sirolimus and Prostatic-Intraepithelial-Neoplasia

PTEN is one of the most frequently mutated tumor suppressor genes in human cancers. The role of PTEN in carcinogenesis has been validated by knockout mouse models. PTEN heterozygous mice develop neoplasms in multiple organs. Unfortunately, the embryonic lethality of biallelic excision of PTEN has inhibited the study of complete PTEN deletion in the development and progression of cancer. By crossing PTEN conditional knockout mice with transgenic mice expressing a tamoxifen-inducible Cre-ER(T) under the control of a chicken actin promoter, we have generated a tamoxifen-inducible mouse model that allows temporal control of PTEN deletion. Interestingly, administration of a single dose of tamoxifen resulted in PTEN deletion mainly in epithelial cells, but not in stromal, mesenchymal or hematopoietic cells. Using the mT/mG double-fluorescent Cre reporter mice, we demonstrate that epithelial-specific PTEN excision was caused by differential Cre activity among tissues and cells types. Tamoxifen-induced deletion of PTEN resulted in extremely rapid and consistent formation of endometrial in situ adenocarcinoma, prostate intraepithelial neoplasia and thyroid hyperplasia. We also analyzed the role of PTEN ablation in other epithelial cells, such as the tubular cells of the kidney, hepatocytes, colonic epithelial cells or bronchiolar epithelium, but those tissues did not exhibit neoplastic growth. Finally, to validate this model as a tool to assay the efficacy of anti-tumor drugs in PTEN deficiency, we administered the mTOR inhibitor everolimus to mice with induced PTEN deletion. Everolimus dramatically reduced the progression of endometrial proliferations and significantly reduced thyroid hyperplasia. This model could be a valuable tool to study the cell-autonomous mechanisms involved in PTEN-loss-induced carcinogenesis and provides a good platform to study the effect of anti-neoplastic drugs on PTEN-negative tumors.

Topics: Adenocarcinoma; Alleles; Animals; Disease Models, Animal; Dose-Response Relationship, Drug; Endometrial Neoplasms; Endometrium; Epithelial Cells; Everolimus; Female; Gene Deletion; Humans; Hyperplasia; Integrases; Male; Mice; Mice, Knockout; Precancerous Conditions; Prostatic Intraepithelial Neoplasia; Prostatic Neoplasms; PTEN Phosphohydrolase; Recombination, Genetic; Sirolimus; Stromal Cells; Tamoxifen; Thyroid Gland; Thyroid Neoplasms

To evaluate the activation level of the mammalian target of rapamycin (mTOR) signalling pathway in Chinese patients with prostate cancer, as this pathway is over-activated in many human cancers and is an attractive target for cancer therapy.. We used immunohistochemistry to investigate the activation level of five important markers of the mTOR pathway, including PTEN, p-Akt, p-mTOR, p-p70S6K and p-4E-BP1, in tissues from 182 patients with prostate cancer, 20 with benign prostatic hyperplasia (BPH) and 10 with high-grade prostatic intraepithelial neoplasia (HGPIN). The expression levels of these five markers were associated with patient clinical and pathological characteristics.. Expression levels of p-Akt, p-mTOR, p-4E-BP1 and p-p70S6K were significantly higher in prostate cancer tissues than in BPH and HGPIN tissues. In 182 patients with prostate cancer the p-mTOR expression level significantly and positively correlated with its upstream p-Akt and downstream p-4E-BP1 and p-p70S6K expression levels. The cancer Gleason score was significantly correlated with p-Akt and p-mTOR expression level but not with p-4E-BP1 and p-p70S6K expression level. However, the p-4E-BP1 and p-p70S6K expression levels in primary cancer lesions were statistically significantly correlated with patient T stage and distant metastases.. Most patients with prostate cancer have at least one component of the mTOR signalling pathway activated. The activation of the mTOR pathway might be involved in prostate cancer development and progression. The association between activation of mTOR pathway and patient clinicopathological variables suggested that not all patients are equally amenable to treatment strategies targeting the mTOR pathway.

