eTable 1. Comparison of month 6 IL17-F in patients with (A) less vs more disease activity, (B) no vs some disease activity, (C) clinically defined responders vs nonresponders
eTable 2. Comparison of change of IL17-F to month 6 in patients with (A) less vs more disease activity, (B) no vs some disease activity, (C) clinically defined responders vs nonresponders
eFigure. Baseline IL-17F vs number of new MRI lesions
Hartung H, Steinman L, Goodin DS, Comi G, Cook S, Filippi M, O’Connor P, Jeffery DR, Kappos L, Axtell R, Knappertz V, Bogumil T, Schwenke S, Croze E, Sandbrink R, Pohl C. Interleukin 17F Level and Interferon Beta Response in Patients With Multiple
Sclerosis. JAMA Neurol. 2013;70(8):1017-1021. doi:10.1001/jamaneurol.2013.192
High serum levels of interleukin 17F (IL-17F) at baseline have been associated with suboptimal
response to interferon beta in patients with relapsing-remitting multiple sclerosis.
To further investigate the role of IL-17F in predicting treatment response to interferon beta-1b
in patients with relapsing-remitting multiple sclerosis using the Singulex Erenna IL-17F
Design, Setting, and Patients
Serum samples were analyzed from 239 randomly selected patients treated with interferon beta-1b,
250 μg, for at least 2 years in the Betaferon Efficacy Yielding Outcomes of a New Dose
Treatment with interferon beta-1b, 250 μg, for at least 2 years.
Main Outcome Measures
Levels of IL-17F at baseline and month 6 as well as the difference between the IL-17F levels at
month 6 and baseline were compared between the following: (1) patients with less disease activity vs
more disease activity; (2) patients with no disease activity vs some disease activity; and (3)
responders vs nonresponders.
Levels of IL-17F measured at baseline and month 6 did not correlate with lack of response to
treatment after 2 years using clinical and magnetic resonance imaging criteria. Relapses and new
lesions on magnetic resonance imaging were not associated with pretreatment serum IL-17F levels.
When patients with neutralizing antibodies were excluded, the results did not change. All patients
with levels of IL-17F greater than 200 pg/mL were associated with poor response with some clinical
or radiological activity.
Conclusions and Relevance
An increase of IL-17F before and early after treatment with interferon beta-1b was not associated
with poor response. These data do not support the value of IL-17F as a treatment response indicator
for therapy of patients with multiple sclerosis with interferon beta, although high levels of IL-17F
greater than 200 pg/mL may predict nonresponsiveness.
Interleukin 17F (IL-17F) is one of the signature cytokines of helper T 17 cells that play a key role in the defense against pathogens and autoimmunity.1- 3 Helper T 17 cells have been invoked as key determinants of aberrant immune responses in multiple sclerosis.3,4,
Interleukin 17F has been shown to function in part differently from another member of the IL-17 cytokine family, IL-17A.2 Interleukin 17F is a central mediator of cellular immunity governing the expression of critical cytokines that exert proinflammatory effects.3 High serum levels of IL-17F at baseline have been associated with suboptimal response to interferon beta-1b in patients with relapsing-remitting multiple sclerosis.5,6 To further investigate the role of IL-17F in predicting treatment response to interferon beta-1b, we used the Singulex Erenna IL-17F immunoassay to analyze serum samples collected at baseline and after 6 months of treatment from 239 patients who participated in the Betaferon Efficacy Yielding Outcomes of a New Dose (BEYOND) Study.
Serum samples were analyzed from 239 randomly selected patients treated with interferon beta-1b, 250 µg, for at least 2 years in the BEYOND Study.7 In these patients, IL-17F was measured at baseline and after 6 months of treatment using the Singulex Erenna immunoassay. The Erenna IL-17F immunoassay kit makes use of a microparticle-based single-molecule counting technology.8 Human serum samples were diluted in assay buffer and concentrations of IL-17F were determined using a reference and standard curve. The limit of detection of the assay was 0.5 pg/mL. The expected median value of IL-17F in human serum from healthy control subjects was 20 pg/mL. The low limit of quantification was 1.6 pg/mL (≤20% coefficient of variation and ±20% recovery). The short-term effects of interferon beta were confirmed by measuring 2 known interferon-inducible proteins (interferon gamma–induced protein 10 and interferon-inducible T-cell α chemoattractant) using the Human Discovery MAP250 version 1.0 Luminex-based, multianalytes profiling assay (Myriad RBM).
