PURPOSE: We analyzed the expression of genes to identify reliable molecular markers in the diagnosis and progression of prostate cancer. EXPERIMENTAL DESIGN: Gene expression profiling was done using HG-U133 set microarrays in 32 prostate cancer and 8 benign tissues of patients with cancer. Expression levels of 11 genes were selected for quantitative real-time PCR evaluation in 52 prostate cancer and 20 benign tissues. Further, to assess transcriptional inactivation, we analyzed the promoter methylation of genes by quantitative methylation-specific PCR in 62 tumor and 36 benign tissues. RESULTS: Our results showed a significant down-regulation in the mRNA expression levels of PRIMA1, TU3A, PDLIM4, FLJ14084, SVIL, SORBS1, C21orf63, and KIAA1210 and up-regulation of FABP5, SOX4, and MLP in prostate cancer tissues by TaqMan real-time PCR. Quantitative methylation-specific PCR of PDLIM4, SVIL, PRIMA1, GSTP1, and PTGS2 detected prostate carcinoma with a sensitivity of 94.7%, 75.4%, 47.4%, 89.5%, and 87.7%, and a specificity of 90.5%, 75%, 54.2%, 95.8%, and 90.2%, respectively. Using this panel of methylation markers in combination, we were able to distinguish between prostate cancer and adjacent benign tissues with sensitivities and specificities of about 90% to 100%. Our data provide evidence of transcriptional repression of the putative tumor suppressor gene PDLIM4 by hypermethylation. CONCLUSIONS: Our analysis revealed differential expression of eight down-regulated and three up-regulated genes, implicating their role in prostate cancer development and progression. We further showed that the hypermethylation of PDLIM4 gene could be used as a sensitive molecular tool in detection of prostate tumorigenesis.