Tumour necrosis factor-α levels are elevated in adolescent patients with juvenile idiopathic arthritis on etanercept therapy

The use of etanercept, a tumor necrosis factor (TNF) inhibitor, has revolutionized the treatment of juvenile idiopathic arthritis (JIA). TNF is a key cytokine implicated in the pathogenesis of inflammatory arthritis and etanercept, which is a soluble TNF receptor fusion protein, binds and inactivates TNF-α and lymphotoxin-A.


Introduction
The use of etanercept, a tumor necrosis factor (TNF) inhibitor, has revolutionized the treatment of juvenile idiopathic arthritis (JIA). TNF is a key cytokine implicated in the pathogenesis of inflammatory arthritis and etanercept, which is a soluble TNF receptor fusion protein, binds and inactivates TNF-α and lymphotoxin-A.

Objectives
The aim of this study was to profile serum levels of TNF-α in a large cohort of adolescent patients with JIA.

Methods
Serum TNF-α was measured in samples derived from 200 adolescent and young adult patients with JIA attending the adolescent and young adult rheumatology clinic at University College London Hospital using a commercial enzyme linked immunosorbent assay (ELISA) kit (eBioscience). Samples were tested in duplicate. Median age at sampling and median disease duration were 18 years and 8 years 9 months, respectively. Male:female ratio was 1:1.2. Equal numbers of patients with polyarticular (n=64) and enthesitis related arthritis (ERA, n=64) were tested in addition to 48 with oligoarticular, 16 systemic onset, and 8 psoriatic arthritis. Erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) measurements were also collected. Furthermore, an L929 cell viability bioassay was used to determine if the addition of etanercept abrogates the cytotoxic effects of TNF-α in L929 cells.

Results
Surprisingly, TNF-α serum levels were shown to be markedly elevated in patients on etanercept (median TNF-α on etanercept= 134.2pg/ml, .1], median not on etanercept = 4.2pg/ml, IQR [1.4-11.0], p<0.0001). TNF-α levels were also higher in patients on etanercept compared to those on other biologics (adalimumab, infliximab, abatacept, or tocilizumab, median= 4.4pg/ml, IQR [1.8-9.1]) or disease modifying anti-rheumatic drugs alone (median = 4.2 pg/ml, IQR [1.1-12.9]), p<0.0001. In addition, ESR and CRP levels had a negative correlation with high TNF-α levels in patients on etanercept (p=0.0018 and p=0.0034 respectively). Etanercept was included at its therapeutic serum concentration (2.4ug/ml) to ensure there was no cross reactivity with the assay. Finally, we showed that the addition to TNF-α to human serum leads to cytotoxicity in a TNF-α sensitive cell line, while adding etanercept at its therapeutic concentration along with TNF-α significantly reduces cell death (p = 0.0277).

Conclusion
Patients treated with etanercept have higher levels of TNF-α. As the majority of patients with elevated TNF-α on etanercept were in remission, it is likely that this circulating TNF is biologically inactive. This is supported by our in vitro experiments in which the cytotoxic effect of TNF-α was abrogated upon addition of etanercept. Our hypothesis is that etanercept prolongs the half-life of circulating TNF-α. Further studies are needed to confirm these findings and dissect the mechanisms involved. As the association between high TNF-α and etanercept Arthritis Research UK Centre for Adolescent Rheumatology at University College London, Great Ormond Street Hospital and UCLH, University College London, London, United Kingdom treatment is so strong, we hypothesise that it may be possible to measure TNF-α levels as a surrogate marker of adherence to this drug in this cohort of patients where adherence to medication can be a significant problem. This is a hypothesis that warrants further investigation.

Disclosure of interest
None declared.