Mechanism of microRNA-30b-5p enhancing the response of MDA-MB-231 cells to cisplatin by regulating RAP1B expression

The present study investigated the role of hsa-microRNA (miR)-30b-5p in potentiating cisplatin-induced cytotoxicity in triple-negative breast cancer (TNBC) MDA-MB-231 cells via Ras-related protein Rap-1b (RAP1B) regulation. Expression levels of hsa-miR-30b-5p and RAP1B were examined in multiple breast cell lines, including normal breast epithelial cells (MCF-10A) and breast cancer cell lines (MCF-7, MDA-MB-231 and MDA-MB-468). MDA-MB-231 cells were divided into control, miR-30b-5p mimic, cisplatin (DDP), miR-30b-5p mimic + DDP and miR-30b-5p mimic + overexpressed RAP1B + DDP groups. Reverse transcription-quantitative PCR (RT-qPCR) demonstrated significantly lower miR-30b-5p expression in MDA-MB-231 compared with MCF-10A normal breast epithelial cells (P<0.001), which was rescued by miR-30b-5p mimics. Cell Counting Kit-8 assays revealed that miR-30b-5p overexpression or DDP alone suppressed cell viability (P<0.001), with combined treatment showing synergistic inhibition (P<0.001). Flow cytometry demonstrated increased apoptosis in miR-30b-5p mimic and DDP groups, further amplified by their combination (P<0.001). Luciferase reporter and RNA pull-down-qPCR assays confirmed miR-30b-5p directly targets RAP1B 3'-untranslated region (UTR) via sequence-specific binding. RNA pull-down demonstrated molecular association between miR-30b-5p and RAP1B mRNA, while the luciferase reporter assay confirmed 3'-UTR-specific interaction. Western blotting and RT-qPCR showed miR-30b-5p mimics downregulated RAP1B (P<0.001), reversed by RAP1B overexpression. In the rescue experiment, RAP1B overexpression partially reversed the pro-apoptotic effect of miR-30b-5p under cisplatin treatment, supporting the hypothesis that RAP1B mediates the effect of miR-30b-5p on the cisplatin response (P<0.001). These findings indicate that miR-30b-5p potentiates cisplatin-induced cytotoxicity in TNBC by suppressing RAP1B, suggesting a novel therapeutic strategy to overcome chemoresistance.