Tumor and Stem Cell Biology YAP-InducedResistanceofCancerCells toAntitubulinDrugs Is Modulated by a Hippo-Independent Pathway

Although antitubulin drugs are used widely to treat human cancer, many patients display intrinsic or acquired drug resistance that imposes major obstacles to successful therapy. Mounting evidence argues that cancer cell apoptosis triggered by antitubulin drugs relies upon activation of the cell-cycle kinase Cdk1; however, mechanistic connections of this event to apoptosis remain obscure. In this study, we identified the antiapoptotic protein YAP, a core component of the Hippo signaling pathway implicated in tumorigenesis, as a critical linker coupling Cdk1 activation to apoptosis in the antitubulin drug response. Antitubulin drugs activated Cdk1, which directly phosphorylated YAP on five sites independent of the Hippo pathway. Mutations in these phosphorylation sites on YAP relieved its ability to block antitubulin drug-induced apoptosis, further suggesting that YAP was inactivated by Cdk1 phosphorylation. Notably, we found that YAP was not phosphorylated and inactivated after antitubulin drug treatment in taxol-resistant cancer cells. Our findings suggest YAP and its phosphorylation status as candidate prognostic markers in predicting antitubulin drug response in patients. Cancer Res; 74(16); 1–11. 2014 AACR. Introduction Antitubulin drugs, including paclitaxel (taxol), docetaxel, and vinblastine, have widely been used for the treatment of a variety of human cancers such as breast and ovarian cancers, non–small cell lung cancer, and head and neck cancer (1–3). These drugs can directly bind to b-tubulin and either promote or reduce b-tubulin polymerization, which changes the stability ofmicrotubules (4). The change inmicrotubule stability can subsequently exert a mitotic block by altering kinetochore tension, which can activate the spindle assembly checkpoint, resulting in mitotic delay and subsequent apoptosis (5). Although antitubulin drugs kill cancer cells and can sometimes effectively suppress tumor growth in patients with cancer, a significant proportion of tumors is either intrinsically resistant to antitubulin drugs or later develop resistance after primary therapy, leading to disease relapse and patientmortality, which is a major obstacle to successful cancer treatments (2, 6). Therefore, identification and characterization of cellular genes or signaling pathways responsible for antitubulin drug resistance are critical for successful treatment of cancers. In addition, although tremendous progress has been made toward identifying the genes involved in antitubulin drug response, surprisingly, the signaling pathways by which antitubulin drugs cause mitotic arrest and cell death are poorly defined. YAP (Yes-associated protein) and TAZ (transcriptional coactivator with PDZ-binding motif) are paralogs of WW domain-containing transcriptional coactivators, which share 60% similarity and activate many transcriptional factors important for the development of various tissues in mammals (7, 8). Recently, TAZ and YAP have been identified as oncogenes and major components of an emerging Hippo signaling pathway that plays important roles in regulating animal size, cell proliferation, apoptosis, tumorigenesis, stem cell renewal and differentiation, neuronal growth, andmechanotransduction in both Drosophila and mammals (8–12). In this novel pathway, mammalian homologs of Drosophila Hippo, Mst1, and Mst2 phosphorylate and activate tumor suppressors and kinases LATS1 and LATS2, which in turn inactivate YAP and TAZ by phosphorylating multiple sites (five sites for YAP and four sites for TAZ) with consensus sequence of HxH/R/KxxS/T (H, histine; x, any amino acid; R, arginine; K, lysine; S, serine; T, threonine; Mst1/2!LATS1/2!YAP/TAZ). These core components also interact withmany upstream (e.g., NF2, RASSF1A, and FAT4) and downstream (e.g., CTGF, Cyr61, and BMP4) cellular genes, forming a complex Hippo signaling pathway. Recently, we and others have identified YAP and TAZ as important players in response of breast cancer cells to apoptosis induced by antitubulin drug taxol (13, 14). It has been shown that overexpression of YAP or TAZ in MCF10A mammary cells causes transformation, epithelial–mesenchymal transition (EMT), and resistance to taxol, whereas knockdown of TAZ in MDA-MD231 breast cancer cells sensitize these cells Authors' Affiliations: Department of Pathology and Molecular Medicine, Queen's University, Kingston; Centre for Biologics Research, Biologics and Genetic Therapies Directorate, Health Canada, Ottawa, Ontario; Rosalind & Morris Goodman Cancer Research Centre, McGill University, Montreal, Canada Note: Supplementary data for this article are available at Cancer Research Online (http://cancerres.aacrjournals.org/). Corresponding Author: Xiaolong Yang, Queen's University, 88 Stuart Street, Kingston, Ontario K7L 3N6, Canada. Phone: 613-533-6000; Fax: 613-533-2907; E-mail: [email protected] doi: 10.1158/0008-5472.CAN-13-2712 2014 American Association for Cancer Research. Cancer Research www.aacrjournals.org OF1 Research. on April 15, 2017. © 2014 American Association for Cancer cancerres.aacrjournals.org Downloaded from Published OnlineFirst May 8, 2014; DOI: 10.1158/0008-5472.CAN-13-2712