1-25(oh)2-16-ene-23-yne-d3 and Osteosarcoma

The secosteroid hormone 1alpha,25-dihydroxyvitamin D3 [1alpha,25(OH)2D3] is metabolized in its target tissues through modifications of both the side chain and the A-ring. The C-24 oxidation pathway, the previously well established main side chain modification pathway, is initiated by hydroxylation at C-24 of the side chain. The C-3 epimerization pathway, the newly discovered A-ring modification pathway, is initiated by epimerization of the hydroxyl group at C-3 of the A-ring. The end products of the metabolism of 1alpha,25(OH)2D3 through the C-24 oxidation and the C-3 epimerization pathways are calcitroic acid and 1alpha,25-dihydroxy-3-epi-vitamin-D3 respectively. During the past two decades, numerous noncalcemic analogs of 1alpha,25(OH)2D3 were synthesized. Several of the analogs have altered side chain structures and as a result some of these analogs have been shown to resist their metabolism through side chain modifications. For example, two of the analogs, namely, 1alpha,25-dihydroxy-16-ene-23-yne-vitamin D3 [1alpha,25(OH)2-16-ene-23-yne-D3] and 1alpha,25-dihydroxy-16-ene-23-yne-20-epi-vitamin D3 [1alpha,25(OH)2-16-ene-23-yne-20-epi-D3], have been shown to resist their metabolism through the C-24 oxidation pathway. However, the possibility of the metabolism of these two analogs through the C-3 epimerization pathway has not been studied. Therefore, in our present study, we investigated the metabolism of these two analogs in rat osteosarcoma cells (UMR 106) which are known to express the C-3 epimerization pathway. The results of our study indicate that both analogs [1alpha,25(OH)2-16-ene-23-yne-D3 and 1alpha,25(OH)2-16-ene-23-yne-20-epi-D3] are metabolized through the C-3 epimerization pathway in UMR 106 cells. The identity of the C-3 epimer of 1alpha,25(OH)2-16-ene-23-yne-D3 [1alpha,25(OH)2-16-ene-23-yne-3-epi-D3] was confirmed by GC/MS analysis and its comigration with synthetic 1alpha,25(OH)2-16-ene-23-yne-3-epi-D3 on both straight and reverse-phase HPLC systems. The identity of the C-3 epimer of 1alpha,25(OH)2-16-ene-23-yne-20-epi-D3 [1alpha,25(OH)2-16-ene-23-yne-20-epi-3-epi-D3] was confirmed by GC/MS and 1H NMR analysis. Thus, we indicate that vitamin D analogs which resist their metabolism through the C-24 oxidation pathway, have the potential to be metabolized through the C-3 epimerization pathway. In our present study, we also noted that the rate of C-3 epimerization of 1alpha,25(OH)2-16-ene-23-yne-20-epi-D3 is about 10 times greater than the r

Topics: Animals; Calcitriol; Chromatography, High Pressure Liquid; Gas Chromatography-Mass Spectrometry; Magnetic Resonance Spectroscopy; Osteosarcoma; Oxygen; Rats; Time Factors; Tumor Cells, Cultured

We compared the separate effects of 1alpha,25-dihydroxyvitamin D3 (1alpha,25(OH)2D3) and its analog, 1alpha,25-dihydroxy-16ene,23yne-vitamin D3 (1alpha25(OH)2-16ene,23yne-D3), as well as their interactions with 17-beta estradiol (E2) in our human osteosarcoma SaOS-2 cell models representing two stages of differentiation, the SaOS+DEX and SaOS-DEX cells. SaOS+DEX cells have been previously shown to express higher PTH-stimulated adenylate cyclase (PTH-AC) and basal alkaline phosphatase (ALP) activities compared with SaOS-DEX cells. ALP: In SaOS+DEX cells, 0.1 nmol/L analog, but not 1alpha,25(OH)2D3, increased ALP activity 1.7-fold (p < 0.05). Instead, 1 nmol/L 1alpha,25(OH)2D3 increased ALP 1.4-fold (p < 0.05). In these cells, E2 enhanced 1alpha,25(OH)2D3-stimulated ALP activity (ANOVA, F = 51.22, p <0.0001), while inhibiting the effect of the analog. [3H]-Thymidine uptake: In SaOS+DEX cells, 1alpha,25(OH)2D3 had biphasic effects (ANOVA, F = 13.08, p < 0.0001), which were not altered by E2. In contrast, the analog was stimulatory only with E2 (ANOVA, F = 3.59, p < 0.025). Osteocalcin (OC): 1alpha,25(OH)2D3 and its analog stimulated OC production in SaOS-DEX cells with smaller effects in SaOS+DEX cells. In SaOS-DEX cells, E2 enhanced the effect of 1alpha,25(OH)2D3, but not that of the analog. PTH-AC: In SaOS-DEX cells, 100 nmol/L analog inhibited PTH-AC activities by 50% (p < 0.01), whereas 1alpha,25(OH)2D3 had little effect. In SaOS+DEX cells, both compounds inhibited PTH-AC approximately 35%. E2 inhibited the effect of the analog in SaOS-DEX cells, but enhanced the effects of both compounds in SaOS+DEX cells. These results show that the analog 1alpha,25(OH)2-16ene,23yne-D3 was effective in regulating osteoblastic function; its effects were modulated by E2 and dependent upon the stage of osteoblast differentiation.

Topics: Adenylyl Cyclases; Alkaline Phosphatase; Calcitriol; Cell Differentiation; Dihydroxycholecalciferols; Estradiol; Humans; Osteoblasts; Osteocalcin; Osteosarcoma; Parathyroid Hormone; Thymidine; Tumor Cells, Cultured