Prior chemotherapy deteriorates T-cell quality for CAR T-cell therapy in B-cell non-Hodgkin’s lymphoma

Immunotherapies, such as CAR T cell therapy, depend on T cells that are genetically modified to recognize and attack cancer cells. Their effectiveness thus hinges on the quality of a patient’s own T cells. We studied blood samples from B-NHL patients collected both before any treatment (pre-therapy) and following at least two lines of chemotherapy (post-therapy) to evaluate the potential negative effects of prior exposure to chemotherapy on T cell quality. Utilizing advanced multi-parameter flow cytometry and single-cell RNA sequencing (scRNAseq), we discovered notable alterations in T cell composition and gene expression after therapy, such as increasingly differentiated phenotypes and elevated expression of exhaustion markers. Additionally, scRNAseq and DNA methylation analyses revealed gene expression and epigenetic changes associated with diminished T cell function in post-therapy samples. In vitro cytotoxicity studies demonstrated superior killing efficacy of CAR T cells derived from T cells of treatment-naïve patients compared to T cells of patients with treatment history. These findings corroborate that using T cells collected prior to chemotherapy exposure could enhance the effectiveness of CAR T cell therapies  and offer new strategies for improving patient outcomes in B-NHL treatment. The Illumina Methylation EPIC Bead Chip was used to compare DNA methylation patterns of B-NHL patient samples. T cells were isolated before and after patient exposure to chemotherapy and methylation was compared on T cells at time of apheresis (n=22) as well as on the CAR T cell product (n=9).