Fully human antibody v H domains to generate mono and bispecific CAR to target solid tumors

Abstract

Background Chimeric antigen receptor (CAR) T cells are effective in B-cell malignancies. However, heterogeneous antigen expression and antigen loss remain important limitations of targeted immunotherapy in solid tumors. Therefore, targeting multiple tumor-associated antigens simultaneously is expected to improve the outcome of CAR-T cell therapies. Due to the instability of single-chain variable fragments, it remains challenging to develop the simultaneous targeting of multiple antigens using traditional single-chain fragment variable (scFv)-based CARs. Methods We used Humabody V H domains derived from a transgenic mouse to obtain fully human prostate-specific membrane antigen (PSMA) V H and mesothelin (MSLN) V H sequences and redirect T cell with V H based-CAR. The antitumor activity and mode of action of PSMA V H and MSLN V H were evaluated in vitro and in vivo compared with the traditional scFv-based CARs. Results Human V H domain-based CAR targeting PSMA and MSLN are stable and functional both in vitro and in vivo. V H modules in the bispecific format are capable of binding their specific target with similar affinity as their monovalent counterparts. Bispecific CARs generated by joining two human antibody V H domains can prevent tumor escape in tumor with heterogeneous antigen expression. Conclusions Fully human antibody V H domains can be used to generate functional CAR molecules, and redirected T cells elicit antitumoral responses in solid tumors at least as well as conventional scFv-based CARs. In addition, V H domains can be used to generate bispecific CAR-T cells to simultaneously target two different antigens expressed by tumor cells, and therefore, achieve better tumor control in solid tumors.