Dynamic changes in the bone marrow niche following CD19-targeted CAR-T cell infusion in diffuse large B cell lymphoma (DLBCL).

Abstract

Abstract Introduction CAR T cells have changed the treatment landscape of relapse/refractory (RR) DLBCL, offering a curative option. About half of the patients will not be cured with CAR-T and will experience poor outcomes. One potential mechanism of CAR-T resistance involves T-cell dysfunction and an immunosuppressive tumor microenvironment (TME). CAR-T products and blood biomarkers have been extensively studied in that regard. We postulate that the bone marrow niche is essential for T-cell functionality and circulation and may impact CAR-T responses. Here, we present the role of the dynamic immune changes in the bone marrow during CAR-T treatment. Methods To characterize the dynamic changes in the bone marrow niche in patients receiving CAR T cell (axicabtagene ciloleucel) treatment, we obtained bone marrow 3-5 weeks pre–CAR T infusion and 30 to 60 days post-CAR T infusion. We used single-cell RNAseq 10X sequencing with TCR enrichment to characterize both gene expression and TCR sequences. We characterized a total of 15 samples (9 pre- and six post-CAR) from 9 patients, with six patients having paired bone marrow samples before and after CAR treatment. We also performed profiling in normal bone marrow as a control. In addition to comparing treatment outcomes, we associated the immune population with ferritin levels (500 ng/mL) at the time of Leukocyte apheresis and 1 day before lymphodepletion. Results We characterized 91418 cells, of which 26106 cells were CD3-expressing T-cells. Our data did not reveal the presence of gamma retrovirus sequences, suggesting the presence of non-transduced T-cells. TCR analysis revealed clonal expansion of non-CAR-T CD3+ T-cells following CAR T infusion, indicating a possible endogenous immune response, as well as that the bone marrow serves as a circulating niche for these T-cell populations. In patients with DLBCL who responded to CAR T therapy, the bone marrow was enriched with gamma delta-like T cells undergoing proliferation. Interestingly, three of the four patients also had high ferritin levels at the time of lymphodepletion, with a decrease in ARG1+ neutrophils and GMP-like cells. Altogether, the result highlighted that maintenance of inflammation is associated with favorable responses. Conclusion Our study revealed dynamic changes in innate and effector immune cells before and after treatment, providing critical insights that will enable a better understanding of the immunological factors governing CAR efficacy, as well as future optimization of CAR T-cell therapies in the DLBCL setting.