Abstract #SY09-3: Inactivation of failsafe programs by twist oncoproteins in EMT

Abstract

An intimate crosstalk between failsafe program escape and EMT Senescence and apoptosis are often described as tumor suppressor mechanisms, or failsafe programs, activated in premalignant lesions to negate the proliferative effect of activated mitogenic oncogenes. Accordingly, failsafe program escape is essential for malignant transformation. The p53 tumor suppressor protein is at the crux of senescence and apoptosis signaling pathways hence its mutation over 50% of human cancers. This fraction significantly increases when considering additional mechanisms of p53 neutralization such as activation of native negative regulators. Recent works have identified the overexpression of Twist proteins as novel routes toward wt p53 inactivation outside the canonical ubiquitin ligases (Ansieau et al., 2008; Valsesia-Wittmann et al., 2004; Maestro et al., 1999). Twist1 and Twist2 belong to the bHLH transcription factor family and are essential for proper embryonic development. Undetectable in most healthy adult tissues, both TWIST genes are actively expressed in a large panel of human cancers including carcinomas, sarcomas, melanomas, gliomas and neuroblastomas where they constitute markers of poor prognosis. The main oncogenic properties of Twist proteins rely on their dual ability to override oncogene induced failsafe programs and to promote cell motility. Indeed, we showed that Twist proteins cooperate with oncogenes such as N-Myc or the activated version of Ras (Ras^V12) to fully transform murine fibroblasts and provide them with a tumorigenic potential in nude mice (Ansieau et al., 2008; Valsesia-Wittmann et al., 2004). The cooperation is based on the ability of Twist proteins to override associated apoptosis or senescence by disrupting both p53- and Rb-dependent signaling pathways. Twist proteins negatively regulate the p53 pathway in multiple ways, by preventing ARF activation and thereby p53 stabilization, p53 activation by post-translational modifications, by directly interacting with p53 and thereby disrupting its DNA binding activity and by inhibiting p53 target gene activation through their ability to titrate its co-activator CBP/p300 (Puisieux et al., 2006, for review). By blocking both CIP1/WAF1 and INK4A gene activation by p53 and Ras^V12 respectively, Twist proteins additionally enhance Rb phosphorylation thereby promoting cell proliferation (Ansieau et al., 2008). Additionally, by comparing the tumorigenic and metastatic potential of isogenic murine cell lines, the Weinberg laboratory associated Twist1 expression with metastatic capabilities (Yang et al., 2004). Specifically, by down-regulating epithelial markers (such as the E-cadherin) in favor of mesenchymal markers (such as N-Cadherin), Twist1 expression in epithelial cells promotes an epithelio-mesenchymal transition (EMT) and thereby cell motility. Reminiscent of its functions during development, Twist1 might therefore favor cancer cell dissemination. Interestingly, in agreement with data from the Weinberg laboratory, we also recently reported that EMT triggers stem-like properties, presumably favoring both primary tumor and metastatic growth (Morel et al., 2008; Mani et al., 2008). While these dual properties were originally depicted as unrelated, cooperation assays performed in epithelial cells instead demonstrated that Twist proteins promote cell growth or survival by overriding oncogene-induced failsafe programs and concomitantly cooperate with these oncogenes to promote a complete EMT. This double-barreled cooperation presumes that re-activation of TWIST genes might be sufficient to promote both malignant transformation and early cancer cell dissemination, a hypothesis that we currently investigate using various mouse models. These functions of Twist provide a route to a novel theory regarding cancer metastasis. Cancer progression has been depicted as the result of sequential waves of Darwinian selection and metastatic dissemination is generally presented as the ultimate step. Our results support the hypothesis that, at least in some cases, cancer cell dissemination might already be initiated from premalignant lesions. In line with this hypothesis, the Klein laboratory recently demonstrated, using a MMTV-ErbB2 transgenic mouse model, that ErbB2-expressing cells positive for the epithelial cell marker cytokeratin (CK⁺) become detectable in bone marrow at the stage when early stage atypical ductal hyperplasia (ADH) or ductal carcinoma in situ (DCIS) was found in the mammary tissues. Interestingly, ADHs express the transcription factor Twist1 strengthening a role of Twist in promoting cell dissemination early on in transformation (Husemann et al., 2008). Consistently, CK⁺ cells isolated from the bone marrow of breast cancer patients display a low number of chromosomal abnormalities (Schardt et al., 2005). As the primary tumors of these patients displayed large chromosomal changes detectable by CGH, this observation further supports that the disseminated tumor cells detected without CGH aberrations must have disseminated at the early stage of hyperplasia. 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Citation Information: In: Proc Am Assoc Cancer Res; 2009 Apr 18-22; Denver, CO. Philadelphia (PA): AACR; 2009. Abstract nr SY09-3.