Stable expression of human VDR in murine VDR-null cells recapitulates vitamin D mediated anti-cancer signaling

Abstract

Mammary tumor cells derived from vitamin D receptor (VDR) knock-out (KO) mice were engineered to stably express wild-type (WT) or mutated VDR for characterization of the mechanisms by which 1,25-dihydroxyvitamin D (1,25D), the VDR ligand, mediates growth regulation. Although KO cells were completely resistant to 1,25D, introduction of WT human VDR restored gene expression and growth inhibition in response to 1,25D and a variety of structural analogs. Pdgfb, Vegfa, and Nfkbi were identified as genomic targets of both human and murine VDR signaling in this cell model. KO cells expressing hVDRs containing point mutations (W286R, R274L) that reduce or abolish ligand binding did not exhibit changes in gene expression or growth in response to physiological doses of 1,25D but did respond to higher doses and more potent analogs. KO cells expressing hVDR with the G46D point mutation, which abrogates VDR binding to DR3 response elements, exhibited partial growth inhibition in response to 1,25D and synthetic vitamin D analogs, providing proof of principle that VDR signaling through alternative genomic or non-genomic mechanisms contributes to vitamin D mediated growth effects in transformed cells. We conclude that the 1,25D-VDR signaling axis that triggers anti-cancer effects is highly conserved between the murine and human systems despite differences in VDR protein, cofactors, and target genes and that these actions are not solely mediated via canonical VDRE signaling.

Modeling vitamin D actions in triple negative/basal-like breast cancer

“…For the studies described here, WT145 and KO240 cells were adapted to growth in DMEM/F12 medium supplemented with 5% charcoal-stripped FBS to eliminate exposure to serum vitamin D metabolites. In previous work [14] we confirmed that stable expression of hVDR into KO240 cells using a plasmid vector (pSG5-hVDR-hygro) recapitulated 1,25D mediated anti-cancer signaling, however, sensitivity to 1,25D was lost over time in these clones. To overcome this problem, we created a retroviral hVDR vector (pBABE-puro-hVDR, Fig 1) and generated a new series of hVDR expressing cells which we characterized in the present studies.…”

“…Recently we provided proof of principle that expression of VDR in KO240 cells using a plasmid vector recapitulated 1,25D mediated anti-cancer signaling [14] however, these VDR expressing cells lost sensitivity over time. For these follow-up studies, we created a retroviral hVDR expression vector (pBABE-hVDR-Puro, Fig 1A) and selected multiple KO240 clonal cell lines with stable expression of hVDR protein.…”

Genomic instability in human cancer: Molecular insights and opportunities for therapeutic attack and prevention through diet and nutrition

“…There are no known interactions for the other compounds, except selenium, which inhibits growth in some cases while inducing it in others [356] , [357] . In a related characteristic of cancer cells, evasion of antigrowth signaling, all compounds are able to inhibit growth except vitamin B, which shows no relationship, and carotenoids, which have mixed results [255] , [358] , [359] , [360] , [361] , [362] , [363] , [364] , [365] , [366] . Similarly, all of the compounds are able to prevent replicative immortality by impairing telomerase activity or inducing senescence [364] , [367] , [368] , [369] , [370] , [371] , [372] , [373] , [374] , [375] , [376] and increasing cell death except for vitamin B, which has no known relationship to apoptosis [377] , [378] , [379] , [380] , [381] , [382] .…”

Vitamin D Receptor Signaling and Pancreatic Cancer Cell EMT

“…Conversely, downregulation of VDR expression in ALVA-31 prostate cancer cells attenuated the ability of 1,25D to inhibit cell growth (47). Keith et al (48) argued that the percentage of VDR-positive cells rather than the absolute level of cellular VDR expression in a heterogenous tumor would be the best predictor of growth inhibition induced by vitamin D compounds. Overall, these studies demonstrated that VDR is required for the antiproliferative effect of 1,25D in cancer cells.…”