Characterization of novel dual tandem CD19/BCMA chimeric antigen receptor T cells to potentially treat multiple myeloma

Abstract

Background: Treatment with chimeric antigen receptor (CAR)-engineered T cells directed against the B-cell maturation antigen (BCMA) promoted transient recovery from multiple myeloma (MM). However, the absence of this antigen on immature plasma cells may limit the efficacy of this modality and facilitate relapse. The purpose of this study is to characterize a novel CAR that includes both a single-chain variable fragment (scFv)-BCMA and an scFv-CD19 in tandem orientation (tan-CAR) in an attempt to target both BCMA and CD19 expression on MM cells. Method: The scFv sequences from the anti-CD19 antibody FMC63 and the anti-BCMA antibody C11D5.3 were ligated in tandem with transmembrane and T-cell signaling domains to generate the tan-CAR construct. Specificity and efficacy of activated tan-CAR T cells were analyzed using in vitro proliferation, cytokine release, and cytolysis assays. We also evaluated the in vivo efficacy with a xenograft mouse model that included target tumor cells that expressed CD19 or BCMA and compared the results to those obtained with conventional CAR T cells. Results: The in vitro studies revealed specific activation of tan-CAR T cells by K562 cells that overexpressed CD19 and/or BCMA. Cell proliferation, cytokine release, and cytolytic activity were all comparable to the responses of single scFv CAR T cells. Importantly, in vivo studies of tan-CAR T cells revealed specific inhibition of tumor growth in the mouse xenograft model that included cells expressing both CD19 and BCMA. Systemic administration of tan-CAR T cells resulted in complete tumor remission, in contrast to the reduced efficacies of BCMA-CAR T and CD19-CAR T alone in this setting. Conclusion: We report the successful design and execution of novel tan-CAR T cells that promote significant anti-tumor efficacy against both CD19 and BCMA antigen-positive tumor cells in vitro and in vivo . The data from this study reveal a novel strategy that may help to reduce the rate of relapse in the treatment with single scFv-CAR T cells.

Physalin B inhibits cell proliferation and induces apoptosis in undifferentiated human gastric cancer HGC‐27 cells

“…Data are expressed as the mean ± SD based on at least three independent experiments analyzed by GraphPad Prism software (Version 6.0, San Diego, CA, USA). 26 The significance of differences between groups was assessed by one-way analyses of variance (ANOVA) followed by Tukey's test for multiple comparisons using SPSS software 16.0 (Chicago, IL, USA). A two-tailed p-value <0.05 and a p-value <0.1…”

The antigen‐binding moiety in the driver's seat of CARs

“…TandemCARs encode multiple antigen‐binding domains in a single CAR molecule (Figure 4.V). This has been described with multiple scFvs, 140,143–147 but a general risk with these TandemCARs, is the potential cross‐pairing between the V L and V H chains of different scFvs. Therefore, TandemCARs incorporating combined scFv/muteins, 148 scFv/natural ligands, 146 multiple VHHs, 104 and combined protein scaffolds (see below) 149 have been reported.…”