seocalcitol and calcipotriene

Multiple genetic and environmental steps may underpin the development of human colorectal neoplasia, and experimental evidence suggests that promoters of colorectal cancer also induce colorectal epithelial cell hyperplasia. In vitro crypt cell production rate (CCPR) was measured to determine the effect of calcium, epidermal growth factor (EGF), vitamin D3 metabolites and synthetic analogues on human rectal epithelial cell proliferation. In a double-blind trial of oral calcium supplementation, CCPR was reduced by 49% in patients with familial adenomatous polyposis (FAP), but there was no effect on established neoplasia. In control tissue, the active form of vitamin D3 (1,25(OH)2D3) reduced rectal CCPR by 57% at 1 microM, 55% at 10 nM and 45% at 100 pM. Likewise, in tissue taken from patients with FAP, 1,25(OH)2D3 reduced CCPR by 52%. Vitamin D3 has profound effects on calcium metabolism, but synthetic analogues can avoid these. The effects of a synthetic analogue (MC-903) on human rectal CCPR were therefore studied. MC-903 (10(-7) M) reduced CCPR in control tissue by 51%, and in FAP tissue by 52% at 10(-6) M and 51% at 10(-7) M. In addition, MC-903 and a related analogue, EB 1089, produced a clear-cut dose-dependent inhibition of both HT-29 and Caco2 colorectal cancer cells maintained in culture. Hence, vitamin D3 and its analogues can reduce the rate of cell proliferation in normal, premalignant and malignant colorectal epithelial cells and might therefore have future therapeutic uses as chemoprotective or chemotherapeutic agents. Lastly, EGF increases CCPR by 102% in FAP tissue that expresses the EGF receptor. Thus, human colorectal cell proliferation is influenced by a variety of luminal and humoral agents and a greater understanding of these actions should help plan future treatments.

Topics: Adenomatous Polyposis Coli; Adolescent; Adult; Aged; Antineoplastic Agents; Calcitriol; Calcium; Cell Division; Double-Blind Method; Epithelium; Female; Humans; In Vitro Techniques; Male; Middle Aged; Rectum; Vitamin D

Vitamin D receptor (VDR) activities have been noted for a number of B cell malignancies which showed varying sensitivities to vitamin D3 (1,25-dihydroxyvitamin D3, VD3, calcitriol) and its synthetic analogs. The objective of this study was to address the potential effects of VD3 and vitamin D3 analogs (VDAs) on the growth of Hodgkin's lymphoma (HL), a malignant pathology of B cell origin, in vitro.. Immunofluorescence staining showed the expression of VDR by primary Hodgkin's (H) and Reed-Sternberg (RS)-HRS-tumor cells in HL histological sections. Western blot analyses revealed expression of VDR in the HL cell lines Hs445, HDLM2, KMH2, and L428. One-way analysis of variance (ANOVA) on data obtained from water-soluble tetrazolium 1 (WST-1) cell proliferation assay showed decreased cell growth in HDLM2 and L428, 72 h after treatment with 10 µM of either VD3 of VDAs. Western blot analyses showed that treatment of L428 cells with the VDAs (calcipotriol and EB1089) resulted in modest increases in nuclear accumulation of VDR (nuVDR) compared to either dimethyl sulfoxide (DMSO) or VD3 treatments. nuVDR for DMSO control and VD3 was comparable. These results suggest that VD3 or VDAs may affect growth of HL.

Topics: Calcitriol; Cell Line, Tumor; Cell Proliferation; Cholecalciferol; Dimethyl Sulfoxide; Gene Expression Regulation, Neoplastic; Hodgkin Disease; Humans; Receptors, Calcitriol; Vitamin D

The plethora of actions of 1alpha,25(OH)(2)D(3) in various systems suggested wide clinical applications of vitamin D nuclear receptor (VDR) ligands in treatments of inflammation, dermatological indication, osteoporosis, cancers, and autoimmune diseases. More than 3000 vitamin D analogues have been synthesized in order to reduce the calcemic side effects while maintaining the transactivation potency of these ligands. Here, we report the crystal structures of VDR ligand binding domain bound to two vitamin D agonists of therapeutical interest, calcipotriol and seocalcitol, which are characterized by their side chain modifications. These structures show the conservation of the VDR structure and the adaptation of the side chain anchored by hydroxyl moieties. The structure of VDR-calcipotriol helps us to understand the structural basis for for the switching of calcipotriol to a receptor antagonist by further side chain modification. The VDR-seocalcitol structure, in comparison with the structure of VDR-KH1060, a superagonist ligand closely related to seocalcitol, shows adaptation of the D ring and position of C-21 in order to adapt its more rigid side chain.

