ABSTRACT
A common treatment of advanced prostate cancer involves the deprivation of androgens. Despite the initial response to hormonal therapy, eventually all the patients relapse. In the present study, we sought to determine whether dietary polyunsaturated fatty acid (PUFA) affects the development of castration-resistant prostate cancer. Cell culture, patient tissue microarray, allograft, xenograft, prostate-specific Pten knockout and omega-3 desaturase transgenic mouse models in conjunction with dietary manipulation, gene knockdown and knockout approaches were used to determine the effect of dietary PUFA on castration-resistant Pten-null prostate cancer. We found that deletion of Pten increased androgen receptor (AR) expression and Pten-null prostate cells were castration resistant. Omega-3 PUFA slowed down the growth of castration-resistant tumors as compared with omega-6 PUFA. Omega-3 PUFA decreased AR protein to a similar extent in tumor cell cytosolic and nuclear fractions but had no effect on AR messenger RNA level. Omega-3 PUFA treatment appeared to accelerate AR protein degradation, which could be blocked by proteasome inhibitor MG132. Knockdown of AR significantly slowed down prostate cancer cell proliferation in the absence of androgens. Our data suggest that omega-3 PUFA inhibits castration-resistant prostate cancer in part by accelerating proteasome-dependent degradation of the AR protein. Dietary omega-3 PUFA supplementation in conjunction with androgen ablation may significantly delay the development of castration-resistant prostate cancer in patients compared with androgen ablation alone.
Subject(s)
Dietary Fats, Unsaturated/pharmacology , Fatty Acids, Omega-3/pharmacology , Fatty Acids, Unsaturated/pharmacology , PTEN Phosphohydrolase/deficiency , Prostatic Neoplasms/drug therapy , Animals , Cell Line, Tumor , Cell Proliferation/drug effects , Cells, Cultured , Dietary Fats, Unsaturated/metabolism , Drug Resistance, Neoplasm , Epithelial Cells/drug effects , Epithelial Cells/metabolism , Fatty Acid Desaturases/metabolism , Fatty Acids, Omega-3/metabolism , Fatty Acids, Omega-6/metabolism , Fatty Acids, Unsaturated/metabolism , Gene Knockdown Techniques/methods , Gene Knockout Techniques , Humans , Inflammation/drug therapy , Inflammation/metabolism , Male , Mice , Mice, Knockout , Mice, Nude , Mice, Transgenic , Neovascularization, Pathologic/drug therapy , Neovascularization, Pathologic/metabolism , Orchiectomy , PTEN Phosphohydrolase/genetics , PTEN Phosphohydrolase/metabolism , Prostatic Neoplasms/genetics , Prostatic Neoplasms/metabolism , Prostatic Neoplasms/surgery , Proteasome Endopeptidase Complex/drug effects , Proteasome Endopeptidase Complex/metabolism , Proteolysis/drug effects , Proto-Oncogene Proteins c-akt/metabolism , Receptors, Androgen/genetics , Receptors, Androgen/metabolismABSTRACT
Vitamin D(3) is a promising preventative and therapeutic agent for prostate cancer, but its implementation is hampered by a lack of understanding about its mechanism of action. Upon treatment with 1α,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3), vitamin D(3)], the metabolically active form of vitamin D(3), adult prostate progenitor/stem cells (PrP/SC) undergo cell-cycle arrest, senescence, and differentiation to an androgen receptor-positive luminal epithelial cell fate. Microarray analyses of control- and vitamin D(3)-treated PrP/SCs revealed global gene expression signatures consistent with induction of differentiation. Interestingly, one of the most highly upregulated genes by vitamin D(3) was the proinflammatory cytokine interleukin-1α (IL-1α). Systems biology analyses supported a central role for IL-1α in the vitamin D(3) response in PrP/SCs. siRNA-mediated knockdown of IL-1α abrogated vitamin D(3)-induced growth suppression, establishing a requirement for IL-1α in the antiproliferative effects of vitamin D(3) in PrP/SCs. These studies establish a system to study the molecular profile of PrP/SC differentiation, proliferation, and senescence, and they point to an important new role for IL-1α in vitamin D(3) signaling in PrP/SCs.
