Analysis of small RNA expression in tumors from doxorubicin sensitive and resistant MDA-MB-231 xenografts

Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer. While chemotherapy is the mainstay therapy for the treatment of TNBC patients, many patients exhibit either de novo chemoresistance or ultimately develop resistance to chemotherapy, leading to significantly high mortality rates. Here we performed whole transcriptome sequencing (both RNA and small RNA-sequencing), coupled with network simulations and patient survival data analyses to build a novel miRNA-mRNA interaction network governing chemoresistance in TNBC. We report that fibronectin (FN1) is a central chemoresistance driver gene in our miRNA-mRNA interaction network, and overexpressing miR-326 reverses FN1-driven chemoresistance by targeting FN1 receptor, ITGA5. miR-326 is downregulated by increased hypoxia/HIF1A and ECM stiffness in chemoresistant tumors leading to upregulation of ITGA5 and activation of the downstream FAK/Src signaling pathways. Overexpression of miR-326 or inhibition of ITGA5 overcomes FN1-driven chemotherapy resistance in vitro by inhibiting FAK/Src pathway and potentiates the efficacy of chemotherapy in vivo. Importantly, lower expression of miR-326 or higher levels of predicted miR-326 target genes is significantly associated with worse overall survival in chemotherapy-treated TNBC patients. In chemoresistant tumors, hypoxia and resulting ECM stiffness repress the expression of the tumor suppressor miRNA, miR-326. Hence, re-expression of miR-326 or inhibition of its target ITGA5 stand as attractive therapeutic approaches to enhance chemotherapy response in TNBCs.