MiR-195 and Its Target SEMA6D Regulate Chemoresponse in Breast Cancer

Abstract

Background: poor prognosis primary breast cancers are typically treated with cytotoxic chemotherapy. However, recurrences remain relatively common even after this aggressive therapy. Comparison of matched tumours pre- and post-chemotherapy can allow identification of molecular characteristics of therapy resistance and thereby potentially aid discovery of novel predictive markers or targets for chemosensitisation. Through this comparison, we aimed to identify microRNAs associated with chemoresistance, define microRNA target genes, and assess targets as predictors of chemotherapy response. Methods: cancer cells were laser microdissected from matched breast cancer tissues pre- and post-chemotherapy from estrogen receptor positive/HER2 negative breast cancers showing partial responses to epirubicin/cyclophosphamide chemotherapy (n = 5). MicroRNA expression was profiled using qPCR arrays. MicroRNA/mRNA expression was manipulated in estrogen receptor positive/HER2 negative breast cancer cell lines (MCF7 and MDA-MB-175 cells) with mimics, inhibitors or siRNAs, and chemoresponse was assessed using MTT and colony forming survival assays. MicroRNA targets were identified by RNA-sequencing of microRNA mimic pull-downs, and comparison of these with mRNAs containing predicted microRNA binding sites. Survival correlations were tested using the METABRIC expression dataset (n = 1979). Results: miR-195 and miR-26b were consistently up-regulated after therapy, and changes in their expression in cell lines caused significant differences in chemotherapy sensitivity, in accordance with up-regulation driving resistance. SEMA6D was defined and confirmed as a target of the microRNAs. Reduced SEMA6D expression was significantly associated with chemoresistance, in accordance with SEMA6D being a down-stream effector of the microRNAs. Finally, low SEMA6D expression in breast cancers was significantly associated with poor survival after chemotherapy, but not after other therapies. Conclusions: microRNAs and their targets influence chemoresponse, allowing the identification of SEMA6D as a predictive marker for chemotherapy response that could be used to direct therapy or as a target in chemosensitisation strategies.