Updated project metadata. Nonaggressive prostate cancer (NAG-PCa) with Gleason score of 6 is considered as a low-risk disease and does not require clinical interventions. However, current assessment of aggressiveness of PCa is invasive using needle biopsies. Urine is an appealing biospecimen for noninvasive detection of aggressive PCa. Using urine, particularly from easily obtainable from pre-DRE urine specimens, to identify AG-PCa-associated molecular markers is critical for clinical risk stratification. Herein, we acquired quantitative mass spectrometry data for glycoproteins from pre-DRE and post-DRE urine specimens from patients underwent PCa diagnosis with biopsies. Compared with urinary glycoproteins identified from post-DRE urine samples, we confirmed that three previously reported AG-PCa-associated glycoproteins identified in post-DRE urine specimens were also found to be significantly associated with AG disease in pre-DRE urine samples. In addition, new AG-PCa-associated glycoproteins were identified in pre-DRE urine specimens. Our study provides a foundation for further studies of AG-PCa biomarkers using pre-DRE urine specimens.