title: Autophagy fails to alter withaferin A-mediated lethality in human breast cancer cells.
abstract: We have shown previously that withaferin A (WA), which is a highly promising anticancer constituent of Ayurvedic medicine plant Withania somnifera, inhibits viability of cultured breast cancer cells in association with reactive oxygen species (ROS)-dependent apoptosis induction. Because ROS production is implicated in induction of autophagy, which is an evolutionary conserved process for bulk degradation of cellular components including organelles (e.g., mitochondria) and considered a valid cancer chemotherapeutic target, we questioned whether WA treatment resulted in autophagy induction. Indeed exposure of MDA-MB-231 and MCF-7 human breast cancer cells as well as a spontaneously immortalized and non-tumorigenic normal human mammary epithelial cell line (MCF-10A) to pharmacologic concentration of WA resulted in autophagy as evidenced by transmission electron microscopy, processing of microtubuleassociated protein 1 light chain 3 isoform B, and/or acridine orange staining. Inhibition of MDA-MB-231 xenograft growth in vivo by WA administration was also associated with a significant increase in level of LC3 protein in the tumor. However, WA-mediated inhibition of MDA-MB-231 and MCF-7 cell viability was not compromised either by pharmacological suppression of autophagy using 3-methyl adenine or genetic repression of autophagy by RNA interference of Atg5, a critical component of the autophagic machinery. Finally, Beclin1 was dispensable for WA-mediated autophagy as well as inhibition of MDA-MB-231 cell viability. Based on these observations we conclude that autophagy induction fails to have any meaningful impact on WA-mediated lethality in breast cancer cells, which may be a therapeutic advantage because autophagy serves to protect against apoptosis by several anticancer agents.
authors.affiliation: 2.32A Hillman Cancer Center Research Pavilion, 5117 Centre Avenue, Pittsburgh, PA 15213, USA.
authors.forename: Eun-Ryeong
authors.initials: ER
authors.lastname: Hahm
authors.forename: Shivendra V
authors.initials: SV
authors.lastname: Singh
mesh_terms: D000818:Animals; D000972:Antineoplastic Agents, Phytogenic / Q000009:adverse effects / Q000494:pharmacology / Q000627:therapeutic use*; D017209:Apoptosis / Q000187:drug effects*; D051017:Apoptosis Regulatory Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D001943:Breast Neoplasms / Q000188:drug therapy* / Q000378:metabolism / Q000648:ultrastructure; D002460:Cell Line; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D020834:Ethnopharmacology; D005260:Female; D006801:Humans; D042361:Mammary Glands, Human / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D008515:Medicine, Ayurvedic; D008819:Mice, Nude; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D009363:Neoplasm Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D021381:Protein Transport / Q000187:drug effects; D047368:Tumor Burden / Q000187:drug effects; D054358:Withanolides / Q000009:adverse effects / Q000494:pharmacology / Q000627:therapeutic use*; D023041:Xenograft Model Antitumor Assays
chemical_list: D000972:Antineoplastic Agents, Phytogenic; D051017:Apoptosis Regulatory Proteins; C477012:MAP1LC3A protein, human; D008869:Microtubule-Associated Proteins; D009363:Neoplasm Proteins; D054358:Withanolides; C009684:withaferin A