Prostate cancer (PC) is the most common and the second leading cause of cancer-related death among American men. Early diagnosis is a prerequisite to improving therapeutic benefits. However, the current clinical biomarkers for PC do not reliably decipher indolent PC from other urogenital disorders. Thus, effective clinical intervention necessitates development of new biomarkers for early detection of PC. The present study aimed to identify the miRNA signature in organ-confined (Gleason Score 6) prostate tumors. MicroRNA (miRNA/miR) array analysis identified 118 upregulated and 73 downregulated miRNAs in microdissected tumors in comparison to matched neighboring normal prostate epithelium. The miRs-Plus-A1083, -92b-5p, -18a-3p, -19a-3p, -639, -3622b-3p, -3189-3p, -155-3p, -410, -1179, 548b-5p, and -4469 are predominantly expressed (7-11-fold), whereas miRs-595, 4490, -3120-5p, -1299, -21-5p, -3677-3, -let-7b-5p, -5189, 3-121-5p, -4518, -200a-5p, -3682-5p, -3689d, -3149 represent the most downregulated (12-113-fold) miRNAs in microdissected prostate tumors. The array expression profile of selected miRNA signature and their potential mRNA targets was validated by qRT-PCR analysis in PC cell lines. Integrated in silico and computational prediction analyses demonstrated that the dysregulated miRNA signature map to key regulatory factors involved in tumorigenesis, including cell cycle, apoptosis, and p53 pathways. The newly identified miRNA signature has potential clinical utility as biomarkers, prognostic indicators, and therapeutic targets for early detection of PC. Further studies are needed to assess the functional significance and clinical usefulness of the identified miRNAs. Impact Statement To our knowledge his is the first study of identifying miRNA signatures in microdissected indolent (Gleason score 6) prostate cancer in comparison to matched normal prostate epithelium. By employing in silico and computational prediction analysis, the study provides a landscape of potential miRNA targets and key cellular pathways involved in prostate tumorigenesis. Identification if miRNAs and their relevant targets and pathways pave the way for underpinning their mechanistic role of miRNAs in human prostate tumorigenesis, and possibly other human cancers. Importantly, the outcome of the study has important clinical implications for the management of prostate cancer, including the use of miRNA(s) as biomarkers for early detection of prostate cancer.