alitretinoin and Prostatic-Neoplasms

A number of hormonal ligands and/or the nuclear receptors that mediate their actions have been targeted for prostate cancer therapy. Androgens, the ligands for the androgen receptor (AR), are critical for the growth of prostate cancer. Inhibition of androgen production has been the mainstay of treatment for advanced prostate cancer for decades. Other more recently tested targets include retinoid receptors (RAR and RXR), glucocorticoid receptors (GR), estrogen receptors (ER) and peroxisome proliferator-activated receptors (PPAR). Calcitriol, acting through the Vitamin D receptor (VDR), has many tumor suppressive activities in the prostate, including inhibition of proliferation, induction of apoptosis and/or differentiation, and reduction of cellular invasion. Because of these properties, calcitriol and its less hypercalcemic analogs are being evaluated as agents to prevent or treat prostate cancer. Androgens, retinoids, glucocorticoids, estrogens and agonists of PPAR directly or indirectly impact Vitamin D signaling pathways, and vice versa. In order to design the most effective strategies to use calcitriol to prevent or treat prostate cancer, the interactions of other nuclear receptors and their ligands with the Vitamin D signaling pathway need to be considered.

Topics: Alitretinoin; Androgen Receptor Antagonists; Androgens; Animals; Antineoplastic Combined Chemotherapy Protocols; Calcitriol; Drug Interactions; Glucocorticoids; Histone Deacetylase Inhibitors; Hormones; Humans; Male; PPAR gamma; Prostatic Neoplasms; Proteasome Inhibitors; Receptors, Androgen; Receptors, Cytoplasmic and Nuclear; Retinoids; Signal Transduction; Tretinoin; Vitamin D

Topics: Alitretinoin; Androgens; Angiotensin Receptor Antagonists; Animals; Benzimidazoles; Biphenyl Compounds; Cholecalciferol; DNA-Binding Proteins; Drug Design; Genistein; Humans; Male; Neoplasms, Hormone-Dependent; Prostatic Neoplasms; Receptor, Angiotensin, Type 1; Receptors, Angiotensin; Telomerase; Tetrazoles; Tretinoin

Radiation therapy is a common treatment for prostate cancer, however recurrence remains a problem. MicroRNA expression is altered in prostate cancer and may promote therapy resistance. Through bioinformatic analyses of TCGA and CPC-GENE patient cohorts, we identified higher miR-191 expression in tumor versus normal tissue, and increased expression in higher Gleason scores. In vitro and in vivo experiments demonstrated that miR-191 overexpression promotes radiation survival, and contributes to a more aggressive phenotype. Retinoid X receptor alpha, RXRA, was discovered to be a novel target of miR-191, and knockdown recapitulated radioresistance. Furthermore, treatment of prostate cancer cells with the RXRA agonist 9-cis-retinoic acid restored radiosensitivity. Supporting this relationship, patients with high miR-191 and low RXRA abundance experienced quicker biochemical recurrence. Reduced RXRA translated to a higher risk of distant failure after radiotherapy. Notably, this miR-191/RXRA interaction was conserved in a novel primary cell line derived from radiorecurrent prostate cancer. Together, our findings demonstrate that miR-191 promotes prostate cancer survival after radiotherapy, and highlights retinoids as a potential option to improve radiotherapy response.

Topics: Alitretinoin; Animals; Antineoplastic Agents; Biomarkers, Tumor; Cell Line, Tumor; Chemoradiotherapy, Adjuvant; Disease-Free Survival; Down-Regulation; Gene Expression Regulation, Neoplastic; Gene Knockdown Techniques; Humans; Kallikreins; Kaplan-Meier Estimate; Male; Mice; MicroRNAs; Middle Aged; Neoplasm Grading; Neoplasm Recurrence, Local; Primary Cell Culture; Prognosis; Prostate; Prostate-Specific Antigen; Prostatectomy; Prostatic Neoplasms; Radiation Tolerance; Retinoid X Receptor alpha; Survival Rate; Time Factors; Xenograft Model Antitumor Assays

