The central hypothesis of this proposal is that there are abnormal qualitative and quantitative differences in the expression of the c-erbB proto-oncogene or the epidermal growth factor receptor (EGFR) between normal glial cells and the neoplastic glial cells of malignant gliomas and medulloblastomas. Glioma variant EGFR proteins, already identified in this laboratory, and the intact glioma EGFR will be affinity purified and biochemically analyzed to further characterize those structural and/or functional characteristics of these EGFRS which may confer a growth advantage for gliomas. This proposal is based on the following: 1) The EGFR gene is often amplified and highly expressed in malignant human gliomas; 2) structurally unique EGFRs are present in human gliomas, including one distinctive glioma EGFR, which resembles the v-erbB oncogene; and 3) monoclonal antibodies (Mabs) can be made against the normal and glioma variant EGFRs. These Mabs will then be powerful tools for elucidating the role of the overexpressed, normal EGFR and the molecular variant EGFRs in the growth of these malignant brain tumors.
Our specific aims are: a) To purify and characterize variant EGFR proteins in malignant human gliomas which amplify and rearrange the EGFR gene, with particular focus on the variant EGFR in glioma D- 245, which amplifies a markedlly rearranged EGFR gene and expresses a small EGFR that resembles the truncated EGFR expressed by the viral oncogene erbB; b) to compare the biochemical features of the purified EGFR from gliomas which amplify and rearrange the gene with those of EGFRs from gliomas which amplify but do not rearrange the EGFR gene, gliomas which do not amplify and do not rearrange the EGFR gene, A431 squamous carcinoma which expresses an intact EGFR and normal tissues such as placenta which also express an intact EGFR; c) to produce Mabs directed against normal EGFR and the purified variant EGFRs from malignant human gliomas; d) to define the expression of EGFR and variant EGFRs in human gliomas, normal adult and fetal tissues qualitatively by Mab immunocytochemistry and quantitatively by radioimmunoassay and radioreceptor binding assays; e) to use these Mabs to examine the function of the normal and variant glioma EGFRs in cell-cell interactions, clonagenicity and invasion utilizing in vitro and in vivo assays of cell growth, adhesion and invasiveness; f) to demonstrate the efficacy of Mabs exhibiting broad reactivity within and among human gliomas and minimal normal brain reactivity in radioimmunoimaging and immunotherapeutic studies in the nude mouse- human glioma xenograft model; and g) to utilize the identical approaches and reagents for medulloblastoma, using the best model systems in existence. These studies will thus examine the functional significance of normal and variant EGFRs in the biologic behavior of human gliomas and medulloblastomas and will establish pre-clinical efficacy of radiolabeled anti-EGFR Mabs in glioma radioimmunoimaging and radioimmunotherapy to aid in selecting monoclonal antibodies for clinical trials.
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