Poster Presentation 27th Lorne Cancer Conference 2015

Identifying primary targets of vitamin D in breast cancer (#263)

Lei SHENG 1 , Kathleen I Pishas 1 , Paul H Anderson 2 , Howard A Morris 3 , Deepak J Dhatrak 4 , Grantley P Gill 5 , David F Callen 1
  1. Cancer Therapeutics Laboratory, School of Medicine, University of Adelaide, Adelaide, South Australia, Australia
  2. Musculoskeletal Biology Research Laboratory, School of Pharmacy and Medical Sciences, University of South Australia, Adelaide, SA, Australia
  3. Endocrine Bone Research Laboratory, SA Pathology, Adelaide, SA, Australia
  4. SA Pathology, Adelaide, SA, Australia
  5. Breast, Endocrine and Surgical Oncology Unit, Royal Adelaide Hospital, Adelaide, SA, Australia

Background and Aims: Breast cancer is the second leading cause of cancer associated death in women worldwide. Multiple studies have shown that lower vitamin D levels are associated with an increased risk of breast cancer. By binding to its intracellular vitamin D receptor (VDR), 1 alpha, 25-dihydroxyvitamin D3 (1,25D), the biologically active form of vitamin D3, exerts its effect on a wide range of tissues, resulting in a variety of biological effects including anti-tumor activity. However, the underlying molecular mechanisms of the protective effect of vitamin D remain unclear. The aim of this study is to identify VDR controlled target genes and molecular pathways that mediate the anti-tumor effect of 1,25D using human breast tissue.

Methodology: Ex vivo cultured human malignant breast tissues (n=3), adjacent normal (n=2) as well as reduction mammoplasty (n=1) were treated with 100nM of 1,25D or vehicle control for 24 hours. Total RNA was extracted and subjected to cDNA libraries preparation and Illumina DNA sequencing.

Results: mRNA-Seq revealed 559 significantly differentially expressed genes, 320 of which were upregulated and 239 downregulated in 1,25D-treated breast cancer.GoSeq KEGG pathway analysis revealed significant downregulation of several cellular metabolic pathways in 1,25D-treated breast cancer explants including cellular nitrogen compound/cellular macromolecule/nucleic acid metabolic processes as well as transferase activity and ATP binding. Pathways that were enriched in 1,25D-treated breast cancer explants included cell adhesion molecules (CAMs), ECM-receptor interaction, focal adhesion, and tight junction processes, suggesting that 1,25D can enhance intercellular adhesion. RNA-seq and qRT-PCR analysis also revealed that CYP24A1, CLMN, SERPINB1, and EFTUD1 are upregulated by 1,25D. Following VDR knockdown in non-malignant and malignant breast cell lines (n=3), upregulation of these four target genes was ablated following 1,25D treatment, confirming that induction of these target genes by 1,25D are VDR dependent.

Conclusion: In conclusion, this study successfully identified primary VDR targets and several molecular pathways involved in the anti-tumor activities of vitamin D3 in human breast cancer.