9-cis-Retinoic acid (9cRA), which binds to both retinoic acid receptors and retinoic X receptors, inhibits prostate cancer induction in rats and reduces growth of prostate cancer cells. However, the nature of this growth inhibition and the interactive influence of androgens are not well defined and are the subject of this report. LNCaP and PC-3 cells were cultured and treated with a range of 9cRA concentrations for 3-6 days in the absence or presence of 5α-dehydrotestosterone. 9cRA inhibited cell proliferation in a dose-dependent manner, plateauing at 10 mol/l. Treatment of cells with 10 mol/l 9cRA inhibited 5α-dihydroxytestosterone (DHT)-stimulated proliferation, the effect of which was maximal at 10 mol/l DHT. Treatment of DHT (10 mol/l)-exposed cells with 9cRA caused a dose-dependent increase in prostate-specific antigen in the medium after 6 days, but not 3 days. 9cRA caused a dose-dependent increase in apoptotic cells stained with H33258 after 3 days, but not 6 days; however, on using flow cytometry, apoptosis was apparent at both 3 and 6 days. Flow cytometry also revealed interference of G0/G1 to S phase transition by 9cRA. Inhibition by 9cRA of anchorage-independent growth of PC-3 cells was also found; LNCaP cells did not grow colonies in soft agar. 9cRA inhibited growth and induced differentiation of human LNCaP prostate cancer cells in vitro and inhibited anchorage-independent growth of PC-3 cells. Because 9cRA and 13-cis-retinoic acid, which is retinoic acid receptor-selective, prevent prostate carcinogenesis in rats, and 13-cis-retinoic acid also inhibits growth of human prostate cancer cells, the RAR is a potential molecular target for prostate cancer prevention and therapy.

Topics: Alitretinoin; Cell Differentiation; Cell Line, Tumor; Cell Proliferation; Dose-Response Relationship, Drug; Drug Interactions; Humans; Male; Prostatic Neoplasms; Testosterone; Tretinoin

The retinoids, the natural or synthetic derivatives of Vitamin A (retinol), are essential for the normal development of prostate and have been shown to modulate prostate cancer progression in vivo as well as to modulate growth of several prostate cancer cell lines. 9-cis-retinoic acid and all-trans-retinoic acid are the two most important metabolites of retinol. Gap junctions, formed of proteins called connexins, are ensembles of intercellular channels that permit the exchange of small growth regulatory molecules between adjoining cells. Gap junctional communication is instrumental in the control of cell growth. We examined the effect of 9-cis-retinoic acid and all-trans retinoic acid on the formation and degradation of gap junctions as well as on junctional communication in an androgen-responsive prostate cancer cell line, LNCaP, which expressed retrovirally introduced connexin32, a connexin expressed by the luminal cells and well-differentiated cells of prostate tumors. Our results showed that 9-cis-retinoic acid and all-trans retinoic acid enhanced the assembly of connexin32 into gap junctions. Our results further showed that 9-cis-retinoic acid and all-trans-retinoic acid prevented androgen-regulated degradation of gap junctions, post-translationally, independent of androgen receptor mediated signaling. Finally, our findings showed that formation of gap junctions sensitized connexin32-expressing LNCaP cells to the growth modifying effects of 9-cis-retinoic acid, all-trans-retinoic acid and androgens. Thus, the effects of retinoids and androgens on growth and the formation and degradation of gap junctions and their function might be related to their ability to modulate prostate growth and cancer.

Topics: Alitretinoin; Androgens; Cell Line, Tumor; Connexins; Gap Junction beta-1 Protein; Gap Junctions; Humans; Male; Prostatic Neoplasms; Retinoids; Tretinoin

