The calcitropic hormone 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] also regulates cellular growth and differentiation, and so it has been considered as a treatment for malignancy and psoriasis. Several analogs of 1,25(OH)2D3 are up to 10,000 times more potent that 1,25(OH)2D3 as growth-inhibitory and differentiating agents, but their enhanced activity is not correlated with greater affinity to the nuclear vitamin D receptor (VDR) or with enhanced calcium mobilization. Instead, these activities are correlated with enhanced transcriptional activation of VDR. The objective of this study is to dissect the mechanisms that lead to maximal activation of the VDR by these analogs so that more effective, clinically applicable compounds can be developed. The hypothesis is that the mode of ligand interaction with the VDR can be changed by chemical and stereochemical modifications of 1,25(OH)2D3; these structural changes in the ligand also modulate the levels and the spectrum of VDR-mediated transcriptional activities by changing dimerization and DNA-binding preferences of VDR/ligand complexes. To test the hypothesis the 1,25(OH)2D3-binding site will be mapped by site-directed mutagenesis and its contact points with the VDR will be confirmed by covalent labeling of the ligand-binding site with a 1,25(OH)2D3 derivative that can be photoactivated. The binding activity of 1,25(OH)2D3 to wild-type and mutated VDR will be compared with that of the analogs, so that the effect of chemical and stereochemical modifications in the ligand on its binding requirements can be determined. To determine whether or not analogs can modify VDR interaction with nuclear receptors their effect on dimerization preferences of VDR will be examined in vitro. Bacterially-expressed VDR or RXR will be used to capture ligand-activated VDR complexes, and the latter will be quantified. The effect of 1,25(OH)2D3 and analogs on dimerization interfaces will be examined by mutation analysis of the ligand-dependent dimerization domain. To study the effect of differential ligand interaction on the type of response element used for VDR action, cellular or synthetic VDR-ligand complexes will be incubated with random oligonucleotides. The specific DNA sequences that bind will be extracted from the receptor/DNA complexes, amplified by the polymerase chain reaction, sequenced and tested for transcriptional activity by DNA-transfer methods.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK050583-03
Application #
2905798
Study Section
Special Emphasis Panel (ZRG4-GMB (04))
Program Officer
Margolis, Ronald N
Project Start
1997-05-01
Project End
2000-04-30
Budget Start
1999-05-01
Budget End
2000-04-30
Support Year
3
Fiscal Year
1999
Total Cost
Indirect Cost
Name
University of Texas MD Anderson Cancer Center
Department
Internal Medicine/Medicine
Type
Other Domestic Higher Education
DUNS #
001910777
City
Houston
State
TX
Country
United States
Zip Code
77030
Peleg, Sara; Nguyen, Cuong V (2010) The importance of nuclear import in protection of the vitamin D receptor from polyubiquitination and proteasome-mediated degradation. J Cell Biochem 110:926-34
Klopot, Anna; Hance, Kenneth W; Peleg, Sara et al. (2007) Nucleo-cytoplasmic cycling of the vitamin D receptor in the enterocyte-like cell line, Caco-2. J Cell Biochem 100:617-28
Marks, Hilary D; Fleet, James C; Peleg, Sara (2007) Transgenic expression of the human Vitamin D receptor (hVDR) in the duodenum of VDR-null mice attenuates the age-dependent decline in calcium absorption. J Steroid Biochem Mol Biol 103:513-6
Peleg, Sara; Khan, Farhana; Navone, Nora M et al. (2005) Inhibition of prostate cancer-meditated osteoblastic bone lesions by the low-calcemic analog 1alpha-hydroxymethyl-16-ene-26,27-bishomo-25-hydroxy vitamin D3. J Steroid Biochem Mol Biol 97:203-11
Phokela, Sarabjit S; Peleg, Sara; Moya, Fernando R et al. (2005) Regulation of human pulmonary surfactant protein gene expression by 1alpha,25-dihydroxyvitamin D3. Am J Physiol Lung Cell Mol Physiol 289:L617-26
Fizazi, Karim; Sikes, Charles R; Kim, Jeri et al. (2004) High efficacy of docetaxel with and without androgen deprivation and estramustine in preclinical models of advanced prostate cancer. Anticancer Res 24:2897-903
Kahraman, Mehmet; Sinishtaj, Sandra; Dolan, Patrick M et al. (2004) Potent, selective and low-calcemic inhibitors of CYP24 hydroxylase: 24-sulfoximine analogues of the hormone 1alpha,25-dihydroxyvitamin D(3). J Med Chem 47:6854-63
Posner, Gary H; Crawford, Kenneth R; Yang, Hong Woon et al. (2004) Potent, low-calcemic, selective inhibitors of CYP24 hydroxylase: 24-sulfone analogs of the hormone 1alpha,25-dihydroxyvitamin D3. J Steroid Biochem Mol Biol 89-90:5-12
Ismail, Ayesha; Nguyen, Cuong V; Ahene, Ago et al. (2004) Effect of cellular environment on the selective activation of the vitamin D receptor by 1alpha,25-dihydroxyvitamin D3 and its analog 1alpha-fluoro-16-ene-20-epi-23-ene-26,27-bishomo-25-hydroxyvitamin D3 (Ro-26-9228). Mol Endocrinol 18:874-87
Swamy, Narasimha; Chen, Tai C; Peleg, Sara et al. (2004) Inhibition of proliferation and induction of apoptosis by 25-hydroxyvitamin D3-3beta-(2)-Bromoacetate, a nontoxic and vitamin D receptor-alkylating analog of 25-hydroxyvitamin D3 in prostate cancer cells. Clin Cancer Res 10:8018-27

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