Signal transducer and activator of transcription phosphorylation (pSTAT5) studies showed increased baseline pSTAT5 induction without IL-2 stimulation in patient compared to control.

The patient’s arthritis improved partially with naproxen and steroid joint injections. Given genetic results, subcutaneous anakinra was initiated at 10 mg/kg daily with significant improvement in arthritis, rash, and fatigue. Labs after one month showed resolved anemia, normal inflammatory markers, and high IL-18 (7,824 pg/mL, normal 89-540). He switched to 4.29 mg/kg of canakinumab monthly. After one month, he had mildly active arthritis, occasional fatigue, and stable labs apart from an increase of IL-18 to 15,329 pg/mL. The mother, who is poorly controlled off therapy, has elevated IL-18 (7,176 pg/mL)

Conclusion: We present a family cohort with juvenile-onset arthritis and rash, found to have elevated IL-18, VUS in both NLRC4 and IL2RA, and a chromosomal duplication consistent with a heritable autoinflammatory syndrome. While it appears that both variants are required for symptomatology, his presentation is most consistent with an unrecognized gain of function NLRC4 mutation despite lack of recurrent fevers or enterocolitis. Genetic evaluation led to targeted therapy and improved outcomes, with additional testing underway to further personalize therapy.

The patient's family consented for publication.


1. Canna SW, et al. An activating NLRC4 inflammasome mutation causes autoinflammation with recurrent macrophage activation syndrome. Nature Genetics. 2014;46(10):1140-6.

2. Romberg N, et al. Mutation of NLRC4 causes a syndrome of enterocolitis and autoinflammation. Nature Genetics. 2014;46(10):1135-9.

Disclosure of Interest

None Declared

Table 1 (abstract P1010).

INVITAE Panel Results

Relation to Patient















Maternal Aunt



Maternal Half-Aunt



Maternal Grandfather



Maternal Grandmother



1c.741_742insAlu (p.Leu247fs), heterozygous, ExAC 0

2c.76G>C (p.Asp26His), heterozygous, ExAC 0.1%

3c.179T>C (p.Leu60Pro), heterozygous, ExAC 0

4c.741_742insAlu (p.Leu247fs), possibly mosaic, ExAC 0

*2.8 Mb duplication from 3q28 to 3q29 on chromosomal microarray (maternal aunt's is in process)

P1011 Discontinuation of colchicine therapy in children with familial Mediterranean fever

Yonatan Butbul1, Rawan Silman1, Shafe Fahoum2, Yackov Berkun3
1Department of Pediatrics B, , Rappaport Children's Hospital, Rambam Medical Center, Haifa; 2Department of Pediatrics B, Rappaport Children’s Hospital, Rambam Medical Center, Haifa; 3Department of Pediatrics and FMF Clinic, Hadassah-Hebrew University Medical Center, Mount Scopus, Jerusalem, Israel
Correspondence: Yonatan Butbul

Introduction: Clinical phenotype of FMF exists in some carriers of MEFV mutation. These patients tend to have a mild disease. Prolonged colchicine free remission was reported in a small group of FMF patients.

Objectives: To describe and characterize a group of children with FMF in whom colchicine was discontinued.

Methods: The study cohort consisted of all children with FMF followed at 2 referral centers in Israel in whom colchicine was discontinued following prolonged attack free period.

Clinical presentation, mutations in MEFV gene and disease outcome of patients who successfully ceased colchicine therapy were compared with patients with relapse of FMF attacks.

We performed a retrospective study in two referral centers in Israel of 43patients with FMF with 1 or non-mutated MEFV allele who ceased colchicine therapy following prolonged attack free period. The phenotype of the patients was investigated in detail, and the MEFV mutations, laboratory findings, clinical picture and outcome of 30 (70%) patients that successfully ceased colchicine therapy were compared to 13 (30%) patients whofailed.

Results: 47 patients (55% males), mean age 6±3.2 years at the diagnosis, were enrolled in the study, of them 4patients were excluded due to poor follow up. Fever (93%), abdominal pain (79%), arthralgia (19%) and arthritis (12%) were the most common symptom at attack.The average period free of attacks before enrolment was 11.3±9.2 months. The average follow-up after ceasing colchicine was 5.1± 2.9 years. Thirteenpatients (30.2%) had anattack during follow up with most common symptomsof fever (92%) and abdominal pain (77%) and colchicine therapy was restarted within 10.1 months (1.1-36.4months). There were no differences between the groups of patients that were able to stop colchicine and the group that needed to renew therapy in demographic, genetic and most clinical parameters, including the age (13.4±3.9 vs 11.9±3.7p-0.26), level ofSAA at enrolment (4±3.6 vs 3.3±2.4p-0.7) and time of last attackprior to enrolment (12.6±9.6 vs 8.6±8.2 monthsp-0.08). Myalgia and arthritis were more common among children that required to renew therapy compared to the group that didn’t (31% vs 6.7% p-0.058 and 31% vs 3% p-0.024 respectively).

Conclusion: Cessation ofcolchicine therapy following prolonged remission in selected group of patients who are not homozygous for MEFV mutation could be considered. Patients with arthritis or arthralgia are more likely to have an attack after ceasing colchicine therapy.

Disclosure of Interest

None Declared

P1012 Long-term follow up of a mevalonate deficiency kinase patients cohort

Inmaculada Calvo Penades, Berta Lopez Montesinos, M. Isabel Gonzalez Fernandez, Miguel Marti Masanet, Elena Fernandez De La Puebla
Pediatric Rheumatology Unit, HUIP la Fe, Valencia, Spain
Correspondence: Inmaculada Calvo Penades

Introduction: The syndrome mevalonate kinase deficiency (MKD) is part of the syndromes of periodic fever. With typical clinical manifestations and good response to treatment with IL-1 blockade, but the long-term manifestations are little known

Objectives: To assess the clinical features, treatment and evolution of 11 patients diagnosed with Mevalonate Kinase Deficiency (MKD).

Methods: Baseline demographic and clinical characteristics of the patients were considered, including age at symptoms onset, age at diagnosis, time to follow up, clinical features, laboratory data, results of the MVK gene sequencing study, administered treatments and evolution.

Results: Overall, 11 patients. Gender: M/W 7:4. Onset age: 0.8 m (0-15 years). Age at diagnosis: 9.6 years (4m-15 years), time of follow up: 9.8 years (4m-11 years). Clinical features: Fever 93% of the patients, adenopathy 93%, abdominal pain 66%, oral aphtha 66%, diarrhea 60%, arthritis 60%, amygdalitis 60%, other symptoms: skin rash 34%, headache, 34%, hepatomegaly 27%, splenomegaly 20% and serositis 10%. Three patients presented atypical manifestations: intestinal obstruction (6 episods), intestinal invagination (4 episodis), orquitis (3 episodis) and chylothorax. All patients showed IgD levels > 100 U/mL (100-1500). MVK mutations: 6 patients homozygotes (2: V250I, 4: V377I) and 5 patients double heterozygotes: 4(I268T, V377I), 1(N205D, R388X). The genetic study of the whole family was performed in 7 families. Corticoid treatment used for 100% of the patients. Anakinra in 45% (parcial-complete response), Canakinumab 82% (complete response), Etanercept 9% (no response) and adalimumab 9% (hydradenitis). Long-term manifestations: 2 patients with cutaneous abscess and 1 patient with hydradenitis suppurativa.

Conclusion: In this cohort the high response to canakinumab treatment is shown and late clinical manifestations are described, to our knowledge, for the first time in literature.

Disclosure of Interest

None Declared

P1013 Novel assay to diagnose and monitor cryopyrin associated periodic syndromes (CAPS)

Fortunata Carbone1,2, Luca Cantarini3, Teresa Micillo4, Maria Alessio5, Alma Nunzia Olivieri6, Maria Francesca Gicchino7, Antonella Insalaco8, Maria Cristina Maggio9, Orso Maria Lucherini3, Roberto Scarpioni10, Matteo Piga11, Maria Maddalena Angioni11, Laura Obici12, Antonella Simpatico3, Pietro Leccese13, Rita Consolini14, Raffaele Manna15, Paolo Sfriso16, Sara Bindoli16, Paola Galozzi16, Ida Orlando3, Sabrina Chiesa17, Marco Gattorno17, Giuseppe Matarese18,19
1Istituto per l'Endocrinologia e l'Oncologia Sperimentale-Consiglio Nazionale delle Ricerche (IEOS-CNR), Napoli; 2Unità di Neuroimmunologia, IRCCS Fondazione Santa Lucia, Roma; 3Research Centre of Systemic Autoinflammatory Diseases, Behçet's Disease Clinic and Rheumatology-Ophthalmology Collaborative Uveitis Centre, Department of Medical Sciences, Surgery and Neurosciences, University of Siena, Siena; 4Dipartimento di Biologia, Università di Napoli “Federico II”; 5Department of Translational Medical Sciences, Section of Pediatrics, Federico II University; 6Dipartimento della Donna, del Bambino e di Chirurgia Generale e Specialistica; 7Dipartimento della Donna, del Bambino e di Chirurgia Generale e Specialistica, Università degli Studi della Campania “Luigi Vanvitelli”, Napoli; 8Department of Pediatric Medicine, Division of Rheumatology, Bambino Gesù Children's Hospital, Roma; 9Ospedale dei Bambini “Di Cristina” , Palermo; 10Ospedale AUSL “Guglielmo da Saliceto” , Piacenza; 11Reumatologia, Policlinico Universitario, Cagliari; 12Centro per lo Studio e la Cura delle Amiloidosi, Fondazione IRRCS, Policlinico San Matteo, Pavia; 13Rheumatology Institute of Lucania (IReL), Rheumatology Department of Lucania, “San Carlo” Hospital of Potenza and “Madonna delle Grazie” Hospital of Matera, Potenza; 14Laboratory of Immunology, Division of Pediatrics, Department of Clinical and Experimental Medicine, University of Pisa, Pisa; 15Centro delle febbri periodiche e malattie rare, Policlinico Gemelli, Università Cattolica Roma, Roma; 16Unità di Reumatologia, DIMED, Policlinico Universitario, Padova; 17Dipartimento di Scienze Pediatriche Generali e Specialistiche, IRCCS Istituto Giannina Gaslini, Genova; 18Dipartimento di Medicina Molecolare e Biotecnologie Mediche, Università di Napoli Federico II; 19Istituto per l'Endocrinologia e l'Oncologia Sperimentale, Consiglio Nazionale delle Ricerche (IEOS-CNR), Napoli, Italy
Correspondence: Fortunata Carbone

Introduction: Cryopyrin associated periodic syndromes (CAPS) are rare autoinflammatory disorders associated with dominantly gain-of-function mutations in the NLRP3 gene that result in overactivation of the inflammasome, increased secretion of interleukin (IL)-1beta and IL-18, and systemic inflammation. It has been reported that oligomeric particles of the adaptor ASC (apoptosis-associated Speck-like protein with a caspase-recruitment domain) are released together with IL-1beta and active caspase-1 subunits after activation of the inflammosome complex and that patients with CAPS show an increased serum concentration of ASC+ particles.

Objectives: The diagnosis of CAPS is a critical factor due to both the lack of specific laboratory results and the sharing of similar clinical manifestations with other autoinflammatory diseases, our aim is to develop a simple assay to evaluate the levels of ASC particles in the serum of CAPS patients to provide novel biomarkers facilitating early disease diagnosis and able to monitor treatment responses.

Methods: We developed an ELISA for the quantification of ASC particles in serum and plasma of normal and pathological subjects. We analysed samples from CAPS patients and from patients with autoimmune disorders (Multiple Sclerosis (MS), Type 1 Diabetes (T1D) and juvenile idiopathic arthritis), to confirm that ASC presence in the serum is not due to other chronic inflammatory processes characterizing autoimmunity. In addition, we also evaluated the concentration of ASC in the sera of TNF receptor–associated periodic syndrome (TRAPS) patients to reinforce the concept of specificity of this biomarker in CAPS patients and not in individuals suffering from others inflammatory disorders.

Results: We observed that untreated CAPS patients are characterized by the presence of a significant higher amount of ASC particles when compared with healthy controls (HS) and with patients suffering from MS and T1D. This tendency was also evident in patients with arthritis and TRAPS even if the difference was not statistically significant due to the small number of samples. In addition there is a tendency through a reduction of ASC levels in CAPS patients after pharmacological treatment, which require future investigations.

Conclusion: These data suggest that ELISA quantitation of ASC protein could represent a novel and additional strategy for the diagnosis and monitoring of CAPS.

Disclosure of Interest

F. Carbone: None Declared, L. Cantarini: None Declared, T. Micillo: None Declared, M. Alessio: None Declared, A. N. Olivieri: None Declared, M. F. Gicchino: None Declared, A. Insalaco: None Declared, M. C. Maggio: None Declared, O. M. Lucherini: None Declared, R. Scarpioni: None Declared, M. Piga: None Declared, M. M. Angioni: None Declared, L. Obici: None Declared, A. Simpatico: None Declared, P. Leccese: None Declared, R. Consolini: None Declared, R. Manna: None Declared, P. Sfriso: None Declared, S. Bindoli: None Declared, P. Galozzi: None Declared, I. Orlando: None Declared, S. Chiesa: None Declared, M. Gattorno: None Declared, G. Matarese Grant / Research Support from: Giuseppe Matarese reports research grants from Merck-Serono, Biogen Idec, Novartis and IBSA.

P1014 An Italian family with FCAS

Maria Carrabba1, Marina Zarantonello2, Isabella Ceccherini3, Giovanna Fabio1
1Internal Medicine - Rare Diseases Unit, Fondazione IRCCS Ca’ Granda Ospedale Maggiore Policlinico; 2Department of Clinical Sciences and Community Health , Università degli Studi, Milan; 3UOC Genetica Medica, UOS Diagnostica Molecolare e Malattie Ereditarie, Istituto Giannina Gaslini, Genoa, Italy
Correspondence: Maria Carrabba

Introduction: The prevalence of Cryopyrin-Associated Periodic Syndrome (CAPS) is estimated at about one per million. The Familial Cold Autoinflammatory Syndrome (FCAS) presents in about 95% of the patients by 6 months with cold-induced, urticaria-like rash, fever, and arthralgia. FCAS causes lifelong debilitating effects that restrict patients’ daily activities. Diagnostic delay related to lack of knowledge in Autoinflammatory Diseases is still an important problem.

Objectives: To report an Italian family with FCAS successfully treated with anti-IL1.

Methods: Three subjects, members of the same family, were screened by an experienced doctor. Clinical and laboratory variables, Brain-CT scan, X-ray, audiometry and lung function tests were performed. A targeted review of clinical feature for Autoinflammatory diseases and standardized questioning for CAPS-associated symptoms was conduct. Genetic testing for the NLRP3 mutation was performed.

Results: A 51 years old woman presented with a long history of maculopapular rash after cold exposure starting in early childhood associated with low-grade fever, higher during childhood. During cold months, she needed daily FANS because of fever, and topical steroids for skin rash, prescribed by dermatologists. The patient was in chronic therapy with local steroids because of recurrent ulcerative keratitis. Some corneal scars are present as outcomes. She experienced ocular manifestation on exposure to cold and summertime in contact with air conditioning. She has a 18-years daughter and a 15-years son with a long history of maculopapular rash after cold exposure starting in childhood associated with low-grade fever, fatigue, headache and arthralgia (more severe in the son) lasting less than 24hours. The two children had the same ocular manifestations of the mother and underwent local steroids therapy 4-5 times per year. They all carried the A439V mutation on NPLR3 gene exon 3. Patients were vaccinated (Pneumococcus, HBV, HAV) before starting treatment for anti-IL1 (canakinumab, a fully human anti-IL-1β monoclonal antibody, at a dose of 150 mg subcutaneous every 8 weeks). The Auto-Inflammatory Disease Activity Index (AIDAI, the simplified items scored 0–1 (CAPS range 0–155)) and the Autoinflammatory Disease Damage Index (ADDI) scores were assessed before every treatment. Before treatment, the AIDAI score calculated during a winter and summer month was respectively 59 and 5 for the mother, 30 and 3 for the daughter and 40 and 3 for the son. After 12-months of treatment, the AIDAI score is zero for the mother and the daughter as well as the ADDI score. During treatment mother experienced a urinary tract infection and the daughter two distinct infectious episodes: one of pharyngitis and one of diarrhoea. The son delayed his treatment because of a severe chronic sinusitis that needed surgical treatment. He just received his first dose last month.

Conclusion: The NLRP3 mutation A439V was first entered into the Infevers database in 2002 by Hal Hoffman. It is one of the most common mutations in multiplex families with CAPS and it is considered a CAPS disease-causing mutation, although the overall clinical phenotype is rather mild. Most of the patients reported with FCAS carrying A439V have ocular manifestations (conjunctivitis, keratitis, uveitis) that causes lifelong debilitating effects that restrict patients’ daily activities, as well as rash or low-grade fever or myalgia/arthralgia.FCAS-associated ocular manifestations need treatment with anti-IL1, which typically results in dramatic improvement in clinical and laboratory measures of inflammation, and is well tolerated. Unfortunately, ophthalmologists, dermatologist as well as other specialists do not recognise FCAS and do not appropriately treat this disease nor send patients to Autoinflammatory Diseases experienced Centres.

Consent for publication has been obtained from patient


Disclosure of Interest

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P1015 Scores assessment for clinical management of familial Mediterranean fever

Maria Carrabba1, Marina Zarantonello2, Giovanna Fabio1
1Internal Medicine - Rare Diseases Unit, Fondazione IRCCS Ca’ Granda Ospedale Maggiore Policlinico; 2Department of Clinical Sciences and Community Health, Università degli Studi, Milan, Italy
Correspondence: Maria Carrabba

Introduction: Clinical spectrum of Familial Mediterranean fever (FMF) is heterogeneous, ranging between minimal activity of few affected sites and excellent response to colchicine and large number of frequent, intolerable, treatment-resistant attacks. Frequent and severe FMF attacks may result in serious complications. Severity scoring systems for FMF have been developed and validated in adults (1998 Pras et al., 2005 Mor et al.) to objectively quantify the disease severity for both therapeutic and prognostic purposes. In 2016 the ISSF score has been developed with a consensus-driven methodology by paediatricians and internists, with expertise in this disease, and validated in a large database comprising both children and adults with FMF. Recently, the ADDI score, which was developed based on consensus building, purposes to measure the chronic damage by Autoinflammatory Diseases.

Objectives: This study aims to assess the performance of the existing severity scores (Mor 2005 and ISSF 2016) for FMF in predicting the therapeutic outcome (colchicine dosage and residual attacks); and to assess the relation between the above scores and the ADDI score.

Methods: All the patients with FMF in charge since 2003 have been enrolled. Severity scores have been calculated at the diagnosis and ADDI has been calculated at the latest visit. ROC curves and statistical analysis have been performed.

Results: Forty-five consecutive patients affected by FMF (follow-up 1-41 years) with a median age of 39 years (range 9-89) have been evaluated. Ten patients were diagnosed for FMF in childhood and 35 in adulthood, with a median of diagnostic delay of 12 years (range 1-45). Eight patients are homozygous carries of one mutation on MEFV gene exon 10, twelve patients are heterozygous carriers of two mutations on MEFV gene exon 10, and twenty patients are heterozygous carriers of one mutation on MEFV gene exon 10.

All patients except four are under therapy with colchicine. Nine patients need more than 1mg per day of colchicine. Ten patients reported one or two mild FMF attacks during the year before the last visit. According to the severity score of Mor et al., 15 patients were stratified as mild disease, 20 as intermediate and 10 as severe. According to the ISSF score, 37 patients had a mild disease, eight intermediate and no one severe. Sixteen patients have a positive ADDI score (range 0-4). The ROC analysis shows that the score of Mor performed better than the ISSF (AUC 0.903vs0.661) to identify patients needing more than 1mg of colchicine per day. The ISSF has a good performance (AUC 0.694) to identify patients referring one or two mild attacks in the year before the last visit. The ROC curves analysis shows that both the Mor (AUC 0.677) and the ISSF (AUC 0.657) scores can identify patients with chronic damage (positive ADDI score). ROC analysis of Mor, ISSF and ADDI scores for FMF genetic pattern, showed that only ADDI has a good performance (AUC 0.811) to discriminate the FMF homozygous patients.

Conclusion: The score of Mor et al. can be useful for prediction of FMF patients needing higher colchicine dosage. The ISSF score seems to predict better patients with one or two mild attacks per year. The most of homozygous FMF patients have a positive ADDI score despite therapy.

Disclosure of Interest

None Declared

P1016 Single center experience with 402 familial Mediterranean fever patients

Ozlem Ozdemir Isik, Senem Tekeoglu, Duygu Temiz Karadag, Ayten Yazici, Ayse Cefle
Department of Internal Medicine Division of Rheumatology, Kocaeli University Faculty of Medicine, Kocaeli, Turkey
Correspondence: Ayse Cefle

Introduction: Familial Mediterranean Fever (FMF) is an autosomal recessive genetic disorder that causes recurrent episodes of fever, poliserositis, arthritis, skin eruptions.

Objectives: In this study, we aim to present clinical and demographic features of FMF patients followed in our clinic.

Methods: The clinical, demographic, genetic features and management of 402 FMF patients (fulfilling Tel-Hashomer Diagnostic Criteria) were analyzed.

Results: The mean age was 36,8±11,2 (10-71), the mean diagnosis age was 28±11,9 (3-66) years, and mean duration of disease was 189±125 months. Mean duration between disease onset and treatment was 93.6±104 months. 43% (174) of the patients had positive family history for FMF, 7% (29) had consanguineous marriage in family. 24% of the patients had appendectomy. Fever and abdominal pain both were initial symptoms in 72% of the patients, while 7% of them had chest pain, 4% had only fever, 15% had arthritis, 1% had erysipelas like erythema (ELE) as the first symptom of FMF.

Fever was observed 76% of the patients, abdominal pain in 86%, ELE in 13%, chest pain in 21%, and arthritis in 32%. The frequency of monoarthritis was 21%, oligoarthritis 10,5% and polyarthritis was identified 0,5% of the patients. Ankle arthritis was the most frequent(20%) one among the patients who had monoartritis. 16% of the patients had inflammatory back pain, while 11% of the patients were identified with sacroiliitis. 8 patients (2%) suffered from chronic kidney disease and 2 of them were on dialysis programme. Amyloidosis (diagnosed with biopsy) was identified among 14 patients (3,5%).

At least one mutation of MEFV gene was identified in 78% of the patients. No mutation could be identified in 8%. MEFV gene analyses was not performed in 14% of the patients. The most frequent mutation was M694V mutation and its allel frequency was found to be 54%. For V726A, M680I, E148Q, R761H and A744S allels, mutation frequency were in order of 11%, 7%, 7%, 2%, 1%. The frequency of compound heterozygositywas 38% and the most common genotype wasM694V/R202Q (11%).

There was a significant relationship between M694V mutation and arthritis, ELE, amyloidosis and sacroiliitis (p<0.001). Amyloidosis was more frequent in patients with M694 homozygous mutation. Mean age of disease onset was lower in patient who had M694V homozygous mutation than M694V heterozygous mutation (p<0.001).

Conclusion: The most common mutation in our patients is M694V mutation and it is significantly associated with arthritis, ELE, sacroiliitis, and amyloidosis. Tight control and regular treatment are important in FMF patients to protect from amyloidosis.

Disclosure of Interest

None Declared

P1017 Testicular ischemia in deficiency of adenosine deaminase 2 (DADA2)

Katherine Clarke, Cathy Campbell, Ebun Omoyinmi, Ying Hong, Muthana Al Obaidi, Neil Sebire, Paul Brogan
Paediatric Rheumatology, Great Ormond Street Hospital, London, United Kingdom
Correspondence: Katherine Clarke

Introduction: Deficiency of adenosine deaminase 2 (DADA 2) is a rare autosomal recessive autoinflammatory condition. Recognised features include vasculitis predominantly affecting medium sized vessels, livedoid skin rash, central and peripheral nervous system involvement, variable degrees of immunodeficiency, and marrow failure amongst other clinical presentations. We present the case of a six year old male with DADA 2 who presented with acute testicular ischaemia secondary to vasculitis, the first such description in DADA2.

Objectives: We wish to highlight the ever-expanding phenotype of DADA2, and to emphasise the ongoing controversies regarding management of this lifelong genetic disease. Improvements in our understanding of the pathogenesis will ultimately lead to better biomarkers, better treatments, and ultimately even cure using gene therapy based on the favourable clinical outcomes of patients undergoing allogeneic HSCT thus far.

Methods: A six year old male presented acute right-sided testicular pain. His history included transient infantile neutropenia, resolved hepatosplenomegaly, and longstanding livedo racemosa, leading to screening and confirmation of DADA2 caused by homozygous c.139G>C(p.G47R) mutation of ADA2. As his only clinical feature was that of mild livedo racemosa with normal laboratory parameters at diagnosis, he was being actively monitored prior to starting any treatment. At a routine clinic follow-up a 24 hour history of testicular pain was noted on systems review; he was afebrile, and his only physical signs were that of moderate livedo racemosa, and tenderness of the right testicle. Laboratory parameters revealed C-reactive protein 8mg/L (reference range [RR]<20mg/L); erythrocyte sedimentation rate 28 mm/hr (RR<10); and serum amyloid A 5mg/L (RR<10). Ultrasound-scan of the scrotum revealed significantly reduced perfusion of the right testes, without torsion; and surgical scrotal exploration confirmed testicular ischaemia without torsion. Histology demonstrated ischaemic seminiferous tubules with intervening haemorrhage and acute inflammatory cells, consistent with vasculitis of the testis as the cause. He was treated with high dose intravenous methyl-prednisolone followed by a weaning course of oral prednisolone, and subcutaneous adalimumab (anti-tumour necrosis factor alpha, anti-TNFα). Repeat ultrasound-scan 3 weeks later revealed good testicular perfusion, with a small area of focal infarction. At last follow-up (11 months post-event) he remained asymptomatic, on treatment with adalimumab.

