The family of neurotrophins, related to nerve growth factor (NGF), interact with two types of receptors, the Trk tyrosine kinase receptor and the common neurotrophin receptor p75NTR, to support neuronal development in responsive peripheral neurons (sensory, sympathetic) and central neurons (cholinergic, dopaminergic) and to provide signals for survival of neurons in neurodegenerative disorders. This proposal is aimed at further development of neurotrophin mutants ('muteins') designed to produce preferential receptor specificity. A major hypothesis being addressed is that receptor- and/or signal- selective neurotrophin mutants would be more effective therapeutic reagents in certain disorders than wild type neurotrophins. The receptor-selective neurotrophin 'muteins' are at a stage to be tested in animal models of neurodegenerative diseases.
The Specific Aims of the proposal are: (i) To utilize receptor-selective neurotrophin mutein heterodimers to test specific receptor activation models. Novel neurotrophin heterodimers will be characterized to determine the binding affinity to each receptor, the main cellular responses (survival, apoptosis, or differentiation) and enzymes in key signaling pathways, (ii) To characterize the signaling pathway to apoptosis through p53 tumor suppressor protein/capase-3 -dependent and -independent mechanisms. Neurotrophin muteins will be screened for those that discriminate between p53 -dependent and -independent pathways with p53 temperature sensitive PC12 cells. Several neuronal cell lines and primary cultures will also be studied with select muteins. (iii) To test the therapeutic potential of signal- selective neurotrophin muteins by intranasal inhalation delivery to the central nervous system of rodents. Normal mice, p53 null mice, p75NTR null mice, normal rats, and aged rats will be used. Signaling pathways, including TrkA phosphorylation, MAP kinase, and DNA fragmentation, will be compared immunohistochemically and biochemically in the treated rat and mice brains. The rational design of therapeutic reagents derived from neurotrophins that selectively stimulate signaling pathways may lead to novel treatments for Alzheimer's Disease, Parkinson's Disease, and related neurological disorders - response-selective NGF mutants may ultimately be more effective than wild type NGF in treating neurodegenerative diseases. ? ?
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