Single cell transcriptomic analysis of renal allograft rejection reveals novel insights into intragraft TCR clonality.

Bulk analysis of renal allograft biopsies (rBx) identified RNA transcripts associated with acute cellular rejection (ACR); however, these lacked cellular context critical to mechanistic understanding. We performed combined single cell RNA transcriptomic and TCRα/β sequencing on rBx from patients with ACR under differing immunosuppression (IS): tacrolimus, iscalimab, and belatacept. TCR analysis revealed a highly restricted CD8 T cell clonal expansion (CD8 ), independent of HLA mismatch or IS type. Subcloning of TCRα/β cDNAs from CD8 into Jurkat76 cells (TCR ) conferred alloreactivity by mixed lymphocyte reaction. scRNAseq analysis of CD8 revealed effector, memory, and exhausted phenotypes that were influenced by IS type. Successful anti-rejection treatment decreased, but did not eliminate, CD8 , while CD8 were maintained during treatment-refractory rejection. Finally, most rBx-derived CD8 were also observed in matching urine samples. Overall, our data define the clonal CD8 T cell response to ACR, providing novel insights to improve detection, assessment, and treatment of rejection.