The Prostate 59:243 ^251 (2004) Analysis of Vitamin D-Regulated Gene Expression in LNCaP Human Prostate Cancer Cells Using cDNA Microarrays Aruna V. Krishnan, 1 Rajesh Shinghal, 2 Nalini Raghavachari, 3 James D. Brooks, 2 Donna M. Peehl, 2 and David Feldman 1 * 1 Department of Medicine, Stanford University School of Medicine, Stanford,California 2 Department of Urology, Stanford University School of Medicine, Stanford,California 3 Biochemical Technologies,Corning, Inc., NewYork BACKGROUND. 1,25-dihydroxyvitamin D 3 [1,25(OH) 2 D 3 ] exerts growth inhibitory, pro- differentiating, and pro-apoptotic effects on prostate cells. To better understand the molecular mechanisms underlying these actions, we employed cDNA microarrays to study 1,25(OH) 2 D 3 - regulated gene expression in the LNCaP human prostate cancer cells. METHODS. mRNA isolated from LNCaP cells treated with vehicle or 50 nM 1,25(OH) 2 D 3 for various lengths of time were hybridized to microarrays carrying approximately 23,000 genes. Some of the putative target genes revealed by the microarray analysis were verified by real-time PCR assays. RESULTS. 1,25(OH) 2 D 3 most substantially increased the expression of the insulin-like growth factor binding protein-3 (IGFBP-3) gene. Our analysis also revealed several novel 1,25(OH) 2 D 3 - responsive genes. Interestingly, some of the key genes regulated by 1,25(OH) 2 D 3 are also androgen-responsive genes. 1,25(OH) 2 D 3 also down-regulated genes that mediate androgen catabolism. CONCLUSIONS. The putative 1,25(OH) 2 D 3 target genes appear to be involved in a variety of cellular functions including growth regulation, differentiation, membrane transport, cell– cell and cell–matrix interactions, DNA repair, and inhibition of metastasis. The up-regulation of IGFBP-3 gene has been shown to be crucial in 1,25(OH) 2 D 3 -mediated inhibition of LNCaP cell growth. 1,25(OH) 2 D 3 regulation of androgen-responsive genes as well as genes involved in androgen catabolism suggests that there are interactions between 1,25(OH) 2 D 3 and androgen signaling pathways in LNCaP cells. Further studies on the role of these genes and others in mediating the anti-cancer effects of 1,25(OH) 2 D 3 may lead to better approaches to the prevention and treatment of prostate cancer. Prostate 59: 243 – 251, 2004. # 2003 Wiley-Liss, Inc. KEY WORDS: 1,25-dihydroxyvitamin D 3 ; cDNA arrays; prostate cancer cells; growth inhibition; target genes; androgens; IGFBP-3 INTRODUCTION Prostate cancer is the most common non-cutaneous malignancy in men and is the second leading cause of cancer deaths in American males [1]. Prostate cancer growth is promoted by circulating androgens. The main treatment strategy for advanced prostate cancer in- volves androgen deprivation therapy to which patients initially respond very well. However, most patients eventually fail this therapy and develop androgen- independent prostate cancer and metastatic disease Grant sponsor: NIH (to D.F.); Grant numbers: DK42482, CA92238; Grant sponsor: DOD Prostate Cancer Research Program (to D.F.); Grant number: DAMD17-02-1-0142; Grant sponsor: DOD Prostate Cancer Research Program (to J.D.B.); Grant number: DAMD17-98-1- 8555; Grant sponsor: Doris Duke Clinician Scientist Award (to J.D.B.); Grant number: T98064 and CaPCure (to D.F. and D.M.P.). *Correspondence to: David Feldman, MD, Department of Medicine/ Endocrinology, Stanford University School of Medicine, Room S- 025, Stanford, CA 94305-5103. E-mail: feldman@cmgm.stanford.edu Received 22 May 2003; Accepted 6 October 2003 DOI 10.1002/pros.20006 Published online 22 December 2003 in Wiley InterScience (www.interscience.wiley.com). ß 2003 Wiley-Liss, Inc.