Results: Figure 1: Livedo racemosa noted on both lower limbs.

Figure 2: Doppler study of both testes (transverse views) showing globally reduced (but not absent) perfusion in the right testes compared to the left.

Figure 3: Photomicrograph of testicular biopsy showing patchy areas of tubular necrosis with loss of cellular and nuclear detail and nuclear ‘smudging’ (Right side) with more normal, viable tubules (Left side); H&E original magnification x100

Figure 4: Acute phase markers before, during, and after acute testicular infarction in DADA2. Acute phase responses were completely normal prior to testicular infarction. C-reactive protein (CRP) and serum amyloid A (SAA) remained within the normal range throughout the episode and follow-up. Erythrocyte sedimentation rate (ESR) was normal, but was transiently modestly elevated on the day of tissue infarction (arrowed), remaining normal at follow-up.

Conclusion: The phenotype of DADA2 continues to expand, and we add testicular infarction to the features of DADA2. CRP and SAA cannot be relied on as reliable biomarkers to predict tissue ischaemia and hence who to target for anti-TNFα therapy in DADA2, since these remained steadfastly normal before, during, and after testicular infarction in this case

Consent for publication has been obtained from patient


Disclosure of Interest

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P1018 A novel duplication in the X-linked inhibitor of apoptosis protein gene leading to recurrent hemophagocytic lymphohistiocytosis that is responsive to interleukin-1 blockade

Dilan Dissanayake1, Rebecca Marsh2, Ahmed Naqvi3, Michael Jordan2, Rae Yeung1, Ronald Laxer1
1Paediatric Rheumatology, The Hospital for Sick Children, Toronto, Canada; 2Bone Marrow Transplantation and Immune Deficiency, Cincinnati Children's Hospital, Cincinnati, United States; 3Paediatric Haematology/Oncology, The Hospital for Sick Children, Toronto, Canada
Correspondence: Dilan Dissanayake

Introduction: We present here a 9 year-old male with recurrent episodes of hemophagocytic lymphohistiocytosis (HLH), characterized by fever, cytopenias, transaminitis, hyperferritinemia, hypofibrinogenemia, and elevated levels of soluble CD25 and soluble CD163, in whom we investigated for an underlying genetic defect.

Methods: Peripheral blood samples were obtained from the patient and his family members for analysis using next generation sequencing by gene panels commonly associated with HLH and recurrent fever syndromes, followed by whole exome and whole genome sequencing.The patient’s blood was also assessed using an enzyme-linked immunosorbent assay for serum interleukin (IL)-18. Intracellular X-linked Inhibitor of Apoptosis Protein (XIAP) levels were measured by flow cytometric analysis of peripheral blood mononuclear cells (PBMC).The function of XIAP was assessed by a previously established assay that measures intracellular cytokine staining for tumor necrosis factor alpha (TNFa) production following stimulation of PBMC by muramyl dipeptide (MDP), the ligand for the Nucleotide-binding Oligomerization Domain-containing protein 2 (NOD2) receptor.

Results: The HLH and recurrent fever syndrome gene panels, as well as whole exome sequencing were initially unable to identify a contributory variant in this patient.However, we strongly suspected a primary form of HLH after having measured persistently elevated levels of serum IL-18 to 2524 pg/ml in between episodes of HLH.Furthermore, we discovered a family history of recurrent fevers in the maternal grandfather, which was suspicious for an X-linked condition, such as XIAP deficiency. While awaiting results of whole genome sequencing, we performed flow cytometry for XIAP, which demonstrated reduced protein levels in the patient’s T-lymphocyte, B-lymphocyte and natural killer cell populations.Subsequent results from whole genome sequencing demonstrated a large duplication spanning three exons within the XIAP gene, which was present in the patient, his asymptomatic mother, and his maternal grandfather. In addition to decreased protein expression, MDP stimulation of the patient’s PBMC confirmed a significant functional defect in XIAP function downstream of the NOD2 receptor.While the above testing was being completed, the patient was started on anakinra and successfully weaned off steroids, with no further recurrences of HLH to date.

Conclusion: This case describes the identification of a previously undescribed large duplication within the XIAP gene, which was initially not identified on gene panels and whole exome sequencing.This genetic variant is associated with reduced expression of XIAP in peripheral blood mononuclear cells, as well as decreased function downstream of the NOD2 receptor.Interestingly, while patients with XIAP deficiency are typically treated with hematopoietic stem cell transplantation, this patient has been successfully maintained on anakinra, suggesting an IL-1 dependent link between XIAP and recurrent HLH. We thereby illustrate the utility of combining genetic and molecular tools for the identification of contributors to unexplained autoinflammatory presentations, and present IL-1 blockade as a potential safe and effective alternative for some patients with XIAP deficiency.

Consent for publication has been obtained from patient


Disclosure of Interest

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P1019 10 year prognosis of patients diagnosed with familial Mediterranean fever

Serdal Ugurlu1, Bugra H. Egeli2, Asli E. Soykut2, Bilgesu Ergezen2
1Division of Rheumatology, Department of Internal Medicine, Cerrahpasa Medical Faculty; 2University of Istanbul - Cerrahpasa, Istanbul, Turkey
Correspondence: Bugra H. Egeli

Introduction: In Familial Mediterranean Fever (FMF), other than amyloidosis factors affecting mortality are being debated. In our previous study, we did not observe any atherosclerotic plaque formation in carotid or femoral artery. We thought that the risk of atherosclerosis did not increase in patients diagnosed with FMF.

Objectives: The aim of this study was to assess the 10 year prognosis and comorbidity of patients diagnosed with FMF who have been treated in our rheumatology clinic.

Methods: The sample group is a subset of 2009 study. In 2009, the patients who already had myocardial infarction or cancer diagnosis were excluded. The patients were interviewed with polar questions of whether they were diagnosed with acute myocardial infarction (AMI), cerebrovascular events, cancer, diabetes, and hypertension.

Results: We studied 71 patients (37 males, 34 females; mean age: 49.66±6.91) with FMF, and 59 patients (24 males, 35 females) in healthy control (HC) group. The gender and age difference between two groups was not found significant.

During 10 year follow-up, 8% of FMF patients had either a cardiovascular or cerebrovascular event comparing to 5% in HC (p>0.05). 3% of FMF patients had a cancer diagnosis comparing to 3% in HC (p>0.05). Even though diabetes mellitus diagnosis rate was higher in FMF patients (15% to 10%), results were still not significant (p>0.05). Hypertension diagnosis was 5% higher in FMF group (p<0.05)

Conclusion: Even though there was a significant increase in hypertension, increased diabetes, cancer, and AMI/Stroke ratio was not found significant when compared to the HCs. Therefore, any cardiovascular and malignancy related comorbidities are not associated with FMF.

Disclosure of Interest

None Declared

Table 1 (abstract P1019).

Prognostic Factors of FMF patients compared with Healthy Controls


FMF 2018, n(%)

HC 2018, n(%)

p value


34 (47.89)

35 (59)







6 (8.45)

3 (5.08)



2 (2.82)

2 (3.39)



9 (14.86)

6 (10.17)



25 (33.78)

10 (16.95)






P1020 In a familial Mediterranean fever prevalent region, are familial Mediterranean fever and Behçet’s disease associated?

Ozgur Alparslan1, Bugra H. Egeli2, Yeltekin Demirel3, Serdal Ugurlu4
1Gaziosmanpasa University, Tokat; 2university of Istanbul- Cerrahpasa, Istanbul; 3Sivas Cumhuriyet University, Sivas; 4Division of Rheumatology, Department of Internal Medicine, Cerrahpasa Medical Faculty, University of Istanbul- Cerrahpasa, Istanbul, Turkey
Correspondence: Bugra H. Egeli

Introduction: The co-existence of Familial Mediterranean Fever (FMF) and Behçet’s Disease (BD) has been questioned. There have been a variety of claims on a common pathogenesis.

Objectives: We intended to report the prevalence of Familial Mediterranean Fever (FMF) and Behçet’s disease (BD) and comorbidity ratio of these two diseases in Sivas, Turkey, a city where FMF is known to be very high.

Methods: Seventy-two primary schools in the center of Sivas participated in the study. A total of 14881 randomized sample children from 6th, 7th, and 8th grades, and also 985 of them with their parents (n: 978) were interviewed. During these interviews, the family tree up to second degree relatives was drawn. The presence of a diagnosis of FMF or BD was questioned. The ones who have a diagnosis were confirmed by contacting the medical centers. The ones who were suspected of a disease were further investigated at Sivas Cumhuriyet University Medical Faculty, Family Medicine Outpatient unit. For each disease a disease related history, physical examination, eye examination and pathergy test for BD were performed when needed.

Results: 985 students, 978 mothers, 953 fathers and 1876 relatives (4792 in total) were included in the study. Only 30 (0.6%) of the sample was diagnosed with FMF, and 3 (%0.06) was diagnosed with BD. One of them had concomitant FMF diagnosis.

Conclusion: The prevalence of FMF in Sivas is higher than Turkey’s prevalence; however, BD prevalence was found very low. According to these findings, it is not easy to conclude that these two diseases share a similar background of pathogenesis.

Disclosure of Interest

None Declared

Table 1 (abstract P1020).

FMF symptoms within the last year




Abdominal Pain






Joint Pain



Chest Pain



Muscle Pain



Erysipelas like erythema



P1021 Does testing for SAA is more beneficial than CRP for the follow-up of patients with FMF?

Oguzhan Selvi, Serdal Ugurlu, Bilgesu Ergezen, Huri Ozdogan
Division of Rheumatology, Department of Internal Medicine, Cerrahpasa Medical Faculty, University of Istanbul - Cerrahpasa, Istanbul, Turkey
Correspondence: Bilgesu Ergezen

Introduction: In order to follow subclinical inflammation and adjust the therapy for an optimal disease control, clinicians seek for readily accessible, affordable and reproducible markers. C reactive protein (CRP) is widely used for this purpose. Some suggest that CRP measures are not conclusive in all cases, especially in initial stages of inflammation. It is suggested that Serum Amyloid A (SAA) may be more reliable and sensitive in predicting an ongoing inflammation.

Objectives: It is aimed to evaluate if SAA and CRP is correlated in M694V homozygous FMF cases and if one is superior to the other with regards to early sensitivity.

Methods: In order to evaluate and to compare the sensitivity of SAA and CRP, 234 measurements from 40 FMF patients with M694V homozygous mutation were obtained during a mean follow-up of 5 months.For the analysis, the folds of normal CRP and SAA values were used for correlation.Serum levels of the given markers were measured with nephelometric kits (normal CRP levels <5 mg/L and SAA levels <6,8 mg/L).

Results: All patients were on prophylactic colchicine. Among 40 patients 1 patient was being treated with tocilizumab, 2 patients with adalimumab, 19 patients with anti-IL-1 regimens. There were a total of 234 measurements of CRP and SAA from 40 patients.A similar significant correlation was found when we tested only the values obtained during 202 attack-free occasions (r = 0,863, p < 0,01). Both acute phase reactants were increased in 169 measurements, while in 15 CRP was high but SAA was normal and in 40 SAA was high however CRP was within normal limits. 13 patients has amyloidosis. A number of 86 CRP and SRR measures, these two parameters were correlated in patients with FMF Amyloidosis (r = 0,818, p < 0,01). The mean increase in CRP of the population was 2,82 ± 4,52 fold, whereas mean increase in SAA was 8,47 ± 15,15 fold of the normal.

Conclusion: According to these results, serial testing of SAA does not provide any additional advantages over CRP. Follow-up with CRP measures particularly in patients with amyloidosis might be adequate when it is considered that SAA is not superior to CRP. Readily accessible and affordable bio-marker CRP seems to be sufficient for follow-up of patients with FMF.

Disclosure of Interest

None Declared

P1022 The pregnancy outcomes in FMF patients who are exposed to IL-1 blockade with anakinra

Bilgesu Ergezen, Serdal Ugurlu, Huri Ozdogan
Division of Rheumatology, Department of Internal Medicine, Cerrahpasa Medical Faculty, University of Istanbul - Cerrahpasa, Istanbul, Turkey
Correspondence: Bilgesu Ergezen

Introduction: Some colchicine resistant and/or intolerant FMF cases as well as some patients who experience severe FMF episodes during pregnancy may require alternative therapies during their pregnancies. An Anti-IL-1 agent, Anakinra is being used for this purpose although is still under investigation of researchers.

Objectives: To assess the safety of anakinra in pregnant FMF patients and its effect on fetal and maternal outcomes.

Methods: Thirteen patients who were exposed to anakinra during their pregnancies were monitored closely for disease activity, side effects, fetal USG and pregnancy outcome.

Results: A total of 13 FMF cases followed in our clinic were exposed to Anakinra during the course of their pregnancies due to severe protracted febrile myalgia in 4, thrombocytopenia in 1 and amyloidosis in 1 and severe attacks during pregnancy in 6. One patient had a single injection of anakinra due to pericarditis at 2nd GW and had a spontaneous abortus at 4th GW. Among 13 patients, 2 are still pregnant and both at 8th GW, one started Anakinra at 5th GW, while the other concieved on Anakinra. One of our patients was reported previously by Lachman et al1. We represent detailed data of 9 pregnancies which we have conclusive data regarding the whole pregnancy as well as the follow up period after birth. The relation of the use of the drug and pregnancy are represented in Table 1.

Conclusion: Anakinra seems to be a very effective alternative in the treatment of protracted febrile myalgia non-responsive to colchicine and corticosteroid therapy, even in pregnant FMF patients who have active disease despite colchicine or intolerant and also can be given transiently during pregnancy and successfully stopped after delivery


1) Lachman HJ, et al. Anti IL-1 therapies and pregnancy outcome. Pediatric Rheumatology 2013, 11 (Suppl 1):A269

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Disclosure of Interest

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Table 1 (abstract P1022).

See text for description


Maternal Age at pregnancy

Anakinra Relation to pregnancy


Weeks at delivery

Gender of the baby/fetus

Mode of the delivery

1st minute APGAR

Follow-up duration after birth (months)

Complications after birth



started at 12th Gestational Week (GW) and used until birth


Birth at 40th GW




34 mo




Conceived on Anakinra (have been on Anakinra since 2012), discontinued at 29th GW, reintroduced at 33th GW due to symptom flare.


Birth at 38th GW




48 mo

Methicillin-Sensitive Staphylococcus Aureus incision-site infection (treated with Tygecycline) in mother



started at 15th GW and useduntil birth


Birth at 38th GW




48 mo

thrombocyte count of the baby was low (23,000/mm3) (After three courses of IVIG, it was increased to 95,000/mm3 and a month later to 269,000/mm3)



started at 16th GW, continued for 6 months following the birth at 31th GW


Birth at 31th GW

2 girls


8 for both babies

42 mo




Started at 23th GW used until birth at 37th GW


Birth at 37th GW




42 mo

Injection site reaction



Started at 32nd GW, used until birth 40 th GW


Birth at 40th GW




37 mo




Started at 2013, conceived under Anakinra. Anakinra was stopped at the first month of pregnancy, she flared and it was reintroduced.


Birth at 38th GW




42 mo




Started at 34th GW, used until still birth at 37th GW

Intrauterine death at 37th GW

Pregnancy terminated at 37th GW




10 mo




Started at 6th GW, still using after birth


Birth at 36th GW




9 mo


P1023 Clinical picture of 7 PAPA patients followed in a single pediatric rheumatologic center

Silvia Federici1, Camilla Celani1, Virginia Messia1, Giulia Marucci1, Christoph Kessel2, Fabrizio De Benedetti1, Antonella Insalaco1
1Division of Rheumatology, IRCCS, Ospedale Pediatrico Bambino Gesu’, Rome, Italy; 2Department of pediatric Rheumathology & immunology, University Children’s Hospital, Muenster, Germany
Correspondence: Silvia Federici

Introduction: Pyogenic sterile arthritis, pyoderma and acne (PAPA) syndrome is an autosomal dominant inflammatory disorder caused by mutations in the PSTPIP1 gene primarily affecting joints and skin. The E250K mutation cause the hyperzincaemia/hypercalprotectinemia syndrome termed PSTPIP1-associated-related proteinemia inflammatory (PAMI) syndrome in which a bone marrow involvement is reported

Objectives: To describe the clinical presentation of 7 PAPA patients followed at a single pediatric rheumatology center

Methods: For each patient clinical and laboratory data were collected from medical charts. PSTPIP1 was sequenced through Sanger Sequencing or targeted resequencing using a customized panel and analyzed with the NextSeq® platform (Illumina)

Results: We describe 7 patients from 4 unrelated families with the E250K mutation in a mother and 2 siblings, the A230T variant in a father and his son and the R405C and D266N respectively in the last 2 unrelated patients. Disease onset occurred within the 7thyear of life in all patients. Patients 3 and 4 (table) presented since the 1st year of life recurrent episodes of fever without any cutaneous or articular symptoms. In both patients inflammatory markers were elevated during fever episodes but persistently negative during well-being not requiring any therapy. The variants described in these patients were not previously reported. However their pathogenic role is supported by the detection of markedly high serum calprotectin levels (>10.000 microg/ml). The predominant feature of patients 1 and 2 was articular involvement with recurrent episodes of arthritis associated to acne. Patient 1 was initially treated with prednisone with good clinical response but relapse of arthritis at discontinuation followed bythe development of a sterile muscle abscess. An anti-TNF drug was started in both patients with complete clinical response. Patient 5 reported severe acne and psoriasis, and recurrent episodes of sterile arthritis. She presented a persistent elevation of acute phase reactants with severe anemia and leukopenia not resolving after splenectomy. His son (pts 6) presented with recurrent episodes of sterile arthritis, hepato-splenomegaly, anemia and neutropenia. Zinc and calprotectin serum levels resulted respectively 728 micromol/l and 2600 microg/ml. IL-1 inhibition determined a complete normalization of inflammatory parameters with no effects on anemia and neutropenia. In patient 6 zinchemia decreased to almost normal value after 4 months of therapy. Patient 7 presented at the age of 4 years a sterile lymphnode abscess. She also presented with splenomegaly and neutropenia with persistent elevation of acute phase reactants. Anakinra was proposed but not administered for poor compliance

Conclusion: The clinical picture of patients carrying PSTPIP1 mutation may be heterogeneous. In our cohort TNF-inhibitors were successfully used in PAPA patients preventing new arthritis episodes and resolving cutaneous manifestation where present. In 2 patients the clinical picture was mild not requiring continuous treatment. One PAMI patient had a good response to IL-1 inhibition, which however, had no effect on hematological manifestations

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Disclosure of Interest

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Table 1 (abstract P1023).

See text for description



Clinical features

Laboratory features

Response to therapy



Recurrent fever, acne, cutaneous abscesses, pyogenic arthritis





Acne, pyogenic arthritis





Recurrent fever





Recurrent fever





Acne, psoriasis, pyogenic arthritis, hepatosplenomegaly

←CRP,ESR,SAA, Zinc, serum calprotectin




pyogenic arthritis,hepato- splenomegaly

←CRP,ESR,SAA, Zinc, serum calprotectin

IL-1 inhibition



Limphnode abscess, hepatosplenomegaly



P1024 Clinical presentation, genetic analysis and IFN-score in patients with undefined interferonopathies

Silvia Federici1, Gianmarco Moneta1, Chiara Passarelli1, Claudia Bracaglia1, Luana Raffaele1, Fabrizio De Benedetti2, Antonella Insalaco1
1Division of Rheumatology; 2Ospedale Pediatrico Bambino Gesù, Rome, Italy
Correspondence: Silvia Federici

Introduction: A group of genetic disorders with a disturbance of the homeostatic control of IFN-mediated immune responses, have been identified (type I interferonopathies). An increased expression of type I IFN regulated genes, IFN signature (IS), is reported

Objectives: To evaluate the correlation between clinical presentation, genetic analysis and IFN-score in 10 pts with undefined interpheronopathies

Methods: Patients with suspected interferonopathy based on the presence of typical clinical manifestations (neurological, muco-cutaneous symptoms), laboratory parameters (complement deficiency, low platelet count, presence of autoimmunity), instrumental abnormalities (cerebral calcification), were screened for the IFN-score. Defined IFN-mediated diseases were excluded. Patients with IFN-score above 10 underwent genetic screening by running a panel of 24 genes involved in interferonopathies

Results: 10 pts followed in a pediatric rheumatology center were included. 7/10 presented with recurrent fever (table). Pts 2, 3 and 7 displayed neurological manifestation, respectively epilepsy, epilepsy and mental retardation and progressive hemiplegia. To note epilepsy in pts 2 might be due to a bilateral intraventricular hemorrhage presented at birth. In pts 1,4 gastrointestinal manifestation resembling inflammatory bowel diseases were described while pts 5,7 and 10 suffered from recurrent abdominal pain, diarrhea and patient 10 from hypertransaminasemia. Half of the patients complained arthromyalgia; arthritis developed in pts 2. Cutaneous involvement presented in pts 1,3,6 respectively with widespread panniculitis of trunk and limbs, aspecific vasculitis and Schonlein Henoch purpura (HSP). Other cutaneous manifestation were urticarial rash (pts2) and an erythematous, desquamative confluent eczema (pts 4). Autoimmunity was detected in 2/10 pts. Pts 4 8 had an immunological defect with recurrent infections. The genetic analysis resulted negative in pts 1 and 7 and is ongoing in patients 5,6 and 8. Patients 2,3,4,9 and 10 carried one mutation in at least one IFN correlated gene not confirming the diagnosis. All patients presented an increased IS ranging from 14 to 172.

Conclusion: An elevated IFN-score represent a useful instrument in clinical practice to classify patients with suspected interferonopaties. It may be an important tool to select pts to be genetically screened with a defined panel of interferonopathies correlated genes. In pts in which the genetic analysis results negative, the presence of a positive IFN-score, may guide clinicians in the management of these patients and may support therapeutic decisions

Disclosure of Interest

None Declared

Table 1 (abstract P1024).

See text for description











Systemic symptoms








FeverFever, Hemophagocytic



Cerebral calcification



Epilepsy, mental retardation






Epatosplenomegaly, aspecific IBD


IBD simil-RCU

Abdominal pain, diarrhoea


Abdominal pain




Arthro-myalgia, , panniculitis

Arthro-myalgia, arthritis, urticarial rash

Arthro-myalgia, vasculitis



erythema polymorphe, HSP




28 16


(90-180 mg/dl)










(10-40 mg/dl)












ANA 1:2.560

Anti ds-DNA 1:640

ANA 1:10.240

Anti dsDNA 1:1280


ANA neg








WBC/ mmc3










Hb g/dl









16,5 9,7










150 56











Genetic analysis





IFIH1: p.Arg374His







P1025 Lesson from eurofever registry after the first ten years of enrollment

Martina Finetti, Ilaria Gueli, Joost Frenkel, Seza Ozen, HelenLachmann, Fabrizio De Benedetti, Isabelle Koné-Paut, Carine Wouters, Paul Brogan, Hermann Girschick, Benedicte Neven, Alberto Martini, Nicola Ruperto, Marco Gattorno, on behalf of the Paediatric Rheumatology International Trials Organisation (PRINTO) and the Eurofever Project
UOSD Centro Malattie Autoinfiammatorie e Immunodeficienze, on the behalf of the Paediatric Rheumatology International Trials Organisation (PRINTO) and the Eurofever Project, IRCCS Istituto Giannina Gaslini, Genoa, Italy
Correspondence: Martina Finetti

Introduction: In 2008 the Paediatric Rheumatology European Society (PReS) promoted an International Project for the study of Autoinflammatory Diseases (AIDs) named Eurofever, whose main purpose is to create a web-based registry for the collection of information in AIDs patients.

Objectives: To assess the impact of the Eurofever Registry on scientific community with particular interest in the geographical coverage, diagnostic delay, access to treatment and pubblications.

Methods: The data analyzed in the study were extracted from the Eurofever registry, which is hosted in the PRINTO website.

Results: Up to date 4175 patients have been enrolled from 62 countries (3843 of them with complete demographic data). Most of patients (72%) are resident in Western Europe, 8% in Central-Eastern Europe, 11% in Southern-Eastern Mediterranean, 2% in South America and 7% in other countries. Compared to the first Eurofever report (Toplak et al, 2012) we have observed an increase of enrolled patients from 1388 to 2651 in Western Europe, from 106 to 313 in Central-Eastern Europe, from 294 to 406 in Southern-Eastern Mediterranean. The median onset age is 4 years (range 1 month – 75 years), the median diagnosis age is 8 years (range 1 month – 78 years). The median diagnostic delay observed in 2012 was 7.3 years (range 0.3–76), from patients enrolled after 2012 it was 1.9 years (range 0-57). Comparing the mean diagnostic delay from 1980 to 2018, we have observed an encouraging constant reduction of period between AIDs onset e diagnosis (from a mean diagnostic delay value of 20 years for patients born before 1980, to a mean value of 1 year for patients born after 2011). Complete information on access to treatment were available in 2430 patients. DMARDs were used in 1031 (42%), biologics in 396 (16%) patients. According to the number of enrolled patients, biologics were used in 361/1782 (20%) of Western europeans, 17/342 (5%) of Central-Eastern europeans, 6/259 (2%) of Southern-Eastern Mediterranean patients. Regarding Eurofever impact on Scientific Community, during this first 10 years the Registry provided 12 papers with more than 800 citations. Detailed analysis of clinical features collected in Eurofever database allowed to perform studies with large cohort of patients, to purpose new classification criteria (Federici et al, 2015), to validate damage and activity score (Piram et al, 2014 and N Ter Haar et al, 2017) and to evaluated genotype/phenotype correlation (Papa et al, 2017).

