calcitriol and Osteosarcoma

Osteosarcoma (OS) is the basic bone neoplasm with lower survival and poor prognosis. It is distinguished by its offensive nature and metastatic potential. The fundamental death source in OS patients is lung metastasis. In addition, the proliferation and cell migration are thus essential for cancer progression, especially for intrusion and transformation. Several studies have illustrated that 1,25-Dihydroxyvitamin D (1,25(OH)2D) has a critical role in the growth and differentiation of bone. However, knowledge of the outcome of 1,25(OH)2D on the progression and incursion of osteosarcoma cells is minimal.. The present study aimed to analyze the effect of different concentrations of 1,25(OH)2D on the multiplication, progression, and intrusion of OS cells and verify the effective doses of 1,25(OH)2D that can decrease the intensity of the disease and improving the prognosis in OS patients.. Saos-2 cells were treated with 1,25(OH)2D (0, 50, 100, and 200 nM) for 48, 72, and 96 hours. Proliferation, invasion, and migration were determined by MTT assay, Transwell assay, and Scratch test, respectively. The levels of c-Myc and FOXO1 proteins were determined by Western blotting.. The proliferation, invasiveness, and migration of Saos-2 cells that were treated with 1,25(OH)2D were significantly decreased compared with untreated cells. Although 1,25(OH)2D notably decreased c-Myc protein levels (after 48 and 72 hours), FOXO1 protein levels have been significantly increased after 48 and 72 hours. 1,25(OH)2D and the vitamin D receptor (VDR) suppress c-Myc function through regulating the c-Myc/MXD1 network and thus, providing a molecular basis of 1,25(OH)2D related to the cancer-preventive actions.. Based on the present results, 1,25(OH)2D by targeting c-Myc and FOXO1 expression displays anti-invasive, anti-migration and anti-proliferative effects on OS cells in vitro. Our findings suggest that effective doses of the 1,25(OH)2D may reduce the aggressive potential of the OS cell line. However, further investigation and clinical trials are needed.

Topics: Antineoplastic Agents; Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; Bone Neoplasms; Cell Line, Tumor; Cell Movement; Cell Proliferation; Forkhead Box Protein O1; Humans; Neoplasm Invasiveness; Osteosarcoma; Proto-Oncogene Proteins c-myc; Receptors, Calcitriol; Repressor Proteins; Signal Transduction; Vitamin D

To clarify the molecular mechanism for analog potency and selectivity, we investigated the ability of 1,25(OH)(2)D(3) analogs to recruit coactivators to the vitamin D receptor (VDR) transcriptional complex. Using a modified version of the avidin-biotin complex DNA binding assay, we discovered that 20S-analogs enhance the binding of specific coactivators to the transcriptional complex relative to natural hormone and that the enhanced binding occurs independently of vitamin D response element and cell type. With the exception of two of these coactivators, DRIP205 and DRIP240, all proteins were recruited to the transcriptional complex in a dose-dependent manner. While the results do not provide an explanation for tissue selectivity of 2-methylene-19-nor-(20S)-1,25-dihydroxyvitamin D(3) (2MD), they provide evidence that in the presence of a full-length side chain, the 20S configuration improves binding of specific proteins to the VDR transcriptional complex while modifications at carbon 2 do not.

Topics: Animals; Cell Line; Gene Expression Regulation, Neoplastic; Osteosarcoma; Rats; Receptors, Calcitriol; Transcriptional Activation; Vitamin D

1alpha,25-dihydroxy vitamin D3 has a major role in the regulation of the bone metabolism as it promotes the expression of key bone-related proteins in osteoblastic cells. In recent years it has become increasingly evident that in addition to its well-established genomic actions, 1alpha,25-dihydroxy vitamin D3 induces non-genomic responses by acting through a specific plasma membrane-associated receptor. Results from several groups suggest that the classical nuclear 1alpha,25-dihydroxy vitamin D3 receptor (VDR) is also responsible for these non-genomic actions of 1alpha,25-dihydroxy vitamin D3. Here, we have used siRNA to suppress the expression of VDR in osteoblastic cells and assessed the role of VDR in the non-genomic response to 1alpha,25-dihydroxy vitamin D3. We report that expression of the classic VDR in osteoblasts is required to generate a rapid 1alpha,25-dihydroxy vitamin D3-mediated increase in the intracellular Ca(2+) concentration, a hallmark of the non-genomic actions of 1alpha,25-dihydroxy vitamin D3 in these cells.

