Chimeric Antigen Receptor (CAR)-T cell therapy, a form of genetically engineered T cell therapy, has shown substantial anti-tumor activity against B cell malignancies. Despite this, approximately 30% of non-Hodgkin lymphoma patients still experience tumor relapse after CAR-T treatment. Multiple factors can cause tumor cells to be resistant to CAR-T therapy, including: 1) modulation of the tumor cell’s apoptotic sensitivity to T cell-mediated killing (e.g., increased inhibitors for granzyme-mediated proteolytic activity and reduced expression of pro-apoptotic regulators associated with apoptosis), and 2) loss of tumor antigen (e.g., CD19) expression recognized by CAR-T cells. Therefore, there is a pressing need to develop novel strategies to overcome tumor resistance to CAR-T therapy for better clinical responses.

To address this, we hypothesize that combining antibody therapy with CAR-T could be a feasible solution to overcome the limitations of CAR-T therapy. This hypothesis is based on: 1) antibodies can induce target cell lysis via multiple pathways (e.g., Complement-dependent cytotoxicity (CDC) and antibody-dependent cellular phagocytosis (ADCP)), which are independent of T cell-mediated killing (e.g., Granzyme/perforin pathway). Thus, antibody combinations can sensitize tumor cells to apoptosis even when they acquire resistance to T cell-mediated killing. 2) antibody combinations can also overcome antigen escape after CAR-T therapy by targeting orthogonal tumor antigens different from CD19 (e.g., CD20 and CD22).

We first evaluated rituximab in combination with CAR-T therapy to test our hypothesis. Rituximab is a monoclonal antibody targeting CD20, the standard therapy for treating B-cell lymphoma patients. Upon the addition of rituximab, we observed that tumor cell lysis significantly increased compared to either CAR-T or rituximab alone in vitro and in vivo. This demonstrates that antibody therapy can be an effective combination with CAR-T therapy. However, patients treated with CAR-T may relapse or be refractory to prior therapy that includes rituximab, suggesting that lymphoma cells in these patients could be less sensitive to rituximab treatment. A prior study identified that the upregulation of CD55, a decay-accelerating factor that inhibits the cascade of CDC, in B-cell lymphoma positively correlates with rituximab resistance. Therefore, we further tested a novel antibody targeting CD20 and CD55 (i.e., anti-CD20xCD55 antibody) in combination with CAR-T therapy. Interestingly, the anti-CD20xCD55 antibody dramatically improved the combination effect with CAR-T in the rituximab-resistant cell line. Finally, we tested whether antibody combination can overcome antigen escape in CAR-T therapy. We found that a combination of CD20 targeting antibody with CAR-T completely eradicated both CD19+ and CD19- lymphoma cells, while CAR-T single treatment failed to eliminate CD19- tumors.

In conclusion, our study demonstrates that combining antibody therapy with CAR-T therapy can be a highly effective strategy to improve the anti-tumor activity of CAR-T therapy and can be readily translated into clinical application.