cholecalciferol and Carcinoma--Basal-Cell

Vitamin D3 (VD3) is a seco-steroid that inhibits the Hedgehog (Hh) signaling pathway. Initial studies suggested its anti-Hh activity results from direct inhibition of Smoothened, a seven-transmembrane cell surface receptor that is a key regulator of the Hh signaling cascade. More recently, a role for the Vitamin D Receptor in mediating inhibition of Hh-signaling by seco-steroid has been suggested. Herein, an affinity-based protein profiling study was carried out to better understand the cellular proteins that govern VD3-mediated anti-Hh activity. We synthesized a novel biotinylated VD3 analogue (8) for use as a chemical probe to explore cellular binding targets of the seco-steroidal scaffold. Through a series of pull-down experiments and follow up mass spectrum analyses, heat shock protein 70 (Hsp70) was identified as a primary binding protein of VD3. Hsp70 was validated as a binding target of VD3 through a series of biochemical and cellular assays. VD3 bound with micromolar affinity to Hsp70. In addition, both selective knockdown of Hsp70 expression and pharmacological inhibition of its activity with known Hsp70 inhibitors suppressed Hh-signaling transduction in murine basal cell carcinoma cells, suggesting that Hsp70 regulates proper Hh-signaling. Additional cellular assays suggest that VD3 and its seco-steroidal metabolites inhibit Hh-signaling through different mechanisms.

Topics: Animals; Antineoplastic Agents; Carcinoma, Basal Cell; Cell Proliferation; Cells, Cultured; Cholecalciferol; Dose-Response Relationship, Drug; Drug Screening Assays, Antitumor; Hedgehog Proteins; HSP70 Heat-Shock Proteins; Mice; Molecular Structure; Signal Transduction; Structure-Activity Relationship

Hydroxyderivatives of vitamin D3, including classical 1,25(OH)

Topics: Animals; beta Catenin; Carcinoma, Basal Cell; Carcinoma, Squamous Cell; Cholecalciferol; Cholesterol Side-Chain Cleavage Enzyme; Humans; Low Density Lipoprotein Receptor-Related Protein-2; Mice; Receptors, Calcitriol; Skin Neoplasms; Vitamin D; Zinc Finger Protein GLI1

The effect of UVR on human basal cell carcinoma (BCC) epidemiology is complex-the incidence rises until approximately 30,000 hours of lifetime sunlight exposure and then plateaus. We hypothesize that UVR has opposing effects on BCC carcinogenesis-stimulatory via mutagenesis and inhibitory via production of hedgehog-inhibiting vitamin D

Topics: Administration, Topical; Animals; Carcinogenesis; Carcinoma, Basal Cell; Cell Proliferation; Cholecalciferol; Disease Progression; Female; Gene Deletion; Genotype; Keratinocytes; Male; Mice; Mice, Transgenic; Oxidoreductases Acting on CH-CH Group Donors; Radiation, Ionizing; Sex Factors; Skin; Skin Neoplasms; Ultraviolet Rays

UVR in sunlight causes mutations that drive basal cell carcinomas. However, the incidence of these tumors plateaus with prolonged exposure, but the incidence of other skin cancers increases. Makarova et al. now show that vitamin D

Topics: Animals; Carcinogenesis; Carcinoma, Basal Cell; Cholecalciferol; Mice; Skin; Skin Neoplasms; Sunlight

1α, 25-dihydroxyvitamin D3 (VD3), a secosteriod that has been explored as an anti-cancer agent, was also shown to promote cell survival. Its receptor, the Vitamin D Receptor (VDR), is a direct target of the proto-oncogene ΔNp63α, which is overexpressed in non-melanoma skin cancers. The interconnection between VDR/VD3 signaling and ΔNp63α, led us to examine whether VDR/VD3 signaling promotes keratinocyte proliferation by regulating ΔNp63α levels. Our data demonstrate that VDR regulates ΔNp63α expression at both the transcript and protein level. Interestingly, although low doses of VD3 led to an increase in ΔNp63α protein levels and keratinocyte proliferation, high doses of VD3 failed to increase ΔNp63α protein levels and resulted in reduced proliferation. Increased expression of ΔNp63α by low dose VD3 was shown to be dependent on VDR and critical for the proliferative effects of VD3. VD3-mediated increases in ΔNp63α protein levels occur via activation of both p38 MAPK and Akt kinases. Finally, analysis of samples from patients with squamous cell carcinoma (SCC), basal cell carcinoma and precursors to invasive SCC demonstrated a significant correlation between p63 and VDR levels when compared with healthy normal skin control samples. Delineation of the mechanisms by which VD3 exerts its effect on ΔNp63α and cell proliferation is critical for determining the future of VD3 in cancer therapies.

Topics: Carcinoma, Basal Cell; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Cell Survival; Cholecalciferol; Heterocyclic Compounds, 3-Ring; Humans; Imidazoles; Keratinocytes; Naphthalenes; p38 Mitogen-Activated Protein Kinases; Proto-Oncogene Mas; Proto-Oncogene Proteins c-akt; Pyrazoles; Pyridines; Receptors, Calcitriol; RNA Interference; RNA, Small Interfering; Signal Transduction; Skin Neoplasms; Transcription Factors; Tumor Suppressor Proteins

Constitutive Hedgehog (HH) signaling underlies several human tumors, including basal cell carcinoma (BCC). Recently, Bijlsma and colleagues reported a new biologic function for vitamin D3 in suppressing HH signaling in an in vitro model system. On the basis of that work, we have assessed effects of vitamin D3 on HH signaling and proliferation of murine BCCs in vitro and in vivo. We find that indeed in BCC cells, vitamin D3 blocks both proliferation and HH signaling as assessed by mRNA expression of the HH target gene Gli1. These effects of vitamin D3 on Gli1 expression and on BCC cell proliferation are comparable to the effects of cyclopamine, a known inhibitor of the HH pathway. These results are specific for vitamin D3, because the precursor 7-dehydrocholesterol and the downstream products 25-hydroxy vitamin D3 [25(OH)D] and 1,25-dihydroxy vitamin D3 [1,25(OH)(2)D] are considerably less effective in reducing either Gli1 mRNA or cellular proliferation. Moreover, these effects seem to be independent of the vitamin D receptor (VDR) because short hairpin RNA knockdown of VDR does not abrogate the anti-HH effects of D3 despite reducing expression of the VDR target gene 24-hydroxylase. Finally, topical vitamin D3 treatment of existing murine BCC tumors significantly decreases Gli1 and Ki67 staining. Thus, topical vitamin D3 acting via its HH inhibiting effect may hold promise as an effective anti-BCC agent.

Topics: Animals; Blotting, Western; Bone Density Conservation Agents; Carcinoma, Basal Cell; Cell Differentiation; Cell Proliferation; Cells, Cultured; Cerebellar Neoplasms; Cholecalciferol; Hedgehog Proteins; Immunoenzyme Techniques; Keratinocytes; Kruppel-Like Transcription Factors; Medulloblastoma; Mice; Receptors, Calcitriol; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; RNA, Small Interfering; Steroid Hydroxylases; Vitamin D3 24-Hydroxylase; Zinc Finger Protein GLI1