The central hypothesis of this proposal is that deletion mutant epidermal growth factor receptors (EGFRs) in malignant human gliomas are different quantitatively in level of expression and qualitatively in structure and function compared to the intact EGFR in normal cells. We have identified and sequenced three classes of deletion mutant EGFRs in human glioblastoma. The three mutations segregate into structural classes based on the size and location of the extracellular domain deletion. We found the most common type of mutant EGFR in close to 20% of all glioblastomas, using our anti- synthetic peptide antibody directed against the mutation site. This proposal is based on the following: 1) Amplified and rearranged EGFR genes in human glioma result in high expression of mutant EGFR proteins; 2) Pure deletion mutant EGFR proteins may be utilized in structure-function studies; and 3) Our anti-mutation site antibody may be utilized to detect the mutation for expression and prognostic studies and to explore the role of mutant EGFR in glioma biologic behaviors and differential diagnosis.
The specific aims of this proposal are the following: 1) To biochemically characterize purified tumor-derived and recombinant deletion mutant EGFRs from the three classes of mutation, and to define the functional consequences of these structural EGFR alterations in ligand-binding and kinase activity, 2) To characterize the role of the mutant EGFRs in glioma mitogenic signalling, migration and invasion by investigating oligomerization of the mutant receptors as a mechanism for transducing ligand signals, and by defining the effect of EGF, a monoclonal antibody reactive against the EGF-binding site, and our anti-mutation site antibody on in vitro glioma cell proliferation, adhesion, migration and invasiveness, 3) To assess the expression of the EGFR mutant products of the amplified, rearranged EGFR genes by detecting expression of the mutants using mutation-site specific oligonucleotide probes and antibodies and by quantitating mutant EGFR expression level in tumor xenografts by radioimmunoassay, and 4) To assess the differential diagnostic utility of our anti-mutation site antibody and mutation site oligonucleotide probes in distinguishing glioblastoma from anaplastic astrocytomas and reactive gliosis and to assess the prognostic significance of the mutant EGFRs by comparing survival curves of glioblastoma patients with and without the mutations. These studies will thus examine the structural, functional and clinical significance of deletion mutants EGFRs in malignant human gliomas.
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| Wikstrand, C J; Stanley, S D; Humphrey, P A et al. (1993) Investigation of a synthetic peptide as immunogen for a variant epidermal growth factor receptor associated with gliomas. J Neuroimmunol 46:165-73 |
