516 MICRORNA PROLING IN PATIENT-MATCHED FRESH-FROZEN AND FORMALIN-FIXED TISSUES AND CELL LINES

Abstract

You have accessJournal of UrologyKidney Cancer: Evaluation and Staging I1 Apr 2010516 MICRORNA PROLING IN PATIENT-MATCHED FRESH-FROZEN AND FORMALIN-FIXED TISSUES AND CELL LINES Van Le, David Juan, Derek Beaton, and Lious Liou Van LeVan Le Boston, MA More articles by this author , David JuanDavid Juan Boston, MA More articles by this author , Derek BeatonDerek Beaton Boston, MA More articles by this author , and Lious LiouLious Liou Cambridge, MA More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2010.02.592AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES MicroRNAs (miRNAs) are a class of endogenous gene regulators that play an important role in oncogenesis through the regulation of oncogenes and tumor suppressors. Profiling miRNA expressions may provide new tools for cancer diagnosis and therapy. The majority of the published clinical studies are performed with fresh frozen tissues. Using formalin-fixed, paraffin-embedded (FFPE) tissues are preferable because of their abundance and clinical correlated data are known. However, the challenge in using FFPE tissues for molecular profiling is the volatility of the nucleic acids, although this seems less crucial with miRNA. Given the promise of miRNA profiling in FFPE tissues and the potential alterations in the original molecular signature, we sought to investigate the effect of formalin fixation on miRNA expression in renal cell carcinoma and urothelial cell lines. METHODS Kidney tissues taken at the time of surgeries for each patient were made into FFPE blocks and flash frozen. RNA from FFPE and fresh-frozen tissues of six patients were extracted using RecoverAll Total Nucleic Acid Isolation Kit and mirVana miRNA Isolation Kit (Ambion, California), respectively. All samples were one to three years old. Extracted total RNA was then quantified with NanoDrop ND-1000 Spectrophotometer and reverse transcribed to cDNA using Quantimir. Real-time qRT-PCR was performed on 60 miRNAs of interest in triplicate using the Oncomir kit (Systems Bioscience, California). Htert cell line was grown with KSFM culture media and was fixed with RNALater, 75% ethanol, 10% formalin, and fresh frozen. Real-time qRT-PCR was performed on 384 miRs in triplicate. RESULTS There was a high correlation (Spearman r: 0.78-0.95, mean = 0.88, p-value < 0.01) between kidney patient-matched formalin-fixed and fresh-frozen tumor specimens. Formalin-fixed normal and corresponding frozen tissues did not correlate as well (Spearman r: 0.17-0.75, mean = 0.50, p-value< 0.01). Analysis also suggested that there was a high correlation between cell line samples fixed with 75% ethanol, 10% formalin, and fresh frozen (Spearman r: 0.91-0.94, mean = 0.92, p-value < 0.01). Formalin-fixed Htert had a mean Spearmen r value of 0.63 when compared with frozen and other fixations. CONCLUSIONS Formalin fixation did not significantly alter miRNA expression in kidney and cell line specimens. This suggests retrospective studies of cancers with FFPE blocks are valid. With available corresponding clinical data, miRNA profiling in FFPE tissues can provide a new means in diagnosing and treating cancers. © 2010 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 183Issue 4SApril 2010Page: e204 Advertisement Copyright & Permissions© 2010 by American Urological Association Education and Research, Inc.MetricsAuthor Information Van Le Boston, MA More articles by this author David Juan Boston, MA More articles by this author Derek Beaton Boston, MA More articles by this author Lious Liou Cambridge, MA More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...