Conclusion: In the last years we have observed an encouraging increase of involved Countries, with a greater number of patients coming from geographic area poorly represented in the first epidemiologic study of Toplak et al. Eurofever data analysis has confirmed an improvement of diagnostic ability during the last years, with a significant reduction of mean diagnostic delay. Long-term studies will help understand the efficacy and safety of different treatments used in these rare conditions.

Disclosure of Interest

M. Finetti: None Declared, I. Gueli: None Declared, J. Frenkel: None Declared, S. Ozen: None Declared, H. Lachmann: None Declared, F. De Benedetti: None Declared, I. Koné-Paut: None Declared, C. Wouters: None Declared, P. Brogan: None Declared, H. Girschick: None Declared, B. Neven: None Declared, A. Martini Grant / Research Support from: The Gaslini Hospital, which is the public Hospital where AM worked till 31/dec/2018as a full time public employee, has received contributions from the following industries:Abbvie, Ablynx, Aim Group, Amgen, Astrazeneca, Biogen, BMS, Boehringer, Celgene, Emd Serono, GSK, Janssen, Novartis, Pfizer, R-Pharm. This money has been reinvested for the research activities of the hospital in a fully independent manner without any commitment with third parties., N. Ruperto Grant / Research Support from: The Gaslini Hospital, where NR works as full-time public employee, has received contributions (> 10.000 USD each) from the following industries in the last 3 years: BMS, Eli-Lilly, GlaxoSmithKline, F Hoffmann-La Roche, Janssen, Novartis, Pfizer, Sobi. This funding has been reinvested for the research activities of the hospital in a fully independent manner, without any commitment with third parties.,Speaker Bureau of: NR has received honoraria for consultancies or speaker bureaus (< 10.000 USD each) from the following pharmaceutical companies in the past 3 years: Ablynx, AbbVie, Astrazeneca-Medimmune, Biogen, Boehringer, Bristol Myers and Squibb, Eli-Lilly, EMD Serono, Glaxo Smith and Kline, Hoffmann-La Roche, Janssen, Merck, Novartis, Pfizer, R-Pharma, SanofiServier, Sinergie, Sobi and Takeda., M. Gattorno Grant / Research Support from: MG has received unrestricted grants from Sobi and Novartis

P1026 Clinical study of Japanese patients with fever of unknown origin: investigation of mutations in 22 genes related to autoinflammatory diseases

Kyoko Fujimoto1, Yukiko Hidaka1, Yumi Yoshida1, Makiko Hayashi1, Takuma Koga1, Shinjiro Kaieda1, Satoshi Yamasaki2, Tomoaki Hoshino1, Hiroaki Ida1
1Division of Respirology, Neurology and Rheumatology, Department of Medicine, Kurume University School of Medicine; 2Center for Rheumatology, Kurume University Medical Center, Kurume-shi, Fukuoka, Japan
Correspondence: Kyoko Fujimoto

Introduction: Autoinflammatory diseases cause systemic inflammation mainly by innate immune abnormality. It is important to have a diagnosis of autoinflammatory diseases in patients with fever of unknown origin. Examining gene mutations is valuable for the diagnosis of autoinflammatory diseases; however, the frequency of genetic mutations in patients with fever of unknown origin is not reported in Japan.

Objectives: We comprehensively analyzed genetic mutations related to autoinflammatory diseases and clinical features in patients with fever of unknown origin.

Methods: We analyzed mutations of 22 genes related to autoinflammatory diseases as follows: TNFRSF1A, MEFV, NLRP3, MVK, NOD2, IL1RN, NLRP12, PSTPIP1, PSMB8, PSMB9, PSMA3, PSMB4, POMP, NLRC4, PLCG2, HMOX1, CECR1, COPA, TNFAIP3, FAM105B, RNF31, RBCK1,in 84 patients with fever of unknown origin who introduced to our hospital from May 2017 to June 2018. Genetic analysis was performed by the next generation sequencer. Furthermore, we investigated precise clinical features in 53 cases.

Results: Fifteen genes were identified as having novel or rare variants. The most frequent variants were determined in NLRP12 (8 cases, 6 sites) and NLRP3 (8 cases, 4 sites). In addition, missense mutations including genetic polymorphisms were observed in MEFV(66.7%). In 53 cases in which clinical symptomatic investigations were possible, 40 cases (75.5%) had definite periodic fever. Among cases with periodic fever, joint symptoms were observed in 52.5%, and abdominal symptoms were in 35.0%. Colchicine treatment was effective in 37.5%. Sixty-five percent of patients with periodic fever had MEFVmutations, including genetic polymorphisms. Almost half of them had novel or rare mutations in autoinflammatory diseases related genes other than MEFV. The most frequent variants were determined in NLRP3.

Conclusion: Novel or rare variants in NLRP12and NLRP3were noted in patients with fever of unknown origin. Sixty-five percent of patients with periodic fever had MEFVmutations including genetic polymorphism.

Disclosure of Interest

None Declared

P1027 A case of adenosine deaminase 2 deficiency (DADA2) with an uncommon clinical presentation and response to IV IG

Francesca Garbarino1, Roberta Caorsi2, Stefano Volpi2, Alice Grossi3, Isabella Ceccherini3, Marco Gattorno2
1Università Degli Studi Di Genova; 2Clinica Pediatrica Reumatologica, UOSD Malattie Autoinfiammatorie-Immunodeficienze; 3UOC Genetica Medica e UOSD Genetica e Genomica delle Malattie Rare, Ist. Giannina Gaslini, Genova, Italy
Correspondence: Francesca Garbarino

Introduction: DADA2 is an autoinflammatory disease with autosomal recessive inheritance characterized by a heterogeneous clinical phenotype ranging from multisystemic inflammation (fever, polyarteritis nodosa, cerebral stroke, livedo reticularis, gastro-intestinal involvement, peripheral neuropathy etc.) to immune-dysregulation and immunodeficiency (lymphoproliferation, recurrent infections, cytopenia etc.).

Objectives: To extend the clinical spectrum of DADA2 reporting a case of isolated nonspecific systemic inflammatory syndrome associated with slight signs of immune-dysregulation in a patient with a novel ADA2 mutation.

Methods: In a patient with nonspecific inflammatory phenotype associated to susceptibility to viral infections, Next Generation Sequencing (NGS) panel was performed; mutations detected were confirmed by direct sequencing (Sanger analysis). ADA2 enzymatic activity was analyzed in monocyte isolated from the patient and incubated with adenosine and an ADA1 inhibitor.

Results: The girl, adopted and of Asian origin, began to suffer from nonspecific systemic inflammatory symptoms (high persistent fever and arthralgias) at the age of 6 years. In past history recurrent respiratory infections and impaired immunological response to viruses (CMV related hepatitis, measles after vaccination) were reported. After few months the patient developed clinical and laboratory findings of HLH (Hemophagocytic Lympho-Histocytosis), confirmed on bone marrow samples; treatment with intravenous (IV) high dose (HD) steroids was started, with prompt response. During steroids tapering fever and systemic inflammation reappeared; anti-IL1 treatment (anakinra) was not effective. Immunologic assessment demonstrated mild hypogammaglobulinemia and moderate NK deficiency on lymphocyte subsets. HD IV Immunoglobulins (IG) (2 g/kg every month) allowed to achieve a complete control of the clinical picture; the frequency of administration was progressively reduced to every 4 months due to persistent wellbeing. At the age of 9, after switching IG to the substitutive dosage, the patient experienced an Herpes Zoster virus reactivation (requiring prolonged antiviral treatment), followed by the reappearance of the inflammatory phenotype complicated by HLH with neurological involvement (irritability and lethargy), responsive to HD steroids and IG. A later cerebral MRI evidenced a small gliotic area in left Centrum Ovale. After steroids suspension, monthly HD IV IG administrations maintained clinical remission. Further immunological studies confirmed a reduction of NK cells with normal function. Hereditary HLH, Autoimmune Lympho-Proliferative Syndrome (ALPS) and main primary immunodeficiencies were ruled out. Given the clinical picture, a large NGS diagnostic panel (courtesy by K. Botzug, Vienna) for autoinflammatory diseases and immunodeficiencies was performed revealing the homozygous LEU141PRO ADA2 mutation, confirmed by Sanger analysis. Being this mutation novel, an ADA2 enzymatic activity test was performed revealing a complete loss of ADA2 activity. The parents refused anti-TNF treatment and the patient is still on monthly HD IG with a complete wellbeing after 3 years of follow-up.

Conclusion: The current report enlarges the clinical spectrum associated with DADA2 to a persistent unspecific inflammatory syndrome, complicated by HLH. This case further emphasizes the possibility that NGS could unravel unusual phenotypes of already known inflammatory syndromes. Even if further reports are required, the response to high doses IG observed in the present case it is of interest. Even if anti-TNF is the treatment of choice HD IV IG could be a possible treatment in DADA2, especially during the acute phase.

Consent for publication has been obtained from patient


Disclosure of Interest

None Declared

P1028 Non-amyloid kidney diseases and autoinflammatory diseases: report of 20 cases and literature review

Charlotte Borocco1,2, Isabelle Kone-Paut1,2, Alexandre Belot3, Marine Desjonqueres3, Alexandre Karras4, Corinne Miceli-Richard5, Bruno Moulin6, Tim Ulinski7, Jean-Jacques Boffa8, David Buob9, Gilles Grateau10,11, Sophie Georgin-Lavialle10,11
1Pediatric Rheumatology, Bicetre University Hospital; 2CeReMAIA, Le Kremlin-Bicêtre; 3Pediatric Rheumatology, Lyon University Hospital, Bron; 4Department of Nephrology, Georges Pompidou European Hospital; 5Department of Rheumatology, Cochin University Hospital, Paris; 6Department of Nephrology, Civil Hospital, Strasbourg; 7Pediatric Nephrology, Trousseau University Hospital; 8Department of Nephrology; 9Department of Pathology; 10Department of Internal Medicine, Tenon University Hospital; 11CeReMAIA, Paris, France
Correspondence: Sophie Georgin-Lavialle

Introduction: Autoinflammatory diseases (AID) are associated with abnormalities of innate immunity. Patients display recurrent fever and various signs including digestive disorders, cutaneous rashes and joint features. One of the most severe complication of AID is inflammatory amyloidosis (AA) including renal involvement, that can lead to kidney failure. However, other nephropathies exist and can be misdiagnosed.

Methods: This study was a retrospective, multicentric French study carried out by medical societies and national reference centers of AID. All patients with non-amyloid kidney diseases (NAKD) occurring at pediatric or adult age with a genetically proven or not AID were included.

Results: Twenty patients were included: 13 patients with familial Mediterranean fever (FMF), 3 with mevalonate kinase deficiency (MKD), one with TNF-receptor associated periodic syndrome (TRAPS) and 3 with unclassified AID (uAID). Among the 13 FMF patients, 8 displayed homozygous M694V MEFV mutation, the mean age at the occurrence of nephrological symptoms was 28.6 years old. Renal diagnosis were: Henoch-Schönlein purpura (HSP) nephritis (n=3), minimal change disease (n=3), IgA nephropathy (n=2), diabetic nephropathy (n=2), nephroangiosclerosis (n=2) and idiopathic nephrotic syndrome (n=1). Renal treatments were represented by angiotensin-converting enzyme inhibitors (ACEIs) (n=6), then colchicine (n=4; dose adjustment or reinstatement of treatment), corticosteroids (n=4), antihypertensive medication (n=4), dialysis (n=2), levamisol (n=1), kidney transplantation (n=1) and anakinra (n=1). After treatment, 5 patients were in renal remission, 2 patients had a transient proteinuria, 5 patients a persistent proteinuria, 2 patients had a renal insufficiency, and 1 patient was in remission after kidney transplantation.

Among the 3 MKD patients, the mean age at onset of nephrological symptoms was 17.7 years. Renal biopsies found: pauci immune extra capillary glomerulonephritis (n=1), HSP nephritis (n=1) and segmental focal hyalinosis lesions associated with nephrosclerosis (n=1). One patient received a kidney transplant with a relapse after it, one was in pre-transplant status, one was in remission under treatment. They all were treated with anti IL1 agent.

The TRAPS patient displayed rapidly progressive glomerulonephritis and was treated with steroids, etanercept and then anti IL1 agent.

The 3 uAID patients displayed: C3 depositional glomerulonephritis (n=1) an IgA nephropathy (n=2) at a median age of 36 years old.

In our literature review including our cases, we found 124 cases of FMF with NAKD biopsy proven, 4 MKD patients with NAKD, only 1 TRAPS patient and no cryopyrin associated periodic syndrome.

Conclusion: Non-amyloid nephropathies can occur in AID; the most frequent ones being IgA nephropathy and HSP nephropathy. Kidney biopsy is necessary in case of nephrological signs to optimize the treatment and to limit the evolution towards kidney failure.

Disclosure of Interest

None Declared

P1029 The transition from pediatrics to adulthood in auto-inflammatory diseases: comparison of 2 transition strategies among 72 patients

Sophie Georgin-Lavialle1, Pierre Quartier2, Brigitte Bader-Meunier2, Katia Stankovic Stojanovic3, Virginie Avellino3, Samuel Deshayes3, Isabelle Melki4, Alexandre Belot5, Isabelle Kone-Paut6, Gilles Grateau7, Véronique Hentgen8 on behalf of transition group project, CEREMAIA, FAI2R and Transition Working Group, National Reference Center for Autoinflammatory Diseases and AA Amyloidosis
1Internal Medicine, Tenon hospital, AP-HP, Paris; 2Pediatric Rheumatology, AP-HP, Necker Hospital; 3Internal Medicine, Tenon Hospital; 4Pediatric Rheumatology, AP-HP, PARIS; 5Pediatric Rheumatology, HCL, Hopital Mère Enfant, LYON; 6Pediatric Rheumatology, AP-HP, Kremlin-Bicêtre hospital, Kremlin-Bicêtre; 7Internal Medicine, AP-HP, Tenon hospital, PARIS; 8General pediatry, CH Versailles André mignot, Versailles, France
Correspondence: Sophie Georgin-Lavialle

Introduction: Transition is the period during which the young patient will move from the pediatric team to the adult team for the follow-up of his or her chronic disease. It requires perfect coordination between the two medical teams but also the active involvement of the young person and his or her family. A specific transition program increases the chances of good health and well-being and reduces mortality in this age group. We experienced two different transition strategies from two different pediatric wards to a single adult ward specialized in auto-inflammatory diseases (AIDs) and compare the efficacy of the 2 strategies.

Objectives:To compare the effectiveness of the transition process in the two strategies, evaluated by the lost of sight patients avec a median 5 years of follow up.

Methods: Children from 2 pediatric centers were addressed for transition in a single adult ward. The first strategy concerned 53 patients that were to prepare since the age of 14 years old and at the age of 18, they were presented for transition thru a joint consultation with the pediatrician in the adult clinics. The second strategy concerned 19 patients who came directly from pediatrics without the pediatrician for an adult medical consultation; the pediatrician explained bye mail and/or phone the case and sent the patient with his medical file.

Results: Seventy-two young people (29 girls, 43 boys) passed the 10-year transition step between 2007 and 2017. The AIDs encountered were: familial Mediterranean fever (n=46), PFAPA syndrome, mevalonate kinase deficiency (n=6 of each), Still's disease (n=4), TRAPS syndrome (n=3) or another rare MAI (NRLP12 mutation, CAPS, or other rare) (1 to 2 cases each time). A total of 81% young patients came regularly (about once a year). Only 3 patients are currently out of sight (4%); none had a clearly identified single gene MAI requiring very regular follow-up. The young people interviewed are generally satisfied.

Conclusion: No difference was observed in the transition efficacy process when comparing the two transition strategies (joint consultation or not). Joint consultation is experienced as necessary, but our results show that this strategy is not mandatory for a successful transition. When asked, young people are eager to have written documentation about their illness and to have the telephone number and e-mail address of the doctor to contact in case of problems.

These positive results of the effectiveness of a transition program require close collaboration between the pediatrician and the adult physician regardless of the transition strategy chosen. The adult doctor must invest in taking care of young people who are very connected and in case of problems expect a quick response by email most often. Indeed, more than a joint consultation, the involvement of the adult physician seems essential to the successful transition from pediatrics to adult service.

Disclosure of Interest

None Declared

P1030 The longitudinal eurofever project: an update on enrollment

Ilaria Gueli, Martina Finetti, Fabrizio De Benedetti, Jordi Anton Lopez, Maria Alessio, Joost Frenkel, Luca Cantarini, Romina Gallizzi, Judith Sanchez Manubens, Marco Cattalini, Efimia Papadopoulou-Alataki, Rolando Cimaz, Donato Rigante, Alma Nunzia Olivieri, Pavla Dolezalova, Alberto Martini, Nicola Ruperto, Marco Gattorno, on behalf of the Paediatric Rheumatology International Trials Organisation (PRINTO) and the Eurofever Project
UOSD Centro Malattie Autoinfiammatorie e Immunodeficienze, on the behalf of the Paediatric Rheumatology International Trials Organisation (PRINTO) and the Eurofever Project, IRCCS Istituto Giannina Gaslini, Genoa, Italy
Correspondence: Ilaria Gueli

Introduction: In 2008 the Paediatric Rheumatology European Society (PReS) promoted an International Project for the study of Autoinflammatory Diseases (AIDs) named Eurofever, whose main purpose is to create a web-based registry for the collection of information in AIDs patients

Objectives: To implement the Registry with the new recently described AIDs and to increase the collection of longitudinal data

Methods: The data were extracted from the Eurofever registry, which is hosted in the PRINTO website (http://www. From February 2015 we started the longitudinal collection of follow-up data with particular focus on treatment, modification of the clinical picture, onset of complication/adverse events. We have enrolled patients included in the registry up to 28 September 2018.

Results: Up to date 4175 patients have been enrolled (3843 of them with complete demographic information, 1903 M and 1940 F) from 62 countries. For 3356 (87%) patients also clinical data from onset to diagnosis, collected during the first visit performed at referred pediatric or adult center, are available. For each disease the number of enrolled patients is: FMF 1086 pts (951 with complete clinical data); TRAPS 273 pts (256 complete); CAPS 298 pts (279 complete); MKD 205 pts (190 complete); Blau’s disease 49 pts (26 complete); PAPA 42 pts (41 complete); NLRP-12 mediated periodic fever 13 pts (11 complete); DADA2 14 pts (9 complete); DIRA 3 pts (all complete); SAVI 3 pts (all complete); CANDLE 1 pt (complete) and Majeed 4 pts (all complete). Among multifactorial autoinflammatory diseases: PFAPA 676 pts (551 complete); CNO 581 pts (540 complete); Behcet 214 pts (186 complete), undefined periodic fever 368 pts (292 complete) and Schnitzler syndrome 13 pts (all complete). The median onset age is 4 years (range 1 month – 75 years), the median diagnosis age is 8 years (range 1 month – 78 years). Most of patients (3509, 91%) presented disease onset during pediatric age (<16 years), 334 (9%) during adult age (81 FMF, 31 CAPS, 53 TRAPS, 40 CRMO, 12 Schnitzler syndrome and 90 unknown fever). 405 of 3509 (12%) patients with pediatric onset received diagnosis during adult age. The median diagnostic delay is 5 years; diseases with longer diagnostic delay are: NLRP12 (24 years, range 4-76), CAPS (15 years, range 0-77), PAPA (14 years, range 2-57), TRAPS (12 years, range 0-77). 396 patients have been treated with at least one biologic drug, 1031 with DMARDs, 427 with systemic steroid and 686 with others drugs. The most frequent diseases treated with biologic drugs are: CAPS (38%), multifactorial diseases (22%), TRAPS (14%), MKD (11%), rare monogenic (8%: 1 CANDLE, 2 DIRA, 2 NLRP12, 3 Majeed, 8 DADA2 and 14 PAPA), and FMF (7%). Since February 2015, longitudinal visits have been inserted for 477 (12%) patients, with detailed data on treatment and safety.

Conclusion: The enrollment in Eurofever Registry is still ongoing. The analysis of data will improve our knowledge both on the natural history of the single disease and on the efficacy/safety of treatment commonly used in the clinical practice.

Disclosure of Interest

I. Gueli: None Declared, M. Finetti: None Declared, F. De Benedetti: None Declared, J. Anton Lopez: None Declared, M. Alessio: None Declared, J. Frenkel: None Declared, L. Cantarini: None Declared, R. Gallizzi: None Declared, J. Sanchez Manubens: None Declared, M. Cattalini: None Declared, E. Papadopoulou-Alataki: None Declared, R. Cimaz: None Declared, D. Rigante: None Declared, A. N. Olivieri: None Declared, P. Dolezalova: None Declared, A. Martini Grant / Research Support from: The Gaslini Hospital, which is the public Hospital where AM worked till 31/dec/2018as a full time public employee, has received contributions from the following industries:Abbvie, Ablynx, Aim Group, Amgen, Astrazeneca, Biogen, BMS, Boehringer, Celgene, Emd Serono, GSK, Janssen, Novartis, Pfizer, R-Pharm. This money has been reinvested for the research activities of the hospital in a fully independent manner without any commitment with third parties., N. Ruperto Grant / Research Support from: The Gaslini Hospital, where NR works as full-time public employee, has received contributions (> 10.000 USD each) from the following industries in the last 3 years: BMS, Eli-Lilly, GlaxoSmithKline, F Hoffmann-La Roche, Janssen, Novartis, Pfizer, Sobi. This funding has been reinvested for the research activities of the hospital in a fully independent manner, without any commitment with third parties.,Speaker Bureau of: NR has received honoraria for consultancies or speaker bureaus (< 10.000 USD each) from the following pharmaceutical companies in the past 3 years: Ablynx, AbbVie, Astrazeneca-Medimmune, Biogen, Boehringer, Bristol Myers and Squibb, Eli-Lilly, EMD Serono, Glaxo Smith and Kline, Hoffmann-La Roche, Janssen, Merck, Novartis, Pfizer, R-Pharma, SanofiServier, Sinergie, Sobi and Takeda., M. Gattorno Grant / Research Support from: MG has received unrestricted grants from Sobi and Novartis

P1031 Digital brachial index testing as a noninvasive tool in diagnosing peripheral vascular disease in DADA2

Patrycja M. Hoffmann1, Deborah L. Stone1, Karyl Barron2, Cornelia Cudrici3, Alessandra Brofferio3, Anne Jones1, Tina Romeo1, Ivona Aksentijevich1, Daniel L. Kastner1, Amanda K. Ombrello1
1NHGRI; 2NIAID; 3NHLBI, National Institutes of Health, Bethesda, MD, United States
Correspondence: Patrycja M. Hoffmann

Introduction: Deficiency of adenosine deaminase 2 (DADA2), is an autosomal recessive disease caused by biallelic loss-of-function mutations in the ADA2 gene. Deficiency of ADA2 is the first molecularly described monogenic vasculitis syndrome. In some cases, DADA2-associated peripheral vascular disease (PVD) can be severe enough to require amputation.

Objectives: To examine the results of upper and lower digital brachial index (DBI) and to compare upper and lower DBI tests to corresponding upper and/or lower extremity MRA tests in patients with DADA2 who have clinical features of PVD.

Methods: Three out of 45 patients with DADA2 seen at the NIH were found to have moderate to severe PVD. Complete histories were obtained and physical exams were performed. Upper and/or lower extremity DBI tests and MRA exams were performed.

Results: 28 yo male with DADA2 who had triphasic Raynaud’s syndrome and pustular-like lesions, bone resorption, and necrotic tissue of various digits, three of which eventually required partial amputation. DBI of upper and lower extremities showed decreased waveform and pressure in 1st-5th left upper digits, 1st, 3rd-5th right upper digits, 2nd-5th left lower digits and 2nd-5th right lower digits. MRA findings showed a single patent common artery, supplying blood to 2nd-3rd left upper digits, adequate blood flow in right upper 2nd digit and little to no blood flow in right 1st, 3rd-5th digits. Right and left lower 1st digit blood flow was patent. There were numerous collaterals to all lower extremity digits. Thus DBI and angiography were in agreement. DBI results demonstrated a lack of waveform or pressure in digits that showed poor flow on angiography. Based on this data, DBI can be considered as an alternative to MRA if cost or renal insufficiency is a concern.

26 yo female with DADA2 with polyarteritis nodosum and intermittent bilateral 1st toe swelling and pain. Right and left lower DBI showed no waveform or pressure in 1st-5th digits. MRA of lower extremity was limited to medium vessel arteries which were patent bilaterally. Small vessels of the feet were not well visualized because of MRA limitations so digital blood flow could not be analyzed or compared to abnormal DBI results.

39 yo female with DADA2 with multiple chronic ankle ulcerations and Raynoud’s syndrome. Bilateral lower DBI waveform and pressure was normal in the 1st digits bilaterally but there was decreased pressure and waveform in the 2nd-5th digits bilaterally. MRA of lower extremity was limited to medium vessel arteries which were patent bilaterally. Small vessels were not well visualized because of MRA limitations so digital blood flow could not be analyzed or compared to abnormal DBI results.

Conclusion: The complexity of DADA2 increases the need for expanding diagnostic tests, some of which can be long and invasive. In the NIH cohort of 45 patients, 3 were found to have clinical findings of PVD. The upper and lower extremity DBI test helped diagnose patients with moderate to severe PVD. In our 2nd and 3rd patient, the MRA exam was limited to medium vessel blood flow testing and therefore blood flow in the small vessels of the feet were not well visualized.Based on the noninvasive less expensive DBI diagnostic testing, patients with DADA2 can be screened effectivey and in a timely manner and avoid exposure to contrast, especially in the setting of renal deficiency. The additional close monitoring can help expedite diagnosis and treatment to possibly avoid future amputation as was the case in Patient 1.

