Abstract 5509: The role of WW domain-containing oxidoreductase in methotrexate-induced cell death in human squamous cell carcinoma

Abstract

Abstract Refractory cancer cells to initial chemotherapy in squamous cell carcinomas (SCC) may cause subsequent disease relapse and distant metastasis. We show that the susceptibility of SCC cancer cells to methotrexate (MTX) was associated with higher expression levels of tumor suppressor WW domain-containing oxidoreductase (designated WWOX, FOR or WOX1). Upregulation of WWOX/WOX1 expression by MTX treatment, accompanied by caspase activation and apoptotic cell death, was observed in MTX-sensitive SCC cell lines and tumor biopsies. Ectopically overexpressed WWOX/WOX1 sensitized MTX-resistant SCC-9 cells to MTX-induced apoptosis. Conversely, transient expression of a dominant negative mutant or an siRNA construct targeting WWOX/WOX1 suppressed MTX-mediated cell death in SCC-15 cells. Interestingly, following MTX treatment, significant downregulation of autophagy-related Beclin-1, Atg12-Atg5 and LC3-II protein expression and autophagosome formation was observed in MTX-sensitive SCC-15 cells, while autophagy remained robust in MTX-resistant SCC-9 cells. Increased phosphorylation of mTOR and its downstream substrate p70 S6 kinase (p70S6K) was detected in MTX-treated SCC-15 cells, whereas MTX suppressed the autophagy-inhibitory mTOR signaling in SCC-9 cells. We determined that activation of mTOR and p70S6K by MTX treatment in SCC-15 cells was mediated by WWOX/WOX1. Ectopic overexpression of WWOX/WOX1 in SCC cells induced significant downregulation of Beclin-1, Atg12-Atg5 and LC3-II protein expression, and MTX treatment exacerbated their diminution. Suppression of WWOX/WOX1 by lentiviral shRNA prevented MTX-induced reduction in LC3-II expression in SCC-15 cells. Our findings indicate that WWOX/WOX1 renders SCC cells susceptible to MTX-induced apoptosis by dampening autophagy, and the failure to induce WWOX/WOX1 expression leads to chemotherapeutic drug resistance. Note: This abstract was not presented at the meeting. Citation Format: Li-Jin Hsu. The role of WW domain-containing oxidoreductase in methotrexate-induced cell death in human squamous cell carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5509. doi:10.1158/1538-7445.AM2014-5509