Fulvestrant-Induced Cell Death and Proteasomal Degradation of Estrogen Receptor a Protein in MCF-7 Cells Require the CSK c-Src Tyrosine Kinase

Fulvestrant is a representative pure antiestrogen and a Selective Estrogen Receptor Down-regulator (SERD). In contrast to the Selective Estrogen Receptor Modulators (SERMs) such as 4-hydroxytamoxifen that bind to estrogen receptor a (ERa) as antagonists or partial agonists, fulvestrant causes proteasomal degradation of ERa protein, shutting down the estrogen signaling to induce proliferation arrest and apoptosis of estrogen-dependent breast cancer cells. We performed genomewide RNAi knockdown screenings for protein kinases required for fulvestrant-induced apoptosis of the MCF-7 estrogendependent human breast caner cells and identified the c-Src tyrosine kinase (CSK), a negative regulator of the oncoprotein c-Src and related protein tyrosine kinases, as one of the necessary molecules. Whereas RNAi knockdown of CSK in MCF-7 cells by shRNA-expressing lentiviruses strongly suppressed fulvestrant-induced cell death, CSK knockdown did not affect cytocidal actions of 4-hydroxytamoxifen or paclitaxel, a chemotherapeutic agent. In the absence of CSK, fulvestrant-induced proteasomal degradation of ERa protein was suppressed in both MCF-7 and T47D estrogen-dependent breast cancer cells whereas the TP53-mutated T47D cells were resistant to the cytocidal action of fulvestrant in the presence or absence of CSK. MCF-7 cell sensitivities to fulvestrant-induced cell death or ERa protein degradation was not affected by small-molecularweight inhibitors of the tyrosine kinase activity of c-Src, suggesting possible involvement of other signaling molecules in CSK-dependent MCF-7 cell death induced by fulvestrant. Our observations suggest the importance of CSK in the determination of cellular sensitivity to the cytocidal action of fulvestrant. Citation: Yeh W-L, Shioda K, Coser KR, Rivizzigno D, McSweeney KR, et al. (2013) Fulvestrant-Induced Cell Death and Proteasomal Degradation of Estrogen Receptor a Protein in MCF-7 Cells Require the CSK c-Src Tyrosine Kinase. PLoS ONE 8(4): e60889. doi:10.1371/journal.pone.0060889 Editor: Antimo Migliaccio, II Università di Napoli, Italy Received March 6, 2012; Accepted March 6, 2013; Published April 4, 2013 Copyright: 2013 Yeh et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This research was supported by Susan G. Komen for Cure grants FAS0703860 and KG090515, and AstraZeneca Preclinical Study Support IRUSFULV0066, awarded to T. Shioda. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: [email protected]