Replication factor C4, which is regulated by insulin-like growth factor 2 mRNA binding protein 2, enhances the radioresistance of breast cancer by promoting the stemness of tumor cells.

Abstract

Radiotherapy resistance, is usually caused by enhanced tumor stemness and poses a significant challenge in treating breast cancer (BRCA). In this study, we investigated the molecular regulatory mechanism of radiotherapy sensitivity in BRCA associated with replication factor C4 (RFC4) and insulin-like growth factor 2 mRNA binding protein 2 mRNA Binding Protein 2 (IGF2BP2). RFC4 expression was increased in BRCA cell lines and tissues, and high RFC4 expression in BRCA patients predicted the occurrence of lymphatic metastasis. RFC4-specific short hairpin RNA sequences or RFC4 coding sequences were subsequently cloned and inserted into plasmid vectors to downregulate or upregulate RFC4 expression. Knockdown of RFC4 attenuated stemness, as evidenced by a reduction in sphere formation and the downregulation of CD44 and SOX2. RFC4 silencing inhibited migration and invasion, promoted apoptosis, and improved sensitivity to radiotherapy (4-Gy X-ray). The results were detected by a wound healing assay, a transwell assay, and flow cytometry. The overexpression of RFC4 had the opposite effect on BRCA cells. Like RFC4 expression, IGF2BP2 expression was also increased in the cancer tissues of breast cancer patients. The results of the dual luciferase assay and RIP assay confirmed the binding of IGF2BP2 to the RFC4 mRNA coding sequence. Knockdown of RFC4 eliminated the effects of IGF2BP2 overexpression on increasing cell viability, invasion, the expression of stemness markers and radioresistance, suggesting that the effect of RFC4 on the stemness of BRCA cells was regulated by IGF2BP2. In conclusion, we reported that RFC4, a key regulator of BRCA progression, promoted radioresistance in BRCA and was positively regulated by IGF2BP2.