Withaferin A inhibits expression of ataxia telangiectasia and Rad3‐related kinase and enhances sensitivity of human breast cancer cells to cisplatin

Abstract

Ataxia telangiectasia and Rad3‐related (ATR) is a serine/threonine‐specific kinase that plays an important role in the maintenance of genomic integrity. In this study, we investigated the role of ATR in cell‐cycle arrest by withaferin A (WA), a cancer preventative steroidal lactone derived from Withania somnifera plant abundant in India and surrounding countries. The WA treatment decreased the viability of MCF‐7, MDA‐MB‐231, and SUM159 cells. Exposure of breast cancer cells to WA also resulted in suppression of protein level as well as phosphorylation of ATR and its downstream effector kinase (checkpoint kinase 1; CHK1). Both transcriptional and posttranscriptional mechanisms were involved in the WA‐mediated downregulation of ATR protein. Downregulation of ATR protein expression resulting from WA exposure was not attenuated by overexpression of manganese superoxide dismutase. In contrast, the overexpression of CHK1 attenuated WA‐mediated G2/M arrest and augmented S10 phosphorylation of histone H3, a marker of mitotic arrest. The protein level of ATR was lowered by about 50% in breast tumors of WA‐treated mouse mammary tumor virus‐neu mice when compared with vehicle‐treated controls but the difference was not significant due to small sample size. WA treatment sensitized MDA‐MB‐231 and SUM159 cells to growth inhibition and apoptosis induction by cisplatin. Cisplatin treatment resulted in increased autophosphorylation of ATR (T1989) and CHK1 (S345) phosphorylation that was markedly suppressed in the presence of WA. These results indicate that WA is an inhibitor of ATR in human breast cancer cells.

RNA‐seq reveals novel cancer‐selective and disease subtype‐independent mechanistic targets of withaferin A in human breast cancer cells

“…We have shown previously that WA treatment inhibits ATR/CHK1 signaling to enhance sensitivity to cisplatin in BCC. 24 6 which is the end-product of glycolysis. The plasma level of lactate was also significantly lower in WA-treated rats compared with control in the MNU-rat study.…”

A Comprehensive Review and Perspective on Anticancer Mechanisms of Withaferin A in Breast Cancer

“…The ATR kinase is activated by replication stress during cell division or genotoxic insult and functions at the S-and G 2 -phase of the cell cycle (61,62). Human breast cancer cells (MCF-7, MDA-MB-231, and SUM159) treated with WA showed suppression of protein level as well as phosphorylation of ATR and CHK1 due to both transcriptional and posttranscriptional mechanisms (63). Forced expression of CHK1 abolished the WA-mediated G 2 -M-phase arrest but increased Ser10 phosphorylation of histone H3 (63).…”

“…Human breast cancer cells (MCF-7, MDA-MB-231, and SUM159) treated with WA showed suppression of protein level as well as phosphorylation of ATR and CHK1 due to both transcriptional and posttranscriptional mechanisms (63). Forced expression of CHK1 abolished the WA-mediated G 2 -M-phase arrest but increased Ser10 phosphorylation of histone H3 (63). A trend for a decrease in the protein level of ATR was found in the mammary tumors of WA-treated MMTV-neu mice but the difference was not significant (63).…”

Withaferin A – A natural multifaceted therapeutic compound

“…This decrease in viability of the cell lines by WA was through the induction of ROS mediated paraptosis via down regulation of the paraptosis inhibitor Alix/AIP-1 (Hahm et al, 2011). WA also decreased the viability of SUM159 cells and suppressed ATR(Ataxia telangiectasia and Rad3-related protein) thus causing cell arrest at G2/M phase (Hahm et al, 2019). Further the apoptosis caused in cultured breast cancer cell lines on WA treatment was attenuated by knockdown of multi domain proapoptotic protein Bax and Bak.…”