p27 is a cell cycle suppressor gene, whose protein is a negative regulator of cyclin/cdk complexes. p27 is also a potential target of retinoids in cancer prevention studies. In benign prostate hyperplasia (BPH), and in most carcinomas, p27(Kip1) is down-regulated, suggesting its potential resistance to retinoids. To test this hypothesis, we examined the efficacy of 9-cis retinoic acid (9cRA) to suppress prostate cell proliferation (PECP) and carcinogenesis in p27(Kip1) deficient mice.. p27(Kip1) deficient (-/-), heterozygous (+/-) and homozygous (+/+) mice were treated for 7 days with testosterone, 9cRA, or with both, and cell proliferation in dorsolateral prostate (DLP) was determined by BrdU labeling. Prostate carcinogenesis was induced by N-Methyl-N-Nitrosourea (MNU) and hormone stimulation.. PECP in DLP of two-month-old mice of all genotypes was similar but significantly increased in old p27-/- mice only. Testosterone treatment increased PECP in all three p27 genotypes with the highest values in p27-/- mice. p27(Kip1) deficiency did not affect the response of PEC to 9cRA and to 9cRA+testosterone. The decrease of p27(Kip1) in p27+/- and p27-/- mice progressively increased the incidence and frequency of PIN and tumors. 9cRA suppressed PIN in all three p27 genotypes and this was associated with decreased PECP and increased cellular senescence.. This data indicates that p27(Kip1) deficiency promotes prostate cell proliferation and carcinogenesis but does not affect 9cRA's potential to suppress prostate carcinogenesis, suggesting that patients with PIN and carcinomas lacking or having a low level of p27(Kip1) expression may also benefit from clinical trials with retinoids.

Topics: Alitretinoin; Animals; Antineoplastic Agents; Cell Cycle; Cell Proliferation; Cyclin-Dependent Kinase Inhibitor p27; Disease Models, Animal; Gene Expression Regulation, Neoplastic; Genotype; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Prostatic Neoplasms; Retinoids; Tretinoin

Human PCAN1 (prostate cancer gene 1) is a prostate-specific gene that is highly expressed in prostate epithelial tissue, and frequently mutated in prostate tumors. To better understand the regulation of the PCAN1 gene, a 2.6-kb fragment of its 5' flanking region was obtained by PCR. Its promoter activity was examined via the dual-luciferase reporter assay after it had been cloned into a pGL(3)-basic vector generating pGL(3)-p2.6 kb and transfected into LNCaP cells. pGL(3)-basic and pGL(3)-control were respectively used as the negative and positive controls. Sequence analysis with the MatInspector database showed that some possible binding sites for the transcriptional factors, NKX3.1, P53, SP1, cEBP and the PPAR/RXR heterodimers may locate on a 2.6-kb region upstream of the PCAN1 gene. To examine the relevant regulation of PCAN1, pGL(3)-p2.6 kb was transfected into the prostate cancer cell line LNCaP, which was treated with R1881 (10(-7) approximately 10(-9) mol/l), 17beta-estradiol (17beta-E(2), 10(-7) approximately 10(-9) mol/l), all-trans-retinoic acid (all-trans-RA, 10(-5) approximately 10(-7) mol/l) or 9-cis-retinoic acid (9-cis-RA, 10(-5) approximately 10(-7) mol/l), and eukaryotic expression plasmids of NKX3.1, p53, Sp1, Pten, PPARgamma or cEBPalpha were cotransfected with pGL(3)-p2.6 kb into LNCaP cells. pRL-TK, a Renilla luciferase reporter vector, was cotransfected into all the transfection lines as an internal control. The activities of pGL(3)-p2.6 kb (PCAN1 promoter) were analyzed via the dual-luciferase reporter assay 48 h after transfection. The results showed that 9-cis-RA enhanced the PCAN1 promoter activity in a dose-dependent manner, while R1881, 17beta-E(2) and all-trans-RA had no significant effect on PCAN1 promoter activities. Cotransfection with pGL(3)-p2.6kb and the expression plasmids of NKX3.1, p53, Sp1 or Pten respectively resulted in 1.66-, 2.48-, 2.00-and 1.72-fold 2.6 kb PCAN1 promoter activity increases relative to the controls, which were cotransfected with pcDNA3.1(+), while cotransfection of PPARgamma and cEBPalpha yielded no significant effect on PCAN1 promoter activities. These results could be applied for further study of the function and transcription regulation of the PCAN1 gene in prostate development and carcinogenesis.

