A novel T-cell bispecific antibody platform for efficient T-cell mediated killing of tumor cells with minimal cytokine release.

Abstract

209 Background: Bispecific antibodies that recruit cytotoxic T cells to kill tumor cells are popular due to their targeted mechanism of action. Despite their attractiveness, there are limitations in the clinic due to undesirable toxicities associated with cytokine release. We describe here a platform for generation of a large collection of human anti-CD3 antibodies obtained from custom transgenic rats. Combining these unique anti-CD3 arms with different tumor targeting arms enables creation of bispecific antibodies of varying tumor cell killing capability. These anti-CD3 arms were combined with tumor antigen binding arms, namely, an anti-BCMA arm for multiple myeloma therapy, or an anti-CD22 arm for B-cell acute lymphoblastic leukemia therapy. Methods: Our platform utilizes a discovery approach involving antibody repertoire deep sequencing, high-throughput gene assembly, and recombinant expression, generating a highly diverse panel of antibodies with varying affinities. The CD3 antibodies were tested in in vitro T cell assays using human PBMCs to measure activation and cytokine release. Bispecific antibodies were evaluated for their ability to kill target cell lines upon co-culture with primary human PBMCs. The in vivo efficacy of bispecific antibodies was evaluated in a xenograft mouse model. Results: The in vitro T cell activity of these antibodies as measured by interleukin-2, interferon gamma levels and upregulation of the activation marker CD69 covered a broad spectrum of EC50 values. In co-culture systems with human PBMCs, anti-BCMA or anti-CD22 bispecific antibodies potently killed their respective target expressing cells with varying strengths. Additionally, the cytokine release from T cell activation correlated with the affinity of the anti-CD3 arms. The in vivo efficacy of the bispecific antibodies in a xenograft model with human PBMCs transferred into NSG mice showed striking tumor clearance at a wide range of doses. Conclusions: Our platform is highly suitable for creation of an extensive collection of bispecific antibodies for a variety of disease models by selecting the ideal anti-CD3 arm for each unique tumor antigen binding arm.