RNA sequencing to predict response to TNF-α inhibitors reveals possible mechanism for nonresponse in smokers

ABSTRACT

Background: Several studies have employed microarray-based profiling to predict response to tumor necrosis factor-alpha inhibitors (TNFi) in rheumatoid arthritis (RA); yet efforts to validate these targets have failed to show predictive abilities acceptable for clinical practice.

Methods: The eighty most extreme responders and nonresponders to TNFi therapy were selected from the observational BiOCURA cohort. RNA sequencing was performed on mRNA from peripheral blood mononuclear cells (PBMCs) collected before initiation of treatment. The expression of pathways as well as individual gene transcripts between responders and nonresponders was investigated. Promising targets were technically replicated and validated in n = 40 new patients using qPCR assays.

Results: Before therapy initiation, nonresponders had lower expression of pathways related to interferon and cytokine signaling, while also showing higher levels of two genes, GPR15 and SEMA6B (p = 0.02). The two targets could be validated, however, additional analyses revealed that GPR15 and SEMA6B did not independently predict response, but were rather dose-dependent markers of smoking (p < 0.0001).

Conclusions: The study did not identify new transcripts ready to use in clinical practice, yet GPR15 and SEMA6B were recognized as candidate explanatory markers for the reduced treatment success in RA smokers.

KEYWORDS:

• mRNA
• prediction
• qPCR
• response
• rheumatoid arthritis
• RNA sequencing
• TNF-alpha

Acknowledgments

We thank Katja Coeleveld, Kim van der Wurff-Jacobs and Dorien van de Berg, for bio-banking, Karin A.L. Schrijvers-te-Brake, Annemiek Sloeserwij, Joke Nijdeken and Marieke Vianen who facilitated in the acquisition of data, Cornelis Bekker for his technical assistance, Aridaman Pandit for his aid in statistical analyses and the Society for Rheumatology Research Utrecht (SRU) for including patients. We are grateful to all of the patients who have participated in this study.

Contributing SRU hospitals and their contact persons: University Medical Center Utrecht, dr. J. Tekstra; Antonius Hospital Nieuwegein/Utrecht, dr. E.J. ter Borg; Diakonessen Hospital Utrecht, drs. Y. Schenk Meander Medical Center Amersfoort, dr. S. Linn-Rasker; Sint Maartenskliniek Woerden, dr. W.H. van der Laan; Hospital St. Jansdal Harderwijk, drs. D.G. Kuiper-Geertsma; Tergooi Hospital Hilversum, dr. M. Nabibux; Flevo Hospital Almere, dr. C.M. Verhoef.

Authors’ contributions

Bart Cuppen collected clinical data, performed experiments, performed statistical analyses and drafted the manuscript, Marzia Rossato coordinated the experiments, aided in statistical analyses and interpretation of the results and drafted the manuscript, Ruth Fritsch-Stork included patients, coordinated the study and helped to draft the manuscript, Arno N. Concepcion performed qPCR experiments and helped to draft the manuscript, Suzanne Linn-Rasker included patients and helped to draft the manuscript, Jaap van Laar included patients and helped to draft the manuscript, Floris P.J.G. Lafeber conceived and coordinated the study and helped to draft the manuscript, Johannes W.J. Bijlsma conceived and coordinated the study, included patients and helped to draft the manuscript, and Timothy R.D.J. Radstake conceived the study and helped to draft the manuscript. All authors read and approved the final manuscript.

Declaration of interest

JWJ Bijlsma received research grants (to his department) and consultancy fees from AbbVie, BMS, Crescendo, MSD, Mundipharma, Pfizer, Roche, Sun and UCB. TRD Radstake received grants and consultancy fees from AbbVie, BMS, Pfizer, Roche, Lilly and UCB. JM van Laar received grants from Roche, MSD, Eli Lilly and Pfizer. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.

Reviewer disclosures

Peer reviewers on this manuscript have no relevant financial or other relationships to disclose.

Supplementary Material

Supplemental data for this article can be accessed here.

Additional information

Funding

This work was supported by AbbVie, ZonMW: The Netherlands Association for Health Research and Development under Grant number 152001008, Foundation Friends of the University Medical Center Utrecht based on a private gift and unrestricted grants from Pfizer, BMS, UCB, Roche and Schering-Plough. Marzia Rossato was funded by IEF Marie Curie fellowship under Grant number 622811 (MicroSCAP). These funders provided financial support to this research but did not have any additional role in study design, data collection and analysis, or preparation of the manuscript.