Abstract
Chimeric antigen receptor (CAR) T-cell therapy has produced remarkable anti-tumor responses in patients with B-cell malignancies. However, clonal kinetics and transcriptional programs that regulate the fate of CAR-T cells after infusion remain poorly understood. Here we perform TCRB sequencing, integration site analysis, and single-cell RNA sequencing (scRNA-seq) to profile CD8+ CAR-T cells from infusion products (IPs) and blood of patients undergoing CD19 CAR-T immunotherapy. TCRB sequencing shows that clonal diversity of CAR-T cells is highest in the IPs and declines following infusion. We observe clones that display distinct patterns of clonal kinetics, making variable contributions to the CAR-T cell pool after infusion. Although integration site does not appear to be a key driver of clonal kinetics, scRNA-seq demonstrates that clones that expand after infusion mainly originate from infused clusters with higher expression of cytotoxicity and proliferation genes. Thus, we uncover transcriptional programs associated with CAR-T cell behavior after infusion.
Highlights
Chimeric antigen receptor (CAR) T-cell therapy has produced remarkable anti-tumor responses in patients with B-cell malignancies
We examine the T cell receptor beta (TCRB) repertoire and lentiviral integration sites of CD8+ CAR-T cells isolated from the infusion products (IPs) and from blood of patients treated with CD19-targeted CAR-T cell immunotherapy
To better understand changes in the composition of CAR-T cells after infusion, we studied a cohort of patients (n = 10) who received CD19-specific CAR-T cells manufactured from bulk CD4+ T cells and CD8+ central memory-enriched (TCM) cells, which were infused in a 1:1 ratio of CD4+:CD8+ CAR-T cells
Summary
Chimeric antigen receptor (CAR) T-cell therapy has produced remarkable anti-tumor responses in patients with B-cell malignancies. We previously reported in a small subset of patients with acute lymphoblastic leukemia (ALL) that CAR-T cells were polyclonal, both in the IP and at the peak of expansion in the recipient[4] Another group reported one patient in whom a single CD8+ CAR-T cell clone, in which the transgene had integrated into the TET2 locus, dominated at the peak of in vivo expansion[19]. These highly disparate patterns suggest variability in the clonal composition of infused CAR-T cells and potential differences in the ability of individual CAR-T cell clones to expand after adoptive transfer. Using single-cell RNA sequencing (scRNA-seq), we identify transcriptionally distinct clusters of infused CD8+ CAR-T cells that differ in their contribution to the CAR-T cell repertoire in blood after infusion
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