EXPANSION OF ISLET REACTIVE MEMORY CD4 T CELLS IN DIFFERENT STAGES OF T1D AND AFTER THERAPY

Islet antigen reactive memory CD4 (IARM-CD4) T cells contribute to the development of type 1 diabetes (T1D), and are detected in peripheral blood before and after diagnosis. We used a CD154 activation assay in conjunction with single cell RNAseq of IARM-CD4 T cells to identify novel characteristics of circulating IARM-CD4 T cells during disease progression and response to therapy in order to determine their relationship to T1D pathogenesis. A cross sectional cohort of healthy controls (non-risk, n=8), new onset T1D (stage 3, n=8), and longstanding T1D (stage 4, n=9) were studied, including longitudinal samples from a subset of stage 4 T1D, as well as subjects with stage 3 T1D treated with alefacept (LFA-3-Fc, n=8). We detected a significant clonal expansion of IARM-CD4 T cells in stage 3 and 4 T1D subjects compared with healthy controls, suggesting in vivo expansion during disease progression. Expanded clonotypes were stable over time in some subjects, even after treatment with alefacept. However, there was a reduction in IARM-CD4 T cell frequency and a trend towards increased diversity of T cell receptor (TCR) sequences (decreased expansion) following alefacept treatment. The majority of expanded TCR sequences were private, although several TCR beta chain CDR3 junctions matched sequences found in secondary lymphoid tissue of T1D donors. Re-expression of expanded TCRs identified IGRP as a major antigenic target. Our results suggest IARM-CD4 T cells are expanded prior to disease onset and may persist during disease progression. T cell-targeted therapy may alter the frequency and diversity of IARM-CD4 T cells.