calcitriol and Adenocarcinoma

MiR-100 and miR-125b are lost in many cancers and have potential function as tumor suppressors. Using both primary prostatic epithelial cultures and laser capture-microdissected prostate epithelium from 45 patients enrolled in a vitamin D3 randomized trial, we identified miR-100 and -125b as targets of 1,25-dihydroxyvitamin D3 (1,25D). In patients, miR-100 and -125b levels were significantly lower in tumor tissue than in benign prostate. Similarly, miR-100 and -125b were lower in primary prostate cancer cells than in cells derived from benign prostate. Prostatic concentrations of 1,25D positively correlated with these miRNA levels in both prostate cancer and benign epithelium, showing that patients with prostate cancer may still benefit from vitamin D3. In cell assays, upregulation of these miRNAs by 1,25D was vitamin D receptor dependent. Transfection of pre-miR-100 and pre-miR-125b in the presence or absence of 1,25D decreased invasiveness of cancer cell, RWPE-2. Pre-miR-100 and pre-miR-125b decreased proliferation in primary cells and cancer cells respectively. Pre-miR-125b transfection suppressed migration and clonal growth of prostate cancer cells, whereas knockdown of miR-125b in normal cells increased migration indicates a tumor suppressor function. 1,25D suppressed expression of previously bona fide mRNA targets of these miRNAs, E2F3 and Plk1, in a miRNA-dependent manner. Together, these findings show that vitamin D3 supplementation augments tumor suppressive miRNAs in patient prostate tissue, providing evidence that miRNAs could be key physiologic mediators of vitamin D3 activity in prevention and early treatment of prostate cancer.

Topics: Adenocarcinoma; Adult; Aged; Apoptosis; Biomarkers, Tumor; Blotting, Western; Case-Control Studies; Cell Cycle Proteins; Cell Movement; Cell Proliferation; Cells, Cultured; Clinical Trials, Phase II as Topic; E2F3 Transcription Factor; Follow-Up Studies; Gene Expression Profiling; Humans; Laser Capture Microdissection; Male; MicroRNAs; Middle Aged; Oligonucleotide Array Sequence Analysis; Polo-Like Kinase 1; Prognosis; Prostate; Prostatic Neoplasms; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Real-Time Polymerase Chain Reaction; Receptors, Calcitriol; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; RNA, Small Interfering; Vitamin D

Epidemiological studies suggest an inverse correlation between dietary calcium (Ca(2+)) and vitamin D intake and the risk of colorectal cancer (CRC). It has been shown in vitro that the active vitamin D metabolite, 1,25-dihydroxyvitamin D3 (1,25-D3) can upregulate expression of the calcium-sensing receptor (CaSR). In the colon, CaSR has been suggested to regulate proliferation of colonocytes. However, during tumorigenesis colonic CaSR expression is downregulated and we hypothesized that the loss of CaSR could influence the anti-tumorigenic effects of Ca(2+) and vitamin D. Our aim was to assess the impact of CaSR expression and function on the anti-neoplastic effects of 1,25-D3 in colon cancer cell lines. We demonstrated that in the healthy colon of mice, high vitamin D diet (2500 IU/kg diet) increased expression of differentiation and apoptosis markers, decreased expression of proliferation markers and significantly upregulated CaSR mRNA expression, compared with low vitamin D diet (100 IU/kg diet). To determine the role of CaSR in this process, we transfected Caco2-15 and HT29 CRC cells with wild type CaSR (CaSR-WT) or a dominant negative CaSR mutant (CaSR-DN) and treated them with 1,25-D3 alone, or in combination with CaSR activators (Ca(2+) and NPS R-568). 1,25-D3 enhanced the anti-proliferative effects of Ca(2+) and induced differentiation and apoptosis only in cells with a functional CaSR, which were further enhanced in the presence of NPS R-568, a positive allosteric modulator of CaSR. The mutant CaSR inhibited the anti-tumorigenic effects of 1,25-D3 suggesting that the anti-neoplastic effects of 1,25-D3 are, at least in part, mediated by the CaSR. Taken together, our data provides molecular evidence to support the epidemiological observation that both, vitamin D and calcium are needed for protection against malignant transformation of the colon and that their effect is modulated by the presence of a functional CaSR. This article is part of a Special Issue entitled '17th Vitamin D Workshop'.