Using the entire patient group (n = 239), IL-17F levels at baseline and month 6 as well as the difference between IL-17F at month 6 and baseline (ΔIL-17F) were compared between the following: (1) patients with less disease activity (no relapse and ≤4 lesions on magnetic resonance imaging [MRI]) vs more disease activity (≥1 relapse or >4 MRI lesions) within 2 years of treatment; (2) patients with no disease activity (no relapse and no MRI lesion) vs some disease activity (≥1 relapse or ≥1 MRI lesion); and (3) responders (no relapse and no confirmed progression on the Expanded Disability Status Scale) vs nonresponders (≥1 relapse and confirmed progression on the Expanded Disability Status Scale). Additional group comparisons excluded patients with neutralizing antibodies to interferon beta. Outcomes of patients having baseline IL-17F concentrations greater than 50 pg/mL or greater than 200 pg/mL were described in line with previous studies that found patients with pretreatment IL-17F values greater than 50 pg/mL to be clinical nonresponders.5 Finally, correlation of IL-17F serum levels with the number of new MRI lesions and the annualized relapse rate was determined for the entire population.
Nonparametric methods were used for all analyses: Wilcoxon rank sum test for group comparisons and Spearman ρ for correlation. Values were compared graphically by means of scatterplots. P values were not corrected for multiple comparisons.
Baseline characteristics and disease course during the first 2 years of interferon beta-1b treatment were similar between the patient group used for IL-17F measurements and the entire BEYOND Study cohort (Table 1). There was no statistically significant difference between IL-17F levels at baseline and month 6 (median, 16.5 vs 16.8 pg/mL, respectively; P = .22). However, levels of interferon gamma–induced protein 10 and interferon-inducible T-cell α chemoattractant increased significantly from baseline to month 6. There was a 2.4-fold increase of median interferon gamma–induced protein 10 levels (P < .001) and a 2.4-fold increase of median interferon-inducible T-cell α chemoattractant levels (P < .001) (data not shown).
When clinical and MRI criteria were used to measure response to treatment, 115 patients (48.1%) had less disease activity (no relapse and ≤4 MRI lesions) and 124 (51.9%) had more disease activity (≥1 relapse or >4 MRI lesions). Fifty-six patients (23.4%) had no disease activity (no relapse and no MRI lesion) and 183 (76.6%) had some disease activity (≥1 relapse or ≥1 MRI lesion). Using clinical response criteria, 124 patients (51.9%) were responders (no relapse and no confirmed progression on the Expanded Disability Status Scale), while 27 (11.3%) were nonresponders (≥1 relapse and confirmed progression on the Expanded Disability Status Scale).
Levels of IL-17F at baseline and month 6 as well as ΔIL-17F did not differ significantly between patient groups (Table 2, and eTable 1 and eTable 2 in Supplement). Similar results were obtained for the 149 patients who did not develop neutralizing antibodies during the entire study (data not shown).
The frequency of high (>50 pg/mL) or very high (>200 pg/mL) IL-17F values at baseline was similar for the different definitions of active vs inactive disease used in our study (Table 2). Of note, we found that no patients without any clinical or radiological disease activity with interferon beta-1b treatment demonstrated baseline IL-17F values higher than 200 pg/mL, whereas 8 (4.4%) of the patients with at least some disease activity showed such values.
There was no significant correlation between baseline IL-17F level and the annualized relapse rate (ρ = 0.08; P = .21) and the number of new MRI lesions observed within the first 2 years of interferon beta-1b treatment (ρ = −0.13; P = .06) (eFigure in Supplement). Finally, there was no significant correlation between the month 6 IL-17F level and the annualized relapse rate (ρ = 0.01; P = .91), between ΔIL-17F and the annualized relapse rate (ρ = −0.02; P = .70), between the month 6 IL-17F level and the number of new MRI lesions observed within the first 2 years of interferon beta-1b treatment (ρ = −0.10; P = .16), or between ΔIL-17F and the number of new MRI lesions observed within the first 2 years of interferon beta-1b treatment (ρ = 0.01; P = .92).
