Combinatorial impact of sanguinarine on doxorubicin induced HSPA5 mediated ER stress in triple-negative breast cancer cells

Doxorubicin (DOX) is a widely used chemotherapeutic for incredibly aggressive triple-negative breast cancer (TNBC). However, its clinical utility is partially limited due to multiorgan toxicities and development of resistance. This study investigated the combinatorial impact of sanguinarine (SAN) on the DOX-induced ER stress and mRNA expression patterns in TNBC cells at sub-optimal concentration. This study evaluated dose-response profile on individual and combination treatments. The potentiated combination was explored to perform different functional assays, RNA-Seq and RT-qPCR analysis and HSPA5 expression analysis. The results showed that the duo combination induced a synergistic cellular toxicity after 48 h of exposure at sub-optimally lower concentrations on MDA-MB-231 cells. The synergistic combination inhibited viability, proliferation and cell cycle, and significantly induced cell death, ER stress, and ROS generation, with elevated caspase 3 activity. Gene ontology (GO) analysis of data from the RNA-Seq analysis revealed that downregulated genes were mapped mainly to protein modifications in the ER, cell cycle, metabolic pathways, and ubiquitin-mediated proteolysis. In contrast, upregulated genes were mapped to mitophagy, the MAPK pathway, and apoptosis, and showed differential expression of prominent ER stress-related genes. RT-qPCR analysis further confirmed the downregulation of HSPA5 and ATF4 and upregulation of XBP1 and DDIT3. Furthermore, combination treatment reduced the total and csHSPA5 protein expression. These insights highlight a novel mechanism by which the SAN-DOX combination synergistically promotes ERS mediated cell death in TNBC.