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• W4385764938 abstract "Abstract Introduction: Molecular subtyping is integral in the classification of gliomas and increasingly guides the oncologic and neurosurgical management of these tumors. In high-grade gliomas, the presence or absence of the isocitrate dehydrogenase-1 (IDH1) mutation has major prognostic and diagnostic implications for surgical and therapeutic decision-making. The wait time for molecular studies typically ranges from 1-3 weeks and is not currently available as an intraoperative assay. We aimed to assess a novel assay for IDH1 status detection to aid with rapid intraoperative decision-making for glioma management. Methods: A novel IDH1 detection system based on loop-mediated isothermal amplification (LAMP) was developed. A rapid Perioperative-IDH1 Loop-mediated isothermal (Perioperative IDH1-LAMP) assay utilizing fluorescent and colorimetric measurements refined for in vitro use. This assay was first calibrated in vitro on validated glioblastoma (U87, IDH1 WT/WT) and grade 3 oligodendroglioma (BT-142, IDH1 mutant/-) cell lines in replicate. The assay was subsequently used to prospectively evaluate five surgically resected, patient-derived human glioma tumor samples tested directly from the operating room at the USC Keck Hospital. The assay evaluation was performed in a blinded fashion and compared to results from CLIA-certified diagnostic IDH1 assays, immunohistochemistry studies, and Sanger sequencing data. Results: The perioperative IDH1-LAMP assay successfully detected IDH1 mutation in the BT-142 cell line with a minimum DNA concentration of 0.28 ng/μl with 100% likelihood, and remained negative when testing the U87 WT-IDH1 cell line in vitro . 25 ng DNA extracted from BT-142 cell line was amplified and reached the threshold within 29 minutes. Also results showed that 0.255 ng of mutant-IDH1-R132 DNA in WT-IDH1 DNA background were required to correctly indicate IDH1 status via the naked eye and fluorescent signaling within 35 minutes, while 25.5 ng of mutant-IDH1-R132 DNA in WT-IDH1 DNA background determined a positive IDH1 status via monitoring qPCR amplification curves that reached threshold within 30.1 minutes. Additionally, results showed that 1mm 3 of tissue was required to avoid false positive results. In fresh patient-derived glioma samples, the assay correctly detected the IDH1 mutation in one of five samples, showing 100% specificity and sensitivity and full concordance with CLIA-certified molecular testing. The time for intraoperative detection of IDH1 status was 35 minutes. Conclusion: A novel perioperative IDH1-LAMP assay can rapidly indicate IDH1 mutation status through changes in pH and fluorescence detection, and the suppressive probe inhibited the amplification of WT-IDH1 samples from glioblastoma. Our assay matched the results of clinical molecular testing in all prospective, blinded cases. The translational impact of this assay includes rapid notification of IDH1 status, which will increasingly guide intraoperative surgical goals and perioperative clinical decision making for existing and emerging therapies for glioma." @default.
• W4385764938 created "2023-08-12" @default.
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• W4385764938 date "2023-08-11" @default.
• W4385764938 modified "2023-09-27" @default.
• W4385764938 title "Rapid Perioperative IDH1 Mutation Detection in High-Grade Gliomas using Novel LAMP assay" @default.
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• W4385764938 doi "https://doi.org/10.21203/rs.3.rs-3207603/v1" @default.
• W4385764938 hasPublicationYear "2023" @default.
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