We found that serum concentrations of IL-17F alone did not predict response to interferon beta-1b therapy in patients with relapsing-remitting multiple sclerosis. Baseline IL-17F levels in the group of patients with higher disease activity were in the same range as the group having lower disease activity. Relapses and new MRI lesions occurring in the first 2 years of interferon beta-1b exposure did not correlate with pretreatment IL-17F levels. Negative results were also obtained for the comparison of responders and nonresponders defined on the basis of stringent clinical criteria. Similar results were obtained for IL-17F measured at month 6 and for ΔIL-17F. When patients with neutralizing antibodies were excluded, the results did not change. Finally, IL-17F levels in patient serum did not differ during administration of interferon beta-1b, in contrast to large and significant changes in levels of acute interferon beta-1b bioactivity markers interferon gamma–induced protein 10 and interferon-inducible T-cell α chemoattractant.
Of note, the proportions of patients with pretreatment IL-17F values higher than 50 pg/mL, which had been the threshold for nonresponse in the study by Axtell et al,5 were similar in our patient strata. When focusing on patients with even higher pretreatment IL-17F values, we found that all patients with values higher than 200 pg/mL had at least some clinical or radiological disease activity while being treated with interferon beta-1b. Analyses of extreme outcomes are often illuminating because such analyses often eliminate patients who may be misclassified.9,10 However, considering all our results makes it difficult to draw a robust conclusion from this finding.
Conceptual and methodological differences may have contributed to the discordance of findings between the study by Axtell et al5 and this study. Axtell and colleagues collected samples from a single center, and patients were treated with interferon beta-1a or interferon beta-1b. Our study was multicenter, and patients were treated with interferon beta-1b exclusively. The IL-17F serum concentrations were determined in our study using an assay optimized to measure IL-17F level in human serum, which is different from the multiplex assay used by Axtell et al.5 The Singulex IL-17F assay is recognized to be one of the most accurate, reproducible, and sensitive assays available for measuring IL-17F concentrations in human serum.11 Although Axtell et al5 found a positive correlation between pretreatment IL-17 serum concentrations and nonresponse to interferon beta, these results were based on a much smaller patient cohort (n = 26) and exclusively used clinical criteria for nonresponse. In a recent study, Bushnell et al12 were also unable to correlate IL-17F serum levels with treatment response to interferon beta. However, in their study, IL-17F values did not exceed 50 pg/mL, which makes it difficult to compare their results with those reported by Axtell et al5 and those from our study.
Because the biology of interferon beta is complex, it is not unexpected that serum IL-17F concentration by itself is inadequate to predict treatment response. It has also been demonstrated in mice that IL-17F and IL-17A as single factors do not contribute vitally to the development of autoimmune neuroinflammation.13 Measurement of a combination of other cytokines in concert with IL-17F may be required. Although predictors of response to interferon beta remain elusive, much progress has been made in this area.14- 20 Recognizing the synergistic interactions between IL-17F and other potent mediators on immune regulation, identification of predictors of response to interferon beta treatment may still be possible. Given the multifaceted pathophysiology associated with disease progression and response to treatment by patients with relapsing-remitting multiple sclerosis, using extreme patient cohorts in combination with immune-based biomarker signatures may actually be the most efficient way of initially identifying response markers. One approach might be to focus first on subjects with the highest levels of IL-17F (>200 pg/mL) in future studies.
Accepted for Publication: January 22, 2013.
Corresponding Author: Hans-Peter Hartung, MD, Department of
Neurology, Heinrich-Heine-Universität, Moorenstrasse 5, D-40225 Düsseldorf, Germany
Published Online: June 3, 2013. doi:10.1001/jamaneurol.2013.192
Author Contributions: Drs Sandbrink and Pohl
contributed equally to this work.
Study concept and design: Hartung, Steinman, Comi, Filippi, Jeffery, Kappos,
Axtell, Knappertz, Bogumil, Croze, Sandbrink, and Pohl.
Acquisition of data: Hartung, Steinman, Goodin, Comi, Filippi, Jeffery, Kappos,
Croze, and Sandbrink.
Analysis and interpretation of data: Hartung, Steinman, Comi, Cook, Filippi,
O’Connor, Jeffery, Kappos, Knappertz, Bogumil, Schwenke, Croze, Sandbrink, and Pohl.