Topics: Calcitriol; Crystallography, X-Ray; Ligands; Models, Molecular; Molecular Structure; Nuclear Proteins; Receptors, Calcitriol; Structure-Activity Relationship

The beta-catenin signaling pathway is deregulated in nearly all colon cancers. Nonhypercalcemic vitamin D3 (1alpha,25-dehydroxyvitamin D(3)) analogues are candidate drugs to treat this neoplasia. We show that these compounds promote the differentiation of human colon carcinoma SW480 cells expressing vitamin D receptors (VDRs) (SW480-ADH) but not that of a malignant subline (SW480-R) or metastasic derivative (SW620) cells lacking VDR. 1alpha,25(OH)2D(3) induced the expression of E-cadherin and other adhesion proteins (occludin, Zonula occludens [ZO]-1, ZO-2, vinculin) and promoted the translocation of beta-catenin, plakoglobin, and ZO-1 from the nucleus to the plasma membrane. Ligand-activated VDR competed with T cell transcription factor (TCF)-4 for beta-catenin binding. Accordingly, 1alpha,25(OH)2D(3) repressed beta-catenin-TCF-4 transcriptional activity. Moreover, VDR activity was enhanced by ectopic beta-catenin and reduced by TCF-4. Also, 1alpha,25(OH)2D(3) inhibited expression of beta-catenin-TCF-4-responsive genes, c-myc, peroxisome proliferator-activated receptor delta, Tcf-1, and CD44, whereas it induced expression of ZO-1. Our results show that 1alpha,25(OH)2D(3) induces E-cadherin and modulates beta-catenin-TCF-4 target genes in a manner opposite to that of beta-catenin, promoting the differentiation of colon carcinoma cells.

Topics: Active Transport, Cell Nucleus; Adenocarcinoma; Antineoplastic Agents; beta Catenin; Cadherins; Calcitriol; Cell Adhesion Molecules; Cell Differentiation; Cell Membrane; Cholecalciferol; Colonic Neoplasms; Cytoskeletal Proteins; Gene Expression Regulation, Neoplastic; Humans; Ligands; Macromolecular Substances; Phenotype; Protein Binding; Receptors, Calcitriol; RNA, Messenger; Signal Transduction; TCF Transcription Factors; Trans-Activators; Transcription Factor 7-Like 2 Protein; Transcription Factors; Transfection; Tumor Cells, Cultured; Vitamin D

This study examined the action of 9-cis retinoic acid and 1,25-dihydroxyvitamin D3 analogues (KH 1060, EB 1089 and MC 903) on the release of calcitonin (CT) and calcitonin gene-related peptide (CGRP) in the rat C cell line CA-77. This cell line mainly secretes CGRP. Using radioimmunoassays (RIAs) for CT and CGRP, we measured the release of both peptides in the culture medium as well as the amount of these proteins contained in the CA-77 C cells. 9-cis retinoic acid decreased the release of both CGRP and CT dose-dependently in the range between 1 nM and 1 microM. The half-effective dose was 10 nM. The treatment of CA-77 C cells with 0.1 microM calcitriol alone only slightly decreased the release of both CT and CGRP. The increase in the amount of CT and CGRP released by the action of 1 microM dexamethasone was reduced by 1 microM 9-cis retinoic acid, and this effect was enhanced by the addition of 0.1 microM calcitriol or KH 1060, EB 1089 and MC 903. When the C cells were continuously stimulated by dexamethasone, after 6 days of exposure to the combined treatment with calcitriol analogues + 9-cis retinoic acid, there was a greater decrease in the amount of CGRP contained in the C cells than after treatment with 9-cis retinoic alone. Our data suggested that combined treatment with retinoic acid and calcitriol analogues exerted a stronger inhibition on the amounts of the two peptides either contained in the cells or released in the medium than each hormone alone.

