Practical Bioinformatic DNA-Sequencing Pipeline for Detecting Oncogene Amplification and EGFRvIII Mutational Status in Clinical Glioblastoma Samples

Publication date: Available online 15 April 2019Source: The Journal of Molecular DiagnosticsAuthor(s): Michael L. Miller, Jessica Tome-Garcia, Aneta Waluszko, Tatyana Sidorenko, Chitra Kumar, Fei Ye, Nadejda M. TsankovaGlioblastoma is a malignant brain tumor with dismal prognosis. Oncogenic mutations in glioblastoma frequently affect receptor tyrosine kinase pathway components that are challenging to quantify because of heterogeneous expression. EGFRvIII, a common oncogenic receptor tyrosine kinase mutant protein in glioblastoma, potentiates tumor malignancy and is an emerging tumor-specific immunotarget, underlining the need for its more accessible and quantitative detection. We used normalized next-generation sequencing data from 117 brain and 371 reference clinical tumor samples to detect focal gene amplifications across the commercial Ion AmpliSeq Cancer Hotspot Panel version 2 and infer EGFRvIII status based on relative coverage dropout of the gene's truncated region within EGFR. In glioblastomas (n = 45), amplification of EGFR [18 (40%)], PDGFRA [3 (7%)], KIT [2 (4%)], MET [1 (2%)], and AKT1 [1 (2%)] was detected. With respect to EGFR and PDGFRA amplification, there was near-complete agreement between next-generation sequencing and in situ hybridization. Consistent with previous reports, this method detected EGFRvIII exclusively in EGFR-amplified glioblastomas [8 (44%)], which was confirmed using long-range PCR. Our study offers a practical method for detectin...
Source: The Journal of Molecular Diagnostics - Category: Pathology Source Type: research

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Source: Archives of Pathology and Laboratory Medicine - Category: Laboratory Medicine Authors: Tags: Arch Pathol Lab Med Source Type: research
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