Vitamin D analogue TX 527 down‐regulates the NF ‐κ B pathway and controls the proliferation of endothelial cells transformed by K aposi sarcoma herpesvirus

Background and Purpose The K aposi sarcoma (KS)‐associated herpesvirus GPCR (v GPCR ) is a key molecule in the pathogenesis of KS , where it increases NF ‐κ B gene expression and activates the NF ‐κ B pathway. We investigated whether the less calcemic vitamin D analogue TX 527 inhibited the proliferation of endothelial cells transformed by v GPCR by modulation of the NF ‐κ B pathway. Experimental Approach Endothelial cells transformed by v GPCR ( SVEC ‐v GPCR ) were treated with TX 527. Proliferation was measured by 3‐(4,5‐dimethylthiazol‐2‐yl)‐5‐(3‐carboxymethoxyphenyl)‐2‐(4‐sulfophenyl)‐2 H ‐tetrazolium, inner salt ( MTS ) and cell cycle by flow cytometry. m RNA and protein levels were measured by real‐time quantitative reverse transcriptase‐PCR (q RT ‐ PCR ) and immunoblot analysis respectively. Key Results TX 527, similar to bortezomib (0.5 nM), a proteasome inhibitor that inhibits the activation of NF‐κB , reduced proliferation and induced G 0/ G 1 cell cycle arrest in SVEC ‐v GPCR . TX 527 like 1α,25( OH ) 2 D 3 , biological active form of vitamin D , decreased the activity of NF ‐κ B comparable with the effect of bortezomib. Time‐response studies showed that TX 527 significantly decreased NF ‐κ B and increased I κ Bα m RNA and protein levels. The increase of I κ Bα was accompanied by a reduction in p65/ NF ‐κ B translocation to the nucleus. These responses were abolished when vitamin D receptor ( VDR ) expression was suppressed by stable transfection of sh RNA against VDR . I n parallel with NF‐κB inhibition, there was a down‐regulation of inflammatory genes such as IL ‐6, CCL 2/ MCP and CCL 20/ MIP 3α. Conclusions and Implications These results suggest that the anti‐proliferative effects of the vitamin D analogue TX 527 in SVEC ‐v GPCR occur by modulation of the NF ‐κ B pathway and are VDR dependent.