Topics: Alitretinoin; Base Sequence; Cell Line, Tumor; Cloning, Molecular; Gene Expression Regulation; Humans; Luciferases; Male; Molecular Sequence Data; Neoplasm Proteins; Promoter Regions, Genetic; Prostate; Prostatic Neoplasms; Transcription Factors; Transcriptional Activation; Transfection; Tretinoin

Parathyroid hormone-related protein (PTHrP) increases the growth and osteolytic potential of prostate cancer cells, making it important to control PTHrP expression in these cells. We show that 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) and its non-hypercalcemic analog, EB1089, decrease PTHrP mRNA and cellular protein levels in the androgen-dependent human prostate cancer cell line LNCaP and its androgen-independent derivative, the C4-2 cell line. This effect is mediated via a negative Vitamin D response element (nVDREhPTHrP) within the human PTHrP gene and involves an interaction between nVDREhPTHrP and the Vitamin D receptor (VDR). The retinoid X receptor (RXR) is a frequent heterodimeric partner of the VDR. We show that RXRalpha forms part of the nuclear protein complex that interacts with nVDREhPTHrP along with the VDR in LNCaP and C4-2 cells. We also show that the RXR ligand, 9-cis-retinoic acid, downregulates PTHrP mRNA levels; this decrease is more pronounced in LNCaP than in C4-2 cells. In addition, 9-cis-retinoic acid enhances the 1,25(OH)2D3-mediated downregulation of PTHrP expression in both cell lines; this effect also is more pronounced in LNCaP cells. Proliferation of LNCaP, but not C4-2, cells is decreased by 9-cis-retinoic acid. Promoter activity driven by nVDREhPTHrP cloned upstream of the SV40 promoter and transiently transfected into LNCaP and C4-2 cells is downregulated in response to 1,25(OH)2D3 and EB1089 in both cell lines. Co-treatment with these compounds and 9-cis-retinoic acid further decreases CAT activity in LNCaP, but not C4-2, cells. These results indicate that PTHrP gene expression is regulated by 1,25(OH)2D3 in a cell type-specific manner in prostate cancer cells.

Topics: Alitretinoin; Calcitriol; Cell Line, Tumor; Cell Proliferation; Gene Expression Regulation, Neoplastic; Humans; Male; Parathyroid Hormone-Related Protein; Prostatic Neoplasms; Receptors, Calcitriol; Retinoid X Receptors; RNA, Messenger; Time Factors; Tretinoin; Tumor Cells, Cultured; Vitamin D Response Element

To study the regulatory effects of 9-cis retinoic acid (RA) on the expression of human homeobox gene NKX3.1 in prostate cancer cell line LNCaP.. Flow cytometry, reverse transcriptase polymerase chain reaction and Western blotting were performed to evaluate the effects of 9-cis RA on NKX3.1 expression and cell cycle of LNCaP cells. To identify a regulatory region within the NKX3.1 promoter contributing to the regulation induced by 9-cis RA, we have constructed an NKX3.1 promoter-reporter plasmid, pGL3-1040bp, and its 5'-deletion mutants, which were transfected into LNCaP cells with treatment of 9-cis RA in indicated concentrations.. With the treatment of 9-cis RA, the NKX3.1 promoter activity was increased in reporter gene assay and NKX3.1 expression was enhanced at both mRNA and protein levels in LNCaP cells. We found that the region between -936 and -921 in the upstream of NKX3.1 gene involved the inducible regulation by 9-cis RA treatment. In flow cytometry, 9-cis RA treatment caused accumulation of cells in the G(1) phase of the cell cycle and a fewer cells pass through to G(2)/M.. Our results demonstrated that 9-cis RA as a differentiating agent can arrest prostate cancer cells in G(1) phase and reduce cell mitosis, and upregulate the expression of human homeobox gene NKX3.1, which is thought to play an important role in prostate differentiation and to act as a tumor suppressor gene in the prostate.