Consent for publication has been obtained from patient


Disclosure of Interest

None Declared

P1032 The expanding clinical and laboratory spectrum of PAPA syndrome: the NIH cohort

Patrycja Hoffmann1, Amanda K. Ombrello1, Deborah L. Stone1, Karyl Barron2, Anne Jones1, Tina Romeo1, Michele Nehrebecky1, Jae Chae1, Ivona Aksentijevich1, Daniel L. Kastner1
1NHGRI; 2NIAID, National Institutes of Health, Bethesda, MD, United States
Correspondence: Patrycja Hoffmann

Introduction: The dominantly inherited PAPA syndrome is caused by mutations in PSTPIP1. It is one of the least understood of the known monogenic autoinflammatory diseases, both from a pathogenic and treatment perspective. Symptoms include arthritis, cystic acne, and pyoderma gangrenosum. Therapy includes corticosteroids, interleukin-1 receptor antagonists, and tumor necrosis factor inhibitors.

Objectives: To review noninfectious epiglottis and sterile osteomyelitis, rare complications not previously published in PAPA syndrome, and to compare LDH and aldolase levels in patients with PAPA syndrome vs patients with mutation-negative PAPA-like phenotype.

Methods: The NIH cohort of 20 patients with PAPA syndrome and 10 patients with PAPA-like phenotype were reviewed. Comprehensive evaluation was performed. MRI, CBC with differential, LDH, aldolase, CK, CRP and ESR were examined. Patients’ labs were drawn during times of flare and no flare. CK was drawn to ensure there was no inflammatory muscle involvement.

Results: One patient with PAPA syndrome developed severe sore throat. Clinical and radiographic evaluation showed piriform sinus swelling consistent with supraglottitis. CRP and ESR were elevated. WBC was normal. Treatment was started with IV clindamycin and ceftriaxone; doxycycline was added later. Severe sore throat continued and repeat radiographic findings showed ongoing swelling. Clindamycin was stopped.Methylprednisolone and a scheduled dose of golimumab were initiated with significant symptomatic improvement and normalization of radiographic findings, CRP and ESR.

A second patient with PAPA syndrome developed severe right wrist pain and swelling. MRI revealed distal right radial epiphyseal marrow abnormalities consistent with osteomyelitis without synovitis. ESR and CRP were elevated. WBC was normal. The patient was treated with clindamycin without symptomatic benefit or improvement in MRI findings. Clindamycin was discontinued. Methylprednisolone and anakinra were initiated which resulted in decreased pain and improvement in marrow inflammation on MRI. CRP and ESR improved.

Of the 20 patients with PAPA syndrome, LDH, aldolase, and CK levels were drawn. LDH was elevated in 19/20 patients. Aldolase was elevated in 19/19 patients. CK was elevated in 5/20 patients. Of the 10 patients with PAPA-like phenotype, LDH was elevated in 1/10 patients. Aldolase was elevated in 2/10. CK was elevated in 1/10. MRIs done on two patients with PAPA syndrome did not show muscle inflammation during a flare.

Upon further analysis of LDH isoenzymes I-V tested in 12/20 patients with PAPA syndrome, 12/12 had elevated LDH isoenzyme V, a skeletal muscle isoenzyme.

Conclusion: Our findings indicate an expanding clinical spectrum in PAPA syndrome that includes aseptic supraglottitis and non-bacterial osteomyelitis. Improvement in clinical symptoms, MRI findings, and acute phase reactants as well as a normal WBC on methylprednisolone support an inflammatory rather than infectious process.

We do not have a clear understanding as to the relevance of elevation in LDH and aldolase in patients with PAPA syndrome and overall normal LDH and aldolase in the PAPA-like phenotype during periods of flare and no flare. LDH isoenzyme V and aldolase are markers of skeletal muscle involvement. In looking at muscle MRIs and obtaining CK levels, we did not see any muscle inflammation. Therefore, in addition to pursuing further testing to rule out muscle damage, additional etiologies for elevation of LDH and aldolase should be considered.

Disclosure of Interest

None Declared

P1033 Pseudodominant inheritance of Behçet-like autoinflammatory disease associated with TNFAIP3 (A20) and MEFV mutations in a Turkish family with familial Mediterranean fever

Nobuyuki Horita1, Ahmet Gul2, Ivona Aksentijevich1, Daniel Kastner1, Elaine Remmers1
1NIH, Bethesda, United States; 2Istanbul University, Istanbul, Turkey
Correspondence: Nobuyuki Horita

Introduction: Familial Mediterranean Fever (FMF) is an auto-inflammatory disorder that causes recurrent fevers and painful serosal inflammation in the abdomen and pleura as well as synovial inflammation in the joints. Missense M694V, V726A, M694I, M680I and other mutations in exon 10 of the MEFV gene are well known to cause FMF, usually with a recessive mode of inheritance. The MEFV gene encodes pyrin, a key component of the pyrin inflammasome. FMF-associated mutations in pyrin cause dysregulated activation of the inflammasome, leading to activation of caspase, which converts pro-interleukin-1 (IL-1) beta to its active cleaved form.

Objectives: We obtained a Turkish kindred with multiple cases with Familial Mediterranean Fever (FMF) and Behçet's disease (BD)-like manifestations. The index case and her two daughters, all with Behçet-like disease, were previously found to have a TNFAIP3 frameshift mutation. The high frequency of affecteds could be consistent with a dominantly inherited inflammatory disease in this family, although other individuals had clinical features consistent with recessively inherited FMF. We sequenced DNA from members of this family to determine whether the TNFAIP3 frameshift mutation and MEFV variants could explain this autoinflammatory disease pedigree.

Methods: Patients were clinically diagnosed to have FMF or BD. Sanger sequence targeting TNFAIP3 exon 5 and MEFV exon 10 was carried out.

Results: A maternal uncle of the index case and the mother of the index case had compound heterozygous FMF-associated MEFV exon 10 mutations, M680I and R761H. Two daughters of the maternal uncle also had compound heterozygous FMF-associated MEFV mutations, M680I and V726A. The index case and her two daughters had a TNFAIP3 exon 5 frameshift mutation (TNFAIP3 c.799delG, p.Pro268Leufs*19), which is consistent with their HA-20 diagnosis, and also carried a single allele (heterozygous) of the MEFV R761H mutation.

Conclusion: Segregation of BD-like manifestations in a single Turkish family with multiple FMF patients could be explained by co-inheritance of a heterozygous exon 10 MEFV variant and one TNFAIP3 mutation, and contribution of MEFV variants on the BD-like manifestations of HA20 requires further studies. 

Consent for publication has been obtained from patient


Disclosure of Interest

None Declared

P1034 Systematic review of biological treatment of deficiency of interleukin-36 receptor antagonist (DITRA) in children and adolescents

Toni Hospach1, Fabian Glowatzki1, Friederike Blankenburg1, Dennis Conzelmann1, Christian Stirnkorb1, Sandra Müllerschön2, Peter von den Driesch2, Lisa Koehler3, Meino Rohlfs3, Christoph Klein3, Fabian Hauck3
1Olgahospital Stuttgart; 2Klinikum Stuttgart, Stuttgart; 3Dr. von Haunersches Kinderspital, Universität München, München, Germany
Correspondence: Toni Hospach

Introduction: Deficiency of interleukin-36 receptor antagonist (DITRA) is a life threatening autoinflammatory disease caused by autosomal recessive mutations of the IL36RN gene leading to recurrent episodes of generalized pustular psoriasis with systemic inflammation and fever. For this disease no standardized treatment guidelines do exist.

Objectives: To systematically review and analyze the data of biologically treated pediatric DITRA patients.

Methods: For systematic research we made a “pubmed” research using the term “Deficiency of interleukin-36 receptor antagonist”, “Deficiency of interleukin-36 antagonist”, “IL36RN mutation” and “DITRA” with age restriction to 18 years.

Results: Our literature research revealed 13 pediatric patients with DITRA and biolocial treatment. Ten patients were homozygous including six with the p.Leu27Pro, three with the p.Arg10 Argfs* and one with the p.Thr123Met mutation and four were compound heterozygous. We add an unreported DITRA patient with a compound heterozygous IL36RN p.Pro76Leu/pSer113Leu mutation. In total 29 flares in 14 patients were treated with biological agents- targeting IL-1/R, IL-17, IL-12/23 and TNF-α. Complete response was achieved in 15 (52%), partial in 4 (14 %), and no response in 10 (34 %) of the flares. Response rates were heterogeneous among the different agents. While complete/partial/no response with inhibition of TNF-alpha could be achieved in 6 (46%)/3 (23%)/4 (31%), the inhibition of IL-17 and of IL-12/23 led in each 4 flares to a 100 % complete response. IL-1/R inhibition led to complete/partial response in each 1 (13 %) and was not effective in 6 (75%) flares. Of note, the unreported patient was successfully treated with weekly dosed adalimumab.

Conclusion: DITRA is a rare disease that has to be considered in patients with generalized pustular psoriasiswith systemic inflammation and fever. It can be effectively treated with specific biological inhibition of TNF-alpha, IL-12/23 and IL- 17, while anti-IL-1/R treatment seems less effective. Weekly dosed adalimumab appears to be a novel treatment option for pediatric patients. Further reports and studies of biological treated pediatric DITRA patients are warranted for evaluation of optimal treatment.

Disclosure of Interest

None Declared

P1035 Familial Mediterranean fever in Slovakia – clinical and genetic characteristics of Slovak cohort

Milos Jesenak1,2,3, Lenka Kapustova1, Katarina Hrubiskova4, Tomas Dallos5, Peter Banovcin1
1Centre for Periodic Fever Syndromes, Department of Pediatrics; 2Centre for Periodic Fever Syndromes, Department of Pulmonology and Phthisiology, Jessenius Faculty of Medicine, Comenius University in Bratislava; 3Department of Clinical Immunology and Allergology, University Teaching Hospital in Martin, Martin; 4Centre for Periodic Fever Syndrome, 5th Department of Internal Medicine, Comenius University in Bratislava, Faculty of Medicine, University Teaching Hospital; 5Department of Pediatrics, National Institute of Children’s Diseases, Comenius University in Bratislava, Faculty of Medicine, Bratislava, Slovakia
Correspondence: Milos Jesenak

Introduction: Familial Mediterranean fever (FMF) is considered to be a rare disease in the region of Central Europe (CE). True prevalence is still not known and awareness of FMF is very low.

Objectives: Since the data about FMF prevalence are missing, we aimed to find all the FMF cases from the whole area of Slovakia. Then we aimed to analyse the clinical and genetic characteristics of Slovakian FMF patients’ cohort.

Methods: We performed the questionnaire based survey in outpatient clinics for clinical immunology and rheumatology and hospital departments in Slovakia. We collected all the genetically-confirmed FMF patients and invite them for clinical examination in the National Centre for Periodic Fever Syndromes in Martin. They underwent complex clinical and laboratory examination and detailed history analysis.

Results: All together, we detected 53 FMF patients (males 23, 42%), aged 33.64±17.33 years (males 28.93±17.42 years, females 35.39±17.34 years). The age of first symptoms was 12.90±13.37 years and the age of FMF confirmation was 31.30±18.18 years, so the average diagnostic delay was almost 19 years. All the patients had recurrent abdominal pain (100%) which was accompanied by recurrent fever in 50 pts (94.3%). Other reported symptoms associated with FMF were: fatigue (77.4%), arthralgia/arthritis (66.0%), chest pain (56.6%), cervical lymphadenopathy (32.1%), tonsillitis (28.3%), headache (26.4%) and skin rash during flares (15.1%). Pleuritis was confirmed in 18.9%, pericarditis in 11.3% and ascited in 22.6%. 3 patients suffered from renal amyloidosis. 3 pts (5.7%) were homozygotes for pathogenic mutation, 10 (18.9%) compound heterozygotes, 28 (52.8%) with pseudo-AD inheritance and 12 (22.6%) patients carried mutations of unknown or possible benign origin. 79% pts were treated by colchicine (mean dose was 1.03±0.51 mg/day), 10 with anakinra (3 regular, 7 on demand) and 9 by canakinumab. 4 patients were intolerant to colchicine. Isolated elevation of serum amyloid A without concomitant elevation of C-reactive protein between the flares before the initiation of the treatment was observed in 34% pts. In 5 of them (9.4%), the IgD elevation was found. In 7 patients, the origin from FMF endemic regions was confirmed.

Conclusion: This the first complex report about the epidemiology of FMF in Central European Country with presumed low incidence. We confirmed much higher prevalence as was expected. The majority of our patients have pseudo-AD form of FMF. All the available therapeutic options were applied. It is necessary to raise the awareness of FMF and to shorten the diagnostic delay.

Disclosure of Interest

None Declared

P1036 Diagnostic criteria for proteasome-associated autoinflammatory syndromes (PRAASS) including Nakajo-Nishimura syndrome, JMP syndrome and CANDLE syndrome

Nobuo Kanazawa1, Hiroaki Ida2, Noriko Kinjo3, Tomoaki Ishikawa4, Ryuta Nishikomori5
1Department of Dermatology, Wakayama Medical University, Wakayama; 2Department of Medicine, Division of Respirology, Neurology, and Rheumatology, Kurume University School of Medicine, Kurume; 3Department of Pediatrics, University of the Ryukyus Graduate School of Medicine, Nishihara; 4Department of Pediatrics, Nara Medical University, Kashihara; 5Department of Pediatrics, Kyoto University Graduate School of Medicine, Kyoto, Japan
Correspondence: Nobuo Kanazawa

Introduction: Nakajo-Nishimura syndrome (NNS) was described originally as “secondary osteoperiostosis with pernio” in Japanese by Nakajo in 1939 and by Nishimura in 1950, and then reported in English as “a syndrome with nodular erythema, elongated and thickened fingers, and emaciation” in 1985 and as “hereditary lipo-muscular atrophy with joint contracture, skin eruptions and hyper-gamma-globulinemia” in 1993. It was in 2011 that similar syndromes have first been reported from outside Japan, which included “joint contractures, muscular atrophy, microcytic anemia and panniculitis-induced lipodystrophy (JMP)” syndrome and “chronic atypical neutrophilic dermatitis with lipodystrophy and elevated temperature (CANDLE)” syndrome. As PSMB8 mutations have commonly been identified in NNS and these two syndromes, they are now collectively called as “proteasome-associated autoinflammatory syndromes (PRAASs)”. Recently, their responsible genes are expanding to other proteasomal genes and, on the other hand, a similar but distinct proteasome-associated entity “proteasome-associated autoinflammation and immunodeficiency disease (PRAID)” has been proposed. In Japan, NNS is now registered as an officially-recognized intractable disease diagnosed by officially-approved criteria.

Objectives: To define the diagnostic criteria of PRAASs.

Methods: So far reported 30 NNS, 3 JMP and 21 CANDLE/PRAAS cases are reviewed and the temporal diagnostic criteria of NNS are verified for these cases.

Results: Among 8 points of 1) autosomal recessive inheritance (parental consanguinity or familial occurrence), 2) pernio-like purplish rash in hands and feet (appearing in winter since infancy), 3) haunting nodular erythema with infiltration and induration (sometimes circumscribed), 4) repetitive spiking fever (periodic, not necessarily), 5) Long clubbed fingers and toes with joint contractures, 6) progressive partial lipomuscular atrophy and emaciation (marked in the upper part of body), 7) hepatosplenomegaly, and 8) basal ganglia calcification, more than 5 are required for temporal clinical diagnosis of NNS. 80% of NNS, 100% of JMP and 67% of CANDLE/PRAAS cases meet these criteria, while most of NNS cases reported before 1990 and CANDLE/PRAAS cases without any description on pernio-like rash do not meet the criteria. If the point 7) is changed to hepatomegaly and 2 points of 9) microcytic anemia and 10) hyper-gamma-globulinemia are added and more than 6 of the final 10 points are required for the diagnosis, positivity of the new criteria reaches 93% of NNS, 100% of JMP and 76% of CANDLE/PRAAS cases. PRAID cases do not meet these criteria. Genetically, 11 cases of NNS, 3 cases of JMP and 11 cases of CANDLE/PRAAS cases have homozygous or compound heterozygous PSMB8 mutations, while other digenic (PSMB8 + PSMA3, PSMB8 + PSMB4, PSMB9 + PSMB4), compound heterozygous PSMB4 or heterozygous POMP mutations are observed in 8 CANDLE/PRAAS patients. PRAID cases have distinct heterozygous POMP or PSMB9 mutations.

Conclusion: We propose a new diagnostic criteria for PRAASs: clinically, at least 6 points are required among above-mentioned 10 points, and genetically, “definite” when disease-associated proteasomal gene mutation(s) are identified and “probable” even if such mutation(s) are not identified, but when other diseases are differentiated.

Disclosure of Interest

None Declared

P1037 Short term follow-up results of children with familial Mediterranean fever after cessation of colchicine: is it possible to quit?

Ayşe Tanatar, Hafize Emine Sönmez, Şerife Gül Karadağ, Mustafa Çakan, Nuray Aktay Ayaz
Pediatric Rheumatology, Health Sciences University Kanuni Sultan Süleyman Training and Research Hospital, Istanbul, Turkey
Correspondence: Şerife Gül Karadağ

Introduction: To define the characteristics of children with familial Mediterranean fever (FMF) whose colchicine treatment was discontinued and then to compare these features of the patients whose colchicine was restarted with the ones not restarted.

Methods: Sixty-four out of 1786 children with FMF whom colchicine was stopped by the physician or patients/parents own decision were enrolled. These patients were grouped into two as: group 1; children whose colchicine was re-started and group 2; children whose colchicine was not re-started. The demographic, clinical and genetic data were collected and compared between group1 and group 2.

Results: Colchicine was stopped in 59.4% (38/64) by the physician and 40.6% (26/64) of them had stopped colchicine by patients/parents will. Colchicine was ceased at a median of 10.6 (2.120.5) years of age, and attack- and inflammation-free periods of 18.2 (6-148) months. The median follow-up of 64 patients after colchicine cessation was 37.4 (6.4-154.7) months. It was re-started in seventeen patients due to attacks (n=11) or elevated acute phase reactants (n=6), while remaining 47 patients did not require colchicine. The age at cessation of the colchicine was lower (p= 0.04) and duration of colchicine treatment until its cessation was shorter (p= 0.007) in group 1 than group 2.

Conclusion: Even though the results of our study are not satisfactory enough to endorse the hypothesis that colchicine may be discontinued by close follow-up; older age and long duration of colchicine treatment before cessation may be two important features that should be considered in the future studies

Disclosure of Interest

None Declared

P1038 Periodic fever syndromes: a year follow-up of a tertiary pediatric rheumatology outpatient clinic in Turkey

Nuray Aktay Ayaz, Şerife G. Karadağ, Hafize Emine Sönmez, Ayşe Tanatar
Pediatric Rheumatology, Health Sciences University Kanuni Sultan Süleyman Training and Research Hospital, Istanbul, Turkey
Correspondence: Şerife G. Karadağ

Introduction: To define the frequency of patients who were suspected to have periodic fever syndromes (PFS) and their final diagnosis.

Methods: We prospectively evaluated the patients who were initially referred to our department with suspicion of PFS in a year period. These findings cover only ten months results as a preliminary study.

Results: A total of 2317 new patients (1142 male/1175 female) were seen. Among them, 724 patients were referred to evaluate for the presence of PFS. Finally, 553 patients were classified as having PFS. Of those, 444 patients were diagnosed with familial Mediterranean fever, 43 with periodic fever with aphthous stomatitis, pharyngitis, and adenitis, 2 with cryopyrin-associated periodic fever syndromes, and 1 with hyper-immunoglobulin D syndrome. Genetic analyses are still in progress in the remaining 63 patients. Most common MEFV variant was M694V in our patients. The rest of the patients who were suspected to have PFS were diagnosed as follows: gastrointestinal disorders (n= 161), infections (n=6), dysmenorrhea (n=2), immunodeficiency (n=1).

Conclusion: Diagnosing PFS requires a careful evaluation. As our study shows nearly one third of patients referred to our center were not accepted as PFS.A detailed evaluation of the patient’s signs and symptoms and also taking the recurrency of these features into account will help to exclude unnecessary referrals to pediatric rheumatology units. Although recommendations are present for rheumatologists, generating some clear-cut algorithms about PFS may provide a practical approach for general pediatricians while they are evaluating and referring these patients.

Disclosure of Interest

None Declared

P1039 Assessing French liberal pediatricians awareness and referral for reccurent fever syndromes: the fireville survey

Valerian Koskas1, Remy Assathiany2, Sylvie Hubinois3, Corinne Levy4, Marc Koskas5, Isabelle Koné-Paut6
1Pediatric Rheumatology, APHP, University of Paris Sud, Le Kremlin Bicêtre; 2Pediatrics, Liberal Exercise, Issy les Moulineaux; 3AFPA (Association Française de Pédiatrie Ambulatoire), Saint Germain en Laye; 4Association Clinique et Thérapeutique Infantile du Val de Marne; 5Liberal Pediatrician, Saint Maur; 6Pediatric rheumatology, APHP, Bicetre Hospital, Le Kremlin Bicêtre, France
Correspondence: Valerian Koskas

Introduction: Patients with recurrent fevers almost always refer initially to their family (liberal) paediatrician, their general physician and/or the emergency hospital departments. If in most cases, benign recurrent viral infections are in causes, it may underlie rare well–defined disorders such as systemic auto inflammatory diseases (SAID). At present few is known on the degree of awareness of liberal paediatricians on SAID and nothing on how they deal with these patients in terms of further investigations, treatment and referral. FIREVILLE study tries to better understand why patients with SAID undergo complex medical journey before appropriate referral in our country.

Objectives: To survey, liberal paediatricians on theirs knowledge and practices regarding children with recurrent fevers

Methods: We took the facilities of the AFPA (French Association of Ambulatory Paediatrics) to email 1248 active liberal paediatricians between June and September 2018, to answer a Survey-Monkey type questionnaire. The survey, included 36 questions divided into 3 parts; i.e.Socio-demographic characteristics of paediatricians and their knowledge on recurrent fever syndromes (RFS), clinical case analysis, then more general evaluation of knowledge on and analysis of city-hospital networks.


360 paediatricians (28.8%) answered the survey after 4 email reminders. 77% were women aged 44 to 60 years with a completion rate of 79%. 22% had part time of hospital practice. The top 3 regions were: Paris ile de France: 24.9%, Auvergne-Rhône-Alpes: 18.3% and Provence-Alpes-Côte d'Azur f 9,9%. 69% of paediatricians considered their knowledge on HRF, and 83% on SAID. PFAPA was the best known (46%), on the basis of the frequency (92%) and the regularity (89%) of fever. Only 66% considered the duration of fever as an added diagnostic value. The clinical case was a PFAPA chart. 92% of paediatricians required whole blood cell count and 91.8% a CRP. 70% of paediatricians ruled out radiologic evaluations. They were 80.6% to suspect PFAPA and 14% to suspect FMF. 57% asked second opinion, in a paediatric rheumatology unit (46%) and in general paediatrics (35%). Youngest paediatricians chose more significantly a paediatric rheumatology unit (p = 0.009).Only 44% of responders knew a SAID referral centre in their area. Finally, 90% were interested in joined city-hospital care.

Conclusion: This is the first study surveying liberal paediatricians on their knowledge and practices with a child with supposed HRF. In spite of their thought insufficient knowledge, their answers were accurate in most cases, however the questionnaire revealed insufficient knowledge of the dedicated resources and network for SAID.

Disclosure of Interest

None Declared

P1040 Crimean Tatars is new target nationality for the familial Mediterranean fever

Olga Zhogova1, Natalya Lagunova1, Sergey Ivanovskiy1, Evgeny Suspitsin2,3, Mikhail Kostik2
1V.I. Vernadskiy, Crimean Federal University, Simferopol; 2Saint-Petersburg State Pediatric Medical University; 3N.N. Petrov Institute of Oncology, Saint-Petersburg, Russian Federation
Correspondence: Mikhail Kostik

Introduction: Familial Mediterranean Fever (FMF) is a monogenic autoinflammatory disease with high prevalence in some nationalities, such as Jew, Armenians, Turkish, Arabians and other nationalities with Mediterranean origin. Before 2016 there were no data about FMF distribution in the Crimea region, but the first 15 new cases of FMF were diagnosed in the last 2 years.

Objectives: The aim of our study is the evaluation of the distribution of FMF in the Crimea region.

Methods: Our cohort consists of 13 children and 2 adults, among them were 2 parents and 6 kids from 1 family. All belong to Crimea Tatar nationality. This nationality is close to Turkish. The diagnosis of FMF was based on the Tel-Hashomer criteria and later was confirmed by MEFV gene sequence.

Results: 10 children with FMF have M694V heterozygous mutation, 3 have M694V homozygous mutations, and 2 adults (parents) have M694V homo (father) and heterozygous (mother) mutations. Ten children and 2 adults are treated with colchicin. 2 kids and 1 adult (all M694V/M694V) received canakinumab due to inefficacy and 1 child (M694V/N) due to intolerance of increased doses. Clinical characteristic of the studied population are in the table.

Conclusion: Crimean Tatars is a new nationality with an increased prevalence of FMF with typical high penetrance mutations. Further epidemiological studies required about MEFV alleles distribution in the healthy population and in the FMF patients.

Disclosure of Interest

None Declared

Table 1 (abstract P1040).

