Anti-CD38-CAR IPS-Derived Cytotoxic T Lymphocytes Efficiently Eliminate Myeloma Cells Treated with Daratumumab and Isatuximab in Vitro and In Vivo

Patients with myeloma have improved prognosis with proteasome inhibitor (PI), immunomodulatory drugs (IMiDs), and antibodies, especially anti-CD38 antibodies such as daratumumab and isatuximab. However, even introduction of these novel therapeutics still gives dismal prognosis in relapsed and refractory myeloma patients. Recently, although anti-BCMA-CAR-T therapy is clinically available for these patients, this option has profoundly problems including long-time preparation of CAR-T cells and limited number of suitable patients. We previously developed anti-CD38-CAR-T cells and reported that these CAR-T cells efficiently eliminated myeloma cells. To use anti-CD38-CAR-T cells as off-the-shelf, we tried to develop anti-CD38-CAR iPS-derived cytotoxic T lymphocytes (iPS-rCTLs). Firstly, we investigated whether CD38 express on iPS-rCTLs because CD38 is induced or enhanced on T cells in activation for viral transduction. Interestingly, iPS-rCTLs never expressed CD38 even in the case of activation. Thus, we never needed addition of anti-CD38 antibody in the culture medium for blockade of interaction of CD38 and anti-CD38-CAR or knockdown/knockout of CD38 on T cells. Accordingly, we retrovirally transduced iPS-rCTLs with anti-CD38-CAR and then we obtained anti-CD38-CAR iPS-rCTLs with >70% of transduction efficiency. Secondly, we performed co-culture experiments of anti-CD38-CAR iPS-rCTLs and myeloma cells in vitro. Using flow cytometric analysis, these cells efficiently abrogated myeloma cells in time-dependent and in cell number-dependent manner (48-hour incubation of both cells at E:T ratio of 1:2 showed >90 % of cytotoxicity with anti-CD38-CAR iPS-rCTLs). xCELLigence cell analyzer also got results which were almost consistent with those of flow cytometry. Next, we tried to investigate the cytotoxicity of anti-CD38-CAR iPS-rCTLs in the xenograft model of NOG mice inoculated with RPMI8226 cells. Following inoculation of RPMI8226 cells (1x106), anti-CD38-CAR iPS-rCTLs were intravenously injected into mice (1 x 106) twice. Intriguingly, anti-CD38-CAR iPS-rCTLs clearly inhibited myeloma growth in xenografted mice until day 30. Next, we investigated their efficacy to myeloma cells treated with daratumumab or isatuximab. Surprisingly, anti-CD38-CAR iPS-rCTLs efficiently killed myeloma cells in the presence of daratumumab or isatuximab in vitro. Now that, we are investigating the cytotoxicity of anti-CD38-CAR iPS-rCTLs in xenografted mice model of myeloma in vivo. In conclusion, we obtained the cytotoxic results of anti-CD38-CAR iPS-rCTLs against myeloma cells in vitro in the presence of daratumumab or isatuximab. And anti-CD38-CAR iPS-rCTLs definitively inhibited the myeloma growth in vivo. These results suggest that the epitope recognizable by anti-CD38-CAR iPS-rCTLs was different from that of daratumumab and isatuximab. Anti-CD38-CAR iPS-rCTLs could shed light on myeloma patients as an off-the-shelf, leading to the increased number of the cutting edged therapeutic option.