Topics: Animals; Genome; Osteosarcoma; Rats; Receptors, Calcitriol; RNA, Small Interfering; Vitamin D

We recently identified 1alpha,25-dihydroxy-3-epi-vitamin D3 [1alpha,25(OH)2-3-epi-D3] as a metabolite of 1alpha,25-dihydroxyvitamin D3 [1alpha,25(OH)2D3] produced in rat osteosarcoma cells (UMR 106). We now report the isolation of 24R,25-dihydroxy-3-epi-vitamin D3 [24R,25(OH)2-3-epi-D3] as a metabolite of 24R,25-dihydroxyvitamin D3 [24R,25(OH)2D3] by high-performance liquid chromatography (HPLC) with chiral column and its structure assignment by proton nuclear magnetic resonance (1H-NMR) and liquid chromatography-mass spectrometry (LC-MS) analysis. We also demonstrated the production of 24R,25(OH)2-3-epi-D, in two other cell lines [human colon carcinoma cells (Caco-2) and porcine kidney cells (LLC-PK1)] which were previously shown to convert 1alpha,25(OH)2D3 into 1alpha,25(OH)2-3-epi-D3. It can be seen that the production of 24R,25(OH)2- 3-epi-D3 from 24R,25(OH)2D3 is lower than that of 1alpha,25(OH)2-3-epi-D3 from 1alpha,25(OH)2D3 in all the cells studied. 24R,25(OH)2-3-epi-D3 was found to be inactive in terms of its ability to bind to the vitamin D receptor (VDR), in inhibiting proliferation and in inducing differentiation of human promyelocytic leukemia cells (HL-60). Thus, our study indicates that the C-3 epimerization pathway is common to both 1alpha,25(OH)2D3 and 24R,25(OH)2D3 and may play an important role in modulating the concentration and the biological activity of these two major vitamin D3 metabolites in target tissues.

Topics: 24,25-Dihydroxyvitamin D 3; Angiogenesis Inhibitors; Animals; Bone Neoplasms; Caco-2 Cells; Chromatography, High Pressure Liquid; Gas Chromatography-Mass Spectrometry; Humans; Isomerism; Magnetic Resonance Spectroscopy; Osteosarcoma; Swine; Tumor Cells, Cultured; Vitamin D

Methotrexate is commonly used in the treatment of rheumatoid arthritis. An osteopathy has been described in children treated with methotrexate for leukaemia, consisting of bone pain, osteoporosis and fractures. Animals given short-term high-dose and long-term low-dose methotrexate have both reduced bone formation and increased resorption on histomorphometry. As patients with rheumatic diseases have numerous risk factors for osteoporosis, possible additional risk from low-dose methotrexate is of relevance to the rheumatologist. To investigate further the mechanism of osteoporosis in animals and man, in vitro studies were carried out on an osteoblast cell line, using concentrations found in patients with rheumatic disease. UMR 106 rat osteoblast-like osteosarcoma cells were incubated with methotrexate, and also with sulphasalazine, an anti-rheumatic drug with no known effect on bone, for comparison. A dose-dependent toxic effect of methotrexate on the cell line was observed using concentrations found in patients with rheumatic disease. This was not observed with sulphasalazine. The reduced bone formation observed in animals and man may be due to a direct effect of methotrexate on the osteoblast.

Topics: Alkaline Phosphatase; Animals; Antirheumatic Agents; Bone Neoplasms; Cell Count; Cell Division; Cell Survival; Dose-Response Relationship, Drug; Endothelium, Vascular; Humans; Leucovorin; Methotrexate; Osteosarcoma; Rats; Sulfapyridine; Sulfasalazine; Tumor Cells, Cultured; Vitamin D

Aromatic compounds 2a-c, analogs of 1 alpha, 25-dihydroxyvitamin (calcitriol, 1), and heteroaromatic compounds 4a-c and 5a-c, analogs of 19-nor-1 alpha, 25-dihydroxyvitamin D3 (3), were designed to simulate the topology of their biologically potent parent compounds while avoiding previtamin D equilibrium. Convergent and facile total syntheses of the analogs (+)-2b, (+)-2c, (-)-4b, and (-)-5b were achieved via carbonyl addition of regiospecifically formed organolithium nucleophiles to the enantiomerically pure C,D-ring ketone (+)-17, characteristic of natural calcitriol (1). Likewise, hybrid analogs 20a-c were prepared to determine whether incorporation of a known potentiating side chain would lead to increased biological activity. Preliminary in vitro biological testing showed that aromatic analogs (+)-2b, (+)-2c, and 20a-c as well as heteroaromatic analogs (-)-4b and (-)-5b have very low affinities for the calf thymus vitamin D receptor but considerable antiproliferative activities in murine keratinocytes at micromolar concentration. No biological advantage was observed in this keratinocyte assay for the doubly modified hybrid analogs 20a-c over the singly modified parent (+)-2b. Analog (+)-2b, but surprisingly not the corresponding analog 20b differing from (+)-2b only in the side chain, showed considerable activity in nongenomic opening of calcium channels in rat osteosarcoma cells.

Topics: Animals; Calcitriol; Calcium Channel Blockers; Cattle; Cell Division; Drug Design; Keratinocytes; Magnetic Resonance Spectroscopy; Mice; Molecular Structure; Osteosarcoma; Rats; Receptors, Calcitriol; Thymus Gland; Vitamin D