Topics: Adenocarcinoma; Aniline Compounds; Animals; Caco-2 Cells; Calcium; Cell Differentiation; Cell Proliferation; Cell Transformation, Neoplastic; Colon; Colonic Neoplasms; Dietary Supplements; Gene Expression Regulation, Neoplastic; HT29 Cells; Humans; Male; Mice; Mice, Transgenic; Mutation; Phenethylamines; Propylamines; Receptors, Calcium-Sensing; Receptors, G-Protein-Coupled; RNA, Messenger; Signal Transduction; Transfection; Vitamin D

The active metabolite of vitamin D3, 1,25-dihydroxyvitamin D3 (calcitriol) has antiproliferative effects in non-aggressive prostate cancer, however, its effects in more aggressive model systems are still unclear. In these studies, effects of calcitriol and a less-calcemic vitamin D analog, QW-1624F2-2 (QW), were tested in vivo, using the aggressive autochthonous transgenic adenocarcinoma of mouse prostate (TRAMP) model. To study prevention of androgen-stimulated prostate cancer, vehicle, calcitriol (20 µg/kg), or QW (50 µg/kg) were administered to 4 week-old TRAMP mice intraperitoneal (i.p.) 3×/week on a MWF schedule for 14 weeks. Calcitriol and QW slowed progression of prostate cancer as indicated by reduced urogenital tract (p = 0.0022, calcitriol; p = 0.0009, QW) and prostate weights (p = 0.0178, calcitriol; p = 0.0086, QW). However, only calcitriol increased expression of the pro-differentiation marker, cadherin 1 (p = 0.0086), and reduced tumor proliferation (p = 0.0467). By contrast, neither vitamin D analog had any effect on castration resistant prostate cancer in mice treated pre- or post-castration. Interestingly, although vitamin D showed inhibitory activity against primary tumors in hormone-intact mice, distant organ metastases seemed to be enhanced following treatment (p = 0.0823). Therefore, TRAMP mice were treated long-term with calcitriol to further examine effects on metastasis. Calcitriol significantly increased the number of distant organ metastases when mice were treated from 4 weeks-of-age until development of palpable tumors (20-25 weeks-of-age)(p = 0.0003). Overall, data suggest that early intervention with vitamin D in TRAMP slowed androgen-stimulated tumor progression, but prolonged treatment resulted in development of a resistant and more aggressive disease associated with increased distant organ metastasis.

Topics: Adenocarcinoma; Animals; Blotting, Western; Cell Differentiation; Cell Proliferation; Disease Models, Animal; Humans; Immunoenzyme Techniques; Incidence; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Prostatic Neoplasms; Prostatic Neoplasms, Castration-Resistant; Tumor Cells, Cultured; Vitamin D

