The utility of characterizing and detecting epidermal growth factor receptor mutations in developing personalized medical strategies toward glioblastoma multiforme

Glioblastoma multiforme (GBM) is the most common primary brain cancer of adults. GBM develops rapidly and has complex molecular genetics, resulting in ineffective treatment and high mortality. In GBM, the epidermal growth factor (EGF) receptor is frequently amplified and/or overexpressed, and up to half of these cases also involve receptor mutations. EGF receptor expression status is linked to GBM development and may serve a causative role. In the upcoming era of personalized medicine, EGF receptor expression and mutation may serve as an attractive target for developing therapeutics and to establish diagnosis and treatment surveillance using rapid proteomic microarrays.;A GBM-relevant EGF receptor mutation featuring an in-frame, tandem duplication of the kinase domain (TKD) was expressed and characterized in B82L mouse fibroblast cells and U87MG GBM cells. The TKD possessed elevated basal levels of autophosphorylation/activation, attributable to the C-terminal kinase domain, and localized strongly to the intracellular compartment of cells. TKD basal activity was resistant to extracellular antibody inhibitors, but remained susceptible to intracellular small molecule TKIs. Finally, the elevated basal activity of the receptor led to downstream activation of several members along the MAPK signaling axis as well as the PLCgamma1 pathway. These molecular events may explain how the TKD receptor mutation leads to GBM tumorigenesis.;This study also reports progress in developing microarray technology to rapidly detect and quantify EGF receptor. Purified, radiolabeled, wild type EGF receptor was captured by anti-EGF receptor antibody-coated polydimethylsiloxane (PDMS) and polystyrene (PS) surfaces, transferred to high-energy surfaces, and quantified. While PDMS capture of receptor was reproducible and specific, and displayed flexibility in concentrations and antibody-immobilization, the sensitivity was surpassed by PS surfaces. Transfer of EGF receptor from PDMS surfaces to the high-energy surfaces proved highly efficient (up to 75% transfer). The successful capture of EGF receptor by PDMS and subsequent transfer to high-energy surfaces makes this attractive technology for developing rapid microarrays for liquid crystal-based readouts.