Topics: Alitretinoin; Base Sequence; Cell Cycle; Cell Differentiation; Cell Line, Tumor; DNA Primers; Flow Cytometry; Gene Expression Regulation, Neoplastic; Homeodomain Proteins; Humans; Male; Promoter Regions, Genetic; Prostatic Neoplasms; Reverse Transcriptase Polymerase Chain Reaction; Transcription Factors; Tretinoin; Up-Regulation

The efficacy of MDI-301, a non-toxic novel synthetic retinoid, was found to be equivalent to the natural 9-cis-retinoic acid (RA) in vitro against estrogen-dependent MCF7 and T47D breast cancer cell lines which express RA receptor (RAR) alpha. Both retinoids also showed similar efficacy against established PC-3 prostate carcinoma xenografts. MCF7 tumor xenografts showed a reduction in tumor growth of 48% without systemic side-effects upon treatment with MDI-301 compared with MCF7 controls. Tumor xenografts derived from MDA-MB-231, an estrogen-independent breast cancer cell line that expresses low levels of RARalpha, were unresponsive. This study demonstrates that MDI-301 is as efficacious as 9-cis-RA against cancer cells with RARalpha, with no signs of toxicity in vivo, making it a potential candidate for cancer therapy.

Topics: Alitretinoin; Animals; Antineoplastic Agents; Breast Neoplasms; Cell Line, Tumor; Female; Humans; Immunohistochemistry; Male; Mice; Mice, Nude; Prostatic Neoplasms; Receptors, Retinoic Acid; Retinoic Acid Receptor alpha; Retinoids; Tretinoin; Xenograft Model Antitumor Assays

1,25-Dihydroxyvitamin D(3) [1,25-(OH)(2)D(3)] is an effective agent for inhibiting the growth of prostate cancer cells including LNCaP and PC-3 cell lines. However, the extent of growth inhibition in these cell lines differs because LNCaP cells are much more responsive than PC-3 cells. Previous studies in LNCaP cells have shown that 1,25-(OH)(2)D(3) treatment results in G(0)/G(1) cell cycle accumulation, loss of Ki67 expression, and induction of apoptosis. One difference between the two cell lines is that PC-3 cells lack functional p53, a protein that plays roles both in cell cycle regulation and induction of apoptosis. In this study, the role of p53 in 1,25-(OH)(2)D(3) action was examined using the p53-negative PC-3 cells and a line of LNCaP cells, called LN-56, in which p53 function was shut off using a dominant negative p53 fragment. We found that treatment with 1,25-(OH)(2)D(3) extensively inhibits growth of LN-56 prostate cancer cells lacking p53, but in contrast to the parental LNCaP cells, the LN-56 cells recover rapidly. Moreover, in prostate cancer cells, the synergism between 1,25-(OH)(2)D(3) and 9-cis retinoic acid appears to be dependent on the presence of functional p53; however, 1,25-(OH)(2)D(3)-mediated induction of G(1) cell cycle accumulation and induction of apoptosis is not.

Topics: Alitretinoin; Apoptosis; Calcitriol; Cell Division; Cyclin-Dependent Kinase Inhibitor p21; Cyclins; Drug Synergism; G1 Phase; Humans; Ki-67 Antigen; Male; Prostatic Neoplasms; Proto-Oncogene Proteins c-bcl-2; Resting Phase, Cell Cycle; Tretinoin; Tumor Cells, Cultured; Tumor Suppressor Protein p53

The vitamin D(3) receptor, which is the nuclear receptor for 1alpha,25-dihydroxyvitamin D(3) (VD(3)), forms a heterodimer with the retinoid X receptor (RXR), which is the nuclear receptor for 9-cis-retinoic acid (9-cis-RA). The heterodimer binds to a specific response element consisting of two directly repeated pairs of motifs, AGGTGA, spaced by three nucleotides [direct repeat (DR) 3] and modulates the expression of VD(3)-responsive genes. Telomerase activity, which is seen in most immortal cells and germ cells, is a complex of enzymes that maintain the length of telomeres. One of the major components of human telomerase, human telomerase reverse transcriptase (hTERT), is the catalytic subunit, and the expression of hTERT might correlate most strongly with telomerase activity. We found that the sequence of 5'-AGTTCATGGAGTTCA-3' (DR3') is similar to that of DR3 in the promoter region of hTERT. Our results showed that the combination of VD(3) and 9-cis-RA inhibited telomerase activity through direct interaction of the heterodimer of vitamin D(3) receptor and RXR with the DR3' sequence in the hTERT promoter as well as the combination of VD(3) and selective RXR ligand did. Also, in vivo data showed that the growth of xenografts in nude mice was inhibited by VD(3) and 9-cis-RA. The results of the present study provide evidence on the molecular mechanism of the inhibition of cell growth by these agents, and they could be novel therapeutic agents for prostate cancer.

