INTRODUCTION: Diffusion-weighted magnetic resonance (MR) imaging (DWI) might be able to fulfill the need to accurately identify high-grade prostate carcinoma, in patients initially selected for active surveillance in the Prostate Specific Antigen (PSA) screening era based on transrectal ultrasound-guided biopsy Gleason score. We aimed to determine whether DWI is able to correctly identify those patients with a biopsy Gleason score of =3 + 3 = 6, but harboring Gleason 4 and/or 5 components in their radical prostatectomy (RP) specimen. MATERIALS AND METHODS: Whole-mount RP specimens were used to identify regions of interest corresponding with tumor on the DWI-derived apparent diffusion coefficient (ADC) maps in 23 patients with a Gleason =3 + 3 = 6 on biopsy. ADC values were correlated with RP Gleason grades. Statistical analysis was performed by calculating area under the receiver operating characteristic curve for identification of prostate cancer with Gleason 4 and/or 5 components using DWI, and Mann-Whitney U testing was performed to detect differences in median ADC values for tumors with presence of Gleason grade 4 and/or 5 versus a highest Gleason grade of =3 on RP. RESULTS: A diagnostic accuracy of median ADC values for identifying patients subject to transrectal ultrasound-guided biopsy undergrading with an area under the receiver operating characteristic curve of 0.88 was established using RP Gleason score as a reference. In patients harboring a Gleason 4 and/or 5 component, the median ADC was 0.86 � 10(-3) mm /s (standard deviation � 0.21), whereas patients harboring no Gleason 4 and/or 5 component displayed a median ADC of 1.16 � 10(-3) mm /s (standard deviation � 0.19) for the single tumor slice with the lowest median ADC (P < 0.002). CONCLUSIONS: DWI is able to predict the presence of high-grade tumor in patients with a Gleason =3 + 3 = 6 on biopsy, providing important information for treatment decisions.