Topics: Alitretinoin; Animals; Calcitonin; Calcitonin Gene-Related Peptide; Calcitriol; Carcinoma, Medullary; Dexamethasone; Dose-Response Relationship, Drug; Drug Interactions; Kinetics; Rats; Thyroid Neoplasms; Tretinoin; Tumor Cells, Cultured

The antitumoral potency of 1,25-dihydroxyvitamin D3 and its synthetic derivatives MC903, EB1089, and KH1060 was investigated on a tumoral Bowen-like epidermis reconstructed from an immortalized human keratinocyte cell line transfected by expression vectors coding for E6 and E7 of human papillomavirus 16. Treatment of skin equivalents by vitamin D derivatives (10(-9) M or 10(-12) M) was performed during (from day 1 to day 15 of culture) or after tissue reconstruction (from day 15 to day 30). Pharmacologic effects were evaluated by morphologic and immunohistologic analysis and compared with those of controls (vehicle alone) and with treatment of skin equivalents derived from normal keratinocytes. When performed during epidermal reconstruction, treatment of tumoral skin equivalents induced only minor morphologic and immunohistologic changes. Conversely, when performed after epidermal reconstruction, treatment with 1,25-dihydroxyvitamin D3, MC903, and EB1089 clearly improved the phenotype of treated tissues. Morphologic analysis showed reorganization of epidermal layers with the appearance of a distinct basal layer and of stratified orthokeratotic stratum corneum. Immunohistochemical analysis demonstrated that the terminal differentiation markers profilaggrin and cytokeratin 10 were re-expressed in the treated tissues while absent in controls. Overall, the results indicate that 1,25-dihydroxyvitamin D3, MC903, and EB1089 can induce a partial reversion of the tumoral phenotype in this in vitro model.

Topics: Antineoplastic Agents; Calcitriol; Humans; Phenotype; Skin; Skin Neoplasms

We have examined the in vitro effects of 1,25 dihydroxy-vitamin D3 [1,25(OH)2D3] and of two side-chain modified analogs of 1,25(OH)2D3 (EB1089 and MC903) on cell growth and parathyroid hormone related peptide (PTHRP) production in immortalized (HPK1A) and neoplastic (HPK1A-ras) keratinocytes. Cell proliferation was strongly inhibited by 1,25(OH)2D3 and its analogs in HPK1A cells, and in this system EB1089 was 10-100 times more potent than 1,25(OH)2D3 or MC903. A similar effect on cell proliferation was observed in HPK1A-ras cells; however, 10-fold higher concentrations of 1,25(OH)2D3 or its analogs were required. We also observed a strong and dose-dependent inhibitory effect of these compounds on PTHRP secretion and gene expression. In both immortalized and neoplastic keratinocytes, EB1089 was 10-100 times more potent than 1,25(OH)2D3 or MC903 on inhibiting PTHRP production. However, although effective in HPK1A-ras cells, 10-fold higher concentrations of 1,25(OH)2D3 or its analogs were required to produce similar actions in this neoplastic model. These studies therefore demonstrate that a 1,25(OH)2D3 analog with low calcemic potency in vivo (EB1089) can inhibit keratinocyte proliferation and PTHRP production by such cells with greater potency than 1,25(OH)2D3. The observed effects of such analogs in neoplastic keratinocytes predicts their potential usefulness in vivo in inhibiting squamous cancer growth and its associated hypercalcemia.

