title: Colorimetric TMPRSS2-ERG Gene Fusion Detection in Prostate Cancer Urinary Samples via Recombinase Polymerase Amplification.
abstract: TMPRSS2 (Exon 1)-ERG (Exon 4) is the most frequent gene fusion event in prostate cancer (PC), and is highly PC-specific unlike the current serum prostate specific antigen (PSA) biomarker. However, TMPRSS2-ERG levels are currently measured with quantitative reverse-transcription PCR (RT-qPCR) which is time-consuming and requires costly equipment, thus limiting its use in clinical diagnostics. Herein, we report a novel rapid, cost-efficient and minimal-equipment assay named "FusBLU" for detecting TMPRSS2-ERG gene fusions from urine. TMPRSS2-ERG mRNA was amplified by isothermal reverse transcription-recombinase polymerase amplification (RT-RPA), magnetically-isolated, and detected through horseradish peroxidase (HRP)-catalyzed colorimetric reaction. FusBLU was specific for TMPRSS2-ERG mRNA with a low visual detection limit of 10(5) copies. We also demonstrated assay readout versatility on 3 potentially useful platforms. The colorimetric readout was detectable by naked eye for a quick yes/no evaluation of gene fusion presence. On the other hand, a more quantitative TMPRSS2-ERG detection was achievable by absorbance/electrochemical measurements. FusBLU was successfully applied to 12 urinary samples and results were validated by gold-standard RT-qPCR. We also showed that sediment RNA was likely the main source of TMPRSS2-ERG mRNA in urinary samples. We believe that our assay is a potential clinical screening tool for PC and could also have wide applications for other disease-related fusion genes.
authors.affiliation: 1. Centre for Personalized Nanomedicine, Australian Institute for Bioengineering and Nanotechnology (AIBN), The University of Queensland, QLD 4072, Australia;
authors.forename: Kevin M
authors.initials: KM
authors.lastname: Koo
authors.affiliation: 1. Centre for Personalized Nanomedicine, Australian Institute for Bioengineering and Nanotechnology (AIBN), The University of Queensland, QLD 4072, Australia;
authors.forename: Eugene J H
authors.initials: EJ
authors.lastname: Wee
authors.affiliation: 1. Centre for Personalized Nanomedicine, Australian Institute for Bioengineering and Nanotechnology (AIBN), The University of Queensland, QLD 4072, Australia;; 2. School of Chemistry and Molecular Biosciences, The University of Queensland, QLD 4072, Australia.
authors.forename: Matt
authors.initials: M
authors.lastname: Trau
mesh_terms: D003124:Colorimetry / Q000379:methods*; D006801:Humans; D008297:Male; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D011471:Prostatic Neoplasms / Q000175:diagnosis* / Q000473:pathology; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics / Q000652:urine*; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000235:genetics*; D045522:Recombinases / Q000378:metabolism; D012697:Serine Endopeptidases / Q000032:analysis / Q000235:genetics*; D000071230:Transcriptional Regulator ERG / Q000032:analysis / Q000235:genetics
chemical_list: C072312:ERG protein, human; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D045522:Recombinases; D000071230:Transcriptional Regulator ERG; D012697:Serine Endopeptidases; C421305:TMPRSS2 protein, human
keywords: FusBLU; TMPRSS2-ERG