See text for description


The # of patients

Big criteria

 1. Typical recurrent seizures fever with serositis


 2. Peritonitis


 3. Pleurisy


 4. Monoarthritis (hip, knee, ankle joints)


Small criteria

 1. Stomach


 2. Thorax


 3. Joint


 4. Load pain in legs


 5. A good response to colchicine therapy.


Supporting criteria

 1. The presence of FMF cases in the family history


 2. Belonging to the relevant ethnic group


 3. Age of onset of the disease d 20 years


 4. heavy bedridden


 5. spontaneous resolution attack


 6. the presence of bright gaps


 7. increase in laboratory markers of inflammation


 8. episodes of proteinuria/hematuria


 9. unproductive laparotomy or removal of “white”appendix


 10. parental blood marriage


P1041 New variant in the IL1RN-gene associated with late onset and atypical presentation of DIRA – follow-up

Jasmin B. Kuemmerle-Deschner1, Kerstin Reicherter2, Susanne Schlipf3, Sandra Hansmann1, Anton Hospach4, Ilias Tsiflikas5, Xiao Liu6, Susanne Benseler7, Alexander Weber6
1Department of Pediatrics, Division of Pediatric Rheumatology, University Hospital Tuebingen; 2CEGAT, Tuebingen; 3Kinderarztpraxis Dr. Lakner, Schwäbisch Gmünd; 4Zentrum für Pädiatrische Rheumatologie, Klinikum Stuttgart, Olgahospital, Stuttgart; 5Pediatric Radiology, Department of Radiology, University Hospital Tuebingen; 6Department of Immunology, University of Tuebingen, Tuebingen, Germany; 7Rheumatology, Department of Pediatrics, University of Calgary, Calgary, Canada
Correspondence: Jasmin B. Kuemmerle-Deschner

Introduction: Deficiency of the Interleukin-1 receptor antagonist (DIRA) is an autoinflammatory disease characterized by severe systemic inflammation with bone and skin involvement present in the first days of life. Here we report on diagnosis, treatment and follow up of a novel variant in the IL1RN-gene associated with late onset and atypical phenotype of DIRA.

Objectives: Here we report on diagnosis, treatment and follow up of a novel variant in the IL1RN-gene associated with late onset and atypical phenotype of DIRA.

Methods: A 3 year-old boy presented with recurrent monthly episodes of fever and fatigue, associated with lymphadenopathy, pericarditis, pleuritis, pancreatitis, and arthritis involving sacroiliac, hip, knee and ankle joints in the absence of any skin involvement. Symptoms had started at age one and had progressed over time to life-threatening episodes requiring intensive care therapy. Throughout, inflammatory parameters including ESR, CRP, SAA, S100A8/9, leukocytes and platelet counts were highly elevated. Treatment with colchicine and steroids improved symptoms but did not prevent flares. Typical immune deficiencies were ruled out; genetic testing for FMF, CAPS, TRAPS, HIDS and DITRA did not reveal variants in the associated genes.

Results: Whole exome sequencing detected a novel homozygous stop variant c.62C>G; p.Ser21* in the ILRNgene (NM_173842.2). Mother, father and brother were heterozygous for the same variant. In addition, three variants of unknown significance were identified in the patient's PCGF5, CPA1and SPTA1genes. Functional studies revealed only marginal secretion of IL-1RA in the patient’s unstimulated leukocytes and after stimulation with IL-1βand LPS, confirming the disease-causing nature of the variant.

IL-1 inhibition with anakinra at 2 mg/kg/d was started and resulted in complete resolution of clinical symptoms and signs of inflammation on MRI, in normal inflammatory markers, and dramatically improved energy levels. Intolerance to daily subcutaneous injections prompted a switch to canakinumab at 4 mg/kg/4 weeks. However, as per the patient’s and mother’s assessment of disease activity, canakinumab was inferior to anakinra. After four months a flare occurred, prompting re-start of anakinra and resulting in sustained and complete resolution of symptoms in an observation period of 14 months.

Conclusion: This is the first report of the novel c.62C>G; p.Ser21* variant in the IL1RN-gene primarily causing severe serositis in a homozygous carrier, while heterozygous family members were completely symptom-free. Skin disease, one of the most prominent features in other patients with DIRA was not observed in this patient, while IL-1 inhibition was likewise effective.

The different phenotype in the patient reported here, may be due to the selective loss of secreted IL1RN. Unlike one report of successful canakinumab treatment of one patient with late onset of DIRA [1], our patient did not respond favourably to canakinumab, but treatment with anakinra lead to sustained remission.

1 Ulusoy et al. J Med Case Reports 2015; 9:145

Consent for publication has been obtained from patient


Disclosure of Interest

J. Kuemmerle-Deschner Grant / Research Support from: Novartis, SOBI,Consultant for: Novartis, SOBI, K. Reicherter Employee of: CEGAT, S. Schlipf: None Declared, S. Hansmann: None Declared, A. Hospach Consultant for: Novartis, Chugai-Roche, SOBI, I. Tsiflikas: None Declared, X. Liu: None Declared, S. Benseler Consultant for: Novartis, SOBI, abbvie, A. Weber: None Declared

P1042 Efficacy of anti-IL-1 treatment in familial Mediterranean fever: a single-center experience

Tuba Kurt, Halide O. Basaran, Fatma Aydın, Nermin Uncu, Banu Celikel Acar
Pediatric Rheumatology, Ankara, Child Health Hematology and Oncology Education and Research Hospital, Ankara, Turkey
Correspondence: Tuba Kurt

Introduction: In 5%–10% of patients with familial Mediterranean fever (FMF), colchicine is not effective in preventing inflammatory attacks. Furthermore, another 5%–10% of patients are intolerant to effective doses of colchicine and experience serious side effects. In recent years, it has been shown that interleukin-1 (IL-1) plays a central role in the pathogenesis of FMF.Several reports have pointed out the efficacy of IL-1 blockade in colchicine resistant FMF.

Objectives: To review the patients followed with FMF who received anti-IL-1 treatment, in terms of outcome and side effects

Methods: 18 FMF patients who were treated with anti-IL-1 treatment were retrospectively reviewed with regard to indication, effect on disease activity and acute phase response, adverse events and AIDAI score and patient global assessment. Colchicine resistance was defined as at least one attack per month for three consecutive months and elevated erythrocyte sedimentation rate or C-reactive protein in-between attacks despite taking adequate dose of colchicine.

Results: There were 18 patients with FMF (9 M/ 9 F) who were treated with Anakinra and Canakinumab for various indications (colchicine resistant recurrent febrile attacks in 16, colchicine related side effects in 1, subclinic inflamation in 1). 11 patients were treated with Anakinra while were 15 patients with Canakinumab. All patients except 5 had homozygous or compound heterozygous exon ten mutations. The mean age of onset of anti-IL-1 treatment was 15± 2 (11-18 years) years. The mean duration of the disease was 11.1 ± 4.34 years. All patients were taking adequate dose of colchicine for their age before treatment with a median dosage of 0,03 ±0,012 mg/kg/day before anti-IL-1 treatment (0.03–0.06 mg/kg/day). 11 patients were treated with anakinra with a median duration of 29,7 months (8- 60 months), but 6 of them switched to canakinumab because of noncompliance and side affects (2 headache, 1 urticerial rash), all responded.After the initiation of Anakinra treatment 6 patients became attack-free, 2 patients reported more than 50% decrease, and 3 patients no change in the frequency of the attacks.

15 patients were treated with Canakinumab but 2 (%13,3) of them swiched to anakinra because had increase in frequency of the attacks. All of patients complete respondend and any of them were no side effects.

A significant decrease was observed in the mean CRP (from 8.5± 8.7 mg/dLto 0.68 ± 0.75 mg/dL), WBC (from 10335 ± 3206 mm³ to 7047 mm³± 2108mm³) and ESR levels (from 44.35 ± 18.7 mm/h to 9.5 ± 7.6 mm/h), respectively (referance range for CRP: 0–0.3mg/dL).

AIDAI score decreased from 25±17,5 to 0,33±1 and mean pyscian’s global assessment 7,4±1,5to 1,25±1,1 respectively, under anti-IL-1 treatment treatment.

As for the adverse events, 1 patients (%9) had allergic reactions under Anakinra treatment (severe disseminated rash in 1 patient ) and 2 patients (%18,1) had headache which necessitated termination of treatment in 3 patients. There were no adverse events in the remaining all patients during the course of treatment.

Conclusion: Based on the results of our study, anti-IL-1 therapy seems to be a safe and effective alternative for patients with FMF who do not respond to or cannot tolerate colchicine, however approximately one fourth of the patients stop anakinra for insufficient response and injection site reaction. The treatment should be modified and decided for each patient on an individual basis.

Disclosure of Interest

None Declared

P1043 Erysipelas-like erythema as the primary gripe of FMF

Omer Kuru1, Ahmet Ki̇vanc Cengi̇z2
1Physical Medicine and Rehabilitation, Division of Rheumatology, University of Heath Sciences, Istanbul Okmeydani Research and Training Hospital, Istanbul; 2Physical Medicine and Rehabilitation-Rheumatology, 19 Mayis University Faculty of Medicine, Samsun, Turkey
Correspondence: Omer Kuru

Introduction: Familial Mediterranean Fever (FMF) is a monogenic autoinflammatory disease characterized by recurrent polyserositis attacks, mainly involving peritoneum, pleura and synovium. Erysipelas-like erythema (ELE) is an aseptic neutrophilic dermatose defined as well-demarcated, warm, tender, erythematous and infiltrated plaques which resolves spontaneously in 2-3 days. ELE is a pathognomic cutaneous manifestation of FMF. The reported frequency of ELE is 5-30 % in different populations. It typically developes on the extensor surface of lower extremities especially on dorsum of ankles and feet. It is usually unilateral and associated with more severe FMF clinical phenotype and M694V homozygosity.

Objectives: Here we would like to present a 12 year old girl admitted to the emergency service with an erythematous, painful rash at the dorsum of her foot. This was her fourth admission in approximately 9 monthstime with a similar skin lesion at dorsum of the same foot. In her medical records previous diagnosis for the described lesion were insect bite, contact dermatitis and cellulitis. Oral and topical antibiotics, topical steroids were prescribed in her previous visits. She told that regardless of the medication used the lesions resolved spontaneously in 3-4 days time. She had also noticed that the lesions usully occured after her volleyball matches. Exercise and long time standing usually has caused pain in her legs and if she insists on the activity despite this pain, the lesions occured. She had no prominant abdominal or pleural pain. She didn’t have arthritis but had episodes of fever lasting 36-48 hours accompanying the skin lesions. Her family history was unremarkable except FMF in one of her cousins. Her laboratory tests revealed leukocytosis, elevated crp and fibrinogen levels. She did not have proteinuria.

Methods: Case report

Results: The recurrence of the typical lesion, spontaneous recovery in 3-4 days time, accompanying fever and positive family history reminds the diagnosis of FMF. Genetic analysis was performed and compound heterozygous genotype (M694V/V726A) was detected. Colchicine was prescribed. She is 14 years old now and did not have any typical serositis attack yet. Under colchicine treatment the ELE attacks did not disappear completely but their frequency and severity decreased prominently and fever did not accompany her ELE attacks any more.

Conclusion: Spontaneous recovery, recurrence of the attacks, accompanying fever and elevated acute phase reactants during the attacks points out FMF especially in inhabitants of the Mediterranean basin. Occurence of ELE as the first manifestation of FMF is rare but this possibility must also be kept in mind.

Consent for publication has been obtained from patient


Disclosure of Interest

None Declared

P1044 Analysis of new referrals to a specialist UK adult autoinflammatory disease service

Serdal Ugurlu1, Philip N. Hawkins2, Charalampia Papadopoulou2, Tamer Rezk2, Dorota Rowczenio2, Helen J. Lachmann2
1Division of Rheumatology, Department of Internal Medicine, Cerrahpasa Medical Faculty, University of Istanbul - Cerrahpasa, Istanbul, Turkey; 2National Amyloidosis Centre, Centre for Amyloidosis and Acute Phase Proteins, Division of Medicine, Royal Free Campus, UCL, London, United Kingdom
Correspondence: Helen J. Lachmann

Introduction: Diagnosis of the SAIDs requires a high index of suspicion and previous series has suggested that there are often long diagnostic delays, particularly in TRAPS and MKD.

Objectives: To look at the case mixed referred to a single adult clinic in London specialising in assessment of potential SAIDS over the course of the year of 2017.

Methods: All new referrals were accepted for clinical assessment. At the first visit patients had a full history and examination, genetic testing – varying from single gene to a 20 gene panel depending on clinical features, and laboratory testing including fortnightly blood draws for serial analysis of the hepatic acute phase response proteins, CRP and SAA over a 3 month period.

Patients with a non suggestive history, non contributory genetic testing and no evidence of inflammation accompanying symptoms were felt not to have SAIDS and referred back to their local hospitals for further management. Other cases were diagnosed based on full clinical assessment, other investigations – for example ferritin in AOSD, genetic testing results, serial monitoring of CRP and SAA and therapeutic trials, for example colchicine in presume FMF and anti IL-1 therapies in CAPS and Schnitzler’s syndrome.

Results: 273 new patients were referred. Median age at referral was 37.4 years, the oldest patient was 84.3 years old and 59% were female. 174 (64%) were of northern European ancestry, 68 (25%) were eastern Mediterranean, west Asian or southern European ancestry, 19 (7%) were of south or east Asian ethnicity and 4% were of African or Afro-Caribbean ancestry. 76% of referrals were from hospital specialities. The referral source was: rheumatology 38%, general practitioner 24%, dermatology 8%, immunology 8%, gastroenterology 6%, infectious diseases 3%, clinical genetics 3%, nephrology 2%, haematology 2%, gynaecology 2%, emergency department 1%, respiratory 1%, other 2%.

After work up 135 (49.5%) were felt not to have a SAID as the cause of their symptoms. Of the remaining 138 patients who did have evidence of a SAID the diagnoses made were: FMF 33%, uncharacterised SAID 26%, CAPS 9%, AOSD 8%, recurrent idiopathic pericarditis 6%, Schnitzler’s Syndrome 5%, TRAPS 4%, variant PFAPA 4%, DADA2 1%, MKD 1%, CRMO 1%, Behcets 1%, Cattleman’s disease 1%.

The median interval between reported symptom onset and diagnosis were as follows: 16 yrs for FMF, 28.1 yrs for CAPS, 5.0 years for recurrent idiopathic pericarditis, 4.5 yrs for Schnitzler’s Syndrome, 5.7 yrs for TRAPS, 20.5 yrs for variant PFAPA,12.5 yrs for DADA2, 17 yrs for MKD and 2 years for CRMO.

Conclusion: This series suggests that recognition and diagnosis of the SAIDS remains a challenge. More than 1/3 of referrals were from rheumatology, referrals from primary care were almost exclusively from patients with a known family history of one the inherited syndromes. The wide variety of referring specialities reflects the diverse nature of SAIDS and the importance of almost all specialities considering the possibility of SAIDS. Only just over 50% referrals had evidence of diseases falling within the recognised SAID spectrum and 26% of these have currently uncharacterised disease with non diagnostic genetic testing. Of those in whom a diagnosis could be made there are significant diagnostic delays fortunately despite late initiation of treatment no patients had evidence of systemic AA amyloidosis.

Disclosure of Interest

None Declared

P1045 Application of autoinflammatory disease damage index (ADDI) to other autoinflammatory diseases in a tertiary referral hospital

Mireia Lopez, Estefania Moreno
Pediatric Rheumatology, Hospital Vall d’Hebron, Barcelona, Spain
Correspondence: Mireia Lopez

Introduction: Autoinflammatory diseases (AIDs) cause systemic inflammation that can be chronic and result in damage to multiple organs. Recently, the autoinflammatory disease damage index (ADDI) has been developed and validated to the four most common monogenic AIDS, cryopyrin-associated periodic syndrome (CAPS), familial Mediterranean Fever (FMF), mevalonate kinase deficiency (MKD) and tumor necrosis factor receptor-associated periodic fever syndrome (TRAPS). ADDI could be useful in other AIDs different than the four reported.

Objectives: The aim of the study is to asses the application of ADDI to patients with AIDs followed in our hospital with the four most common monogenic diseases and other AIDs, determine the reliability and point out encountered comments about the scoring.

Methods: All patients with AIDs followed in Transitional Care unit or Pediatric Rheumatology specialized in AIDs consult from Hospital Universitari Vall d’Hebron were identified. A cross- sectional, descriptive study was performed applying ADDI by two pediatric rheumatologists (EM, ML). Laboratory test including C-reactive protein (CRP) mg/dl, amyloid protein (AP) mg/L, erythrocyte sedimentation rate (ESR) mm/h and protein/creatinine rate (mg/g Cr) were performed at the moment ADDI was applied. Variables related with disease duration, current treatment and accumulated corticosteroids treatment was assessed. The continuous data are presented as mean and standard deviation (SD). Categorical variables are presented by percentages.

Results: A total of 41 patients with AIDs were included, of whom 61% were female, with a median age of 20 years (SD 11.9) at inclusion. Disease duration has a mean of 11 years (SD 8.2). AIDs included were 11 patients with FMF (26.8%), TRAPS n=4 (9.8%), MKDn=3 (7.3%), CAPS n= 2 (4,9%), Blau syndrome n= 7 (17.1%), SAVI syndrome n=3 (7.3%), CRMO n=4 (9.8%), PFAPA n=2 (4.9%), APLAID n=1 (2.4%), Stickler syndrome n=1 (2.4%), and 3 unknown AIDs with genetic test negative n=3 (7.3%). Current treatment is variable among patients, 6 (15.8%) are taking disease-modifying antirheumatic drugs (DMARDs), 9 (23.7%) Colchicine, 8 (21.1%) Anakinra, 13 anti-TNF therapy (34.2%), 1 (2.6%) Ruxolitinib and 1 (2.6%) Abatacept.Just 6 patients were receiving corticoids with mean prednisone dose of 7.5 mg/day.

ADDI mean score was 2.3 (SD 2.2) for all patients. Regarding the eight different items evaluated, musculoskeletal domain was the highest punctuated with a mean of 1.02 points, followed by ocular domain with 0.42 points. Laboratory test results were mean ESR 27.2 mm/h (SD 26.7), CRP 0.7 mg/dl (SD 1.3), AP 13.9 mg/L (SD 18.6). Proteinuria was present in 2 patients with mean 286.5 mg/g (SD 246.1). EM and ML applied ADDI in 5-10 minutes average.

Conclusion: ADDI is a feasible index suitable to measure damage in a single patient. Despite it was performed to the four most common AIDs it could be applied to other diseases. In our cohort mean ADDI was low with musculoskeletal item as the highest punctuated. This result could be explained by the correct control of the disease with the current treatment.Laboratory tests also support this finding. Despite good response to medication, it is difficult to reverse bone deformities or joint restriction. Nevertheless, some organ systems are not represented like respiratory, cardiovascular or cutaneous damage, important in some syndromes included in our cohort. Knowing the difficulties of conducting an unified index for all diseases, ADDI must be applied in longitudinal cohorts.

Disclosure of Interest

None Declared

P1046 Vasculitis in autoinflammatory diseases in a tertiary hospital

Rosa M. Alcobendas, Sara M. Loza, Pablo F. Fraga, Clara U. Gascon, Catarina Fervenza, Agustin Remesal
Pediatric Rheumatology, University Hospital La Paz, Madrid, Spain
Correspondence: Sara M. Loza

Introduction: Autoinflammatory diseases with vasculitis and interpheronopaties have been recently described and may present with variable clinical signs. Such entities are considered rare diseases and potentially life-threatening. Clinicians should be aware of the existence of autoinflammatory vasculitis to diagnose and treat them correctly.

Objectives: To describe the initial manifestations, laboratory findings and therapeutic approaches of patients diagnosed with autoinflammatory vasculitis during the last 5 years in a pediatric rheumatology unit of a tertiary hospital.

Methods: Retrospective chart review. Inclusion criteria were: children under 18 years diagnosedwith monogenic autoinflammatory vasculitis in a tertiary hospital.

Results: During the study period, 5 patients were identified (2 boys and 3 girls). The main clinical manifestations were severe cutaneous lesions (5/5), intermittent fever (4/5) and neurological involvement (4/5) (see table 1).The only patient without neurologic manifestations had relatives with psychomotor retardation and sensorineural symptoms of unknown etiology. All patients were diagnosed of with another rheumatology condition,with no response to conventional treatment.

Conclusion: Autoinflammatory vasculitis are extremely rare entities but potentially fatal. The early age of onset, the intense skin involvement as well as the presence of affected relatives may be suggestive data. Clinical suspicion is important in order to establish an early and adequate treatment.

Disclosure of Interest

None Declared

Table 1 (abstract P1046).

Clinical characteristics of the patients


Diagnosis and gene

Age of onset and clinical manifestations

Laboratory & inmunology

Previous diagnosis

Treatment & response

Patient 1



Neonatal period:

Fever, chondritis, aseptic meningitis, joint limitations, erythematous plaques,lipodystrophy, lymphadenopathy, splenomegaly and hepatomegaly

Anemia, ESR 122 mm/h, CRP 100 mg/L, AST 55 U/L, ALT 51 U/L,


Anakinra (no)

Tocilizumab (no)

Baricitinib (yes)

Patient 2

ISG15 defficiency (ISG15)

6 months:

Intermittent fever, severe cutaneous ulcerations skin, chorioretinitis, brain calcifications

CRP 75 mg/L, HLA B51 (+), ANA (-), ANCA (-), AST 80 U/L, ALT 65 U/L

Early onset Behçet disease

Corticosteroids (yes)

Etanercept (partial)

Anakinra (no)

Adalimumab (no: ADA)

Tocilizumab (partial)

Patient 3


6 years: Raynaud, amputation of phalanx, inflammatory arthritis and chilblains

Normal blood counts,

Inmunology (-)


Etanercept (no)

Tofacitinb (yes)

Patient 4



7 months:

intermittent fever,

livedo reticularis,

paralysis of VI craneal nerve,


CRP 16 mg/L, IgG 567 mg/dl, IgA 22 mg/dlIgM 32 mg/dl, Total lymphocytes 950/μL

ANA and ANCA (-)



Etanercept (yes)

Patient 5



5 years:

Intermittent fever,

livedo resticularis,

paralysis of VI craneal nerve,

painful nodular lesions, arthromyalgia, abdominalgia, Raynaud, peripheral neuropathy

ESR 27 mm/h; RCP 87.6 mg/L ANA and ANCA (-)



Corticosteroids (yes)

Cyclophosphamide (no)

Etanercept (unknown)


CRP: C-reactive protein, ESR: erytrosedimentation rate, ADA: antidrug antibodies, FCL: familiar chilblain lupus, JIA: Juvenile Idiopathic Arthritis, PAN: polyarteritis nodosa, ANA: antinuclear antibodies, ANCA: antineutrophilic antibodies, ASA: acetylsalicylic acid

P1047 Diagnosis and treatment of periodic fevers: a single centre experience

Peter McNaughton1,2, Jane Peake1,3, Ben Whitehead1,3, Su Han Lum2, Kahn Preece4
1Queensland Children's Hospital, Brisbane, Australia; 2Great North Children’s Hospital, Newcastle upon Tyne, United Kingdom; 3University of Queensland, Brisbane; 4John Hunter Hospital, Newcastle, Australia
Correspondence: Peter McNaughton

Introduction: Diagnosis of periodic fever syndromes is difficult due to atypical presentations and overlap in inflammatory symptoms. Treatment of suspected periodic fevers varies widely due to the lack of established clinical guidelines.The utility of genetic testing in identifying monogenic periodic fever syndromes is also unclear due to high frequency of variants of uncertain significance, somatic mutations and heterozygous mutations in genes associated with autosomal recessive conditions.

Objectives: This study aims to evaluate the diagnosis, treatment and use of genetic testing of patients diagnosed with periodic fever syndromes at a single tertiary paediatric hospital.

Methods: We retrospectively reviewed the clinical history of patients diagnosed with periodic fever syndromes at Queensland Children’s Hospital, Brisbane, Australia between November 2014 and June 2018.

Results: 43 patients were diagnosed based on their clinical presentation with periodic fever syndromes.10 patients were diagnosed with PFAPA, 9 with TRAPS, 6 with CAPS, 4 with MKD and 14 unspecified.Median age of onset of symptoms was 24m (range: birth-96m) and median age of diagnosis was 60m (9m-180m).Median time to diagnosis from onset of symptoms was 24m (0-149m).

Multiple medications were used in 15 patients.The medications used varied widely (prednisolone (22), anakinra (9), etanercept (5), tofacitinib (2), tocilizumab (2), cimetidine (2)).

Genetic testing of between 1-26 genes was performed in 26 patients (60%).1-3 genes were tested in 13 patients, targeted panels in 10 patients, SNP only in 1 patient.Genetic variants were identified in 9 patients (34% of those tested) however only 2 of these variants were clearly pathogenic (7.7% of those tested).

Clinical diagnosis and the Eurofever classification criteria were in agreement for patients diagnosed with CAPS (p=0.046) and TRAPS (p=0.025) but not for patients diagnosed with MKD (p=0.47).10 of the patients where clinical diagnosis and Eurofever classification criteria were in agreement had 2 diagnoses positive on the classification score.Two patients diagnosed with CAPS were exclusively positive for CAPS on the classification score and none of the patients diagnosed with TRAPS were exclusively positive for TRAPS.6 of the 7 TRAPS patients had a positive eurofever score for at least one PFS diagnosis.

Conclusion: As reported in previous studies there was a significant delay between onset of symptoms and diagnosis.This reflects an ongoing need to raise awareness of these conditions with primary care providers. The large number of patients treated with multiple medications and the broad range of medications used reflects the lack of established treatment protocols and varied response to treatments in this group of patients.

The clinical diagnosis and diagnostic score showed agreement for CAPS and TRAPs however many patients had a diagnostic score positive for more than one diagnosis meaning that a combination of clinical score and clinical judgement is required to make a diagnosis.

Consistent with previous studies many patients with heterozygous mutations in genes associated with periodic fevers were identified.The significance of these variants is not clear and the diagnostic yield from genetic testing in this cohort was low. Further improvements in availability of next generation sequencing and molecular understanding of autoinflammatory conditions will hopefully improve this yield and allow more confident diagnoses and targeted therapies.

