The main challenge for clinical management of prostate cancer is to distinguish tumors that will progress faster and will show a higher tendency to recur from the more indolent ones. We have compared expression profiles of 18 prostate cancer samples (seven with a Gleason score of 6, eight with a Gleason score of 7, and three with a Gleason score of <8) and five nonneoplastic prostate samples, using the Affymetrix Human Array GeneChip Exon 1.0 ST. Microarray analysis revealed 99 genes showing statistically significant differences among tumors with Gleason scores of 6, 7, and <8. In addition, mRNA expression of 29 selected genes was analyzed by real-time quantitative RT-PCR with microfluidic cards in an extended series of 30 prostate tumors. Of the 29 genes, 18 (62%) were independently confirmed in the extended series by quantitative RT-PCR: 14 were up-regulated and 4 were down-regulated in tumors with a higher Gleason score. Twelve of these genes were differentially expressed in tumors with a Gleason score of 6 to 7 versus <8. Finally, IHC validation of the protein levels of two genes from the 12-gene signature (SEC14L1 and TCEB1) showed strong protein expression levels of both genes, which were statistically associated with a high combined Gleason score, advanced stage, and prostate-specific antigen progression. This set of genes may contribute to a better understanding of the molecular basis of prostate cancer. TCEB1 and SELC14L1 are good candidate markers for predicting prognosis and progression of prostate cancer. © 2012 American Society for Investigative Pathology.