AGING & NEURONAL DEATH--FIRST GENERATION OF NGF ANALOG

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One of the primary pathological processses occurring in the aging nervous system is neuronal death. For example, in Alzheimer's disease (AD) cholinergic neurons in the basal forebrain, which are thought to function in memory and learning, are lost. Neurotrophic factors are proteins which interact with cell surface receptors to promote the survival and growth of neurons. There is considerable evidence that neurotrophic factors such as nerve growth factor (NGF) can prevent neuronal death or damage and that they may be able to enhance the function of compromised neurons. Basal forebrain cholinergic neurons have NGF receptors and respond to NGF; therefore, NGF may be relevant to understanding and perhaps minimizing the loss of neurons in AD. A significant limitation to both understanding the physiological role of endogenous NGF in vivo and the potential therapeutic application of NGF is the lack of stable, low molecular weight agonists and antagonists of NGF which can effectively reach the central nervous system and penetrate the brain parenchyma. The long term objectives of this work are: i) to begin to define structural features of NGF (NGF's active site) which will allow it to interact with its receptor and ii) to develop a first generation of low molecular weight synthetic NGF peptide analogs which can mimic or inhibit the neurotrophic effect of NGF in vitro and eventually in vivo. The general strategy of this proposal is to map the active site(s) of NGF by synthesizing short peptides corresponding to potential active sites. These peptides will be assayed for their ability to block or mimic several measures of NGF bioactivity in vitro. By systematically varying which residues of NGF are includes in each peptide and by making peptides of different lengths, one can begin to map an active site. Active peptides will then be modified by conventional techniques to form a first generation of NGF analogs. This synthetic peptide strategy has been used many times to define active sites and produce active analogs of many proteins, most of which are considerably more complex than NGF. Indeed, our initial studies with NGF peptides suggest that this method will be successful with NGF as well. The specific aims during the proposed grant period will be: 1) To synthesize and purify NGF peptides and analogs derived from peptides which are active. 2) To test these peptides and analogs for their ability to mimic or inhibit three well-characterized activities of NGF in vitro. 3) To test the most potent peptides for their ability to compete with NGF's binding at its receptor.