Disclosure of Interest

None Declared

P1048 Multifocal osteomyelitis revealing a PSTPIP1- associated myeloid-related proteinemia inflammatory (PAMI) syndrome: case report and review of the literature

Manel Mejbri, Katerina Theodoropoulou, Michael Hofer
Femme-Mère-Enfant, Centre Hospitalier Universitaire Vaudois CHUV, Lausanne, Switzerland
Correspondence: Manel Mejbri

Introduction: PAMI syndrome is a recently described condition, previously known as Hyperzincemia/Hypercalprotectinemia(Hz/Hc) syndrome. It is a very rare auto-inflammatory disorder characterized by a chronic systemic inflammation, cutaneousand osteo-articular manifestations, hepatosplenomegaly, anemia and neutropenia. Increased blood levels of MRP 8/14(S100A8/A9 or calprotectin) and zinc distinguish this condition. Specific pathogenic mutations in PSTPIP1 gene (p.E250K andp.E257K) were identified as the genetic cause of this condition.

Objectives: Case report and review of the literature of PAMI syndrome

Methods: We report a case of 13 months age female referred to our unity for recurrent episodes of osteoarthritis.Physical examination showed hepatosplenomegaly. Blood work revealed a systemic inflammation, a microcytic anemia and neutropenia. A complete workup for metabolic disorders, oncologic processes and uncommon infections was negative.Because of history of recurrent osteoarthritis, a whole-body MRI was performed and confirmed a multifocal osteomyelitis.

Whole exome sequencing identified the missense p.E250K in the PSTPIP1 gene.

A literature search on PAMI syndrome was performed until the15 October 2018. Pubmed was screened using a combination of the following terms: Hyperzincemia, Hypercalprotectinemia, E250K mutation, PSTPIP1 mutation, PAPA with E250K mutation.

Results: We identified 20 cases of PAMI syndrome in the literature. PAMI syndrome is an early onset inflammatory diseasewith a median age of 2.4 years. Clinical manifestations include Osteo-articular manifestations (80%), skin lesions (71%), splenomegaly (89%), hepatomegaly (68%), lymphadenopathy (42%), growth failure (58%) and hemorragic diasthesis withrecurrent epistaxis and/or haematoma tendency in 5 patients. All cases had relevant abnormalities in hematologic parameters:mild to severe neutropenia and anemia (100%). Thrombocytopenia (42%). Systemic inflammation was confirmed in 94% using the monitoring of CRP, ESR or SAA. Zinc and MRP 8/14 blood concentrations were markedly elevated in all tested patients. Genetic analyses of PSTPIP1 gene revealed the two specific identified mutations (p.E250K and p.E257K) in all patients. Response to the treatment was variable with no consistently effective therapy. Most common therapeutic optionswere AINS, Corticosteroids (n=9), Anakinra (n=9), Anti-TNF (n=6) and Cyclosporine A (n=4).

Conclusion: PAMI syndrome is a rare auto inflammatory condition which should be considered in patients with undefined systemic inflammation and neutropenia, even without skin or osteo-articular manifestations. Zinc and serum MRP 14/8measurement may be helpful tools for the diagnostic orientation in these cases.

Consent for publication has been obtained from patient


Disclosure of Interest

None Declared

Monogenic autoinflammatory diseases (basic science)

P1049 Investigation of inflammasome components in the process of cell migration in FMF patients

Tayfun H. Akbaba1, Z. Yeliz Akkaya-Ulum1, Selcan Demir2, Zeynep Tavukcuoglu1, Seza Ozen2, Banu Balci-Peynircioglu1
1Medical Biology; 2Pediatric Nephrology and Rheumatology, Hacettepe University, Ankara, Turkey
Correspondence: Tayfun H. Akbaba

Introduction: Autoinflammatory diseases, periodic fever syndromes, are a group of diseases that develop recurrent inflammatory response in the absence of infection. Familial Mediterranean fever, which is the most common disease among autoinflammatory diseases, is caused by mutant pyrin production due to mutations in MEFV gene. Pyrin protein is thought to be involved in inflammatory pathways by means of protein-protein interactions. There are studies supporting the role of pyrin as a proinflammatory regulator in the literature and involved in pathways associated with cell migration.

Objectives: The aim of this study was to investigate the effect of pyrin inflammasome on cell migration in FMF patients, in CAPS patient as disease control and healthy controls.

Methods: Within the scope of the thesis; pyrin inflammasome was examined in mononuclear cells isolated from the blood samples of 11 FMF patients homozygous for M694V mutation, 1 CAPS patient with somatic mosaicism and 7 healthy individuals in the process of inflammatory cell migration. Lipopolysaccharide was used to induce the activation of inflammasome in the cells, while arachidonic acid was used for the inhibition of inflammasome. IL-1 beta secretion was analyzed by western blot as an indicator of inflammasome activation, and transwell filter assay was performed in order to examine the differences in cell migration.

Results: We demonstrated that the inhibition of inflammasome in the cells of FMF patients compared to the controls was less under the same application conditions. Following inflammasome activation and inhibition, filter experiments were performed and fetal bovine serum was used to induce inflammatory cell migration. In both inflammasome activation and suppression conditions; we observed a statistically significant increase in the ratio of cell migration of the mononuclear cells of the FMF patients compared to CAPS patient and control indivuals.

Conclusion: These findings support the idea of increased cell migration ratio in patients with FMF due to the more active pyrin inflammasome seen in patients. Although the involvement of other inflammasome components remains to be defined, this study has made significant contributions to illuminate the role of pyrin protein in inflammatory cell migration through the structure of inflammasome.

Keywords: FMF, pyrin inflammasome, cell migration.

This thesis was supported by Hacettepe University Scientific Research CoordinationUnit (Project Number: TYL-2018-17354)

Disclosure of Interest

None Declared

P1050 Cell migration defect in hyperimmunoglobulin D syndrome

Tayfun H. Akbaba1, Selcan Demir2, Z. Yeliz Akkaya-Ulum1, Seza Ozen2, Banu Balci-Peynircioglu1
1Medical Biology; 2Pediatric Nephrology and Rheumatology, Hacettepe University, Ankara, Turkey
Correspondence: Tayfun H. Akbaba

Introduction: Hyper-IgD syndrome or Mevalonate Kinase Deficiency(HIDS/MKD, MIM #260920) is a ultra rare, autosomal recessive hereditary autoinflammatory syndrome caused by compound heterozygous or homozygous mutations in the mevalonate kinase(MVK) gene. It is characterized by recurrent febrile episodes with abdominal pain, lymphadenopathy and an increased serum immunoglobulin D (IgD) level.Anti-IL-1 and anti-TNF treatment are applied to alleviate the symptoms of the disease and to reduce the number of attacks.

Objectives: In this study, we aimed to analyze cell migration, an important process of inflammation,in HIDS patients and compare with FMF patients and controls in inflamasome activated and inhibited conditions.

Methods: Peripheral blood mononuclear cell isolation from HIDS patients, FMF patients and controls was performed. LPS was used to induce inflammasome in isolated cells while arachidonic acid was used for inhibition of inflammasome.After activation and inhibition of inflammasome, transwell filter experiments were performed with isolated cells from patients. As a result of the experiment, the migrating cells were stained with calcein-AM and analyzed by Image J programme.

Results: We did not observe cell migration in HIDS patients’ mononucleer cells under normal conditions, after LPS stimulation. We observed increased cell migration in FMF patients and normal rate in controls as a positive control of these experiments.Considering that 2 HIDS and 1 FMF patients involved in these experiments were receiving anti-IL-1 treatment, it was thought that the drug used by the patients had no effect in cell migration pattern that we observed.

Conclusion: Our preliminary data is the first study identified cell migration defect seen in HIDS. Due to the mutant protein produced as a result of MVK gene mutations, the deficiency in the function of the mevolanate kinase enzyme observed in HIDS leads to the depletion of geranylgeranyl pyrophosphate, a key substrate for protein prenylation.Considering the close relationship between protein prenylation and cytoskeleton elements related with cell migration, a possible decrease in prenylation may be a potential explanation of cell migration defect seen in HIDS patients.

Keywords: HIDS, cell migration defect, prenylation

This project was supported by Hacettepe University Scientific Research Coordination Unit (Project Number: TYL-2018-17354)

Disclosure of Interest

None Declared

P1051 Possible regulatory effects of miRNAs in the pathogenesis of systemic auto inflammatory diseases, from the perspective of familial Mediterranean fever

Z. Yeliz Akkaya-Ulum1, Zeynep Tavukcuoglu1, Ezgi Deniz Batu-Akal2, Tayfun Hilmi Akbaba1, Hafize Emine Sonmez2, Banu Balci-Peynircioglu1, Seza Ozen2
1Medical Biology; 2Pediatric Nephrology and Rheumatology, Hacettepe University, Ankara, Turkey
Correspondence: Tayfun Hilmi Akbaba

Introduction: Systemic Auto Inflammatory Diseases (SAID) is a group of rare and hereditary periodic fever syndromes with recurrent inflammatory involvement developing in the absence of infection. Among same SAID patients, phenotypic heterogeneity is common, and modifier mechanisms such as epigenetic factors may be considered as one of the reason of these variations. MicroRNAs (miRNAs), a type of epigenetic mechanisms, are regulated in most of the biological processes like inflammation.

Objectives: This study aimed to explore the potential effect of miRNAs in the auto inflammation mechanism seen in SAID from the perspective of Familial Mediterranean fever (FMF) which is the most common auto inflammatory disorder.

Methods: The expression levels of miRNAs were analyzed by miRNA array, performed on whole blood RNA samples from healthy controls, homozygous FMF patients with severe phenotype, homozygous FMF patients with mild phenotype, carriers who displayed the disease phenotype, healthy carriers and other rare SAIDs, in pediatric term. The raw data was analyzed by Multi Experiment Viewer (MeV) and TAC programs. Then we performed pathway analyses using DAVID v6.8. and Panther analysistools. Candidate miRNAs shown to be related with inflammatory pathways by bioinformatics analysis, are further studied functionally for their possible effect on expression levels of inflammatory genes, caspase I activation and cell migration (transwell and wound healing experiments) in SW982 (synovial fibroblast) cell lines.

Results: The four patient groups were compared in between and miR-30e-3p, miR-374b-5p, miR-329-3p, miR-29c-3p, miR-25-5p were found to be significantly down regulated in the patient groups. The expression levels of these miRNAs were validated with qRT-PCR. All these miRNAs were found to be known regulators in TGF-beta and Toll-like receptor signaling pathway, apoptosis and actin cytoskeleton regulation by bioinformatics. After pre-miR transfection of miR-30e-3p, miR-374b-5p, miR-29c-3p; expression levels of inflammatory genes (IL-1β, IL-18, TNF-α, TGF-β) were decreased, caspase I activation and cell migration ratio were significantly decreased (p<0.05).

Conclusion: Functional results of this study showed that these miRNAs may possibly have an anti-inflammatory effect and can regulate the expression of genes found in inflammatory pathways. The investigation of potential target genes of these miRNAs by bioinformatics tools and 3’ UTR luciferase assay is underway. The results of this study will be informative for understanding and investigating the possible effect of miRNAs in other auto inflammatory diseases.

This project has been funded by E-RARE-3 project (INSAID, grant 003037603) and The Technical and Scientific Research Council of Turkey (TUBITAK), Grant number: TUBITAK 1001-SBAG 315S096.

Keywords: SAID, FMF, epigenetics, miRNA, inflammation.

Disclosure of Interest

None Declared

P1052 MIR-197 regulates inflammation in monocytes and synovial fibroblasts by targeting IL1R1

Yeliz Z. Akkaya Ulum1, Zeynep Tavukcuoglu1, Tayfun Hilmi Akbaba1, Engin Yilmaz2, Banu Balci-Peynircioglu1
1Medical Biology, Hacettepe University, Ankara; 2Medical Biology, Acibadem Mehmet Ali Aydinlar University, Istanbul, Turkey
Correspondence: Yeliz Z. Akkaya Ulum

Introduction: Familial Mediterranean Fever (FMF); is an autosomal recessively inherited autoinflammatory disease. FMF is caused by the mutations in the Mediterranean Fever (MEFV) gene which encodes the pyrin protein. In many studies, pyrin has been implicated in important cellular processes like apoptosis, cytoskeleton dynamics, signal transduction, and inflammation. Recent studies have shown that epigenetic control mechanisms, particularly non-coding RNAs, may play a role in the pathogenesis of autoinflammation. microRNAs (miRNAs) are small non-coding RNAs that play critical roles in regulating host genome expression at the post-transcriptional level. Dysregulated miRNA expression patterns have been documented in a broad range of diseases including cancer, inflammatory, and autoimmune diseases.

Objectives: Phenotypic heterogeneity seen in FMF disease indicated that FMF is not a simple monogenic disease. Therefore it has been suggested that epigenetic factors can be one of the reason for the variations.

Methods: Previously we identified miR-20a-5p and miR-197-3p as significantly differentially expressed among healthy controls (-/-) and patients (M694V/M694V). The validation of differentially expressed miRNAs was done by quantitative reverse transcription polymerase chain reaction (qRT-PCR). miRNA target genes were determined in miRWalk, the database on predicted and validated miRNA targets. Then pathway analysis was performed in DAVID. For functional assays, SW982 (synovial fibroblast) and THP-1 (monocyte) cell lines were cultured and transfected with miR-197 mimic and scramble control. After transfection, cytokines expression levels (MEFV, IL-1b, IL-18, TNF-α, TGF-β), caspase I activity, apoptosis and cell migration rate were determined. Migration was analyzed in two ways by Transwell migration chamber and wound healing assays. For target gene studies, 3’UTR lusiferase activity assay was done.

Results: qRT-PCR analysis confirmed the results of the miRNA profiling for 2 miRNAs. From two of them; miR-20a showed induction and the other one; miR-197 showed reduction in the homozygote group compared to controls (Akkaya-Ulum et al., 2017).

For functional studies, after pre-miR-197 transfection, the expression levels of cytokines were decreased, cells showed less migration and caspase 1 activity. There was no effect in apoptosis. These results showed that miR-197 could have an anti-inflammatory effect by causing less migration and cytokine secretion. miR-197 was found to bind to the interleukin-1beta (IL-1β) receptor, type I (IL1R1), which is one of the key molecules of the inflammatory pathways.

Conclusion: These findings provide evidence that miR-197 may play role in FMF pathogenesis. Therefore, this study may contribute to understand the inflammatory process seen in FMF disease, as well as to development of the new drug targets and biomarkers in addition to the existing colchicine and similar treatments.

As miRNA-based therapeutics are promising approaches for treating autoinflammatory diseases, we are planning to continue to study on potential therapeutic usage of miR-197 in mouse model of FMF disease.

This study was supported by The Scientific and Technological Research Council of Turkey TUBITAK 1001-SBAG Project Number: 214S106 and Hacettepe University Scientific Research Projects Coordination Unit, Thesis Support, PhD Project Number: TDK-2017-16253 and BAP Comprehensive Project Number: 013D05101005 and Turkey Rheumatology Association.

Keywords: Familial Mediterranean Fever, inflammation, microRNA, miR-197, IL1R1.


Akkaya-Ulum YZ, Balci-Peynircioglu B, Karadag O, Eroglu FK, Kalyoncu U, Kiraz S, et al. Alteration of the microRNA expression profile in familial Mediterranean fever patients. Clin Exp Rheumatol. 2017 Nov-Dec;35 Suppl 108(6):90-94.

Disclosure of Interest

None Declared

P1053 Comparison of FMF patients with age of onset before 20 versus 40 years and over

Okan Aydin, Serdal Ugurlu, Huri Ozdogan
Division of Rheumatology, Department of Internal Medicine, Cerrahpasa Medical Faculty, University of Istanbul - Cerrahpasa, Istanbul, Turkey
Correspondence: Okan Aydin

Introduction: Familial Mediterranean fever (FMF) isa disease withan onset before 20 years of age in 90% of the patients.However late onset FMF defined as age of onset over 40 years is being recognised more frequently.

Objectives: To better define patients with FMF who had their first attack before age 40 and compare them with early onset patient group in Turkish population

Methods: The files of 2180 FMF patients followed in a single center between 2008-2017 who have fulfilled Tel-Hashomer criteria, were reviewed with regard to age of onset 40 years and over (index patients, Group 1).For control purposesfiles before and after the index patients were browsed and

first patients with an onset before age 20 years (Group 2) were included. The demographic, clinical and geneticcharacteristics are compared between these 2 subgroups.

Results: Patients with an onset after 40 years consisted 2.7% of our FMF population. 50 of the 59 patients with an onset 40 yearsor over were re-evaluated and compared with early onset group consisting of 100 patients (Table 1).The delay in diagnosis, and disease durationwere significantly longer and number of patients with M694V homozygosity and M694V allele frequencywere significantly more frequent among group 2. In general, phenotypes of both onset groups were similar, the only significant differences being the frequency of fever and myositis which were less common amonggroup 1. Also response to colchicine was more pronouncedingroup 1. One other interesting observation was the low incidence ofamyloidosis in a group with such a significant delay in diagnosis and thus treatment.

Conclusion: FMF should be included among the differential diagnosis of patients over 40 years of age with recurrent autoinflammatory manifestations. Less than 3% of FMF patients experience their first attacks after 40 years of age. The frequency of M694V is significantly less in the late onset group, pointing out a milder disease.

Disclosure of Interest

None Declared

Table 1 (abstract P1053).

Demographic, clinical and genetic features of the study groups


≥40 years


≤20 years

n= 100


Sex (F:M); present age (mean±SD) (yr)

32:18; 57.2±7.9

62:38; 31.8±9.1

NS; <0.001

Mean age at onset,(mean±sd) (yr)




Mean age at diagnosis (mean ±sd) (yr)




Delay in diagnosis (mean ±sd) (yr)




Mean disease duration (mean ±sd) (yr)




Abdominal pain, n (%)




Chest pain, n (%)


27 (27.0)


Fever, n (%)




Arthritis, n (%)




Myalgia, n (%)




Amyloidosis, n (%)




Positive family history, n (%)


62 (65.2)


Response to colchicine, n (%)


93 (94.8)


M694VHomozygous, n (%)




N of M694Vallelles

24( 48 )

82 (82)


No mutation, n (%)




P1054 What history of appendectomy will tell us about the course of familial Mediterranean fever in adulthood?

Erdal Bodakçi1, Nazife S. A. Bilge1, Nuh Ataş2, Berkan Armağan3, Hasan Satış2, Alper Sarı3, Hakan Babaoğlu2, Gözde K. Yardımcı3, Reyhan B. Salman2, Levent Kılıç3, Mehmet A. Öztürk2, Berna Göker2, Seminur Haznedaroğlu2, Umut Kalyoncu3, Abdurrahman Tufan2, Timuçin Kaşifoğlu1
1Deparment of Internal Medicine, Division of Rheumatology, Eskişehir Osmangazi University Faculty of Medicine, Eskişehir; 2Deparment of Internal Medicine, Division of Rheumatology, Gazi University Faculty of Medicine; 3Deparment of Internal Medicine, Division of Rheumatology, Hacettepe University Faculty of Medicine, Ankara, Turkey
Correspondence: Erdal Bodakçi

Introduction: Peritonitis attacks of FMF usually requires emergency medical admissions and it’s hard to distinguish a typical attack from surgical causes of acute abdomen. Therefore, unrevealing abdominal surgery history, particularly appendectomy, is very common in patients with FMF. However, history of appendectomy might also give some clues about the disease course of FMF.

Objectives: to determine whether history of appendectomy is associated with disease course of FMF in adulthood.

Methods: All patients recruited from FMF in Central Anatolia (FiCA) cohort, comprising 970 (mean age 35.3±12 years, 61.5%female) adult subjects. All patients fulfilled Tel Hashomer criteria. Demographic data, FMF disease characteristics, co-morbid conditions, past medical history including surgeries, disease complications were meticulously questioned and laboratory features and genotype data (if available) were recruited from patient files. Disease severity and FMF associated damage were assessed with International Severity Scoring System (ISSF) for FMF and Autoinflammatory Disease Damage Index (ADDI), respectively.

Results: Appendectomy history was evident in 240 (24.7%) subjects.Peritonitis (4.4±6.7 vs 2.9±4.3 attacks/per year, p<0.001) and pleuritis (3.9±5.2 vs 2.8±4.4 attacks/per year, p=0.03) attacks were more frequent in appendectomy performed (AP) group than appendix intact (AI) group. However, there were no difference between AP and AI groups for the attack frequencies of musculoskeletal and skin components. Considering all types of attacks, AP group had more attacks (6.4±8.2 vs 4.3±6.6 attacks, p<0.001), despite they had used more higher doses of colchicine (1.43±0.6 mg/day vs 1.27±0.5 mg/day, p=0.002). ISSF scores were also higher in AP group 3.13±1.68 vs 2.73±1.48, p=0.001). Colchicine nonresponse were more prevalent in AP group as well (15.1% vs 6.7%, pearson X2=20.2, p<0.001). However, ADDI damage scores were similar between AP and AI groups.

Conclusion: Appendectomy history is common in FMF patients and associated with frequent serositis attacks in adulthood. These patients require more colchicine doses with a lower response rate. Hence, history of appendectomy would be a worthy clue for the management of FMF patients.

Disclosure of Interest

None Declared

P1055 Complement endorse the pathogenesis in autoinflammation

Juergen Brunner1, Wilfried Posch2, Doris Wilflingseder2
1Pediatrics; 2Division of Hygiene and Medical Microbiology, Medical University Innsbruck, Innsbruck, Austria
Correspondence: Juergen Brunner

Introduction: The complement system represents a major part of the innate immune system, consisting of more than 30 different proteins in plasma and on cell surfaces and can be activated through three different pathways.Inflammasomes are also part of the innate immune system.A group of disorders in inflammasomes have been associated with autoinflammatory diseases (AIDs). Familial cold autoinflammatory syndrome (FCAS), Muckle-Wells syndrome (MWS) and chronic infantile neurological, cutaneous and articular syndrome/neonatal onset multisystem inflammatory disease (CINCA/NOMID) were originally described as three distinct diseases. After the identification of their common genetic origin, i.e. mutations in the NLRP3 gene on chromosome 1q44, they are perceived as a continuum of one disease entity and labelled cryopyrin-associated periodic syndromes (CAPS).

Objectives: Aim of this preliminary study in a patient with MWS was to find a correlation between the complement system and a disorder of autoinflammation.

Methods: PBMCs (peripheral blood mononuclear cells) were isolated from blood of a healthy donor and of an individual suffering from MWS by density gradient centrifugation using a Ficoll Paque Premium (GE Healthcare). After washing, PBMCs were incubated with anti-human CD14 Magnetic Beads (BD) to obtain CD14+ monocytes. These were stimulated by addition of cytokines (IL-4 and GM-CSF) for five days to generate immature moDCs (iDCs), which were used for cytokine ELISAs and flow cytometric analyses. IL-6 and IL-1β cytokine ELISAs were performed according to the manufacturer (Biolegend) following stimulation of cells using either LPS or differentially complement opsonized HIV-1. Phenotypical characterization of pathogen-exposed DCs was performed by analyzing characteristic surface markers (CD11c, DC-SIGN, CD86) by multi-color flow cytometry.

Results: IL-1β production of iDCs is higher in the patients cells than in the cells of the healthy donor. However, the most significant difference was shown in complement opsonized iDCs. DC-SIGN is higher expressed in complement opsonized iDCs in patient cells compared to cells of a healthy donor (37,12% v28,64%). DC-SIGN is also higher expressed in the iDCs of the MWS patient after stimulation with LPS.

Conclusion: The complement system may play an important role in the development of an proinflammatory milieu in patients with disorders of autoinflammation. The phenomenon shown in a patient with MWS has to be reproduced in more MWS patients a well as in patients with other disorder of autoinflammation.

Disclosure of Interest

None Declared

P1056 Generation of ADA2 genetic knockout in a myeloid cell line using CRISPR/CAS9 genome editing: an in vitro cell line model to study DADA2

Marina S. Casimir, Ying Hong, Paul Brogan, Despina Eleftheriou
Infection, Inflammation and Rheumatology, UCL Great Ormond Street Institute of Child Health, London, United Kingdom
Correspondence: Marina S. Casimir

Introduction: Deficiency of adenosine deaminase type 2 (DADA2) is an autosomal recessive genetic disease causing systemic inflammation and vasculitis resembling polyarteritis nodosa, caused by loss of function mutations in the ADA2 gene. We are curently exploring an alternative treatment strategy of curing DADA2 using gene therapy. An important first step for this line of work was to develop a cell line model, with abrogation of ADA2 protein and enzyme expression and to determine whether the immunophenotype and characteristics of this cell line mimic the phenotype of primary cells derived from DADA2 patients.

Objectives: To develop an ADA2 knockout (KO) monocyte cell line (THP-1) derived using Clustered Regularly-Interspersed Small Palindromic Repeats (CRISPR)/CAS9 and to confirm that this KO has comparable immunophenotype to monocytes from DADA2 patients (reduced ADA2 expression; M1/M2 skewing and pro-inflammatory cytokine production).

Methods: A plasmid delivered CRISPR/CAS9 system was used to generate the ADA2 KO THP-1 cell line. ADA2 enzyme activity was assessed using a modified commercially available assay and ADA2 protein expression using immunoblotting. Cells were treated with PMA to induce differentiation into macrophages before polarisation into M1 through LPS/ INF-γ stimulation; and M2 polarisation through IL-4, IL-10 and IL-13 stimulation. Immunophenotyping was assessed using gene expression analysis by qPCR. We performed similar experiments in monocyte-derived macrophages (MDM) from 4 DADA2 patients using the same protocol as for the cell line. Cytokine production was evaluated using MSD electrochemiluminescence.

Results: We first confirmed effective knockout of ADA2 at the genetic and protein level and a complete loss of ADA2 enzyme activity in culture supernatants when compared to scramble control THP-1 cells (p<0.01).M1 polarised ADA2 KO THP1 cells exhibited increased activation of pro inflammatory markers such as TNF-α (p<0.001), CXCL-10 (p<0.001), STAT-1 (p<0.01) and IL-1β (p<0.001) when compared to control THP-1 cells. The M1/M2 ratio was reversed in the ADA2 KO THP-1 cell line compared to the wild type THP-1 cells. In MDM cells from DADA2 patients, we observed similar induction of the pro-inflammatory pathway as indicated by increased TNF-α and IL-1β gene expression. Release of TNF-α in culture supernatants from M1 polarised cells was also enhanced for both the ADA2 KO THP-1 cell line and in cells derived from DADA2 patients.

