PROTEIN TYROSINE KINASE GROWTH FACTOR RECEPTORS IN TUMORIGENESIS OF PNET

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Medulloblastomas are malignant cerebellar tumors with histologic features resembling those of undifferentiated neuroectodermal cells of the germinal matrix, and hence are also referred to as "primitive neuroectodermal tumors" (PNETs). The Children's Hospital of Philadelphia is a major referral center for the diagnosis and care of children with these tumors. Though histologically undifferentiated, the majority of PNETs show immunochemical and molecular evidences of early differentiation along neuronal or astroglial lineages. In particular, more than half of primary PNETs and all but one established PNET line express neurofilament peptides, a cytoskeletal feature of neuroblasts. Proliferation, differentiation, and eventual exit from the cell cycle of cells derived from CNS germinal matrix and destined to be neurons are regulated by cytokines which employ members of the protein tyrosine kinase (PTK) family of receptors for trans-membrane signalling. The hypothesis we plan to test is that genetic alterations of members of the PTK receptor family contribute to pathogenesis of human CNS PNETs. We plan the following experiments: 1. Members of the PTK family of growth factor receptors involved in neuroblastic differentiation of primitive neuroepithelium will be identified by two approaches: a) subtractive hybridization and cloning from a human tumor line that can be differentiated into postmitotic neurons by treatment with retinoic acid; and b) probing an existing human fetal brain expression library with an anti-phosphotyrosine monoclonal antibody. Established human PNET cell lines and primary human PNET tumors will then be examined for expression of these PTK transcripts, and results will be compared with those obtained with samples of normal cerebellum obtained at autopsy from immature and adult humans. 2. The following genetic alterations of these PTK genes will be sought in CNS PNETs: amplification, insertion, deletion, and point mutation. Each could lead to a dominant phenotype in which neuroblastic differentiation and growth control are perturbed, thus contributing to the pathogenesis of CNS PNETs. The biological significance of PTK receptor gene alterations detected in the PNETs will be assessed by: a) determination of their frequency of occurrence in established CNS PNET cell lines and primary PNET tumor specimens; and b) examination of their functional consequences by transfection assays employing cultures of mouse 3T3 cells and of embryonic rat neuroectodermal cells.