Analysis of the Transcriptome: Regulation of Cancer Stemness in Breast Ductal Carcinoma In Situ by Vitamin D Compounds

Abstract

Ductal carcinoma in situ (DCIS), which accounts for one out of every five new breast cancer diagnoses, will progress to potentially lethal invasive ductal carcinoma (IDC) in about 50% of cases. Vitamin D compounds have been shown to inhibit progression to IDC in the MCF10DCIS model. This inhibition appears to involve a reduction in the cancer stem cell-like population in MCF10DCIS tumors. To identify genes that are involved in the vitamin D effects, a global transcriptomic analysis was undertaken of MCF10DCIS cells grown in mammosphere cultures, in which cancer stem-like cells grow preferentially and produce colonies by selfrenewal and maturation, in the presence and absence of 1a25(OH) 2 D 3 and a vitamin D analog, BXL0124. Using next-generation RNA-sequencing, we found that vitamin D compounds downregulated genes involved in maintenance of breast cancer stem-like cells (e.g., GDF15), epithelial-mesenchymal transition, invasion, and metastasis (e.g., LCN2 and S100A4), and chemoresistance (e.g., NGFR, PPP1R1B, and AGR2), while upregulating genes associated with a basal-like phenotype (e.g., KRT6A and KRT5) and negative regulators of breast tumorigenesis (e.g., EMP1). Gene methylation status was analyzed to determine whether the changes in expression induced by vitamin D compounds occurred via this mechanism. Ingenuity pathway analysis was performed to identify upstream regulators and downstream signaling pathway genes differentially regulated by vitamin D, including TP63 and vitamin D receptor -mediated canonical pathways in particular. This study provides a global profiling of changes in the gene signature of DCIS regulated by vitamin D compounds and possible targets for chemoprevention of DCIS progression to IDC in patients.

The foggy world(s) of p63 isoform regulation in normal cells and cancer

“…Vitamin D upregulated ΔNp63 in keratinocytes through vitamin D receptor (VDR)‐mediated p38 MAPK and Akt activation [169,170], and increased ΔNp63 in mice with local or invasive skin cancer [169]. Moreover, ΔNp63 was the major vitamin D‐induced pathway in MCF10.DCIS basal mammary cells [171]. A role in osteoblast differentiation has also been reported, where 1α,25(OH) 2 D 3 increased ΔNp63, whilst 24R,25(OH) 2 D 3 increased TAp63 [172].…”

Vitamin D and Breast Cancer: Mechanistic Update

“…(31)(32)(33)(34)(35) However, the specific mechanisms and pathways that link the 1,25D-VDR complex to the observed biological effects remain elusive and appear to be highly cell type specific based on mechanistic studies and genomic profiling as described below. (36)(37)(38)(39)(40)(41)(42) It should be noted that the concentrations of 1,25D utilized in vitro (typically 100 nM) have been chosen to maximally activate VDR, but are well above the physiological levels associated with dietary intake or epidermal synthesis. Early studies of 1,25D treatment in animal models of breast cancer rarely achieved anticancer effects without induction of severe hypercalcemia, generating pharmaceutical interest in the development of synthetic vitamin D analogs with activity profiles that favored growth inhibition over calcemic effects.…”

“…More recently, synthetic VDR agonists have been shown to block mammosphere formation and stem cell markers in a model of earlystage basal-like breast cancer (MCF10DCIS.com cells) via downregulation of pathways (such as NOTCH and CD44) that are required for maintenance of breast cancer stem-like cells. (42,101,102) Importantly, treatment of tumor-bearing mice with VDR agonists led to reduction in CD44 signaling in vivo. (101) Inhibitory effects of 1,25D on stem cell populations have also been demonstrated in a WNT1-driven murine mammary tumor model.…”

New Roles for Vitamin D Superagonists: From COVID to Cancer

“…Vitamin D influences carcinogenesis via several mechanisms, including: (a) promotion of differentiation and inhibition of the epithelial mesenchymal transition, (b) regulation of cancer stem cells, (c) reprogramming of gene expression and induction of quiescence in cancer associated fibroblasts (CAFs), and (d) modulation of immune response ( 63 , 114 , 115 ). Observational studies suggest low vitamin D status is associated with an increased risk of cancer incidence and mortality ( 12 , 116 ).…”