Conclusion: We have generated an effective ADA2 genetic KO myeloid cell line using a CRISPR/Cas9 system and confirmed that these cells have a complete loss of ADA2 enzyme activity and a comparable immunophenotype to monocytes from DADA2 patients. This in vitro system can now be used to examine the possibility to reverse the defects associated with DADA2 using gene therapy strategies.

Disclosure of Interest

None Declared

P1057 Promising blood test for diagnosis and treatment options in patients with suspected chronic auto-inflammatory syndromes

Anne-Laure N. Chetaille1, Nathalie Pagé2, Marie-Pier Longchamps2, Louis Bessette2, Laetitia Michou2, Paul Fortin2, Philippe Tessier2, Martin Pelletier2
1Rhumatologie Adulte et Pédiatrique; 2CHU de Québec-Université Laval, Québec, Canada
Correspondence: Anne-Laure N. Chetaille

Introduction: Diagnosis and treatments of Auto-Inflammatory Syndromes (AIS) patients are challenging as clinical symptoms are non-specific especially compared with Systemic Autoimmune Rheumatic Diseases (SARD) and detection rate of genetic mutations in patients with high suspicion for AIS is low. Even if a mutation is found, genotypes don’t correlate with phenotypes neither with the response to treatments. As a consequence, patients can be misdiagnosed, receive inappropriate treatment, leading to severe complications and substantial socio-economic costs.

Objectives: The cytokines abnormally secreted by each individual are unknown. We hypothesized that the secretion of cytokines by peripheral blood mononuclear cells (PBMC) could be an indicator of the disease and could guide the treatment to the most suitable anti-cytokine. Quantification of the secretome of leukocytes may guide diagnosis and treatment of AIS patients.

Methods: Plasma and peripheral blood mononuclear cells (PBMCs) were isolated from healthy controls, suspected AIS patients, and rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE) patients from the CHU de Québec SARD Biobank Repository Database (SBRD). PBMCs were stimulated with well-known immune activators to trigger inflammasome activation, and cytokine levels were analyzed in the plasma and the supernatants by multiplex assays.

Results: Plasma cytokine levels were similar and did not allow to discriminate between suspected AIS patients, SARD patients or healthy volunteers. PBMCs, on the contrary, had a distinct profile of cytokine secretion between groups. PBMCs from AIS patients spontaneously secreted more IL-1a, IL‑12 and IL-18 than cells from SARD patients or healthy control subjects, as well as IFNg in response to NLRP3, AIM2 and pyrin inflammasome activation. PBMCs from some suspected AIS patients secreted excessive IL-1a or IL-1b in the absence of the antagonist IL‑1RA, suggesting blockers of IL-1 could be used as biotherapeutic approach. PBMCs from other suspected AIS patients secreted high levels of IFNg, IL-12 and IL‑18, pointing to the usefulness of treatments such as Janus kinase small molecule inhibitors, and may be the anti-IL‑12/IL-23 p40 antagonist and/or the novel anti-IL-18 in clinical development.

Conclusion: This study demonstrates that analysis of leukocytes’ secretome is reliably more sensitive than serum to reveal cytokine signatures and to predict biologic treatment options in patients with suspected chronic AIS. Quantification of leukocytes’ secretome allows personalized medicine by guiding diagnosis and treatment options of AIS patients.

Disclosure of Interest

None Declared

P1058 Specific dysbiosis associated with familial Mediterranean fever complicated or not with AA amyloidosis

Samuel Deshayes1, Soraya Fellahi2, Jean-Philippe Bastard2, Jean-Marie Launay3, Jacques Callebert3, Thibault Fraisse4, David Buob5, Jean-Jacques Boffa6, Irina Giurgea7, Charlotte Dupont8, Sarah Jegou9, Marjolène Straube9, Alexandre Karras10, Achille Aouba1, Gilles Grateau4, Harry Sokol11, Sophie Georgin-Lavialle4 and AA Amyloidosis Study Group
1Internal Medicine, CHU Côte de Nacre, Caen; 2Biochemistry, Hôpital Tenon; 3Biochemistry, Hôpital Lariboisière; 4Internal Medicine; 5Anatomopathology; 6Nephrology, Hôpital Tenon; 7Medical Genetics, Hôpital Trousseau; 8Reproduction Biology, Hôpital Tenon; 9AP-HP Laboratoire des Biomolécules (LBM), UPMC Université Paris 06; 10Nephrology, Hôpital Européen Georges Pompidou; 11Gastroenterology, Hôpital Saint-Antoine, Paris, France
Correspondence: Samuel Deshayes

Introduction: Familial Mediterranean fever (FMF) can be complicated by inflammatory (AA) amyloidosis, but the reason why only some patients will develop amyloidosis is not completely understood.

Objectives: To assess gut microbiota composition and inflammatory markers in FMF patients complicated or not by AA amyloidosis.

Methods: We included 34 FMF patients without AA amyloidosis, 7 FMF patients with AA amyloidosis, 19 patients with AA amyloidosis from another origin, and 26 controls. The gut microbiota was studied by 16S ribosomal ribonucleic acid gene sequencing on an Illumina MiSeq platform. Associations between bacterial taxa and clinical phenotype were evaluated using the multivariate association with linear models (MaAsLin) statistical method. Blood dosages of interleukin (IL)-1β, IL-6 and tumor necrosis factor-α were carried out by enzyme-linked immunosorbent assay.

Results: Compared to healthy subjects, significant decreases in α-diversity were noted in FMF patients without amyloidosis and in patients with non-FMF-related AA amyloidosis. β-diversity analysis also showed significant differences between healthy controls and two same groups. After multivariate association testing with the MaAsLin statistical method to control for the effects of potential confounding factors (such as age, gender, body mass index and treatments), several operational taxonomic units belonging to the Clostridiales order were associated with FMF. Moreover, 2 operational taxonomic units belonging to the Clostridiales order were overrepresented in FMF-related AA amyloidosis compared to FMF patients without AA amyloidosis.

All studied groups had higher blood levels of IL-1β, IL-6 and tumor necrosis factor-α than controls.

Conclusion: FMF was associated with a gut microbiota dysbiosis characterized by a decreased α-diversity and a significant alteration in composition. Although these results are descriptive, they suggest that the gut microbiota might be involved in the clinical expression of FMF. In FMF patients, amyloidosis was independently associated with a specific alteration in the microbiota composition, suggesting that the gut microbiota may play a role in AA amyloidosis pathogenesis. These data need to be further consolidated in mechanistic and interventional studies.

Disclosure of Interest

None Declared

P1059 Proinflammatory cytokines induced by PBMCS from a patient with a NLRP1 variant showing severe corneal intraepithelial dyskeratosis

Troels Herlin1, Sofie E. Jørgensen2, Christian Høst1, Mette Christiansen3, Dorte A. Larsen4, Trine H. Mogensen5
1Pediatrics, Aarhus University Hospital; 2Biomedicine, Aarhus University; 3Clinical Immunology; 4Ophthalmology; 5Infectious diseases, Aarhus University Hospital, Aarhus, Denmark
Correspondence: Troels Herlin

Introduction: Corneal intraepithelial dyskeratosis is an extremely rare autosomal dominant inherited disorder with the classical form known as hereditary benign intraepithelial dyskeratosis (HBID). Variants of the nucleotide-binding leucine-rich repeat containing purine domain 1, NLRP1gene, have recently been associated with autoinflammatory disorders with arthritis, vitiligo and dyskeratosis, including HBID.

Objectives: In a boy with severe HBID, with a variant p.A59P in the autoinhibitory PYD domain of NLRP1, to examine the level of NF-kB activation and proinflammatory cytokines in stimulated PBMCs.

Methods: \5-year-old boy with severe inflammation of the gingival mucosa with premature loss of several teeth, painful corneal and conjunctival inflammation of both eyes leading to marked photophobia, corneal opacification and loss of vision. He had mild eczema, but no dyskeratosis of the skin. He is the only child of non-consanguineous Danish parents. Grandfather’s brother and father’s grandfather both had hyperkeratotic dermatosis and swollen gingival mucosa but no eye inflammation.

NLRP1Sanger sequencing was performed. Peripheral blood mononuclear cells (PBMCs) from patient and 3 healthy controls were stimulated with NF-kB inducers (TNFa and LPS or left untreated. Phosphorylated IkBa, as a measure of NF-kB activation, was measured by Luminex technology. PBMCs were stimulated with different ligands (TNFa and LPS and muramyl dipeptide (MDP) or left untreated for 16 hrs. Proinflammatory cytokines (IL-1b, IL-6, IL-18, and TNFa) were measured in the supernatants using Mesoscale U-plex multiplex assays.

Results: Sanger sequencing identified a heterozygous variant (c.175G>C, p.A59P) in the autoinhibitory PYD domain of NLRP1. The variant has a high CADD score of 24.1 predicting a high likelihood of deleteriousness and is not present in the gnomAD database. The sequencing revealed other variants (p.L155H, p.V1063M and p.M1188V) with high minor allele frequencies, most likely benign. Family analysis showed that A59P and L155H were carried by the father and the grandfather’s brother but none by the mother.

Patient PBMCs demonstrated significantly increased levels of Ser32/36 phosphorylated IkBa in response to TNFa and LPS stimulation compared to controls, indicating increased NF-kB activation. In response to especially MDP, but also TNFa stimulation, patient PBMCs expressed extremely high levels of IL-1b and IL-6 and increased IL-18 levels in unstimulated PBMCs compared to controls. Clinically and biochemically the patient responded well to monthly treatment with subcutaneous canakinumab (5 mg/kg) with rapid improvement of the gingivitis, and ophthalmologically administration of diluted anakinra (2.5% solution) as 1 eye-droplet three times per day had a dramatic effect on ocular pain and inflammation.

Conclusion: We here report A59P NLRP1 as a novel defect causing inflammasome hyperactivation/autoinflammatory disease and a clinical picture including severe inflammation of the gingival mucosa and keratitis in a 5-year-old boy with a family history suggesting HBID. Patient PBMCs exhibited increased NF-kB activation and elevated levels of IL-1 and IL-6 in response to proinflammatory stimuli. Thus, our results enabled us to select relevant targeted therapy deploying IL-1binhibition with systemic canakinumab and topical anakinra.

Consent for publication has been obtained from patient


Disclosure of Interest

None Declared

P1060 Gain-of-function mutation of NOD2 impairs its ligand specific immune responses in BLAU syndrome patient-derived IPS cells

Naotomo Kambe1, Nhung T. M. Ly1, Megumu K. Saito2, Hiroyuki Okamoto1
1Dermatology, Kansai Medical University, Hirakata, Osaka; 2Clinical Application, CiRA, Kyoto University, Kyoto, Japan
Correspondence: Naotomo Kambe

Introduction: NOD2 is crucial for innate immune response and mainly expressed in hematopoietic lineage cells, especially in monocytic cells. On the recognition of its ligand, muramyl dipeptide (MDP), NOD2 leads to activation of NF-κB pathway, causing upregulation of pro-inflammatory cytokines.

Objectives: Mutations of NOD2 have been associated with Blau syndrome, but the details regarding mechanisms associated mutant NOD2 leads to granuloma formation are still unclear. By using iPS cells derived from the patients, we tried to reveal the molecular mechanism of Blau syndrome.

Methods: The iPS cells with R334W mutation in NOD2 have been established from a Blau patient and we corrected the mutation of the iPS cells into wild type (WT) by using a CRISPR-Cas9 system. These isogenic iPS cells were differentiated into monocytic cell lineages, then transfected with the lentiviral vectors and maintained in the StemPro-34 medium with M-CSF and GM-CSF.

Results: IFNγ induced the same upregulation of NOD2 in iPS-derived monocytes with WT and those with R334W. Without MDP stimulation, pro-inflammatory cytokine production was only found in those with mutant NOD2, suggesting that Blau-associated NOD2 is gain-of-function mutation. However, after stimulating with MDP, the R334W mutant cells showed NF-κB pathway activation and cytokine secretions less than WT cells even with or without IFNγ treatment. On the other hand, both the cell groups showed the comparative immune response to TNFα and LPS treatment, unrelated to NOD2 mutation.

Conclusion: The response to MDP by mutant NOD2 was selectively impaired in Blau syndrome, that may incompletely lead to neutrophilic inflammation but compensatively induce granuloma formation in host defense.

Disclosure of Interest

None Declared

P1061 The role of DNA methylation for disease severity in patients with heterozygous mutations in the Mediterranean fever gene MEFV

Julie Krainer1, Walter Pulverer1, Dirk Foell2, Seza Özen3, Andreas Weinhäusel1
1AIT - Austrian Institute Of Technology, Vienna, Austria; 2Pediatric Rheumatology & Immunology, University Children's Hospital, Muenster, Germany; 3Department of Pediatric Rheumatology, Hacettepe University, Ankara, Turkey
Correspondence: Julie Krainer

Introduction: Familial Mediterranean Fever (FMF) is the most common hereditary SAID, with a high prevalence in patients of eastern Mediterranean decent. It is known as an autosomal dominant disease with mutations in the MEFV gene that encodes for Pyrin, an important innate immunity regulator. However, some heterozygous individuals also show an FMF phenotype, which leads to the assumption that other modifying factors lead to a manifestation of the phenotype1. In recent years, DNA methylation has demonstrated suitable as biomarker and its potential for disease diagnosis by several studies. DNA methylation plays an important epigenetic regulatory effect on gene expression, and aberrances can result in pathological disease states.

Objectives: The main goal of this study was to evaluate the DNA methylation in patients carrying heterozygous mutations in the MEFV gene but show different phenotypes. We hypothesis that alterations in DNA methylation can add important and valuable information about the disease etiopathogenesis.

Methods: The study included 32 patients, 12 of the patients show a typical FMF phenotype and SNP analysis showed a heterozygous mutation, 9 patients showed similar heterozygous mutations but lack FMF characteristics and are healthy. We also included 12 control samples without any mutation in the MEFV gene.

We performed a genome wide DNA methylation analysis using Illumina’s EPIC BeadArray, interrogating over 850.000 CpG sites at single C resolution covering the first exon of >84% of all Genes2. The results were processed and normalized using the R package Champ3 and group comparisons were performed with limma4.

Results: In total four different group comparisons were calculated including Heterozygous Healthy vs. Heterozygous Disease, Heterozygous Healthy vs. Control, Heterozygous Disease vs. Control and Heterozygous (Disease + Healthy) vs. Control. Each comparison revealed over 30000 significant cpgs (p<0.05) where between 28 and 75 cpgs showed at least 15% differences in mean methylation between the groups. Eighty of the significant cpgs were present in every group comparison covering 62 unique genes. During hierarchical clustering the three clusters separated visibly, where two Heterozygous Healthy patients cluster within the control samples. A group comparison between the two Heterozygous groups revealed 71 differentially methylated sites (p<0.05, difference ≥ 15%), able to separate the two groups.

Conclusion: During our experiments we were able to detect differentially methylated sites separating Heterozygous Healthy and Heterozygous Disease patients. We hypothesize that the identified CpG sites can contribute to provide information to the etiopathogenesis of FMF in general.


[1] Sönmez et al.. Familial Mediterranean fever. Current perspectives. In: Journal of Inflammation Research 9; 2016. S. 13–20. DOI: 10.2147/JIR.S91352.

[2] Pidsley et al. “Critical evaluation of the Illumina MethylationEPIC BeadChip microarray for whole-genome DNA methylation profiling.” Genome Biology 17 (1), S. 208; 2016. DOI: 10.1186/s13059-016-1066-1.

[3] Morris et al. “Champ: 450k chip analysis methylation pipeline.” Bioinformatics, 30(3), 428-30; 2014. DOI: 10.1093/bioinformatics/btt684.

[4] Ritchie ME et al. “limma powers differential expression analyses for RNA-sequencing and microarray studies.” Nucleic Acids Research, 43(7), e47; 2015. DOI: 10.1093/nar/gkv007

Disclosure of Interest

None Declared

P1062 Differential kinetic of actin polymerization in neutrophil from patients with familial Mediterranean fever

David Poghosyan1, Anush Martirosyan1, Nune Mkrtchyan2,3, Sona Margaryan1, Susanna Ghonyan1, Gayane Amaryan2,3, Gayane Manukyan1
1Laboratory of Molecular and Cellular Immunology, Institute of Molecular Biology; 2Arabkir Medical Centre, Institute of Child and Adolescent Health, National Pediatric Centre for Familial Mediterranean Fever; 3Yerevan State Medical University, Yerevan, Armenia
Correspondence: Gayane Manukyan

Introduction: Seemingly unprovoked trafficking of neutrophils to the serosal/synovial membranes is a key event in acute inflammatory attacks of familial Mediterranean fever (FMF). Migratory events behind abnormal trafficking of the cells remain largely unknown.

Objectives: The aim of the present study is to analyze kinetic of actin polymerization and migration rate of neutrophils from colchicine naïve and colchicine receiving FMF patients.

Methods: FMF patients in acute attack (colchicine naïve A-FMF), FMF patients in remission period (receiving colchicine, R-FMF), and healthy controls (HD) were enrolled in the study. We have measured the amounts of F-actin in untreated and pre-treated with colchicine neutrophils induced by fMLP at 5s, 15s, 30s, 60s, 120s, and 180s time points in vitro. Actin polymerization was investigated by flow cytometry using FITC-Phalloidin. Neutrophils migration to fMLP was assessed using Transwell cell migration assay.

Results: Neutrophils from A-FMF displayed higher migration rate compared to R-FMF and HD (P< 0.05). R-FMF patients, who were receiving colchicine, displayed the lowest actin polymerization activity in neutrophils at all-time points. The cells from FMF patients in A-FMF stimulated with fMLP displayed a maximal F-actin polymerization earlier (at 5s) than in R-FMF (at 30s) and HD (at 15s) (P< 0.05). In opposite, plateau levels of F-actin content were reached earlier in HD neutrophils (after 1 min), while plateau levels in A-FMF and R-FMF cells were reached later (after 2 min). Colchicine-pretreated cells in all studied groups reached maximal F-actin polymerization later than in untreated with colchicine cells, namely at 15s in A-FMF and 30s in R-FMF and HD groups.

Conclusion: We have demonstrated actin dysfunctions in neutrophils from FMF patients which might cause defects in neutrophil functioning. The results suggest that colchicine may affect actin polymerization in the cells due to unknown mechanism.

Disclosure of Interest

None Declared

P1063 CARD15 mutations in an Indian cohort of Blau syndrome

Amit Rawat1, Deepti Suri1, Rajni Kumrah1, Sagar Bhattad2, Sandesh Guleria1, Vignesh Pandiarajan1, Ankur Jindal1, Anju Gupta1, Isabelle Ceccherini3, Marco Gattarno4, Surjit Singh1
1Pediatrics, Postgraduate Institute of Medical Education and Research, Chandigarh; 2Pediatric Immunology and Rheumatology, Aster CMI Hospital, Bengaluru, India; 3Genetica Medica; 4U.O.C. Pediatria II - Reumatologia, Istituto Giannina Gaslini, Genoa, Italy, Genoa, Italy
Correspondence: Amit Rawat

Introduction: Blau syndrome is an autosomal dominant disorder resulting from genetic variants in the CARD15 gene previously known as NOD2. It is characterized by a clinical triad of arthritis, uveitis and skin rash. Classic features also include skin lesion with non-caseating granulomas, boggy synovitis or tenosynovitis and camptodactyly. CARD15 gene comprises eleven exons encoding for a 1044 amino acid protein. The protein has N-terminal caspase recruitment domains (CARDs) linked to a nucleotide binding domain (NBD) and C- terminal leucine-rich repeats (LRRs).

Objectives: The objective of the study was to determine the underlying genetic variants in our cohort of patients with Blau syndrome and to establish a genotype-phenotype correlation if any.

Methods: Genetic variants in CARD15 were determined in 11 of our patients with Blau syndrome. Targeted next- generation sequencing was employed to detect these variants in six patients at the Istituto Giannina Gaslini, Genoa, Italy. In rest of the 5 patients, conventional Sanger sequencing was used. Since most of the variants have been detected in Exon 4, the exon4 was amplified by polymerase chain reaction. Four pairs of specific primers described in Resource of Asian Primary Immunodeficiency Diseases Database were used for amplification of the Exon 4 and the PCR amplifications generated were sequenced using conventional Sanger sequencing.

Results: Single nucleotide substitutions resulting in missense variants were detected in all of the eleven patients. Nine of the eleven patients had a recurrent, previously reported missense variant in Exon 4 which encodes for the nucleotide binding domain (NBD). This variant resulting from the substitution of Thymine for cytosine at c.1000; c.1000C>T causes the replacement of Arginine with Tryptophan at codon 344; p.R344W. This variant has been reported in the Infevers database. Three of the nine patients with this variant were related, an affected mother with her son and daughter. Rest of the six were from unrelated families. However, six of the nine patients were from the north Indian state of Haryana, and one was from the adjoining northern state of Punjab. The remaining two patients were from the eastern part of India. Two patients had another missense variant in the Exon 4; p.G299D with replacement of Glycine with Aspartic acid at codon 299. This variant has not been reported in the Infevers database or the Human Gene Mutation Database (HGMD), but it has been reported in the Exome Aggregation Consortium (ExAC).

Conclusion: The detection of a similar previously reported missense variant in 9 of the 11 patients might denote a founder mutation especially given the fact that seven of the nine patients with the p.R344W variant were from the same geographical locale in North India. This recurrent variant also provides an opportunity for development of an amplification-refractory mutation system for its detection without resorting to sequencing.

Disclosure of Interest

None Declared

P1064 Differential activation of the Pyrin inflammasome in monocytes and macrophages predicts the pathological significance of MEFV variants in familial Mediterranean fever (FMF) patients

Takeshi Shiba1, Takayuki Tanaka1, Hiroaki Ida2, Misa Watanabe3, Haruna Nakaseko4, Mitsujiro Osawa5, Hirofumi Shibata1, Kazushi Izawa1, Takahiro Yasumi1, Yuri Kawasaki5, Megumu K. Saito5, Junko Takita1, Toshio Heike6, Ryuta Nishikomori1
1Department of Pediatrics, Kyoto University Graduate School of Medicine, Kyoto; 2Department of Respiratory, Neurology, and Rheumatology, Kurume University School of Medicine, Fukuoka; 3Department of Pediatrics, Toho University School of Medicine, Tokyo; 4Department of Infection and Immunology, Aichi Children’s Health and Medical Center, Aichi; 5Department for Clinical Application, Center for iPS cell Research and Application, Kyoto University, Kyoto; 6Department of Pediatrics, Hyogo Prefectural Amagasaki General Medical Center, Hyogo, Japan
Correspondence: Takeshi Shiba

Introduction: While numerous MEFV sequence variants have been reported, the impact of these variants on pyrin inflammasome functions remains unknown.

Objectives: To determine the effect of MEFV variants on IL-1β production by monocytes and monocyte-derived macrophages from Familial Mediterranean Fever (FMF) patients and to extend this approach to the functional evaluation of rare MEFV variants.

Methods: Freshly isolated monocytes and monocyte-derived macrophages from patients with typical FMF and healthy donors were analyzed for IL-1β secretion in response to the pyrin inflammasome activator Clostridium difficile toxin A (TcdA). Induced pluripotent stem cell (iPSC) clones derived from FMF patients and healthy donors were differentiated into macrophages and analyzed for pyrin inflammasome activation. Rare MEFV variants were expressed in iPSC-derived macrophages.

Results: IL-1β secretion by FMF monocytes in response to TcdA was comparable to that of healthy donors, and colchicine inhibited IL-1β secretion from healthy donors, but not from FMF monocytes. FMF macrophages secreted significantly higher levels of IL-1β than healthy donor macrophages, and their IL-1β secretion could be inhibited with colchicine. The characteristics observed in monocyte-derived macrophages were recapitulated in iPSC-derived macrophages. Finally, iPSC-derived macrophages transgenically expressing the N679H variant were functionally similar to those expressing the pathogenic MEFV variant M694I, while cells expressing the T577N variant had a similar phenotype as control cells.

Conclusion: While FMF monocytes carrying the typical MEFV mutations were distinct from healthy monocytes in their unresponsiveness to colchicine, FMF macrophages showed hyperactivation of the pyrin inflammasome and were sensitive to colchicine inhibition. Our novel approach may be useful for identifying MEFV variants that cause the typical FMF phenotype according to their capacity to induce pyrin inflammasome activation.

Disclosure of Interest

None Declared

P1065 Cross-talk between type I IFN pathway and TLR sensing in DNase2 mutated fibroblasts

Alessandra Tesser1, Elisa Piscianz2, Valentina Boz2, Giulia Piperno3, Federica Benvenuti3, Alberto Tommasini1
1IRCCS Burlo Garofolo, Trieste, Italy; 2University of Trieste, Italy; 3ICGEB, Trieste, Italy
Correspondence: Alessandra Tesser

Introduction: Cytoplasmic sensing of nucleic acids via cGAS-TBK1 pathway leads to type I interferon (IFN) production. DNase2 deficiency impairs DNA digestion in phagosomes resulting in cGAS-dependent IFN hyper-production (1). However, other sensors like TLRs can stimulate this pathway, as recently shown in animal models (2).

Objectives: To investigate the interaction between cGAS and TLRs stimulation in DNAse2-deficient fibroblasts.

Methods: Fibroblasts from a subject with DNase2 deficiency and from healthy controls were treated with different Multiplicity Of Infection (MOI) of E. coli and concentration of bacterial LPS, 2’3’-cGAMP and Poly(I:C) for 1 hour. After stimulation, phosphorilated-TBK1 was measured by flow-cytometry with intracellular staining.

