Effect of targeting c-Met/CD47 CAR-T cells on ovarian cancer cells

doi:10.19803/j.1672-8629.20240339

Abstract

Abstract: Objective To investigate the effect of knocking down the expression of mesenchymal-epithelial transition factor (c-Met) on proliferation, invasion, migration and apoptosis of ovarian cancer cells in vitro, prepare chimeric antigen receptor T (CAR-T) targeting c-Met and observe its ability to kill ovarian cancer cells. Methods The expression of c-Met in three ovarian cancer cell lines was detected by real-time fluorescence quantitative polymerase chain reaction and Western blotting. The c-Met was knocked down by transfection, and cell proliferation was detected by cell counting kit-8 (CCK-8) and cell cloning. Cell invasion and cell scratch were used to detect cell invasion and migration ability. The cell cycle, apoptosis and the positive rate of lentivirus infected cells were detected by flow cytometry. CCK-8 was used to detect the proliferation of CAR-T cells stimulated by target cells. Lactate dehydrogenase assay was used to detect the killing rate of cells while the release of interferon-γ (IFN-γ) and interleukin-2 (IL-2) was detected by enzyme-linked immunosorbent assay. A subcutaneous xenograft model of nude mice was established before T, CAR-T, c-Met CAR-T, c-Met/CD47 CAR-T effector cell groups were injected. The tumor weight was measured. Hematoxylin-eosin staining and immunohistochemistry were used to study the therapeutic effect of CAR-T cells against ovarian cancer. Results The expression of c-Met in SKOV3 cell line was high. Compared with the control group, the cell proliferation, invasion, migration ability of the c-Met knockdown group decreased while the apoptosis rate increased. G1 and S phase decreased while G2 phase increased. Flow cytometry showed that the infection efficiency of CAR-T, c-Met CAR-T and c-Met/CD47 CAR-T cells was 97.42%, 97.39% and 97.35%, respectively. CCK-8 assay indicated that the proliferation potential of c-Met/CD47 CAR-T cells to target cells was significantly enhanced. LDH assay suggested that the killing rate of c-Met/CD47 CAR-T cells to target cells was significantly increased. ELISA showed that SKOV3 induced c-Met/CD47 CAR-T cells to release more IFN-γ and IL-2. The weight of transplanted tumor in the c-Met/CD47 CAR-T cell group was lighter than in T, CAR-T and c-Met CAR-T cell groups when SKOV3 cells were injected subcutaneously. HE results showed that there was no physiological structure damage in the tissues and organs of each cell treatment group. Conclusion c-Met/CD47 can be used as a molecular target for CAR-T cell therapy for ovarian cancer, which can target and recognize ovarian cancer cells with higher c-Met expression. The stronger the killing ability, the better the proliferation ability and cytokine release level.

Key words: ovarian cancer, mesenchymal-epithelial transition factor, mesenchymal-epithelial transition factor/cluster of differentiation 47, chimeric antigen receptor T, targetiug, cell

CLC Number:

R979.1

WANG Rumeng, SI Qin. Effect of targeting c-Met/CD47 CAR-T cells on ovarian cancer cells[J]. Chinese Journal of Pharmacovigilance, 2024, 21(10): 1103-1112.

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URL: https://zgywjj.magtechjournal.com/EN/10.19803/j.1672-8629.20240339

https://zgywjj.magtechjournal.com/EN/Y2024/V21/I10/1103

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