104?BCMA-targeting CAR-T cells expanded in IL-15 have an improved phenotype for therapeutic use compared to those grown in IL-2 or IL-15/IL-7

摘要

Background Chimeric antigen receptor-T (CAR-T) cells that target B cell maturation antigen (BCMA-CARs) have emerged as a promising treatment for multiple myeloma (MM). Despite impressive initial responses to BCMA-CAR therapy in clinical trials, relapse is common, signifying a need to improve the in vivo efficacy and persistence of BCMA-CARs. 1 The development of unfavourable differentiation or T cell dysfunction, such as exhaustion and senescence, during the ex vivo expansion of the BCMA-CARs could be limiting their therapeutic potential. For CD19-directed CARs, reduced dysfunction and differentiation and improved anti-tumour responses were achieved by expanding the cells with IL-15 instead of IL-2. 2 Therefore, in this study, our aim was to determine whether expanding BCMA-CARs with IL-15 or IL-15/IL-7 instead of IL-2 alters their levels of exhaustion, senescence, differentiation and activity. Methods T cells stimulated with anti-CD3/anti-CD28-coated beads were supplemented with IL-2, IL-15, IL-15 IL-7 or no cytokine and transduced with ARI2h, a BCMA-CAR with a 4-1BB co-stimulatory domain produced at our institution. 3 Expanded BCMA-CARs were analysed by flow cytometry for markers of T cell dysfunction, or challenged with MM cell line ARP-1 and then tested for cytokine production, cytotoxic ability and activation signals. Results BCMA-CARs cultured in IL-15 or IL-15/IL-7 expanded similarly to those grown in IL-2, with comparable CAR transduction efficiencies, CD4:CD8 ratios and proliferation rates. BCMA-CARs grown in IL-15 had low expression of exhaustion marker LAG-3 and high expression of the costimulatory molecule CD27, which is important for T cell survival and persistence, when compared to BCMA-CARs cultured in IL-2. Moreover, BCMA-CARs grown solely in IL-15 were less differentiated than those supplemented with IL-7, and had higher expression of stem cell memory marker CXCR3 within the na?ve population than those expanded with IL-2. When challenged with MM cell line ARP-1, IL-15-grown BCMA-CARs upregulated activation marker CD69, exhibited strong cytotoxicity and robust production of IFNγ and IL-2. However, in comparison to BCMA-CARs expanded in IL-2 or IL-15/IL-7, those grown with IL-15 had lower mTORC1 activity and p38 MAPK phosphorylation when activated by ARP-1 cells, suggesting differential regulation of key pathways for T cell metabolism and senescence, respectively. Conclusions To summarise, BCMA-CARs expanded with IL-15 alone exhibited the most favourable phenotype for therapeutic use compared those grown with IL-2 or IL-15/IL-7. Future experiments using murine MM models will be critical in understanding the in vivo benefits or drawbacks of culturing BCMA-CARs in IL-15 compared to IL-2 or IL-15/IL-7. Ethics Approval Research involving human material was approved by the Ethical Committee of Clinical Research (Hospital Clinic, Barcelona). Peripheral blood T cells were obtained from healthy donors after informed consent in accordance with the Declaration of Helsinki.