Liarozole acts synergistically with 1alpha,25-dihydroxyvitamin D3 to inhibit growth of DU 145 human prostate cancer cells by blocking 24-hydroxylase activity.
1Alpha,25-dihydroxyvitamin D3 [1,25-(OH)2D3] inhibits the proliferation of many cancer cells in culture, but not the aggressive human prostate cancer cell line DU 145. We postulated that the 1,25-(OH)2D3-resistant phenotype in DU 145 cells might result from the high levels of expression of 25-hydroxyvitamin D-24-hydroxylase (24-hydroxylase) induced by treatment with 1,25-(OH)2D3. As this P450 enzyme initiates 1,25-(OH)2D3 catabolism, we presumed that a high level of enzyme induction could limit the effectiveness of the 1,25-(OH)2D3 antiproliferative action. To examine this hypothesis we explored combination therapy with liarozole fumarate (R85,246), an imidazole derivative currently in trials for prostate cancer therapy. As imidizole derivatives are known to inhibit P450 enzymes, we postulated that this drug would inhibit 24-hydroxylase activity, increasing the 1,25-(OH)2D3 half-life, thereby enhancing 1,25-(OH)2D3 antiproliferative effects on DU 145 cells. Cell growth was assessed by measurement of viable cells using the MTS assay. When used alone, neither 1,25-(OH)2D3 (1-10 nM) nor liarozole (1-10 microM) inhibited DU 145 cell growth. However, when added together, 1,25-(OH)2D3 (10 nM)/liarozole (1 microM) inhibited growth 65% after 4 days of culture. We used a TLC method to assess 24-hydroxylase activity and demonstrated that liarozole (1-100 microM) inhibited this P450 enzyme in a dose-dependent manner. Moreover, liarozole treatment caused a significant increase in 1,25-(OH)2D3 half-life from 11 to 31 h. In addition, 1,25-(OH)2D3 can cause homologous up-regulation of the vitamin D receptor (VDR), and in the presence of liarozole, this effect was amplified, thus enhancing 1,25-(OH)2D3 activity. Western blot analyses demonstrated that DU 145 cells treated with 1,25-(OH)2D3/liarozole showed greater VDR up-regulation than cells treated with either drug alone. In summary, our data demonstrate that liarozole augments the ability of 1,25-(OH)2D3 to inhibit DU 145 cell growth. The mechanism appears to be due to inhibition of 24-hydroxylase activity, leading to increased 1,25-(OH)2D3 half-life and augmentation of homologous up-regulation of VDR. We raise the possibility that combination therapy using 1,25-(OH)2D3 and liarozole or other inhibitors of 24-hydroxylase, both in nontoxic doses, might serve as an effective treatment for prostate cancer.
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Department of Medicine, Stanford University School of Medicine, California 94305-5103, USA.
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Holloway
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Feldman
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D015090:25-Hydroxyvitamin D3 1-alpha-Hydroxylase / Q000037:antagonists & inhibitors*; D002117:Calcitriol / Q000097:blood / Q000494:pharmacology*; D002455:Cell Division / Q000187:drug effects*; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006207:Half-Life; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology*; D008297:Male; D011471:Prostatic Neoplasms / Q000473:pathology*; D018167:Receptors, Calcitriol / Q000378:metabolism; D014407:Tumor Cells, Cultured
D004791:Enzyme Inhibitors; D007093:Imidazoles; D018167:Receptors, Calcitriol; D015090:25-Hydroxyvitamin D3 1-alpha-Hydroxylase; D002117:Calcitriol; C061121:liarozole