Topics: Adult; Aged; Aged, 80 and over; Biomarkers, Tumor; Disease Progression; Humans; Immunohistochemistry; Intracellular Signaling Peptides and Proteins; Male; Middle Aged; Phosphorylation; Prostatic Hyperplasia; Prostatic Intraepithelial Neoplasia; Prostatic Neoplasms; Protein Serine-Threonine Kinases; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases

The tumor suppressor phosphatase and tensin homolog (PTEN) is frequently involved in human prostate carcinoma. PTEN is therefore an attractive target for the development of preclinical animal models. Prostate intraepithelial neoplasia lesions develop in mice with Pten heterozygosity, but disease progression has been reported only in combination with either other tumor suppressor gene alterations or the conditional inactivation of both Pten alleles in prostate epithelial cells. We report that on a C57BL/6 background, in contrast to previous studies on mixed 129 genetic backgrounds, Pten locus heterozygosity is fully penetrant for the development of prostate adenocarcinoma. Grossly observable tumors were detected at 6 months of age, and, by 10 to 12 months, 100% of examined mice developed adenocarcinoma of the anterior prostate. Furthermore, double heterozygotes carrying both Pten and Tsc2-null alleles showed no increase relative to Pten(+/-) heterozygotes in either lesion development or progression. Lesions in both Pten(+/-); Tsc2(+/-), and Pten(+/-) mice exhibited loss of PTEN expression and activation of PI3K signaling. PI3K activation occurred early in prostate intraepithelial neoplasia lesion formation in these animals, consistent with loss of PTEN function, and contributed to the etiology of tumors that developed in Pten(+/-) mice. Furthermore, prostate lesion growth in Pten(+/-) mice was dependent on mTOR, as evidenced by a reduction in both phospho-S6 levels and proliferative index after rapamycin treatment.

Topics: Adenocarcinoma; Animals; Blotting, Western; Disease Progression; Fluorescent Antibody Technique; Gene Expression Regulation, Neoplastic; Humans; Incidence; Loss of Heterozygosity; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Microsatellite Repeats; Phosphatidylinositol 3-Kinases; Prostatic Intraepithelial Neoplasia; Prostatic Neoplasms; Protein Kinases; Proto-Oncogene Proteins c-akt; PTEN Phosphohydrolase; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases; Tuberous Sclerosis Complex 2 Protein; Tumor Suppressor Proteins

In a previous report, we showed that increased activation of Akt, a downstream effector of phosphoinositide 3-kinase (PI3K) together with decreased activation of extracellular-signal-regulated kinase (ERK), a member of the mitogen-activated protein kinase (MAPK) family, predicted poor clinical outcome in prostate cancer (Kreisberg et al. 2004 Cancer Research 64 5232-5236). We now show that Akt activation, but not ERK activation, is correlated with proliferation in human prostate tumors as estimated by the expression of the cell proliferation antigen Ki67. We verified these results in vitro, using the androgen-dependent prostate cancer cell line LNCaP and its androgen-independent clone C4-2 as models of prostate cancer of good and poor clinical outcome, respectively. C4-2 cells expressed higher Akt activation, lower ERK activation and increased proliferation compared with LNCaP cells, similar to cases of poor clinical outcome. The PI3K inhibitor LY294002, but not the MAPK/ERK kinase inhibitor PD98059, induced growth arrest in both cell lines. Transient transfection with constitutively active Akt increased proliferation while dominant negative Akt decreased it, thus showing that Akt plays an important role in prostate cancer proliferation. Akt regulates the expression and activation of the androgen receptor. Androgen receptor inhibition with Casodex induced growth arrest in LNCaP cells, but not in C4-2 cells. Another PI3K downstream effector, p70 S6 kinase, requires prior phosphorylation by mammalian target of rapamycin (mTOR) for complete activation. Activation of p70 S6 kinase was higher in C4-2 compared with LNCaP cells. Rapamycin, an mTOR inhibitor, had a growth-inhibitory effect in C4-2 cells, but not in LNCaP cells. Our data suggest a shift from a Casodex-sensitive proliferation pathway in LNCaP cells to a rapamycin-sensitive pathway in C4-2 cells.

Topics: Androgen Antagonists; Androgen Receptor Antagonists; Androgens; Anilides; Antibiotics, Antineoplastic; Cell Proliferation; Enzyme Inhibitors; Humans; Ki-67 Antigen; Male; Mitogen-Activated Protein Kinases; Neoplasms, Hormone-Dependent; Nitriles; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Phosphorylation; Prostatic Hyperplasia; Prostatic Intraepithelial Neoplasia; Prostatic Neoplasms; Protein Kinases; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; Receptors, Androgen; Ribosomal Protein S6 Kinases, 70-kDa; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases; Tosyl Compounds; Tumor Cells, Cultured