title: Robust detection of EGFR copy number changes and EGFR variant III: technical aspects and relevance for glioma diagnostics.
abstract: Epidermal growth factor receptor (EGFR) is commonly affected in cancer, generally in the form of an increase in DNA copy number and/or as mutation variants [e.g., EGFR variant III (EGFRvIII), an in-frame deletion of exons 2-7]. While detection of EGFR aberrations can be expected to be relevant for glioma patients, such analysis has not yet been implemented in a routine setting, also because feasible and robust assays were lacking. We evaluated multiplex ligation-dependent probe amplification (MLPA) for detection of EGFR amplification and EGFRvIII in DNA of a spectrum of 216 diffuse gliomas. EGFRvIII detection was verified at the protein level by immunohistochemistry and at the RNA level using the conventionally used endpoint RT-PCR as well as a newly developed quantitative RT-PCR. Compared to these techniques, the DNA-based MLPA assay for EGFR/EGFRvIII analysis tested showed 100% sensitivity and specificity. We conclude that MLPA is a robust assay for detection of EGFR/EGFRvIII aberrations. While the exact diagnostic, prognostic and predictive value of such EGFR testing remains to be seen, MLPA has great potential as it can reliably and relatively easily be performed on routinely processed (formalin-fixed, paraffin-embedded) tumor tissue in combination with testing for other relevant glioma markers.
authors.affiliation: Department of Pathology, Nijmegen Center for Molecular Life Sciences, Radboud University Nijmegen Medical Centre, Nijmegen, the Netherlands. j.jeuken@pathol.umcn.nl
authors.forename: Judith
authors.initials: J
authors.lastname: Jeuken
authors.forename: Angelique
authors.initials: A
authors.lastname: Sijben
authors.forename: Cristina
authors.initials: C
authors.lastname: Alenda
authors.forename: Jos
authors.initials: J
authors.lastname: Rijntjes
authors.forename: Marieke
authors.initials: M
authors.lastname: Dekkers
authors.forename: Sandra
authors.initials: S
authors.lastname: Boots-Sprenger
authors.forename: Roger
authors.initials: R
authors.lastname: McLendon
authors.forename: Pieter
authors.initials: P
authors.lastname: Wesseling
mesh_terms: D001932:Brain Neoplasms / Q000175:diagnosis / Q000235:genetics; D066246:ErbB Receptors / Q000235:genetics*; D005784:Gene Amplification; D018628:Gene Dosage; D005910:Glioma / Q000175:diagnosis* / Q000235:genetics*; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D009154:Mutation; D011237:Predictive Value of Tests; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D012680:Sensitivity and Specificity
chemical_list: D066246:ErbB Receptors