In vitro studies revealed a downregulation of Wnt/β-catenin cascade by active vitamin D and TX 527 analog in a Kaposi's sarcoma cellular model

Abstract

The Kaposi´s sarcoma-associated herpesvirus G-protein-coupled receptor (vGPCR) is a key molecule in the pathogenesis of Kaposi´s sarcoma. We have previously demonstrated that 1α,25(OH)2D3 or its less calcemic analogTX 527 exerts antiproliferative effects in endothelial cells stable expressing vGPCR. Since it is well documentedthat vGPCR activates the canonical Wnt/β-catenin signaling pathway, the aim of this study was to evaluate ifWnt/β-catenin cascade is target of 1α,25(OH)2D3 or TX 527 as part of their antineoplastic mechanism. Firstly,Western blot studies showed an increase in β-catenin protein levels in a dose and time dependent manner; andwhen VDR was knockdown, β-catenin protein levels were significantly decreased. Secondly, β-catenin localization, investigated by immunofluorescence and subcellular fractionation techniques, was found increased in thenucleus and plasma membrane after 1α,25(OH)2D3 treatment. VE-cadherin protein levels were also increased inthe plasma membrane fraction. Furthermore, β-catenin interaction with VDR was observed by co-immunoprecipitation and mRNA expression of β-catenin target genes was found decreased. Finally, DKK-1, theextracellular inhibitor of Wnt/β-catenin pathway, showed an initial upregulation of mRNA expression.Altogether, the results obtained by different techniques revealed a downregulation of Wnt/β-catenin cascadeafter 1α,25(OH)2D3 or TX 527 treatment, showing the foundation for a potential chemotherapeutic agent.