MP33-15 COMPARISON OF SERUM miR-371a-3p ASSAY PERFORMANCE BY DIGITAL DROPLET PCR AND REVERSE TRANSCRIPTASE QUANTITATIVE PCR IN MALIGNANT GERM CELL TUMOR PATIENTS

Abstract

You have accessJournal of UrologyCME1 Apr 2023MP33-15 COMPARISON OF SERUM miR-371a-3p ASSAY PERFORMANCE BY DIGITAL DROPLET PCR AND REVERSE TRANSCRIPTASE QUANTITATIVE PCR IN MALIGNANT GERM CELL TUMOR PATIENTS Michelle Nuño, John Lafin, Cinzia Scarpini, Anna Savelyeva, Zhaohui Wang, Jeffrey Gagan, Liwei Jia, Cheryl Lewis, Sarah Murray, Yun Cheng, Vitaly Margulis, Solomon Woldu, Douglas Strand, Nicholas Coleman, James Amatruda, Lindsay Frazier, Matthew Murray, and Aditya Bagrodia Michelle NuñoMichelle Nuño More articles by this author , John LafinJohn Lafin More articles by this author , Cinzia ScarpiniCinzia Scarpini More articles by this author , Anna SavelyevaAnna Savelyeva More articles by this author , Zhaohui WangZhaohui Wang More articles by this author , Jeffrey GaganJeffrey Gagan More articles by this author , Liwei JiaLiwei Jia More articles by this author , Cheryl LewisCheryl Lewis More articles by this author , Sarah MurraySarah Murray More articles by this author , Yun ChengYun Cheng More articles by this author , Vitaly MargulisVitaly Margulis More articles by this author , Solomon WolduSolomon Woldu More articles by this author , Douglas StrandDouglas Strand More articles by this author , Nicholas ColemanNicholas Coleman More articles by this author , James AmatrudaJames Amatruda More articles by this author , Lindsay FrazierLindsay Frazier More articles by this author , Matthew MurrayMatthew Murray More articles by this author , and Aditya BagrodiaAditya Bagrodia More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003266.15AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: MicroRNAs have shown promise as potential germ cell tumor (GCT) biomarkers. In particular, previous work has shown miR-371a-3p alone may serve as a biomarker for GCTs, demonstrating higher sensitivity than current biomarkers. Quantitative reverse transcriptase PCR (RT-qPCR) has been commonly used to measure circulating levels of miR-371a-3p. More recently, miR-371a-3p has also been measured using digital droplet PCR (ddPCR), which has the potential advantage of absolute quantification. We compared the performance of miR-371a-3p as quantified by RT-qPCR and ddPCR. METHODS: Patient samples from the University of Texas Southwestern (UTSW) Medical Center and the University of Cambridge (UoC) were evaluated using both RT-qPCR and ddPCR, as per current protocols (RT-qPCR) or as per standard manufacturer’s recommendations (ddPCR). A range of clinical scenarios (pre-orchiectomy, chemotherapy-naïve RPLND) were intentionally selected. We compared the performance of the two assays using receiver operating characteristic curves and area under the curve values. We also determined an optimal threshold for each procedure by maximizing the Youden Index and compared the corresponding estimated sensitivity and specificity. RESULTS: Data were available for 69 patients. Among these patients, 35 (50.7%) had malignant GCT (MGCT) and 34 (49.3%) had either non-MGCT (n=26) or no tumor (n=8). Patients with non-MCT or with no tumor were considered controls. Cq values were generally lower among patients with MGCT and the number of positive droplets was higher compared with controls. The AUC was 0.96 when using RT-qPCR and 0.82 when using ddPCR to classify patients based on circulating miR-371a-3p. The optimal threshold for ddPCR was determined to be 17.5 positive droplets with a corresponding estimated sensitivity of 71% and 100% specificity. For RT-qPCR, the optimal threshold was determined to be Cq=28.52, with a corresponding estimated sensitivity of 83% and specificity of 100%. CONCLUSIONS: RT-qPCR was more sensitive in identifying MGCT patients in our current cohort when compared with ddPCR. Further investigations are required to optimize PCR methodology, particularly for ddPCR, and determine whether tumor volume or clinical context affects ddPCR performance. Source of Funding: Cancer Prevention and Research Institute of Texas and St. Baldrick's Foundation. © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e456 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Michelle Nuño More articles by this author John Lafin More articles by this author Cinzia Scarpini More articles by this author Anna Savelyeva More articles by this author Zhaohui Wang More articles by this author Jeffrey Gagan More articles by this author Liwei Jia More articles by this author Cheryl Lewis More articles by this author Sarah Murray More articles by this author Yun Cheng More articles by this author Vitaly Margulis More articles by this author Solomon Woldu More articles by this author Douglas Strand More articles by this author Nicholas Coleman More articles by this author James Amatruda More articles by this author Lindsay Frazier More articles by this author Matthew Murray More articles by this author Aditya Bagrodia More articles by this author Expand All Advertisement PDF downloadLoading ...