Subject(s)
Calcitriol/pharmacology , Interleukin-1alpha/physiology , Prostate/cytology , Stem Cells/drug effects , Animals , Cell Cycle/drug effects , Cell Differentiation/drug effects , Cell Division/drug effects , Cells, Cultured/drug effects , Cells, Cultured/metabolism , Cellular Senescence/drug effects , Colony-Forming Units Assay , Gene Expression Regulation/drug effects , Genes, Retinoblastoma , Genes, p16 , Interleukin-1alpha/biosynthesis , Interleukin-1alpha/genetics , Male , Mice , Mice, Knockout , Oligonucleotide Array Sequence Analysis , Prostate/drug effects , RNA, Small Interfering/pharmacology , Receptors, Calcitriol/deficiency , Receptors, Calcitriol/genetics , Stem Cells/metabolismABSTRACT
BACKGROUND: 1-Alpha, 25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3)) inhibits proliferation of multiple cancer cell types including prostate cells and upregulates p21 and/or p27, while loss of Pten and PI3K/AKT activation stimulates survival and downregulates p21 and p27. We hypothesized that inhibition of the PI3K/AKT pathway synergizes with the antiproliferative signaling of 1,25(OH)(2)D(3). METHODS: Viability, cell cycle and senescence of cells were evaluated upon combinational treatment with 1,25(OH)(2)D(3) and pharmacological PI3K/AKT inhibitors. RESULTS: Pharmacological inhibitors of PI3K or Akt and 1,25(OH)(2)D(3) synergistically inhibited growth of DU145, LNCaP, primary human prostate cancer cell strains and Pten null mouse prostatic epithelial cells (MPEC). The inhibitors used included API-2 (Triciribine) and GSK690693 which are currently in clinical trials for treatment of cancer. A novel mechanism for antiproliferative effects of 1,25(OH)(2)D(3) in prostate cells, induction of senescence, was discovered. Combination of 1,25(OH)(2)D(3) and AKT inhibitor cooperated to induce G(1) arrest, senescence, and p21 levels in prostate cancer cells. As AKT is commonly activated by PTEN loss, we evaluated the role of Pten in responsiveness to 1,25(OH)(2)D(3) using shRNA knockdown and by in vitro knockout of Pten. MPEC that lost Pten expression remained sensitive to the antiproliferative action of 1,25(OH)(2)D(3), and showed higher degree of synergism between AKT inhibitor and 1,25(OH)(2)D(3) compared to Pten-expressing counterparts. CONCLUSIONS: These findings provide the rationale for the development of therapies utilizing 1,25(OH)(2)D(3) or its analogs combined with inhibition of PI3K/AKT for the treatment of prostate cancer.
Subject(s)
Antineoplastic Combined Chemotherapy Protocols/pharmacology , Chromones/pharmacology , Enzyme Inhibitors/pharmacology , Morpholines/pharmacology , Phosphoinositide-3 Kinase Inhibitors , Prostatic Neoplasms/metabolism , Proto-Oncogene Proteins c-akt/antagonists & inhibitors , Vitamin D/analogs & derivatives , Animals , Cell Line, Tumor , Cell Proliferation/drug effects , Cellular Senescence/drug effects , Chromones/administration & dosage , Dose-Response Relationship, Drug , Drug Synergism , Enzyme Inhibitors/administration & dosage , Flow Cytometry , Humans , Immunoblotting , Male , Mice , Mice, Knockout , Morpholines/administration & dosage , Oxadiazoles/pharmacology , PTEN Phosphohydrolase/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Prostatic Neoplasms/enzymology , Prostatic Neoplasms/pathology , Proto-Oncogene Proteins c-akt/metabolism , Ribonucleosides/pharmacology , Vitamin D/pharmacologyABSTRACT
Demonstration of the hallmarks of stem cells, self-renewal and multilineage differentiation, is a challenge that has not been met for numerous tissues postulated to possess adult stem cells, including prostate tissue. Using a defined medium, we reproducibly isolated and maintained adult mouse prostatic cells with characteristics of progenitor/stem cells. Clonal populations of cells demonstrated tissue-specific multilineage differentiation by their ability to generate organized prostatic ductal structures in vivo, with luminal and basal cell layers, when grafted under the renal capsules of mice in the presence of fetal rat urogenital mesenchyme. Complete differentiation was demonstrated by the expression and secretion of terminally differentiated prostatic secretory products into the lumens. Self-renewal was demonstrated by serial transplantation of clonal populations that generated fully differentiated ductal structures in vivo. In vitro, undifferentiated cells expressed markers associated with prostate stem cells, including Sca 1 and CD49f, as well as basal cell markers (p63 and cytokeratins 5 and 14) and, at a low level, luminal cell markers (androgen receptor and cytokeratins 8 and 18). When grafted and allowed to differentiate in the presence of fetal urogenital mesenchyme, the cells differentiated into luminal cells and basal cells with more restricted protein expression patterns. These studies are the first to report a reproducible system to assess adult prostatic progenitor/stem cells.