1α,25-Dihydroxyvitamin D3 (1,25-D3) is antiproliferative in preclinical models of lung cancer, but in tumor tissues, its efficacy may be limited by CYP24A1 expression. CYP24A1 is the rate limiting catabolic enzyme for 1,25-D3 and is overexpressed in human lung adenocarcinoma (AC) by unknown mechanisms.. The DNA methylation status of CYP24A1 was determined by bisulfite DNA pyrosequencing in a panel of 30 lung cell lines and 90 surgically resected lung AC. The level of CYP24A1 methylation was correlated with CYP24A1 expression in lung AC cell lines and tumors. In addition, histone modifications were assessed by quantitative chromatin immunoprecipitation-polymerase chain reaction (ChIP-qPCR) in A549, NCI-H460, and SK-LU-1.. Bisulfite DNA pyrosequencing analysis revealed that CYP24A1 gene was heterogeneously methylated in lung AC. Expression of CYP24A1 was inversely correlated with promoter DNA methylation in lung AC cell lines and tumors. Treatment with 5-aza-2'-deoxycytidine (5-Aza) and trichostatin A (TSA) increased CYP24A1 expression in lung AC. We observed that CYP24A1 promoter hypermethylation decreased CYP24A1 enzyme activity in vitro, whereas treatment with 5-Aza and/or TSA increased CYP24A1 enzyme affinity for its substrate 1,25-D3. In addition, ChIP-qPCR analysis revealed specific histone modifications within the CYP24A1 promoter region. Treatment with TSA increased H3K4me2 and H3K9ac and simultaneously decreased H3K9me2 at the CYP24A1 promoter and treatment with 5-Aza and/or TSA increased the recruitment of vitamin D receptor (VDR) to vitamin D response elements (VDRE) of the CYP24A1 promoter.. The expression of CYP24A1 gene in human lung AC is in part epigenetically regulated by promoter DNA methylation and repressive histone modifications. These findings should be taken into consideration when targeting CYP24A1 to optimize antiproliferative effects of 1,25-D3 in lung AC.

Topics: Adenocarcinoma; Aged; Blotting, Western; Chromatin Immunoprecipitation; DNA Methylation; Enzyme-Linked Immunosorbent Assay; Epigenesis, Genetic; Female; Flow Cytometry; Follow-Up Studies; Gene Expression Regulation, Neoplastic; Humans; Lung Neoplasms; Male; Neoplasm Recurrence, Local; Neoplasm Staging; Prognosis; Promoter Regions, Genetic; Real-Time Polymerase Chain Reaction; Receptors, Calcitriol; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Survival Rate; Tumor Cells, Cultured; Vitamin D; Vitamin D3 24-Hydroxylase

Oxidative catabolism of 1α,25-dihydroxyvitamin D(3) [1α,25(OH)(2)D(3)] is mediated by either CYP24A1 or CYP3A4. In this paper, we tested whether induction of CYP3A4 in the LS180 intestinal cell model enhances clearance of 1α,25(OH)(2)D(3) and blunts its hormonal effect on expression of the apical membrane calcium transport protein, TRPV6. Treatment with the hPXR agonist rifampin significantly increased CYP3A4 mRNA content and catalytic activity, but had no effect on CYP24A1 or TRPV6 mRNA content. Pre-treating cells with rifampin for 48h, prior to a 24h 1α,25(OH)(2)D(3) treatment phase, was associated with a subsequent 48% increase in the elimination of 1α,25(OH)(2)D(3) and a 35% reduction of peak TRPV6 mRNA. Introduction of the CYP3A4 inhibitor, 6',7'-dihydroxybergamottin, an active inhibitor in grapefruit juice, reversed the effects of rifampin on 1α,25(OH)(2)D(3) clearance and TRPV6 expression. Over-expression of hPXR in LS180 cells greatly enhanced the CYP3A4 responsiveness to rifampin pretreatment, and elicited a greater relative suppression of TRPV6 expression and an increase in 1α,25(OH)(2)D(3) disappearance rate, compared to vector expressed cells, following hormone administration. Together, these results suggest that induction of CYP3A4 in the intestinal epithelium by hPXR agonists can result in a greater metabolic clearance of 1α,25(OH)(2)D(3) and reduced effects of the hormone on the intestinal calcium absorption, which may contribute to an increased risk of drug-induced osteomalacia/osteoporosis in patients receiving chronic therapy with potent hPXR agonists. Moreover, ingestion of grapefruit juice in the at-risk patients could potentially prevent this adverse drug effect.

