Nerve growth factor (NGF) is a neurotrophic factor known to affect the survival and differentiation of neuroblasts and some neuroblastoma cell lines. The effects of NGF are mediated through a cell surface receptor which has recently been characterized and cloned in our laboratory. Our first Specific Aim is to survey freshly excised neuroblastomas, neuroblastomas placed into short- term culture, and neuroblastoma cell lines for expression of NGF receptor, the affinity and characteristics of the NGF receptor, and the responsiveness of the neuroblastomas to NGF. We will analyze the NGF receptor for these neuroblastomas by immunoperoxidase staining with monoclonal anti-NGF receptor antibodies, by Western blotting and 125I-NGF binding. Differentiation of neuroblastomas will be determined by morphology, 3H-thymidine incorporation, enzyme activities, and Northern blotting. We will correlate these results with the stage of the disease, the clinical prognosis, and the subclass of neuroblastoma. Our second Specific Aim is to determine whether H-myc oncogene which is amplified in about 50% of advanced neuroblastomas alters the expression and molecular characteristics of the NGF receptor and the ability of these cells to respond to NGF. Neuroblastoma cell lines that do not express H-myc will be compared with those that do and with neuroblastoma cell lines that express N-myc as a result of transfection with N- myc oncogene. Our third Specific Aim is to restore expression of NGF receptor in those cell lines which lack the receptor and lo assess whether restored expression results in NGF-inducible differentiation. Expression of NGF receptor will be induced by transfection or with a retrovirus bearing the NGF receptor cDNA. The results of these experiments should provide insight into the mechanism by which activation of oncogenes blocks differentiation in neuroblastoma, and into therapeutic approaches for this disease and other malignancies.
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