Abstract Objective CD19-directed chimeric antigen receptor (CAR) T cells have yielded impressive response rates in refractory/relapse B-cell malignancies. However, loss of or low antigen expression can enable tumor escape and limit the duration of responses achieved with CAR T-cell therapy. Engineering bispecific CAR T cells that target two tumor antigens could overcome antigen-negative escape. CD79b is a critical receptor for successful B cell development that remains highly expressed in several subtypes of B cell lymphoma. The limited efficacy of antibody-drug conjugates (ADC) suggests that a CAR therapy targeting CD79b might improve results. Here, we develop a novel bispecific CAR T-cell product co-targeting CD79b and CD19 to enhance antitumor activity. Methods In this study, we developed novel anti-CD79b and anti-CD19 monoclonal antibodies using hybridoma technology and single B-cell cloning. These in-house-generated scFvs were engineered into four distinct 4-1BB-costimulated CAR constructs (CD79b-CAR, CD19-CAR, CD79b/CD19 loop CAR and CD79b/CD19 tandem CAR). Following lentiviral transduction into primary human T cells, we conducted comprehensive functional characterization through in vitro cytotoxicity assays and in vivo B-cell lymphoma xenograft models. Cellular characteristics and antitumor efficacy were compared through in vitro and in vivo B-cell lymphoma models. Results Our findings demonstrate high CD79b and CD19 expression in B-cell lymphoma cell lines, with a safe expression profile in healthy hematopoietic and non-hematopoietic tissues. We found that CD79b-specific CAR-T cells displayed high affinity and specificity for CD79b on B cells, whereas CD19-specific CAR-T cells showed analogous targeting of CD19. All four engineered CAR-T cell lines exhibited potent anti-tumor activity, effectively eliminating malignant B-cell lymphoma cells. They could be activated and produce inflammatory cytokines in a target-dependent manner. Moreover, in vivo tests showed that the CAR-T cells exhibited significant antitumour effects in a xenotransplanted B-cell lymphoma mouse model. Notably, CD79b/CD19 dual-targeting CAR-T cells showed higher efficacy than single-targeting CAR-T cells, particularly in B-cell lymphoma cases with phenotypically heterogeneous lymphoma populations. Conclusion Our results demonstrate that this novel CD79b/CD19 dual-targeting CAR-T cell therapy exhibits robust antitumor activity against B-cell lymphomas. These findings provide compelling preclinical evidence for the efficacy of bispecific CD79b/CD19 CAR-T cells against B-cell malignancies, supporting their clinical advancement.
Gao et al. (Mon,) studied this question.