Magill, Laura; (2019) Immune-phenotyping of B cells unsing LEGENDScreen(TM) to investigate the role of B cells in immunogenicity, and immune regulation by Bregs, in patients with Rheumatoid Arthritis. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Rheumatoid arthritis (RA) is a multifactorial disease associated with failure of immune tolerance. B cells play a prominent role in the pathogenesis of this disease, via the production of auto-antibodies, pro-inflammatory cytokines, and an impaired regulatory B cell (Breg) response. Biologic drugs have significantly advanced the treatment of RA but are not always effective and indeed the development of anti-drug antibodies (ADA) has been associated with a poorer clinical outcome. Here I set out, firstly, to identify a predictive biomarker that discriminates RA patients who are more likely to develop ADA in response to adalimumab, a human monoclonal antibody against tumor necrosis factor (TNF)α. By taking advantage of an immune-phenotyping platform, LEGENDScreenTM, I measured the expression of 332 cell surface markers on B and T cells in a cross-sectional adalimumab-treated RA patient cohort with a defined ADA response. The analysis revealed seven differentially expressed markers between the ADA+ and ADA- patients. Validation of the differentially expressed markers in an independent prospective European cohort of adalimumab treated RA patients, revealed a significant and consistent reduced frequency of signal regulatory protein (SIRP)α/β-expressing memory B cells in ADA+ versus ADA- RA patients. I also assessed the predictive value of SIRPα/β expression in a longitudinal RA cohort prior to the initiation of adalimumab treatment. I showed that a frequency of less than 9.4% of SIRPα/β-expressing memory B cells predicts patients that will develop ADA, and consequentially fail to respond to treatment, with a receiver operating characteristic (ROC) area under the curve (AUC) score of 0.92. Thus, measuring the frequency of SIRPα/β-expressing memory B cells in patients prior to adalimumab treatment may be clinically useful to identify a subgroup of active RA patients who are going to develop an ADA response and not gain substantial clinical benefit from this treatment. Secondly, further mining of the results from the LEGENDScreenTM has allowed me to identify a signature of RA. Comparison of HCs to RA patients identified 16 differentially expressed markers associated with RA, and not with disease activity or treatment. Validation in an independent cohort determined that the combined expression of CD97, CD170 and CD11c on B cells may identify individuals that are at risk of developing RA. Thirdly, I have identified a novel marker of Bregs. Here I show that CD19hiCD170hi B cells capture the majority of IL-10 producing B cells. This subset and not its negative counterpart (CD19+CD170int/low) suppress IFNγ and IL-17 production by CD4+ T cells, in an IL-10 and CD170 dependent manner. CD19hiCD170hi B cells are numerically reduced and functionally defective in RA patients. Preliminary data suggests that the aberrant production of IL-10 by Bregs in RA patients could be attributed to a defect in CD170 recycling. I propose CD170 may aid future immunological studies of Bregs, and that targeting CD170 therapeutically could improve disease in RA.

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