Interconnected lineage trajectories link conventional and NK-like exhausted CD8+ T cells beneficial in T1D [P452_3]

While distinct NK-like CD57+ and PD-1+ CD8+ exhausted T cell populations (Tex) were both linked to beneficial immunotherapy response in autoimmune Type 1 Diabetes (T1D) patients, relationships between these cell types are poorly understood. We show that PD-1+ and CD57+ Tex populations in this context were epigenetically similar, but CD57+ Tex cells displayed unique increased chromatin accessibility of inhibitory Killer Cell Immunoglobulin-like Receptor (iKIR) and other NK cell genes. PD-1+ and CD57+ Tex also showed reciprocal expression of Inhibitory Receptors (IRs) and iKIRs accompanied by chromatin accessibility of Tcf1 and Tbet transcription factor target sites, respectively. CD57+ Tex showed unappreciated gene expression heterogeneity and shared clonal relationships with PD-1+ Tex, with these cells differentiating along four interconnected lineage trajectories: Tex-PD-1+, Tex-CD57+, Tex-Branching, and Tex-Fluid. Our findings demonstrate new relationships between Tex populations in human autoimmune disease and suggest that modulating common precursor populations may enhance response to autoimmune disease treatment. We performed bulk Assay for Transposase Accessible Chromatin with sequencing (ATAC-seq) on PBMC samples selected from 4 donors with new onset T1D that were responders to alefacept (T1DAL Trial, ITN). Donors were sampled at baseline (Week 0/Visit 0) and 104 wk (Visit 30) post-treatment with alefacept. PBMC samples were stained with a flow cytometry panel (Table S4) and sorted with the FACS Aria II (Becton Dickinson) cell sorter into 3 subsets of non-naïve CD8+ T cells (singlet, live, CD14- CD19- CD56- CD3+ CD8+ CD4-, not CD45RA+ CCR7+): (1) Double Negative non-Tex (DN, KLRG1- TIGIT-), (2) CD57+ Tex (KLRG1+TIGIT+, CD57+), and (3) CD57- Tex (KLRG1+TIGIT+, CD57-). These subsets were compared for downstream differential accesibility analysis.