Diversity of Immunoglobulin Heavy Chain Repertoire in Patients With Rheumatoid Arthritis

Abstract Objectives Rheumatoid Arthritis (RA) is associated with polymorphism in major histocompatibility complex class II genes and dysregulations of CD4+ T cells which cause abnormalities in immune repertoire (iR) expression and intracellular signaling. We monitored nucleotide sequence changes in iR of immunoglobulin heavy chain (IGH), particularly complementarity determining region 3 (CDR3) during the course of treatments in RA patients using massively parallel sequencing technology.Methods CDR3 sequencing was carried out on clinical blood samples from RA patients for disease progress monitoring. The iR of each sample was measured using next generation sequencing (NGS) pipeline. Data analysis was done with a web-based iRweb server. Principal components analysis (PCA) was completed with commercial statistical pipeline. Results Datasets from 14 patients covered VDJ regions of IGH gene. D50 stayed low for all cases (mean D50 = 6.5). A pattern of shared CDR3 sequences was confirmed by a clustering pattern using PCA. Shared profile of 608 CDR3 sequences unique to the disease baseline was identified. D50 analyses revealed clonal diversity would remain low throughout the disease course even after treatment (mean D50 = 11.7 & 8.2 for csDMARD & bDMARD groups respectively) regardless of fluctuated disease activity. PCA has provided a correlation of change in immune diversity along the whole course of RA. Conclusion We have successfully constructed the experimental design, data acquisition, processing, and analysis pipeline of a high throughput massively parallel CDR3 sequences detection to be used to correlate RA disease activity and IGH CDR3 iR during disease progression with or without treatments.