AbstractInactivation of tumor suppressor genes through deletion, mutation and epigenetic silencing has been shown to occur in cancer. In our study, we combined DNA demethylation and histone deacetylation inhibition treatments with suppression subtraction hybridization (SSH) and cDNA microarrays to identify potentially epigenetically downregulated genes in PC‐3 prostate cancer cell line. We found 11 genes whose expression was upregulated after relieving epigenetic regulation. Expression of 3 genes [dual‐specificity phosphatase 1 (DUSP1), serum/glucocorticoid regulated kinase (SGK) and spermidine/spermine N1‐acetyltransferase (SAT)] was subsequently studied in clinical sample material using real‐time quantitative RT‐PCR and immunohistochemistry. TheDUSP1andSGKmRNA expression was lower in hormone‐refractory prostate carcinomas compared to benign prostate hyperplasia (BPH) or untreated prostate carcinomas. BPH, normal prostate and high‐grade prostate intraepithelial neoplasia (PIN) expressed high levels of DUSP1 and SGK proteins. Ninety‐two percent and 48% of the prostate carcinomas showed almost complete lack of DUSP1 and SGK proteins, respectively, indicating common downregulation of these genes. The genomic bisulphite sequencing did not reveal dense hypermethylation in the promoter regions of eitherDUSP1orSGK. In conclusion, the data suggest that downregulation ofDUSP1andSGKis an early event and could be important in the tumorigenesis of prostate cancer. © 2005 Wiley‐Liss, Inc.