Primary melanoma expression of the vitamin D receptor (VDR) is protective for melanoma survival and is associated with increased tumor immune response, decreased Wnt/B-catenin signaling and tumor proliferation

Abstract

VDR loss has been associated with cutaneous melanoma progression (Brozyna et al, 2011) and VDR polymorphisms have been shown to affect melanoma survival (Mocellin et al, 2008) but the genomic basis remains to be explored. We used microarray data from 700 treatment-naïve melanoma primaries (FFPE) from the Leeds Melanoma Cohort (LMC) to perform agnostic bioinformatic analyses that identified candidate pathways/genes associated with VDR expression. The concordance of these In-silico pathways with histopathological measures was assessed, followed by In-vitro and In-vivo experiments to establish cause-effect. We found that VDR gene expression is protective for melanoma survival in the LMC (Haz Ratio=0.51, P=5.7x10-8) independent of AJCC stage (adjusted Haz Ratio=0.77, P=0.002) and also in the TCGA melanoma data (Haz Ratio=0.83, P=0.001). A genome-wide correlation analysis (FDR<5%) produced 2025 positively correlating genes (β>0.2) and 1383 negative correlating genes (β<-0.2), which were then used for functional enrichment analyses (Reactome FIViz, Wu and Stein 2012). The positive correlating genes were enriched for immune function: ECM organization, TNF signaling, IFNγ signaling, IL12-mediated signaling and NFκB signaling. In line with this, VDR expression was higher in tumors with Tumour Infiltrating Lymphocytes (TILs) compared to those without TILs (P=0.02). VDR also correlated positively with imputed immune cells scores (Angelova et al, 2015) particularly central memory CD4 cells, cytotoxic cells, DCs, NK cells, MDSCs, Neutrophils and T cells (P<10-16). Conversely, the negative correlating genes were enriched for proliferation pathways: Mitotic Prophase, Wnt signaling pathway, Mitochondrial translation, TCA cycle and oxidative phosphorylation. In line with this, VDR expression was lower in tumors with increased mitotic cells (P=0.002). Notable among the negatively correlated pathways was Wnt signaling pathway-VDR has been shown to inhibit Wnt/β-catenin signalling in colon carcinoma cells (Larriba et al, 2011) and β-catenin signalling inhibits melanoma immune infiltration (Spranger et al, 2015). Thus we hypothesized that VDR inhibits β-catenin signalling and hence increases tumor immune infiltration. To test this, stable-transfected B16-BL6:VDR mouse melanoma cells were used in an In-vivo metastatic colonisation assay (Speak et al, 2017, ongoing work). Collectively, we conclude that VDR is of prognostic significance in a cohort of 700 primary melanomas and has a pro-immune and anti-proliferative role, based on highly concordant In-silico and histopathological evidence. Taken together, our results provide a novel insight into the effect of VDR in melanoma survival and evidence for the use of VDR as a potentially significant prognostic marker.

Publication
Tumor Biology 2018; 78(Suppl 13):Abstract 5205

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