Topics: Alitretinoin; Animals; Antineoplastic Agents; Calcitriol; DNA-Binding Proteins; Drug Synergism; Humans; Male; Mice; Promoter Regions, Genetic; Prostatic Neoplasms; Reverse Transcriptase Polymerase Chain Reaction; Telomerase; Transcription, Genetic; Tretinoin; Tumor Cells, Cultured

In most previous studies, the incidence and multiplicity of chemically induced prostate tumors have been used as end points for assessing the efficacy of various chemopreventive agents. In this study, we used prostate intraepithelial neoplasia (PIN) in Noble rats as an intermediate end point to examine the chemopreventive efficacy of two retinoids, 9-cis-retinoic acid (9cRA) and 4-(hydroxyphenyl)retinamide, which in previous studies have shown promising inhibitory effects on various carcinogenesis models. We found that 80-100% of Noble rats treated for 36 weeks with testosterone + 17beta-estradiol developed multiple PIN lesions predominantly in the dorso-lateral prostate, which appears relevant to the place of origin of PIN and carcinoma in the human prostate. 9cRA at 50 or 100 mg/kg diet significantly decreased the multiplicity of PIN, whereas 4-(hydroxyphenyl) retinamide at 392 or 784 mg/kg diet, did not have an inhibitory effect on PIN. Thus, we provide for the first time evidence that the testosterone + 17beta-estradiol-induced PIN in Noble rats could be used as a potential intermediate end point in assessing the efficacy of retinoids and possibly of other agents on prostate carcinogenesis, and that 9cRA alone or in combination with other agents may have clinical promise in preventing the development of prostate cancer in men.

Topics: Alitretinoin; Animals; Anticarcinogenic Agents; Body Weight; Estradiol; Fenretinide; Male; Prostatic Intraepithelial Neoplasia; Prostatic Neoplasms; Rats; Testosterone; Tretinoin

A chemoprevention study was conducted to evaluate the activity of 9-cis-retinoic acid (9-cis-RA) as an inhibitor of prostate carcinogenesis in male Wistar-Unilever (HsdCpb:Wu) rats. After pretreatment with a sequential regimen of cyproterone acetate (50 mg/kg/day for 21 days) and testosterone propionate (100 mg/kg/day for 3 days), groups of 40 rats received a single i.v. injection of N-methyl-N-nitrosourea (MNU; 30 mg/kg body weight). Beginning 2 weeks after carcinogen administration, rats received chronic exposure to testosterone administered in s.c. implanted silastic capsules. The study was terminated at 13 months after MNU administration, and prostate cancer incidence was determined by histopathological evaluation of step sections of accessory sex glands. Continuous dietary administration of 9-cis-RA at 100 mg/kg diet or 50 mg/kg diet beginning 1 week before MNU administration reduced cancer incidence in the dorsolateral + anterior prostate from 65% in dietary controls to 18 and 20%, respectively (P < 0.001 for both comparisons). Similarly, these dose levels of 9-cis-RA reduced the incidence of cancer in all accessory sex glands from 79% in dietary controls to 48 and 33% (P < 0.01 for both comparisons), respectively. Chronic dietary administration of 9-cis-RA induced no gross or organ-specific toxicity in any animal and did not suppress group mean body weight gain. The potent anticarcinogenic activity of 9-cis-RA in the rat prostate, when considered with its apparent lack of toxicity in rodents, suggests that this and other ligands for the retinoid X receptor merit consideration for evaluation in clinical prostate cancer chemoprevention trials.

