Enhanced cytotoxicity against solid tumors by bispecific antibody-armed CD19 CAR T cells: a proof-of-concept study

Abstract

Although adoptive cell therapy with chimeric antigen receptor (CAR)-engineered T cells has shown durable clinical efficacy in patients with CD19+ B cell malignancies, the application of this approach to solid tumors is challenging. The goal of this proof-of-concept study was to investigate whether loading of CD19-CAR T cells (CART19) with anti-HER2 or anti-EGFR bispecific antibodies (BiAb) will target HER2+/EGFR+ CD19- targets and signal the intracellular domain of CAR without engaging antigen-specific CD19 ScFv of CAR T cells. We used CART19 armed with anti-CD3 (OKT3)×anti-HER2 BiAb (HER2Bi) or anti-CD3 (OKT3)×anti-EGFR BiAb (EGFRBi) to evaluate the cytotoxicity directed at HER2 or EGFR expressing cancer cell lines compared with unarmed CART19 measured by short-term 51Cr release assay and long-term real-time cell analysis usingxCelligence. We also determined the differences in exhaustion or effector phenotypes and cytokine profiles during the short- and long-term cytotoxicity assays. Specific cytotoxicity was exhibited by CART19 armed with HER2Bi or EGFRBi against multiple tumor cell lines. Armed CART19 and armed activated T cells (ATC) showed comparable specific cytotoxicity that ranged between 10 and 90% against breast, pancreatic, ovarian, prostate, and lung cancer cell lines at 10:1 E/T ratio. Serial killing (repeated killing) by HER2Bi-armed CART19 ranged between 80 and 100% at 10:1 E/T ratio against MCF-7 cells up to 19days (up to 4th round of repeated killing) measured by a real-time cell analysis without CART19 becoming exhausted. HER2Bi- or EGFRBi-armed CART19 exhibited specific cytotoxicity against multiple HER2+/EGFR+/CD19- tumor targets in overnight and long-term serial killing assays. CART19 showed improved survival and were resistant to exhaustion after prolonged repeated exposure to tumor cells.