Effects of 'non-calcaemic' vitamin D analogues on 24-hydroxylase expression in MG-63 osteoblast-like cells

Background: New 'non-calcaemic' analogues of 1,25-dihydroxyvitamin D 3 (1,25(OH) 2 D 3 ) are entering the clinical arena and some of them have been shown to have differential effects in bone. This may have a bearing on the evolution of bone lesions in uraemic patients receiving vitamin D therapies. A potential mechanism for differential effects of analogues lies in their target cell inactivation. Methods: Using a human osteoblastic cell line, MG-63, three analogues, 22-oxacalcitriol (OCT), 19-nor-1,25-dihydroxyvitamin D 2 (paricalcitol) and 1α,25-dihy-droxydihydrotachysterol 2 (1,25(OH) 2 DHT 2 ), were compared with 1,25(OH) 2 D 3 for (1) their affinity for the vitamin D receptor (VDR) by competitive displacement of tritiated 1,25(OH) 2 D 3 from calf thymus VDR; (2) effects on 24-hydroxylase mRNA expression using comparative RT-PCR, and (3) rates of metabolism, using high performance liquid chromatography, over a 24-hour time course. Results: Relative VDR-binding affinities (IC 50 ) were 1,25(OH) 2 D 3 (100%), OCT (25%), paricalcitol (14%) and 1,25(OH) 2 DHT 2 (0.3%). A ≥3-fold increase in 24-hydroxylase mRNA expression was observed for all compounds at 2 h peaking at 7- to 8-fold above control levels by 12 h, with no significant difference between the analogues and 1,25(OH) 2 D 3 . Differences in their rates of metabolism were observed [calculated t1/2 values = OCT (1.2 h) > paricalcitol (2.3 h) > 1,25(OH) 2 D 3 (2.6 h) > 1,25(OH) 2 DHT 2 (3.4 h)], with OCT having a significantly shorter half-life. Conclusion: In MG-63 cells these analogues up-regulate 24-hydroxylase mRNA expression with similar potency, in each case accelerating ligand inactivation, despite significant differences in VDR affinity. VDR affinity did not correspond to either 24-hydroxylase mRNA expression or the rates of ligand disappearance, suggesting cellular metabolism is one of several factors that determine the analogue specificity of these agents in bone.