Vitamin D And Apc Modulate Molecular Crosstalk Between Beta-Catenin/Wnt Signaling And The Vitamin D Receptor In Colon Cancer Cells

1217 Vitamin D is implicated as a protective factor in human colon carcinogenesis. Anti-tumor activity of vitamin D on colonocytes is thought to be mediated by 1,25(OH) 2 D 3 , the active hormonal metabolite of vitamin D. We examined the role of vitamin D on a putative VDR/β-catenin interaction. We first sought to confirm that VDR associates physically with β-catenin regulating critical Wnt signaling events in colon tumorigenesis. Second, we assessed the role of the intact and mutant adenomatous polyposis coli (APC) gene, commonly dysregulated in human colon cancers, as a modulator of the VDR/β-catenin interaction. Third, we investigated the role of germline allele variation in APC as potential commonly occurring gene modifiers of individual susceptibility for the formation of colorectal adenomas. Glutathione-S-transferase (GST) pulldown assays and an in-vitro transcription/translation system to produce β-catenin protein support an interaction between β-catenin and VDR that appears independent of 1,25(OH) 2 D 3 . Further evaluation of this interaction within the cell using a mammalian two hybrid system in transfected HT29-APC, a cell line which allows inducible wildtype APC protein, suggest that APC positively influences the interaction between β-catenin and VDR, and that this association is enhanced at least 6-fold in the presence of 1,25(OH) 2 D 3 . To determine the functional significance of VDR and 1,25(OH) 2 D 3 on the transcriptional activity of β-catenin, a TOPflash assay is being implemented in HT-29 cells. Thus, our biochemical results support the presence of a VDR and β-catenin interaction that occurs independent of 1,25(OH) 2 D 3 and that appears to be significantly enhanced in the presence of both the hormone and intact APC. In an effort to determine if polymorphic variation in APC influenced risk of colorectal neoplasia, haplotype analyses of single nucleotide polymorphisms located in APC were assessed for their relationship with colorectal adenoma recurrence. A strong association (OR = 0.74, 95% CI = 0.59 - 0.92) was observed for a T-A haplotype subgroup and risk of recurrent adenoma. Overall, the data presented herein suggest an APC/VDR binary or ternary complex with β-catenin. This represents a novel crosstalk mechanism whereby 1,25(OH) 2 D 3 -VDR and APC could negatively regulate the Wnt signaling pathway to restrict tumorigenesis. Furthermore, genetic variation in APC may alter the efficiency of the interaction of this complex resulting in variable risk of adenoma recurrence among individuals.