title: miRNA Expression Analyses in Prostate Cancer Clinical Tissues.
abstract: A critical challenge in prostate cancer (PCa) clinical management is posed by the inadequacy of currently used biomarkers for disease screening, diagnosis, prognosis and treatment. In recent years, microRNAs (miRNAs) have emerged as promising alternate biomarkers for prostate cancer diagnosis and prognosis. However, the development of miRNAs as effective biomarkers for prostate cancer heavily relies on their accurate detection in clinical tissues. miRNA analyses in prostate cancer clinical specimens is often challenging owing to tumor heterogeneity, sampling errors, stromal contamination etc. The goal of this article is to describe a simplified workflow for miRNA analyses in archived FFPE or fresh frozen prostate cancer clinical specimens using a combination of quantitative real-time PCR (RT-PCR) and in situ hybridization (ISH). Within this workflow, we optimize the existing methodologies for miRNA extraction from FFPE and frozen prostate tissues and expression analyses by Taqman-probe based miRNA RT-PCR. In addition, we describe an optimized method for ISH analyses formiRNA detection in prostate tissues using locked nucleic acid (LNA)- based probes. Our optimized miRNA ISH protocol can be applied to prostate cancer tissue slides or prostate cancer tissue microarrays (TMA).
authors.affiliation: Department of Urology, Veterans Affairs Medical Center, San Francisco, University of California San Francisco.
authors.forename: Nathan
authors.initials: N
authors.lastname: Bucay
authors.affiliation: Department of Urology, Veterans Affairs Medical Center, San Francisco, University of California San Francisco.
authors.forename: Varahram
authors.initials: V
authors.lastname: Shahryari
authors.affiliation: Department of Urology, Veterans Affairs Medical Center, San Francisco, University of California San Francisco.
authors.forename: Shahana
authors.initials: S
authors.lastname: Majid
authors.affiliation: Department of Urology, Veterans Affairs Medical Center, San Francisco, University of California San Francisco.
authors.forename: Soichiro
authors.initials: S
authors.lastname: Yamamura
authors.affiliation: Department of Urology, Veterans Affairs Medical Center, San Francisco, University of California San Francisco.
authors.forename: Yozo
authors.initials: Y
authors.lastname: Mitsui
authors.affiliation: Department of Urology, Veterans Affairs Medical Center, San Francisco, University of California San Francisco.
authors.forename: Z Laura
authors.initials: ZL
authors.lastname: Tabatabai
authors.affiliation: Department of Urology, Veterans Affairs Medical Center, San Francisco, University of California San Francisco.
authors.forename: Kirsten
authors.initials: K
authors.lastname: Greene
authors.affiliation: Department of Urology, Veterans Affairs Medical Center, San Francisco, University of California San Francisco.
authors.forename: Guoren
authors.initials: G
authors.lastname: Deng
authors.affiliation: Department of Urology, Veterans Affairs Medical Center, San Francisco, University of California San Francisco.
authors.forename: Rajvir
authors.initials: R
authors.lastname: Dahiya
authors.affiliation: Department of Urology, Veterans Affairs Medical Center, San Francisco, University of California San Francisco.
authors.forename: Yuichiro
authors.initials: Y
authors.lastname: Tanaka
authors.affiliation: Department of Urology, Veterans Affairs Medical Center, San Francisco, University of California San Francisco; SainiS@urology.ucsf.edu.
authors.forename: Sharanjot
authors.initials: S
authors.lastname: Saini
mesh_terms: D006801:Humans; D017403:In Situ Hybridization / Q000379:methods; D008297:Male; D035683:MicroRNAs / Q000096:biosynthesis* / Q000235:genetics; D009841:Oligonucleotides / Q000737:chemistry / Q000235:genetics; D016612:Paraffin Embedding; D011379:Prognosis; D011471:Prostatic Neoplasms / Q000235:genetics* / Q000378:metabolism; D015347:RNA Probes / Q000737:chemistry / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods; D016707:Tissue Fixation
chemical_list: D035683:MicroRNAs; D009841:Oligonucleotides; D015347:RNA Probes; C477371:locked nucleic acid