Our observations show that miR-197 and miR-346 contribute to FTC carcinogenesis. Both miRNAs and their target genes might potentially provide for novel molecular markers and act as novel targets for treatment by interference, which could potentially normalize the deregulated profile of many downstream target genes.

Germline mutations in the tumor suppressor gene PTEN (protein phosphatase and tensin homolog located on chromosome ten) predispose to heritable breast cancer. The transcription factor PPARc has also been implicated as a tumor suppressor pertinent to a range of neoplasias, including breast cancer. A putative PPARc binding site in the PTEN promoter indicates that PPARc may regulate PTEN expression. We show here that the PPARc agonist Rosiglitazone, along with Lovastatin, induce PTEN in a dose-and time-dependent manner. Lovastatin-or Rosiglitazoneinduced PTEN expression was accompanied by a decrease in phosphorylated-AKT and phosphorylated-MAPK and an increase in G1 arrest. We demonstrate that the mechanism of Lovastatin-and Rosiglitazone-associated PTEN expression was a result of an increase in PTEN mRNA, suggesting that this increase was transcriptionally-mediated. Compound-66, an inactive form of Rosiglitazone, which is incapable of activating PPARc, was unable to elicit the same response as Rosiglitazone, signifying that the Rosiglitazone response is PPARc-mediated. To support this, we show, using reporter assays including dominant-negative constructs of PPARc, that both Lovastatin and Rosiglitazone specifically mediate PPARc activation. Additionally, we demonstrated that cells lacking PTEN or PPARc were unable to induce PTEN mediated cellular events in the presence of Lovastatin or Rosiglitazone. These data are the first to demonstrate that Lovastatin can signal through PPARc and directly demonstrate that PPARc can upregulate PTEN at the transcriptional level. Since PTEN is constitutively active, our data indicates it may be worthwhile to examine Rosiglitazone and Lovastatin stimulation as mechanisms to increase PTEN expression for therapeutic and preventative strategies including cancer, diabetes mellitus and cardiovascular disease. ' 2006 Wiley-Liss, Inc.

Germline mutations of PTEN (phosphatase and tensin homolog deleted on chromosome 10) are associated with the multihamartomatous disorder Cowden syndrome (CS). Moreover, patients with CS with germline PTEN promoter mutations have aberrant PTEN protein expression and an increased frequency of breast cancer. Here, we examined the downstream effect of five PTEN promoter variants (-861G/T, -853C/G, -834C/T, -798G/C, and -764G/A) that are not within any known cis-acting regulatory elements. Clinically, all five of these patients have been given diagnoses of breast, thyroid, and/or endometrial cancer. We demonstrated that protein binding to the PTEN promoter (-893 to -755) was not altered in the five variants when compared with the wild-type (WT) promoter. However, reporter assays indicated that three of the variants (-861G/T, -853C/G, and -764G/A) demonstrated an ~50% decrease in luciferase activity compared with the WT construct. PTEN messenger RNA (mRNA) levels were not altered in these variants, whereas secondary structure predictions indicated that different PTEN 5' untranslated region transcript-folding patterns exist in three variants, suggesting an inhibition of protein translation. This was confirmed by PTEN protein analysis. These data indicate that variants causing large mRNA secondary structure alterations result in an inhibition of protein translation and a decrease in PTEN protein expression. These data emphasize the importance of PTEN promoter nucleotide variations and their ability to lead to CS progression by a novel regulatory mechanism. Importantly, these patients have a high prevalence of breast, thyroid, and endometrial malignancies; thus, understanding of the mechanism of PTEN dysfunction in these patients will lead to more-sensitive molecular diagnostic and predictive testing and, ultimately, to rational targeted therapies to treat or prevent malignancy.

Germline mutations in the tumor-suppressor gene PTEN predispose to heritable breast cancer. The transcription factor peroxisome proliferator-activated receptor-gamma (PPARgamma) has also been implicated as a tumor suppressor pertinent to a range of neoplasias, including breast cancer. We previously demonstrated that lovastatin may signal through PPARgamma and directly upregulate PTEN expression at the transcriptional level. In our current study, we show that simvastatin, pravastatin and fluvastatin can induce PTEN expression in a dose-dependent manner. This resulted from an increase in PTEN mRNA indicating transcriptional upregulation. In addition, we observed, for the first time, that upregulation of sterol response element-binding protein (SREBP), known to induce PPARgamma expression, can increase PTEN expression. Using reporter assays, we observed that both the statins and SREBP could specifically induce PPARgamma-mediated transcription. However, the statins do not appear to signal through SREBP. Furthermore, our results indicate that SREBP utilizes PPARgamma's transcriptional activity to induce PTEN transcription, whereas the statins signal through PPARgamma's protein activity to upregulate PTEN expression. Overall, our observations suggest that statins signal through another transcription factor, in a PPARgamma-dependent manner, which in turn induces PTEN transcription. We, therefore, studied the full-length PTEN promoter through serial deletion reporter assays and electromobility shift assays and identified a region between -854 and -791 that binds an as-yet-unidentified transcription factor, through which the statins induce PTEN expression. Since PTEN is constitutively active, our data indicate it may be worthwhile to examine statin and SREBP stimulation as mechanisms to increase PTEN expression for therapeutic and preventative strategies in cancer, diabetes mellitus and cardiovascular disease.

Peroxisome proliferator-activated receptor gamma (PPARγ) is a ligand-activated transcription factor, which belongs to the family of nuclear hormone receptors. Recent in vitro studies have shown that PPARγ can regulate the transcription of phosphatase and tensin homolog on chromosome ten (PTEN), a known tumor suppressor. PTEN is a susceptibility gene for a number of disorders, including breast and thyroid cancer. Activation of PPARγ through agonists increases functional PTEN protein levels that subsequently induces apoptosis and inhibits cellular growth, which suggests that PPARγ may be a tumor suppressor. Indeed, several in vivo studies have demonstrated that genetic alterations of PPARγ can promote tumor progression. These results are supported by observations of the beneficial effects of PPARγ agonists in the in vivo cancer setting. These studies signify the importance of PPARγ and PTEN's interaction in cancer prevention.