CAR +and CAR −T cells differentiate into an NK-like subset that is associated with increased inflammatory cytokines following infusion

Abstract

Abstract Chimeric antigen receptor (CAR) T cells have demonstrable efficacy in treating B-cell malignancies. Factors such as product composition, lymphodepletion and immune reconstitution are known to influence CAR +T cell survival and persistence, however the determinants of CAR +T cells’ fate and differentiation and how the patients’ reconstituting immune system influence therapy outcomes remain poorly understood. We applied single cell multi-omics analysis of RNA, protein and clonal profiles of CAR +T and PBMC in patients receiving donor-derived products from a piggybac CAR19 clinical trial (N=8 patients). We used these data to reconstruct a differentiation trajectory, which explained the observed phenotype and fate of CAR +and CAR −T cells within the first 30 days of post-infusion. Following initial lympho-depletion, endogenous CAR −CD8 +, γδ T cells and CAR+ T cells clonally expanded, and differentiated across heterogenous phenotypes. They transitioned from a dominant resting or proliferating state into precursor of exhausted T cells, and finally into a terminal NK-like phenotype. The subset of terminal NK-like CAR +T cells was associated with increased serum concentrations of proinflammatory cytokines including IL-12 and IL-18. Validation on a published cohort (61 patients) showed that this subset was associated with clinical outcome at 6 months. Finally, we analysed in detail one patient that developed CAR T cell induced lymphoma and discussed the impact of early phase therapy in determining this unexpected outcome. These results demonstrate that CAR- and CAR+ T cells share a differentiation profile which may be determined by non-CAR derived signals and which may influence patients’ immune-recovery and outcome. National Health And Medical Research Funding (Australia) (APP1121643, APP1128416) BD Bioscience