Data from Specific targeting of Multiple Myeloma by Dual Split-signaling Chimeric Antigen Receptors T-cells directed against CD38 and CD138

AbstractPurpose: The success of BCMA-specific chimeric antigen receptor (CAR) T-cells illustrates the potential of this novel therapy for multiple myeloma (MM). Nonetheless, broadening CAR T-cell therapy beyond BCMA requires inventive strategies as there are only a few MM- or plasma cell-specific target antigens. We investigated the feasibility of achieving MM specificity by dual-split CD38/CD138 CAR targeting, whereby the stimulatory and costimulatory signals for T-cell activation are split into two separate stimulatory (sCAR) and costimulatory CARs (cCAR). Experimental Design: Using various combinations of CD38 and CD138 sCARs and cCARs with different affinities, we generated several dual-split CAR T-cells and analyzed them for MM-specific effector functions in vitro. The best-functioning CAR T-cells were tested in vivo in a murine xenograft model. Results: We found optimal designs of both CD38sCAR/CD138cCAR and CD138sCAR/CD38cCAR combinations, that effectively lysed MM cells, but spared single CD38 or CD138 positive healthy hematopoietic cells. While the CD38sCAR/CD138cCAR T-cells achieved MM-specific activity solely due to the low affinity of the CD38sCARs, the MM-specific cytotoxicity, cytokine release, and proliferation of CD138sCAR/CD38cCAR T‑cells were established through a true combinatorial stimulatory and costimulatory effect. The most optimal combination comprised a low affinity CD138sCAR combined with a high affinity CD38cCAR. These CD138sCAR/CD38cCAR T-cells also showed dual-antigen specific anti-MM effects in vivo. Importantly, they were also effective against MM cells from daratumumab pre-treated patients with decreased CD38 expression levels. Conclusions: We demonstrate the possibility to specifically target MM cells, even after CD38 targeted therapy, with carefully-designed dual-split CARs directed against CD38 and CD138.