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Objective: To investigate the effects and underlying mechanisms of the recombinant bee venom peptide LSSDR412 on the malignant biological behavior of cervical cancer in vitro and in vivo, and to provide a theoretical basis for its potential clinical translation. Methods: Peptide LSSDR412 was obtained by chemical synthesis. Different concentrations of LSSDR412 were used as treatment groups. The effects of peptide LSSDR412 on the survival and proliferation of cervical cancer cell lines (Hela cells and Caski cells) were detected by cell counting kit-8 (CCK-8) and clone formation assay. Transwell chamber assay was used to detect the effect on cell migration and invasion. Western blot and co-immunoprecipitation assay were used to detect the expression and interaction of epithelial-mesenchymal transition (EMT) and Wnt/β-catenin signaling pathway related proteins. The cervical cancer xenograft model was established in nude mice, and the volume of subcutaneous xenograft tumor was detected after intraperitoneal injection of LSSDR412. The expression of the proliferation associated nuclear antigen (Ki-67) was detected by immunofluorescence. The lung metastasis model of cervical cancer bearing nude mice was established, and HE staining was used to detect the effect of intraperitoneal injection of LSSDR412 on lung metastasis. Results: Compared with the cells without LSSDR412 treatment, the survival rate, clone number, invasive and migratory cell number of Hela and Caski cells were significantly decreased after LSSDR412 treatment (all P<0.05), and showed a concentration-dependent trend. LSSDR412 could inhibit the EMT process of cervical cancer and inhibit the Wnt/β-catenin signaling pathway through Claudin-4 (CLDN4) to play an anti-cervical cancer effect. The results of in vivo experiments showed that the tumor weight of the LSSDR412 group was significantly lower than that of the control group [(0.292±0.073) vs (0.480±0.117) g; t=3.341, P=0.008]. The positive rate of Ki-67 in tumor tissue was significantly lower than that in the control group (21.95%±5.60% vs 87.27%±6.35%; t=26.72, P<0.001), and the proportion of intrapulmonary metastases was significantly lower than that of the control group [25.0% (16.3%, 45.0%) vs 39.1% (31.3%, 48.3%); U=31.00, P=0.044]. There was no significant pathological change in liver and kidney tissue structure in the LSSDR412 group. Conclusions: Peptide LSSDR412 shows significant inhibitory effects on the proliferation, migration and invasion of cervical cancer cells both in vitro and in vivo with no apparent toxic side effects. Its antitumor activity is primarily mediated through the suppression of the CLDN4-mediated Wnt/β-catenin signaling pathway.