Results: Fibroblasts with DNase2 deficiency responded to an overload of bacteria with a greater activation of the IFN pathway compared to healthy controls. DNase2-mutated cells showed an enhanced IFN response even after stimulation with pure E. coli derived-LPS (Tab. 1). Moreover, the combined stimulation of the cGAS-STING pathway and on TLRs (cGAMP + LPS, cGAMP + Poly(I:C)) resulted in a synergic increase of TBK1 phosphorylation compared with any stimulus alone (Tab. 1).

Conclusion: The possibility that the dysregulated IFN pathway in DNase2-deficient fibroblasts may sensitise cells to an increased response to LPS is supported by others preliminary data (not shown) proving that intact STING is necessary for LPS-induced IFN production. Indeed, there are evidences that STING can be activated not only by 2’3’-cGAMP, but also by other molecules collecting the signals from distinct TLRs. Considering these results, we can assume that a dysfunctional IFN pathway could display a “permissive” role towards other stimuli acting on STING-TBK1 signaling. Ongoing experiments are being carried out to clarify the convergence of different TLRs stimuli on the cGAS-TBK1 axis.

1. Rodero MP, et al. Type I interferon-mediated autoinflammation due to DNase II deficiency. Nat Commun. 2017 Dec 19;8(1):2176.

2. Manfredo VS, et al. Translocation of a gut pathobiont drives autoimmunity in mice and humans. Science. 2018 Mar 09:Vol. 359, Issue 6380, pp. 1156-1161.

Disclosure of Interest

None Declared

Table 1 (abstract P1065).

TBK1 phosphorylation assessed by flow-cytometry after infection with different MOI (20, 120, 400) of E. coli, single stimulation with LPS (0.5 ug/ml), cGAMP (20 ug/ml) and Poly(I:C) (20 ug/ml), and combined stimulation of cGAS/TBK1 and TLRs pathways (cGAMP + LPS; cGAMP + Poly(I:C); Poly(I:C) + LPS)



Healthy control fibros


DNase2 mut fibros




E.coli MOI 120



E.coli MOI 400















cGAMP + Poly(I:C)



Poly(I:C) + LPS



NS: not stimulated; MFI: mean fluorescence intensity

P1066 Auto-inflammatory diseases (SAIDS) in Western Switzerland: a descriptive study through the JIRcohorte platform

Lorenzo Tosetti, Manel Mejbri, Aikaterini Theodoropoulou, Michael Hofer
Pediatric Immunology Allergology Rheumatology Unit, University Hospital Lausanne and University Hospital Geneva, Lausanne and Geneva, Switzerland
Correspondence: Lorenzo Tosetti

Introduction: SAIDs are a large and heterogeneous group of inflammatory conditions, including monogenetic and multifactorial diseases, associated with a dysregulation of innate immune system. Early diagnosis and treatment of these conditions are essential to prevent serious complications, in particular the development of amyloidosis. Biological treatments blocking the Interleukins (IL-) 1 and 6 can lead to rapid remission and are expected to improve long-term outcome in these patients. Five of them received an indication to be treated by these medications in Switzerland; therefore, we are interested to evaluate the number of patients who may receive these treatments.

Objectives: To describe and estimate the prevalence of FMF, MKD, TRAPS, CAPS and SoJIA in Switzerland.

Methods: This is a monocentric, prospective and descriptive cohort study, through the JIRcohorte platform. Patients with juvenile-onset sAIDs attending the pediatric rheumatology unit of Western Switzerland in the University Hospitals of Lausanne and Geneva were enrolled at the study until August 2018. AIDs included diagnosis of Systemic onset juvenile idiopathic arthritis (SoJIA), Familial Mediterranean fever (FMF), Cryopyrin-associated periodic syndromes (CAPS), Mevalonate kinase deficiency (MKD) and Tumor necrosis factor receptor-associated periodic syndrome (TRAPS). A projection for Switzerland was made using the population data extracted from the Swiss Federal Statistical Office in 2016.

Results: A total of 123 patients were enrolled, including 67 % females. The median age at last visit was 12.3 years[TA1] . Patients were distributed as follows: 61 patients with SoJIA, 35 with FMF, 15 with CAPS, 8 with MKD and 4 patients with TRAPS. Biologic agents (anti IL-1 or anti IL-6) were used for treatment in 55 % of our patients. The prevalence in our population is 1.4 per 100.000 for sAIDS and 1.5 per 100.000 for SoJIA. The total number of patients with sAIDs is estimated at 139 for monogenic fevers and 131 for SoJIA in Switzerland.

Conclusion: AIDs are rare but not negligible conditions in Switzerland. A national study through the JIRcohorte database is ongoing aiming to evaluate the number of pediatric and adult patients with sAIDs in Switzerland, in order to study clinical presentation, treatments and long term complications.

Disclosure of Interest

None Declared

P1067 Familial Mediterranean fever associated infertility and underlying factors

Nuh Atas1, Berkan Armagan2, Erdal Bodakci3, Timucin Kasifoglu3, Hasan Satis1, Alper Sari2, Nazife S. Y. Bilge3, Hakan Babaoglu1, Gozde K. Yardimci2, Reyhan B. Salman1, Levent Kilic2, Mehmet A. Ozturk1, Berna Goker1, Seminur Haznedaroglu1, Umut Kalyoncu2, Abdurrahman Tufan1
1Department of Internal Medicine, Division of Rheumatology, Gazi University Faculty of Medicine; 2Department of Internal Medicine, Division of Rheumatology, Hacettepe University Faculty of Medicine, Ankara; 3Department of Internal Medicine, Division of Rheumatology, Eskisehir Osmangazi University Faculty of Medicine, Eskisehir, Turkey
Correspondence: Abdurrahman Tufan

Introduction: Familial Mediterranean Fever (FMF) is characterized by recurrent attacks of fever, serositis and arthritis, but some patients may experience long-term complications of disease such as infertility/subfertility.The published data about FMF associated infertility is still limited.

Objectives: The aim of this study is to investigate the frequency and to determine potential factors for FMF associated infertility /subfertility.

Methods: All patients recruited from FMF in Central Anatolia (FiCA) cohort, currently comprising 970 adult subjects. All patients fulfilled Tel Hashomer criteria and all were using colchicine for at least 1 year. Demographic data, FMF disease characteristics and genotype data (if available), disease complications, autoinflammatory damage index (ADDI), laboratory parameters and treatment features were recorded. For this study data on 582 patients (mean age 41±10.6, 65.8% female) who had been willing to have children were used.

Results: Proportions of FMF manifestations were fever 82.3%, peritonitis 90.5%, pleuritis 48.1%, arthritis 41.1% and skin rash 24.7%. MEFV mutations were available in 454 subjects and 79.5% of subjects were harboring M694V mutation (44.5% homozygous for M694V). Among all, 53 (9.1%) patients were colchicine resistant (crFMF). Infertility was present in 64 patients (14.6 % of females and 4 % of males). Multivariate analysis showed female sex (odds ratio, 8.19; 95% confidence interval [CI95%] 2.69-24.97; p<0.05), FMF disease onset <20 years (odds ratio, 9.9; [CI95% 2.06-47.82]; p<0.05), damage accrual (ADDI score) (odds ratio, 2.02; [CI95% 1.65-2.50]; p<0.05) and colchicine nonresponse (odds ratio, 2.07; 95% confidence interval, [CI95% 1.47-2.93]; P<0.05) are the independent predictors of infertility.

Conclusion: Damage accrual (ADDI), FMF disease onset <20 years, colchicine nonresponse and female sex were found to be the independent predictors of infertility. The value of effective therapeutic interventions must be determined to treat infertility in these patients.

Disclosure of Interest

None Declared

P1068 MEFV-mutant induced pluripotent stem-cells have reduced anti-inflammatory properties

Katharina Kessel1,2, Christoph Kessel1, Nadine Ludwig3, Toni Weinhage1, Dirk Foell1, Helmut Wittkowski1
1Pediatric Rheumatology and Immunology, University Children’s Hospital; 2Department of Nuclear Medicine; 3Department of Anesthesiology, Intensive Care and Pain Medicine, University Hospital Münster, Muenster, Germany
Correspondence: Helmut Wittkowski

Introduction: Familial Mediterranean Fever (FMF) is a prototypic autoinflammatory disorder associated with MEFV pyrin-encoding gene mutations, characterized by unprovoked episodes of inflammation. On a molecular level, pyrin inflammasome activation with consecutive increased IL-1beta maturation is an important mechanism of inflammation in FMF.

Objectives: Experiments with human MEFV-mutated cells rely on FMF patient material, but patients are often already treated with an anti-inflammatory therapy, i. e. Colchicine. In order to systematically study functional consequences of MEFV pyrin-encoding gene mutations on a therapy-naïve background we aimed to generate human induced pluripotent stem cells (hiPSCs) from M694V homo- and heterozygous FMF-patients as well as healthy controls and differentiated those to functional human monocytes.

Methods: HiPSCs were generated from MEFV-mutant patients with clinical FMF and from healthy controls by reprogramming of peripheral blood-derived (PB) CD34+ HSCs. After confirmation of MEFV genotypes, HiPSCs were differentiated to monocytes using an embryoid body (EB)-based differentiation protocol. Terminally differentiated cells were phenotyped regarding extra- and intracellular marker expression as well as cell morphology and were subjected to different functional assays. Cells were stimulated with LPS, ATP and S100A12 and cytokine secretion into culture supernatants was quantified by multiplexed bead array assay. For confirmation of observed phenotypes, ex vivo purified primary monocytes from FMF-patients were likewise stimulated and analysed.

Results: Terminally differentiated hiPSC-derived monocytes from both FMF patients and HCs were larger in cell area (mm2) compared to primary human monocytes. On the level of 25F9, CD16a and CD68 expression hiPSC derived monocytes ranged between that detected on primary cells and in vitro differentiated primary monocyte derived macrophages. All cells expressed identical levels of CD14. Similar to our observations on primary monocytes from FMF patients and controls both CD14 and CD16 expression was lowest on p.M694V homozygous hiPSC derived monocytes. On the contrary intracellular S100A9 and A12 expression in both primary as well as hiPSC derived monocytes increased with p.M694V gene dose. Correspondingly, p.M694V homozygous hiPSC-derived monocytes revealed increased S100A8 expression at base line and already spontaneous protein release in stimulation experiments. In contrast, Il1rn expression in p.M694V mutant hiPSC-derived monocytes was low compared to HCs. When induced by IFNa, Il1rn expression was reciprocal according to p.M694V gene dose. In similar lines, p.M694V mutant hiPSC-derived monocytes revealed strongly decreased IL-10 expression on both gene and protein level compared to healthy control cells.

Conclusion: We successfully generated mature monocytes from hiPSC derived from p.M694V hetero- and homozygous FMF-patients as well as healthy individuals. Apart from S100 proteins p.M694V mutant cells did not reveal a pronounced overexpression of inflammatory cytokines but rather demonstrated an intrinsic lack of anti-inflammatory counter-regulation on the level of IL-1Ra and IL-10 expression.

Disclosure of Interest

None Declared

Monogenic autoinflammatory diseases (genetics)

P1069 Three novel cases of late-onset cryopyrin-associated periodic syndromes due to somatic NLRP3 mosaicism

Anna Mensa-Vilaró1, María Teresa Bosque2, Enrique Gómez de la Fuente3, Natalia Palmou4, Luis Martín-Penagos5, Concha Delgado2, Susana Plaza1, Rocío Lara1, María Carmen Antón1, Helios Martinez-Banaclocha6, Juan J. Martinez-García6, Jordi Yagüe1,7,8, Miguel Ángel González-Gay4, Pablo Pelegrin6, Juan I. Arostegui1,7,8
1Immunology, Hospital Clinic, Barcelona; 2Rheumatology, Hospital Universitario Lozano Blesa, Zaragoza; 3Dermatology, Hospital Universitario Fundación Alcorcón, Alcorcón; 4Rheumatology; 5Nephrology, Hospital Universitario Marqués de Valdecilla, Santander; 6Instituto Murciano de Investigación Biosanitaria IMIB-Arrixaca, Murcia; 7Institut d’investigacions Biomèdiques August Pi i Sunyer; 8Universitat de Barcelona, Barcelona, Spain
Correspondence: Juan I. Arostegui

Introduction: Monoallelic gain-of-function NLRP3 mutations are the genetic cause of the dominantly-inherited cryopyrin-associated periodic syndromes (CAPS), which represent the prototypical early-onset, interleukin-1b-mediated disease (1). Post-zygotic NLRP3 mutations leading to somatic mosaicism have been shown as the disease-causing mechanism in a moderate, but increasing number of patients (2-3). Recent reports have also shown that somatic NLRP3 mosaicism may appear later in life, leading to forms of the disease that started during adulthood (4-7).

Objectives: To describe the clinical, analytical and genetic features as well as the outcome of administered treatments in three unrelated Spanish patients with late-onset CAPS carrying somatic NLRP3 mosaicism.

Methods: Clinical and analytical data, and outcome of treatments were collected from patients’ medical charts. Genetic studies were performed using both Sanger and amplicon-based deep sequencing.

Results: The following table summarizes the clinical data of the three enrolled patients.

All patients displayed increased counts of white blood cells, neutrophils and platelets, as well as increased plasma levels of acute-phase reactants.

Genetic studies revealed in each patient a post-zygotic NLRP3 variant: p.Gln306His variant in patient 1 (minor allele frequency [MAF]: 5.1%), p.Ala352Thr variant in patient 2 (MAF: 18.7%) and p.Gln636Glu in patient 3 (MAF: 18.4%). All these NLRP3 variants were classified as pathogenic and gain-of-function variants on the basis of their absence among healthy controls and in public databases, the damaging predictions by using different bioinformatics analyses and additional ex vivo evidences of inflammasome hyperactivation.

All three patients are being treated with anti-IL-1 drugs since several years, resulting in a successful clinical control of the disease, normalization of acute phase reactants and hematological parameters, and improvement of renal function in patient 1 who suffered from AA amyloidosis.

Conclusion: The results here shown add additional evidences of the relevant role of somatic NLRP3 mosaicism as the disease-causing mechanism in patients with late onset, but otherwise typical CAPS. Due to the serious consequences that these data could have with regard to their treatments, this genetic mechanism should be seriously considered in the design of genetic analyses to be done in candidate patients.


1Nat Immunol 2017; 18: 832-842

2Arthritis Rheum 2011; 63: 3625-3632

3Ann Rheum Dis 2015; 74: 603-610

4J Allergy Clin Immunol 2015; 135: 561-564

5Arthritis Rheumatol 2015; 67: 2428-2436

6Arthritis Rheumatol 2016; 68: 3035-3041

7Front Immunol 2017; 8: 1410.

Consent for publication has been obtained from patient


Disclosure of Interest

None Declared

Table 1 (abstract P1069).

See text for description


Patient 1

Patient 2

Patient 3

Sex / Current Age (years)

Female / 63

Female / 66

Male / 67

Age at disease Onset (years)








Urticaria-like Rash




Eye Inflammation

Bilateral Uveitis



Joint Inflammation


Arthralgias / Arthritis

Arthralgias / Arthritis





Hearing Loss




AA-type Amyloidosis




P1070 The NLRP3 p.A441V mutation in cryopyrin-associated periodic syndrome pathogenesis: functional consequences, phenotype-genotype correlations and evidence for a founder effect

Eman Assrawi1, Fawwaz Awad2, Claire Jumeau1, Sylvie Odent3, Veronique Despert4, G. Cam5, Aleth Perdriger6, Camille Louvrier1, Laetitia Cobret1, Bruno Copin1, Phillipe Duquesnoy1, William Piterboth1, Claire Le Jeunne7, Sophie Georgin-Lavialle8, Gilles Grateau8, Marie Legendre1, Irina Giurgea1, Sonia Athina Karabina1, Serge Amselem1
1Sorbonne Université, Inserm UMR_S933; 2Sorbonne Université,Inserm UMR_S933, Paris; 3Centre Hospitalier Universitaire de Rennes, Service de Génétique; 4Centre Hospitalier Universitaire de Rennes, Département de Médecine de L’enfant et de L’adolescent, Rennes; 5Centre Hospitalier de Saint-Malo, Service de Néphrologie, Saint-Malo; 6Centre Hospitalier Universitaire de Rennes, Service de Rhumatologie, Rennes; 7Hôpital Cochin; 8Hôpital Tenon , Service de Médecine Interne, Paris, France
Correspondence: Eman Assrawi

Introduction: Cryopyrin‑associated periodic syndromes (CAPS) are a group of rare monogenic autoinflammatory diseases caused by heterozygous missense gain-of-function mutations in NLRP3 gene, coding for the NLRP3.Upon activation, NLRP3 initiates the formation of a multiprotein complex called inflammasome, which regulates proinflammatory cytokine secretion.

Objectives: To determine the molecular and cellular basis of autoinflammatory syndromes in two unrelated families with two clinically overlapping CAPS phenotypes; a multigenerational French family with Muckle-Wells syndrome and in a patient originating from Portugal with familial cold autoinflammatory syndrome.

Methods: Sanger sequencing of NLRP3 exon 3 was performed in all accessible patients. Microsatellites analysis was used to test the intra-familial segregation of the identified variant and to look for a founder effect. Functional analyses included the study of (i) ASC speck formation in HEK293T cells (stably expressing ASC-GFP and pro-caspase1-FLAG) transiently expressing the wild-type or mutated NLRP3 protein, (ii) IL1β secretion from transfected THP1 cells, and (iii) inflammasome-related gene expression and cytokine secretion from monocytes isolated from patients in crisis (probands from the two families), related patients out of crisis, and from controls.

Results: The same heterozygous mutation (c.1322C>T, p.A441V) in NLRP3 exon 3, segregating with the disease within the first family, was identified in the two families which were shown to share the same mutation-associated NLRP3 haplotype. HEK293T cells transfected with the pNLRP3-A441V construct showed a significantly higher percentage of ASC specks, a common readout of inflammasome activation, as compared to cells transfected with pNLRP3-WT. Transfection of THP1 cells with pNLRP3-A441V led to significantly higher levels of secreted IL1β, a hallmark of inflammasome activation, as compared to cells transfected with pNLRP3-WT. Monocyte inflammasome-related gene expression and cytokine secretion profile was similar in patients out of crisis and in the healthy controls. However, the expression of the inflammasome-related genes in the two probands was different from that of patients without crisis and healthy controls. In addition, this expression pattern was found to be differentially regulated between the two probands, correlating with their phenotypic status.

Conclusion: These molecular and cellular findings, which indicate a founder effect in these two families, clearly demonstrate the pathogenicity of the p.A441V missense mutation in CAPS and point to the interest of studying patients’ primary cells to assess disease activity.

Disclosure of Interest

None Declared

P1071 The analysis of IL-36RA structural dynamics improves pathogenicity predictions for IL36RN variants observed in generalised pustular psoriasis

Camilla Davan-Wetton, Niina Karoliina Hassi, Joseph Chifung Ng, Franca Fraternali, Francesca Capon
King's College London, London, United Kingdom
Correspondence: Francesca Capon

Introduction: Generalised pustular psoriasis (GPP) is a potentially life-threatening autoinflammatory condition. While GPP is associated with mutations of the interleukin-36 receptor antagonist (IL-36Ra encoded by IL36RN), commonly used pathogenicity predictors cannot fully differentiate IL36RN disease alleles from polymorphisms.

Objectives: The aim of this study was to systematically assess the impact of IL-36Ra mutations on the protein three-dimensional structure, in order to identify the computational approaches that best predict in-vitro stability and variant pathogenicity.

Methods: The IL-36Ra 3D structure was derived by homology modelling. The effects of mutations were assessed using the mCSM and RAPSODY programs. Experimental validation was undertaken by western blot analysis of mutagenized constructs.

Results: The results of the mCSM analysis partially correlated with those obtained by western blot, particularly for well-characterised mutations like p.Leu27Pro and p.Ser113Leu. An improved correlation with the western blot data was achieved using RAPSODY, which incorporates information on protein structural dynamics. Of note, this programme also out-performed sequence- based predictors such as CADD.

Conclusion: Computational methods that take into account the dynamic features of the IL-36Ra protein structure show the strongest correlation with experimental data. Extending their implementation to other IL36RN variants (e.g. those that affect the interaction between IL-36Ra and its receptor) will improve our understanding of GPP mutations and enable the development of more accurate pathogenicity predictors to help disease diagnosis.

Disclosure of Interest

C. Davan-Wetton: None Declared, N. K. Hassi: None Declared, J. C. Ng: None Declared, F. Fraternali: None Declared, F. Capon Grant / Research Support from: Boheringer Ingelheim, Consultant for: AnaptysBio

P1072 Spondyloenchondrodysplasia and systemic lupus erythematosus: case report of two sibling

Bulent Kara1, Ayse Cefle2, Ozgur Kasapcopur3, Ayfer Sakarya Gunes1
1Department of Pediatrics Division of Child Neurology; 2Department of Internal Medicine Division of Rheumatology, Kocaeli University Faculty of Medicine, Kocaeli; 3Department of Pediatric Rheumatology, Istanbul University Cerrahpasa Medical School, Istanbul, Turkey
Correspondence: Ayse Cefle

Introduction: Interferons (IFNs) are signalling proteins that are synthesised and released by immune host cells in response to the presence of the several pathogens. Interferonopathies comprise an expanding group of monogenic diseases characterised by disturbance of the homeostatic control of IFN-mediated immun responses. Although differing in the degree of phenotypic expression and severity, the clinical presentation of the diseases shows a considerable degree of overlap, reflecting their common pathogenetic mechanisms.

Objectives: Spondyloenchondrodysplasia with immune dysregulation (SPENCDI) is an immuno-osseous dysplasia combining the typical metaphyseal and vertebral bone lesions of SPENCDI and neurologic involvement.

Methods: 19 years old a male patient presented with arthralgia and rash in 2014. On physical examination short stature and erythematous skin rash were noted. Laboratory studies revealedproteinuria (740 mg/d), lymphopenia (1150/mm3), elevated ESR (51 mm/h), CRP (15 mg/l, N<5) and low C3 level. The C4 level was normal. ANA was homogenously positive, while the anti ds-DNA antibody was also positive. Anticardiolipin IgG and IgM and the lupus anticoagulant were negative. HBsAg, HCV and HIV were negative. IgG, IgA and IgM levels were normal. Skin bipsy revealed perivascular dematitis, while a kidney biopsy showed class II lupus nephritis. The patient was given methyl prednisolone, hydroxycholoroquine and azathioprine. On follow up, the proteinuriadecreased. However, he developed attacks of seizuresand ataxia. The neurologic examination, EEG and EMG, were normal. Cranial tomography revealed calcification of lentiform nucleus, corona radiata, frontal lobe white matter and dentate nuclei. In 2015, he presented with abdominal paine, vomiting and nausea. Abdominal CT revealed edema and inflammation of jejenum and dudenum.The steroid dose was increased. After a few months, the symptoms recurred. The patient was thought to have gastrointestinalinvolvement due to systemic lupus erythematosus (SLE) and rituximab was added to the treatment. Thereafter the patient did not have anysymptoms related toacute abdomen.

Results: A direct raiographic examination of the spine revealed platyspondyly. The parents were consanguineous. His sister was under follow up due to a diagnosis of juvenil onset SLE.Intracranial calcifications, spastic paraparesis, short stature, systemic lupus erythematosus and platyspondyly suggested a diagnosis of SPENCD. Next generation sequencing of the ACP5 gene showed that the patient and his sisters were homozygous for the c.155A C/p.K52T variant. Both parents were heterozygous forthis variant.

Conclusion: SPENCDI is a recessive genetic disease caused by homozygous or compound heterozygous mutation in the ACP5 gene on chromosome 19p13. Immune dysregulation ranges from autoimmunity to immunodeficiency. Neurologic and autoimmune manifestations have been observed in different combinations. Skeletal, nneurologic and immune phenotypes were observed between members of the same family.

Consent for publication has been obtained from patient


Disclosure of Interest

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P1073 ADA2 deficiency without ADA2 mutations explained by a structural homozygous variation in 22Q11.1

Alice Grossi1, Francesca Garbarino2, Roberta Caorsi3, Roberto Cusano4, Marta Rusmini1, Federica Penco3, Francesca Schena3, Rosa Anna Podda5, Paolo Uva4, Isabella Ceccherini1, Marco Gattorno3
1UOC Genetica Medica, IRCCS Istituto Giannina Gaslini; 2Università degli Studi di Genova; 3Clinica Pediatrica Reumatologia e UOSD Centro Malattie Autoinfiammatorie-Immunodeficienze, IRCCS Istituto Giannina Gaslini, Genova; 4Centre for Advanced Studies Research and Development in Sardinia (CRS4), Science and Technology Park Polaris, Pula; 5Clinica Pediatrica,Talassemie e Malattie Rare, Ospedale Brotzu e Università degli studi di Cagliari, Cagliari, Cagliari, Italy
Correspondence: Alice Grossi

Introduction: Adenosine Deaminase 2 deficiency (DADA2) is an autosomal recessive autoinflammatory disease caused by loss of function mutations in the ADA2 gene, located on chromosome 22q11.1. The clinical spectrum of the disease is heterogeneous, ranging from multisystemic inflammation with vascular and multiorgan involvement to immunodeficiency and immunedysregulation.1 A small proportion of patients, despite consistent phenotype and lack of ADA2 enzymatic activity, has an incomplete or negative genotype.

Objectives: To define the genetics underlying DADA2 in a 9 years old girl with a complete clinical phenotype and deficient enzymatic activity but without any mutation in the coding region of the ADA2 gene.

Methods: Whole Genome Sequencing (WGS) was performed in the proband by TruSeq Nano DNA Library Prep kit (Illumina) and HiSeq 3000 Instrument (Illumina) with 150bp paired-end reads.