Previous studies have identified heterogeneity in the karyotype and chemosensitivity of malignant human gliomas, defining one population of BCNU-resistant cells as near-diploid, with an over-representation of chromosomes 7 and 22. These cells secrete PDGF-like peptides. We propose to determine if such cells reside as foci in different parts of the tumor (regional) by analyzing each region of the same tumor by cytogenetic, biochemical and molecular methods. To determine if the aberrant cellular expression of PDGF-like peptides can be used as a marker to detect the resistant populations in vivo, frozen and paraffin sections of tumor will be stained with antisera (polyclonal and monoclonal) generated against normal heterodimer A-B PDGF molecules and homodimer of the B chain of PDGF. After recurrence, resistant cells will be examined, comparing their chemosensitivities and karyotypes to those of the primary tumor. To test the hypothesis that other genes mapped to chromosomes 7 and 22 contribute to the selective growth advantage of this population, it will be determined if the multidrug resistant gene (MDR) mapped to chromosome 7 is amplified and/or overexpressed in BCNU-resistant and sensitive cells. Similarly, amplification or aberrant expression of ornithine decarboxylase (ODC), the rate-controlling enzyme in the biosynthesis of polyamines will be determined. To test the hypothesis that the aberrant expression of the PDGF-like peptides in the BCNU-resistant cells is an autocrine mechanism involving unique phosphotyrosyl substrates or altered levels of phosphotyrosyl substrates, the phosphoprotein patterns of normal glial cells will be compared with those of astrocytomas (AS), anaplastic astrocytomas (AA) and glioblastomas multiforme (GBM) following PDGF binding. Similarly, unique or aberrant expression of phosphoprotein(s) in drug-resistant cells will be determined and characterized as they are enriched in the in vitro assay. In new studies, it will be determined if cisplatin selects an intrinsically resistant cell in vitro in a manner similar to BCNU. Low and high grade gliomas will be dissociated and their karyotypes determined. They will be treated with cisplatin using the colony forming assay. The intrinsically resistant (surviving) cells from each drug dose will be cloned and analyzed using cytogenetic, biochemical and molecular methods. Our long range goal for this project is the application of this information to the design of new therapeutic strategies either as a new treatment or to individualize treatment.
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