Gene re-expression chemoprevention strategies in prostate cancer

Authors
Moray J. Campbell and Nicholas D. James
University of Birmingham

The vitamin D receptor (VDR) is a member of the nuclear receptor super family, which forms a co-ordinated network of ligand activated transcription factors that regulate genes involved in proliferation and differentiation of the prostate gland.  The VDR, when vitamin D₃is bound, targets a number of anti-proliferative target genes (including the cyclin dependent kinase inhibitor p21^(wafI/cipI)) and acutely inhibits proliferation of normal prostate epithelial cells. By contrast, the response is corrupted in prostate cancer cells thus impeding clinical applications of vitamin D₃. Similarly epidemiological evidence and polymorphism studies of the VDR have linked prostate cancer with reduced exposure and sensitivity to the vitamin D_3. Taken together, these data suggest that low exposure to vitamin D₃ or cellular mechanisms that suppress sensitivity are both associated with the malignancy. Dissection of molecular mechanisms that drive cellular insensitivity will generate novel options to identify and treat aggressive prostate cancer.

In the absence of ligand the VDR is part of a large multimeric repressive complex containing co-repressors such as NCoR1, SMRT and Alien and associated histone deacetylases (HDAC). This complex maintains chromatin in a condensed form that suppresses transcription. Vitamin D₃ induces conformational changes in the VDR that release it from this complex and promotes binding of co-activators, associated with histone acetylases (HAT), thereby relaxing DNA-histone interactions and facilitating transcription. We are investigating whether dysregulation of these complexes results in transcriptional silencing of vitamin D₃ antiproliferative target genes. In support of this model we have found;

• elevated co-repressor mRNA expression correlates with reduced sensitivity to the antiproliferative effects of vitamin D₃ in cancer cell line and primary models.
• antiproliferative responses and induction of apoptosis can be restored by co-treatment of vitamin D₃ with minimally active doses of HDAC inhibitors such as Trichostatin A (Figure 1)
• a unique group of antiproliferative target genes, identified by cDNA micro array analysis, that are co-ordinately regulated by vitamin D₃ and HDAC inhibitors.

Control
TSA plus Vitamin D3
Figure 1: loss of mitochondrial membrane integrity and induction of apoptosis in prostate cancer cells treated with vitamin D3 and an HDAC inhibitor

Our data supports a model whereby insensitivity to vitamin D₃ arises from the squelching of VDR action by NCoR/SMRT multimeric complexes, which maintain the promoter region of target genes in a heterochormatic state (Figure 2).

We are currently dissecting the capacity of VDR-co-repressor interactions to suppress VDR transcriptional activity and antiproliferative responsiveness by the use of siRNA, blocking peptides and SAHA (a novel, clinically relevant HDAC inhibitor).  In parallel, prospective analysis of TRUSS biopsies and retrospective analysis of archive material, both by lazar capture microdissection will assess VDR, co-repressors and suppressed target genes as novel biological markers of disease stage. These biomarkers will be used in a neo-adjuvant clinical trial of 1a,25 dihydroxyvitaminD₃ as a prelude to the use of SAHA alone and then the combination of agents.