RBM15 enhances paclitaxel resistance in triple-negative breast cancer by targeting m6A methylation of TNFSF9 and inducing polarization of tumor-associated macrophages to M2 phenotype.

Abstract

Background: Triple-negative breast cancer (TNBC) is one of the breast cancer subtypes with a poor prognosis, and the current main treatment modalities include surgical resection and adjuvant chemotherapy. However, the development of drug resistance in tumor cells to chemotherapeutic agents poses great challenges to anticancer treatment.

Methods: Bioinformatics analysis was used to screen the up-regulated genes in paclitaxel (PTX)-resistant TNBC cells. Cell viability was measured by a CCK-8 kit. TNFSF9 (Tumor necrosis factor receptor superfamily member 9) protein level was detected by Western blot (WB) assay. PTX-resistant TNBC cell lines (MDA-MB-231/PTX, MDA-MB-468/PTX) were constructed and their drug resistance was shown by IC50. The EdU, flow cytometry, Transwell, and other commercial kits were applied to detect the proliferation, apoptosis, migration, invasion, macrophage M2 polarization, and glycolysis of PTX-resistant TNBC cells. RBM15 (RNA binding motif protein 15) levels were measured by RT-qPCR and WB assays. The RIP, MeRIP, and actinomycin D assays were used to analyze the interaction between TNFSF9 and RBM15. The effect of RBM15/TNFSF9 on PTX sensitivity in vivo was verified by xenograft tumor experiments.

Results: TNFSF9 was highly expressed in PTX-resistant TNBC cells. Silencing of TNFSF9 enhanced the sensitivity to PTX. Silencing TNFSF9 induced polarization of macrophages from M2 to M1 phenotype and the release of IL-1β and TNF-α, but decreased the levels of IL-10 and TGF-β. RBM15 targeted the N6-adenylate methylation (m⁶A) modification of TNFSF9, and overexpression of TNFSF9 could reverse the tumor-suppressing effect of silencing RBM15 on PTX-resistant TNBC cells in vitro and transplanted tumors in vivo. Samples from PTX-sensitive and PTX-resistant TNBC patients proved that RBM15 regulated TNFSF9's high expression in PTX-resistant TNBC tissues.

Conclusion: This study demonstrated that RBM15 enhanced PTX resistance in TNBC by promoting m⁶A methylation in TNFSF9 and inducing M2 polarization of tumor-associated macrophages.