Drafting of the manuscript: Hartung, Steinman, Comi, Knappertz, and Croze.
Critical revision of the manuscript for important intellectual content:
Steinman, Goodin, Cook, Filippi, O’Connor, Jeffery, Kappos, Axtell, Knappertz, Bogumil,
Schwenke, Croze, Sandbrink, and Pohl.
Statistical analysis: Steinman, Knappertz, Schwenke, and Croze.
Obtained funding: Knappertz and Sandbrink.
Administrative, technical, and material support: Hartung, Steinman, Comi, Cook,
Filippi, Jeffery, Croze, and Sandbrink.
Study supervision: Hartung, Steinman, Comi, Jeffery, Knappertz, Bogumil, Croze,
Sandbrink, and Pohl.
Conflict of Interest Disclosures: Dr Hartung
received honoraria for consultancy and lectures with approval by the Rector of
Heinrich-Heine-Universität from Bayer Pharma, Biogen Idec, Genzyme, Hoffman–La Roche,
Merck Serono, Novartis Pharma, and Teva–sanofi-aventis. Dr Steinman has served as a consultant
for Bayer, Novartis, MedImmune, sanofi-aventis, Teva, and Biogen Idec. Dr Comi has received
honoraria for consultancy from Bayer, Novartis, Teva, sanofi-aventis, Merck Serono, Actelion, and
Genzyme and has received honoraria for invited lectures from Bayer, Novartis, Teva, sanofi-aventis,
Merck Serono, Biogen, Serono Symposia International Foundation, and Genzyme. Dr Cook has received
research support from Bayer HealthCare and consultation fees from Bayer HealthCare, Merck Serono,
Biogen Idec, Teva, and sanofi-aventis. Dr Filippi serves on scientific advisory boards for Teva and
Genmab, has received funding for travel from Bayer Schering Pharma, Biogen Idec, Genmab, Merck
Serono, and Teva; serves as a consultant to Bayer Schering Pharma, Biogen Idec, Genmab, Merck
Serono, Pepgen Corp, and Teva; serves on speakers’ bureaus for Bayer Schering Pharma, Biogen
Idec, Genmab, Merck Serono, and Teva; receives research support from Bayer Schering Pharma, Biogen
Idec, Genmab, Merck Serono, Teva, Fondazione Italiana Sclerosi Multipla, the Italian Ministry of
Health, and CurePSP; is editor in chief of Journal of Neurology; and serves on the
editorial boards of the American Journal of Neuroradiology, BMC
Musculoskeletal Disorders, Clinical Neurology and Neurosurgery,
Erciyes Medical Journal, Journal of Neuroimaging, Journal
of Neurovirology, Magnetic Resonance Imaging, Multiple
Sclerosis, Neurological Sciences, and Lancet Neurology.
Dr Jeffery has received honoraria for speaking and consulting from Bayer, Biogen Idec, Teva, Serono,
Pfizer, GlaxoSmithKline, Novartis, Acorda, Genzyme, and Questcor and has received research support
from Bayer, Biogen Idec, Teva, Serono, Pfizer, and Novartis. The University Hospital Basel as
employer of Dr Kappos has received and dedicated to research support fees for board membership,
consultancy, or speaking, or grants, in the last 3 years from Actelion, Advancell, Allozyne, Bayer,
Bayhill, Biogen Idec, BioMarin, CSL Behring, Eli Lilly, European Union, Genmab, GeNeuro, Gianni
Rubatto Foundation. Glenmark, Merck Serono, MediciNova, Mitsubishi Pharma, Novartis, Novartis
Research Foundation, Novonordisk, Peptimmune, Roche, Roche Research Foundation, Santhera,
sanofi-aventis, Swiss MS Society, Swiss National Research Foundation, Teva, UCB, and Wyeth. Dr
Sandbrink holds stocks and stock options in Bayer Pharma AG. Dr Pohl holds stock options in Bayer
Funding/Support: This work was supported by
Bayer Pharma AG, Berlin, Germany.
Role of the Sponsor: Bayer Pharma AG was involved in the design and conduct of the
study; collection, management, analysis, and interpretation of the data; and preparation, review,
and approval of the manuscript.
Additional Contributions: Editorial assistance for the development of the manuscript
was provided by S. Dougherty, AxioMed Communications, Inc, and was supported by Bayer Healthcare