[c-Met-targeted chimeric antigen receptor T cells inhibit human serous ovarian cancer cell SKOV-3 <i>in vitro</i>].

The study aimed to construct the second and third generation chimeric antigen receptor T cells (CAR-T) targeting the c-mesenchymal-epithelial transition factor (c-Met) protein, and observe their killing effect on human serous ovarian cancer cell SKOV-3. The expression of <i>MET</i> gene in ovarian serous cystadenocarcinoma, the correlation between <i>MET</i> gene expression and the abundance of immune cell infiltration, and the effect of <i>MET</i> gene expression on the tissue function of ovarian serous cystadenocarcinoma were analyzed by bioinformatics. The expression of c-Met in ovarian cancer tissues and adjacent tissues was detected by immunohistochemical staining. The second and third generation c-Met CAR-T cells, namely c-Met CAR-T(2G/3G), were prepared by lentivirus infection, and the cell subsets and infection efficiency were detected by flow cytometry. Using CD19 CAR-T and activated T cells as control groups and A2780 cells with c-Met negative expression as Non target groups, the kill efficiency on SKOV-3 cells with c-Met positive expression, cytokine release and cell proliferation of c-Met CAR-T(2G/3G) were explored by lactate dehydrogenase (LDH) release, ELISA and CCK-8 respectively. The results showed that <i>MET</i> gene expression was significantly up-regulated in ovarian cancer tissues compared with normal tissues, which was consistent with the immunohistochemistry results. However, in all pathological stages, there was no obvious difference in <i>MET</i> expression and no correlation between <i>MET</i> gene expression and the race and age of ovarian cancer patients. The second generation and third generation c-Met CAR-T cells were successfully constructed. After lentivirus infection, the proportion of CD8⁺ T cells in c-Met CAR-T(2G) was upregulated, while there was no significant change in the cell subsets of c-Met CAR-T(3G). The LDH release experiment showed that the kill efficiency of c-Met CAR-T(2G/3G) on SKOV-3 increased with the increase of effect-target ratio. When the effect-target ratio was 20:1, the kill efficiency of c-Met CAR-T(2G) reached (42.02 ± 5.17)% (<i>P</i> < 0.05), and the kill efficiency of c-Met CAR-T(3G) reached (51.40 ± 2.71)% (<i>P</i> < 0.05). ELISA results showed that c-Met CAR-T released more cytokine compared to CD19 CAR-T and activated T cells (<i>P</i> < 0.05). Moreover, the cytokine release of c-Met CAR-T(3G) was higher than c-Met CAR-T(2G) (<i>P</i> < 0.01). The CCK-8 results showed that after 48 h, the cell number of c-Met CAR-T(2G) was higher than that of c-Met CAR-T(3G) (<i>P</i> < 0.01). In conclusion, both the second and third generation c-Met CAR-T can target and kill c-Met-positive SKOV-3 cells, with no significant difference. c-Met CAR-T(2G) has stronger proliferative ability, and c-Met CAR-T(3G) releases more cytokines.