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The expression of several members of the FOX gene family is known to be altered in a variety of cancers. We show that in breast cancer, FOXF1 gene is a target of epigenetic inactivation and that its gene product exhibits tumor-suppressive properties. Loss or downregulation of FOXF1 expression is associated with FOXF1 promoter hypermethylation in breast cancer cell lines and in invasive ductal carcinomas. Methylation of FOXF1 in invasive ductal carcinoma (37.6% of 117 cases) correlated with high tumor grade. Pharmacologic unmasking of epigenetic silencing in breast cancer cells restored FOXF1 expression. Re-expression of FOXF1 in breast cancer cells with epigenetically silenced FOXF1 genes led to G(1) arrest concurrent with or without apoptosis to suppress both in vitro cell growth and in vivo tumor formation. FOXF1-induced G(1) arrest resulted from a blockage at G(1)-S transition of the cell cycle through inhibition of the CDK2-RB-E2F cascade. Small interfering RNA-mediated depletion of FOXF1 in breast cancer cells led to increased DNA re-replication, suggesting that FOXF1 is required for maintaining the stringency of DNA replication and genomic stability. Furthermore, expression profiling of cell cycle regulatory genes showed that abrogation of FOXF1 function resulted in increased expression of E2F-induced genes involved in promoting the progression of S and G(2) phases. Therefore, our studies have identified FOXF1 as a potential tumor suppressor gene that is epigenetically silenced in breast cancer, which plays an essential role in regulating cell cycle progression to maintain genomic stability., ((c)2010 AACR.)

The promise of retinoids as chemopreventive agents in breast cancer is based on the differentiation and apoptosis induced upon their binding to the retinoic acid (RA) receptor beta (RARbeta). We have previously shown that HOXA5 induces apoptosis in breast cancer cells. In this study, we investigated whether RA/RARbeta and HOXA5 actions intersect to induce apoptosis and differentiation in breast cancer cells. We found that HOXA5 expression can be induced by RA only in RARbeta-positive breast cancer cells. We have, for the first time, identified the RA response element in HOXA5, which was found to be located in the 3' end of the gene. Chromatin immunoprecipitation assays showed that RARbeta binds directly to this region in vivo. Overexpression of RARbeta strongly enhances RA responsiveness, and knocking down RARbeta expression abolishes RA-mediated induction of HOXA5 expression in breast cancer cells. In addition, there is coordinated loss of both HOXA5 and RARbeta expression during neoplastic transformation and progression in the breast epithelial cell model, MCF10A. Knockdown of HOXA5 expression partially abrogates retinoid-induced apoptosis and promotes cell survival upon RA treatment. These results strongly suggest that HOXA5 acts directly downstream of RARbeta and may contribute to retinoid-induced anticancer and chemopreventive effects.

The cytoprotective function of p53 recently has been exploited as a therapeutic advantage for cancer prevention; agents activating the prosurvival activity of p53 are shown to prevent UV-induced damages. To explore the mechanisms of p53-mediated protection from UV-induced apoptosis, we have established stable clones of H1299 lung carcinoma cells expressing a temperature-sensitive p53 mutant, tsp53(V143A). At the permissive temperature of 32 degrees C, the tsp53(V143A)-expressing cells were arrested in G(1) phase without the occurrence of apoptosis; consistent with this is the preferential induction of genes related to growth arrest and DNA damage repair. Previous expression of functional tsp53(V143A) for > or =18 hours inhibited the release of proapoptotic molecules from mitochondria and protected the cells from UV-induced apoptosis; moreover, it suppressed the activation of c-Jun NH(2)-terminal kinase (JNK) signaling and relieved the effect of UV on p53 target gene activation. p53 associated with JNK and inhibited its kinase activity. Using the p53-null H1299 cells, we showed that inhibition of JNK blocked the UV-elicited mitochondrial death signaling and caspase activation. Our results suggest that the ability of p53 to bind and inactivate JNK, together with the activation of the p53 target genes related to cell cycle arrest and DNA damage repair, is responsible for its protection of cells against UV-induced apoptosis.