Topics: Animals; Antineoplastic Agents; Calcitriol; Carcinoma, Squamous Cell; Cell Division; Cell Line, Transformed; Cell Transformation, Viral; Dose-Response Relationship, Drug; Drug Screening Assays, Antitumor; Gene Expression Regulation, Neoplastic; Humans; Hypercalcemia; Keratinocytes; Mice; Mice, Nude; Neoplasm Proteins; Neoplasm Transplantation; Parathyroid Hormone-Related Protein; Protein Biosynthesis; Skin Neoplasms; Tumor Cells, Cultured

Although numerous studies have shown potent antiproliferative and differentiation-inducing effects of 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) and its analogs on cells not directly related to bone metabolism, only few reports focussed on the effects of these analogs on bone. We compared the action of several recently developed analogs with that of 1,25-(OH)2D3 on human (MG-63) and rat (ROS 17/2.8) osteoblast-like cells and on in vitro bone resorption. In MG-63 cells the analogs EB1089 and KH1060 were about 166,000 and 14,000 times more potent than 1,25-(OH)2D3 in stimulating type I procollagen and 100 and 6,000 times more potent in stimulating osteocalcin production, respectively. Also in ROS 17/2.8 cells EB1089 and KH1060 were most potent in inducing osteocalcin synthesis. In vitro bone resorption was 2.3 and 17.5 times more potently stimulated by EB1089 and KH1060, respectively. In MG-63 cells, 1,25-(OH)2D3 and the analogs inhibited cell proliferation, whereas both 1,25-(OH)2D3 and the analogs stimulated the growth of ROS 17/2.8 cells. Differences in potency could neither be explained by affinity for the vitamin D receptor nor by a differential involvement of protein kinase C in the action of the analogs. Together, these data show that also in bone the analogs EB1089 and KH1060 are more potent than 1,25-(OH)2D3 but that the potency of the analogs compared to 1,25-(OH)2D3 is dependent on the biological response. On the basis of these observations it can be concluded that the reported reduced calcemic effect in vivo is not the result of a decreased responsiveness of bone to these analogs. Lastly, in view of eventual clinical application of 1,25-(OH)2D3-analogs, the observed stimulation of in vitro bone resorption and growth of an osteosarcoma cell line warrant in vivo studies to further examine these effects.

Topics: Animals; Antineoplastic Agents; Binding, Competitive; Bone Resorption; Calcitriol; Cell Division; Cells, Cultured; Cholecalciferol; Enzyme Inhibitors; Glyceryl Ethers; Humans; In Vitro Techniques; Osteoblasts; Osteocalcin; Osteosarcoma; Procollagen; Protein Kinase C; Rats; Receptors, Calcitriol; Structure-Activity Relationship; Tumor Cells, Cultured

The nuclear receptor for 1,25-dihydroxyvitamin D3 (VD), VDR, belongs to the nuclear receptor superfamily. This ligand-inducible transcription factor mediates the genomic VD signalling pathways by binding to specific response elements in the promoter region of VD regulated genes. Two types of natural VD response elements are used as models for the VDR-mediated transcriptional activation: one is bound by VDR-homodimers and is found in the human osteocalcin gene promoter, and the other is bound by heterodimers of VDR with retinoid X receptors (RXRs) as in the mouse osteopontin promoter. Here, we demonstrate that the VD analogues MC903, EB1089 and KH1060, previously shown to be potent regulators of proliferation and differentiation, are able to act as ligands for VDR and replace VD as a ligand in both nuclear signalling pathways. We found that they have different potency and sensitivity in their ability to stimulate the hormone-dependent promoter element. MC903 and EB1089 provide about 20% higher induction of gene activity than VD in a gene reporter system, whereas KH1060 was more sensitive, inducing transcription at about 100-fold lower doses than VD. Interestingly, VD and its analogues induce VDR homodimer-mediated gene activity at a 3- to 4-fold lower concentration than that of VDR-RXR heterodimers. This suggests that the ligand concentration is an additional regulatory level in the discrimination between signalling pathways involving homo- and heterodimeric hormone receptors.

Topics: Animals; Base Sequence; Calcitriol; DNA; Gene Expression Regulation; Humans; Ligands; Mice; Molecular Sequence Data; Receptors, Calcitriol; Receptors, Retinoic Acid; Retinoid X Receptors; Transcription Factors; Transfection; Tumor Cells, Cultured