Amplification of the <i>EGFR</i> gene can be maintained and modulated by variation of EGF concentrations in <i>in vitro</i> models of glioblastoma multiforme

<div><p>Glioblastoma multiforme (GBM) is the most common and lethal brain tumor in adults. It is known that amplification of the epidermal growth factor receptor gene (<i>EGFR</i>) occurs in approximately 40% of GBM, leading to enhanced activation of the EGFR signaling pathway and promoting tumor growth. Although GBM mutations are stably maintained in GBM <i>in vitro</i> models, rapid loss of <i>EGFR</i> gene amplification is a common observation during cell culture. To maintain <i>EGFR</i> amplification <i>in vitro</i>, heterotopic GBM xenografts with elevated <i>EGFR</i> copy number were cultured under varying serum conditions and EGF concentrations. <i>EGFR</i> copy numbers were assessed over several passages by quantitative PCR and chromogenic <i>in situ</i> hybridization. As expected, in control assays with 10% FCS, cells lost <i>EGFR</i> amplification with increasing passage numbers. However, cells cultured under serum free conditions stably maintained elevated copy numbers. Furthermore, EGFR protein expression positively correlated with genomic amplification levels. Although elevated <i>EGFR</i> copy numbers could be maintained over several passages <i>in vitro</i>, levels of <i>EGFR</i> amplification were variable and dependent on the EGF concentration in the medium. <i>In vitro</i> cultures of GBM cells with elevated <i>EGFR</i> copy number and corresponding EGFR protein expression should prove valuable preclinical tools to gain a better understanding of <i>EGFR</i> driven glioblastoma and assist in the development of new improved therapies.</p>