Single Cell Multi-Omic Dissection of Response and Resistance to Chimeric Antigen Receptor T Cells Against BCMA in Relapsed Multiple Myeloma

Background: Chimeric antigen receptor (CAR) T cell therapy has revolutionized treatment of relapsed/refractory multiple myeloma (RRMM). Robust variables that predict long-term response are currently missing. Limited data are especially available on the impact of bridging therapies on manufacturing and outcome. We conducted a longitudinal single-cell multi-omics study to identify factors that predict response to BCMA-directed CAR T cells. Changes in the immune microenvironment associated with response were analyzed as well as the impact of prior bridging therapy with bispecific antibodies on subsequent CAR T cell manufacturing and outcome. Methods: Peripheral blood mononuclear cells (PBMCs) were isolated from 29 consecutive MM patients treated with commercially available anti-BCMA CAR T cells on the day of leukapheresis as well as days 30 and 100 after CAR T cell infusion. PBMCs were subjected to single cell RNA, T-cell receptor (TCR) and B-cell receptor (BCR) sequencing. A custom panel of 57 oligonucleotide-coupled antibodies was used to study surface proteomics. Downstream analyses were performed with Seurat. Differences in cellular compositions at all three time points were analyzed with scCODA. To analyze CAR T cell functionality, CAR T cells from peripheral blood were isolated 7 days after infusion and subjected to an in vitro cytotoxicity assay after expansion and stimulation. Patients were grouped based on their best response following CAR T cell infusion (CR: n=12, non CR: n=17). Results: In total, 375,338 cells were sequenced (median 7246 cells/sample, range 1,569-10,972 cells) and 354,878 cells (94.5%) passed quality assessment. Quantitative and qualitative differences in the cellular composition of peripheral blood between CR and non CR patients were detected at the time of leukapheresis as well as on days 30 and 100 following infusion. CR patients harbored significantly more CD8+ effector memory T cells (TEM) at leukapheresis and less NK cells on day 30 after therapy compared to non CR patients. Regulatory T cells isolated at the time of leukapheresis from non CR patients exhibited significantly higher surface protein levels of CXCR3, CD40, CD95 and KLRG1 (p