Topics: Alitretinoin; Animals; Anticarcinogenic Agents; Carcinogens; Cyproterone Acetate; Dose-Response Relationship, Drug; Male; Methylnitrosourea; Neoplasms, Hormone-Dependent; Prostatic Neoplasms; Rats; Rats, Wistar; Testosterone; Tretinoin

We have recently shown that 1alpha,25-dihydroxyvitamin D3 [1,25-(OH)2D3] inhibits proliferation of LNCaP cells, an androgen-responsive human prostate cancer cell line. Also, 1,25-(OH)2D3 increases androgen receptor (AR) abundance and enhances cellular responses to androgen in these cells. In the current study, we have investigated the mechanism by which 1,25-(OH)2D3 regulates AR gene expression and the involvement of AR in the 1,25-(OH)2D3- and 9-cis retinoic acid (RA)-mediated growth inhibition of LNCaP cells. Northern blot analyses demonstrated that the steady-state messenger RNA (mRNA) level of AR was significantly increased by 1,25-(OH)2D3 in a dose-dependent manner. Time-course experiments revealed that the increase of AR mRNA by 1,25-(OH)2D3 exhibited delayed kinetics. In response to 1,25-(OH)2D3, AR mRNA levels were first detected to rise at 8 h and reached a maximal induction of 10-fold over the untreated control at 48 h; the effect was sustained at 72 h. Furthermore, the induction of AR mRNA by 1,25-(OH)2D3 was completely abolished by incubation of cells with cycloheximide, a protein synthesis inhibitor. 1,25-(OH)2D3 was unable to induce expression of an AR promoter-luciferase reporter. Together, these findings indicate that the stimulatory effect of 1,25-(OH)2D3 on AR gene expression is indirect. Western blot analyses showed an increase of AR protein in 1,25-(OH)2D3-treated cells. This increased expression of AR was followed by 1,25-(OH)2D3-induced inhibition of growth in LNCaP cells. Similar to 1,25-(OH)2D3, 9-cis RA also induced AR mRNA expression, and the effect of both hormones was additive. Moreover, 1,25-(OH)2D3 and 9-cis RA acted synergistically to inhibit LNCaP cell growth. These antiproliferative effects of 1,25-(OH)2D3 and 9-cis RA, alone or in combination, were blocked by the pure AR antagonist, Casodex. In conclusion, our results demonstrate that growth inhibition of LNCaP cells by 1,25-(OH)2D3 and 9-cis RA is mediated by an AR-dependent mechanism and preceded by the induction of AR gene expression. This finding, that differentiating agents such as vitamin D and A derivatives are potent inducers of AR, may have clinical implications in the treatment of prostate cancer.

Topics: Alitretinoin; Calcitriol; Cell Division; Cycloheximide; Gene Expression Regulation, Neoplastic; Humans; Male; Prostatic Neoplasms; Protein Synthesis Inhibitors; Receptors, Androgen; Tretinoin; Tumor Cells, Cultured; Up-Regulation

The high incidence and long latent period of prostate cancer make it an ideal target for chemoprevention. We have evaluated a series of agents for chemopreventive efficacy using a model in which hormone-dependent prostate cancers are induced in the Wistar-Unilever (WU) rat by sequential treatment with antiandrogen (cyproterone acetate), androgen (testosterone propionate), and direct-acting chemical carcinogen (N-methyl-N-nitrosourea), followed by chronic androgen stimulation (testosterone). This regimen reproducibly induces prostate cancers in high incidence, with no gross toxicity and a low incidence of neoplasia in the seminal vesicle and other non-target tissues. Dehydroepiandrosterone (DHEA) and 9-cis-retinoic acid (9-cis-RA) are the most active agents identified to date. DHEA inhibits prostate cancer induction both when chronic administration is begun prior to carcinogen exposure, and when administration is delayed until preneoplastic prostate lesions are present. 9-cis-RA is the most potent inhibitor of prostate carcinogenesis identified; a study to determine the efficacy of delayed administration of 9-cis-RA is in progress. Liarozole fumarate confers modest protection against prostate carcinogenesis, while N-(4-hydroxyphenyl)retinamide (fenretinide), alpha-difluoromethylornithine, oltipraz, DL-alpha-tocopherol acetate (vitamin E), and L-selenomethionine are inactive. Chemoprevention efficacy evaluations in the WU rat will support the identification of agents that merit study for prostate cancer chemoprevention in humans.

