Anti-alpha FR CAR-engineered NK-92 Cells Display Potent Cytotoxicity Against alpha FR-positive Ovarian Cancer

摘要

Folate receptor alpha (alpha FR) is overexpressed in 90% of ovarian cancers, one of the most lethal gynecologic cancers. Recent studies have suggested that natural killer (NK) cells may be better chimeric antigen receptor (CAR) drivers because of their favorable innate characteristics, such as directly recognizing and killing tumor cells. However, the therapeutic effects of CAR-engineered NK cells targeting alpha FR in ovarian cancer have not been reported. In this research, 3 generations of anti-alpha FR CAR were constructed, namely alpha FR-zeta (first generation), alpha FR-28 zeta (second generation), and alpha FR-28BB zeta (third generation), and were highly expressed on the surface of NK-92 cells by lentivirus gene transfection. Three anti-alpha FR CAR-engineered NK-92 cells can specifically kill alpha FR-positive tumor cells in vitro, especially ovarian cancer cells with high alpha FR expression. Compared with NK-92 cells expressing alpha FR-zeta or alpha FR-28 zeta, NK-92 cells expressing alpha FR-28BB zeta showed not only higher antigen-specific cytotoxicity and proliferation but also lower antigen-induced apoptosis. Moreover, stronger degranulation and cytokine secretion were detected in NK-92 cells expressing alpha FR-28BB zeta cocultured with alpha FR-positive tumor cells. Real-time cell analysis and live cell imaging recorded the process of NK-92 cells expressing alpha FR-28BB zeta killing ovarian cancer cells in vitro. Furthermore, NK-92 cells expressing alpha FR-28BB zeta can effectively eliminate cancer cells in a mouse xenograft model of ovarian cancer and significantly prolong the survival of tumor-bearing mice. These results demonstrate that the anti-alpha FR CARs redirect NK-92 cells with specific antitumor activity, and the third-generation anti-alpha FR CAR-engineered NK-92 cells display more potent cytotoxicity against alpha FR-positive ovarian cancer, laying the foundation for future clinical research.