CD19-CAR T Cells Develop Exhaustion Epigenetic Programs During a Clinical Response

Abstract

Abstract The goal of this study was to determine the epigenetic landscape of CD19-CAR T cells pre and post infusion in leukemia patients as an initial step to elucidate intrinsic mechanisms that limit CAR T-cell effector functions in humans. A longitudinal analysis of CD8+ CD19-CAR T cell epigenetic changes was performed by whole-genome DNA methylation profiling of CAR T cells during manufacturing and from peripheral blood mononuclear cells (PBMCs) of 15 patients enrolled on our institutional, autologous CD19-CAR T cell therapy study (NCT03573700). CAR T cell expansion and persistence were determined by measuring vector copy numbers in the PBMCs of treated patients. We had previously established novel exhaustion DNA methylation datasets that delineate between progenitor and fully exhausted T cells. These datasets served as a guide for stratifying our post-infusion CAR T cells along the exhaustion developmental trajectory. Our data show that CD19-CAR T cells lose repressive DNA methylation at effector loci (e.g. PRF1, TBET) while gaining methylation at genes associated with memory potential (e.g. LEF1, TCF7). We confirmed these epigenetic changes are coupled to endogenous human T cell effector and memory differentiation by cross-referencing our epigenetic data with publicly available transcriptional profiles for antigen-specific effector and long-lived memory CD8 T cells from individuals vaccinated for yellow fever. Furthermore, we show that CAR T cells were unable to mount an in vivo recall response after relapse of antigen-positive disease or recovery of endogenous B cells. These observations support the conclusion that CD19-CAR T cells acquire stable epigenetic exhaustion programs that limit their protective capacity. This work was supported by the National Institutes of Health (1R01AI114442 to BY and LRP to CCZ), the National Comprehensive Cancer Network Young Investigator Award (to CZ), Alex’s Lemonade Stand Foundation Young Investigator Grant (to CZ), Stand Up to Cancer- SU2C (to BY), the American Lebanese Syrian Associated Charities (ALSAC) to BY, and Assisi foundation to BY.