Selective inhibition of cyclooxygenase-2 (COX-2) by 1 α,25-dihydroxy- 16-ene-23-yne-vitamin D₃, a less calcemic vitamin D analog

Inducible cyclooxygenase-2 (COX-2) has been implicated to play a role in inflammation and carcinogenesis and selective COX-2 inhibitors have been considered as anti-inflammatory and cancer chemopreventive agents. 1α,25-dihydroxyvitamin D₃ (1α,25(OH)₂D ₃), the active hormonal form of vitamin D₃ also has been considered to be a cancer chemopreventive agent in addition to its important role in maintaining calcium homeostasis. Based on these observations, we studied the direct effect of 1α,25(OH)₂D₃ and one of its less calcemic synthetic analogs, 1α,25(OH)₂-16-ene-23-yne- D₃ on the activity of both COX-1 and COX-2 in an in vitro enzyme assay. Preliminary data indicated that both 1α,25(OH)₂D ₃ and 1α,25(OH)₂-16-ene-23-yne-D₃ inhibited selectively the activity of COX-2 with no effect on the activity of COX-1. Out of the two compounds, 1α,25(OH)₂-16-ene-23-yne- D₃ was found to be more effective with an IC₅₀ of 5.8 nM. Therefore, the rest of the experiments were performed using 1α,25(OH)-16- ene-23-yne-D₃ only. 1α,25(OH)₂-16-ene-23-yne-D ₃ inhibited the proliferation of lipopolysaccharide (LPS) stimulated mouse macrophage cells (RAW 264.7) with a reduction in the expression of COX-2 along with other inflammatory mediators like inducible nitric oxide synthase (iNOS) and interleukin-2 (IL-2). Furthermore, 1α,25(OH) ₂-16-ene-23-yne-D₃ also inhibited carrageenan induced inflammation in an air pouch of a rat and effectively reduced the expression of COX-2, iNOS, and IL-2 in the tissues of the same air pouch. In both cases, 1α,25(OH)₂-16-ene-23-yne-D₃ did not show any effect on the expression of COX-1. In summary, our results indicate that 1α,25(OH)₂-16-ene-23-yne-D₃, a less calcemic vitamin D analog, exhibits potent anti-inflammatory effects and is a selective COX-2 inhibitor.