Topics: Adenocarcinoma; Caco-2 Cells; Calcium; Cell Line, Tumor; Colonic Neoplasms; Cytochrome P-450 CYP3A; Dose-Response Relationship, Drug; Enzyme Induction; Humans; Pregnane X Receptor; Receptors, Steroid; Rifampin; Vitamin D

Vitamin D receptor (VDR) mediates the antitumoral action of the active vitamin D metabolite 1alpha,25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3)). VDR expression is lost during colon cancer progression causing unresponsiveness to 1,25(OH)(2)D(3) and its analogs. Previously, Snail1, an inducer of epithelial-to-mesenchymal transition (EMT), was reported to inhibit VDR expression. Here, we show that Snail2/Slug, but not other EMT inducers such as Zeb1, Zeb2, E47 or Twist1, represses VDR gene promoter. Moreover, Snail2 and Snail1 show additive repressing effect on VDR promoter. Snail2 inhibits VDR RNA and protein and blocks the induction of E-cadherin and an adhesive phenotype by 1,25(OH)(2)D(3). Snail2 reduces the ligand-induced VDR transcriptional activation of a consensus response element and of the CYP24 promoter. Concordantly, Snail2 inhibits the induction of CYP24 RNA and p21(CIP1), filamin A and vinculin proteins and the repression of c-MYC by 1,25(OH)(2)D(3). Additionally, Snail2 abrogates beta-catenin nuclear export and the antagonism of the transcriptional activity of beta-catenin-T-cell factor complexes by 1,25(OH)(2)D(3). SNAI2 expression is upregulated in 58% of colorectal tumors and correlates inversely with that of VDR. However, VDR downregulation is higher in tumors coexpressing SNAI2 and SNAI1 than in those expressing only one of these genes. Together, these data indicate that Snail2 and Snail1 cooperate for VDR repression in colon cancer.

Topics: Adenocarcinoma; beta Catenin; Blotting, Western; Cadherins; Colorectal Neoplasms; Fluorescent Antibody Technique; Humans; Immunoenzyme Techniques; Nuclear Proteins; Promoter Regions, Genetic; Receptors, Calcitriol; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Signal Transduction; Snail Family Transcription Factors; Transcription Factors; Transcription, Genetic; Twist-Related Protein 1; Vitamin D; Wnt Proteins

Topics: Adenocarcinoma; Aged, 80 and over; Fibroblast Growth Factor-23; Fibroblast Growth Factors; Humans; Hypophosphatemia; Male; Osteomalacia; Parathyroid Hormone; Prostatic Neoplasms; Syndrome; Vitamin D

To determine if blood levels of 25-hydroxyvitamin D (25-D) or its active metabolite, 1,25-dihydroxyvitamin D (1,25-D), are lower in women at the time of first diagnosis of breast cancer than in comparable women without breast cancer.. This was a clinic-based case-control study with controls frequency-matched to cases on race, age, clinic and month of blood drawing.. University-based breast referral clinics.. One hundred and fifty-six women with histologically documented adenocarcinoma of the breast and 184 breast clinic controls.. There were significant mean differences in 1,25-D levels (pmol ml(-1)) between breast cancer cases and controls; white cases had lower 1,25-D levels than white controls (mean difference +/-SE: -11.08+/-0.76), and black cases had higher 1.25-D levels than black controls (mean difference +/-SE: 4.54+/-2.14), although the number of black women in the study was small. After adjustment for age, assay batch, month of blood draw, clinic and sample storage time, the odds ratio (95% confidence interval, CI) for lowest relative to highest quartile was 5.2 (95% CI 2.1, 12.8) for white cases and controls. The association in white women was stronger in women above the median age of 54 than in younger women, 4.7 (95% CI 2.1, 10.2) vs. 1.5 (95% CI 0.7, 3.0). There were no case-control differences in 25-D levels in either group.. These data are consistent with a protective effect of 1,25-D for breast cancer in white women.

Topics: Adenocarcinoma; Adult; Analysis of Variance; Breast Neoplasms; Case-Control Studies; Female; Humans; Logistic Models; Odds Ratio; Radioimmunoassay; Risk Factors; Vitamin D