Topics: Alitretinoin; Animals; Antineoplastic Agents; Dehydroepiandrosterone; Disease Models, Animal; Immunohistochemistry; Male; Neoplasms, Hormone-Dependent; Prostate; Prostatic Neoplasms; Rats; Rats, Wistar; Treatment Outcome; Tretinoin

1,25 dihydroxyvitamin D3 (1,25D) and retinoids may play an important role in preventing progression of prostate cancer.. We examined the ability of four novel 20-epi-vitamin D3 analogs (CB1093, KH1060, KH1266, and CB1267), either alone or in combination with 9-cis retinoic acid (RA) to inhibit colony growth of a human prostate cancer cell line, LNCaP, using soft agar as well as bone marrow stroma. Also, the effect of these analogs on the cell cycle and expression of Ki-67, p21(waf-1), and p27(kip1) in LNCaP cells was examined.. The analog CB1267 was the most potent, with 8 x 10(-10) M of the analog inhibiting 50% colony growth (ED50) of LNCaP. 9-cis-RA also inhibited colony growth of LNCaP (ED50, 5 x 10(-7) M). Combined, CB1267 and 9-cis-RA synergistically inhibited colony growth and significantly increased the number of LNCaP cells in G0/G1 phase. Cell cycle arrest was associated with increased levels of p21(waf-1) and p27(kip1) and decreased expression of Ki-67 protein. Pulse-exposure to this combination (5 x 10(-8) M) irreversibly inhibited colony growth, both in soft agar and on normal human bone marrow stroma.. Combination of a new vitamin D3 analog (CB1267) and a retinoid (9-cis-RA) potently inhibited colony formation of LNCaP prostate cancer cells in vitro, suggesting further studies in animal models. This combination may afford an interesting therapeutic approach to low-burden prostate cancer.

Topics: Alitretinoin; Antineoplastic Agents; Bone Marrow Cells; Calcitriol; Cell Cycle; Cell Cycle Proteins; Cell Division; Cell Survival; Cyclin-Dependent Kinase Inhibitor p21; Cyclin-Dependent Kinase Inhibitor p27; Cyclins; Drug Synergism; Humans; Male; Microtubule-Associated Proteins; Prostatic Neoplasms; Stromal Cells; Tretinoin; Tumor Cells, Cultured; Tumor Suppressor Proteins

Recent studies have suggested that the active metabolite of vitamin D3, 1,25-dihydroxyvitamin D3, can inhibit the growth and/or induce the differentiation of a variety of cell types and that these characteristics might be useful in the treatment of some cancers. Retinoids also promote the differentiation and inhibit the growth of some cells. That the vitamin D receptor acts as a heterodimer with the retinoid X receptor (RXR) suggests that there may be functional interactions between 1,25-dihydroxyvitamin D3 and retinoids. In this study, we show that the combination of 1,25-dihydroxyvitamin D3 and 9-cis retinoic acid synergistically inhibits the growth of LNCaP prostate cancer cells. That this effect is mediated by RXR rather than retinoic acid receptors was shown using RXR- and retinoic acid receptor-specific ligands. The vitamin D3 analog, EB1089, inhibited growth more effectively than 1,25-dihydroxyvitamin D3 and also acted synergistically with 9-cis-retinoic acid. These treatments caused cells to accumulate in the G1 phase of the cell cycle, suggesting that 1,25-dihydroxyvitamin D3 can regulate one or more factors critical for the G1/S transition.

Topics: Alitretinoin; Calcitriol; Cell Division; DNA-Binding Proteins; Drug Synergism; G1 Phase; Humans; Male; Nuclear Proteins; Prostate; Prostatic Neoplasms; Receptors, Retinoic Acid; Retinoid X Receptors; Transcription Factors; Tretinoin; Tumor Cells, Cultured