AGING & NEURONAL DEATH--FIRST GENERATION OF NGF ANALOG

衰老过程中的主要病理过程之一 神经系统是神经元的死亡。例如，在阿尔茨海默病中 (Ad)基底前脑中的胆碱能神经元，被认为是 在记忆和学习中的功能，都在丧失。神经营养因子是 与细胞表面受体相互作用的蛋白质促进 神经元的存活和生长。有大量证据表明 神经生长因子(NGF)等神经营养因子可以预防 神经元死亡或损伤，它们可能能够增强 受损神经元的功能。基底前脑胆碱能神经元 有NGF受体并对NGF有反应；因此，NGF可能与 了解并可能最大限度地减少AD中神经元的损失。一个 对两者的生理作用的理解都受到显著的限制 体内内源性神经生长因子及其潜在的治疗应用 缺乏稳定的、低分子质量的激动剂和拮抗剂 能有效到达中枢神经系统并穿透的神经生长因子 脑实质。这项工作的长期目标是：i) 开始定义NGF(NGF的活性部位)的结构特征 将允许它与其受体相互作用和ii)发展出第一个 低分子人工合成NGF多肽类似物的制备 能在体外模拟或抑制NGF的神经营养作用 最终在活体内。这项提案的总体战略是绘制 神经生长因子的活性部位(S)通过合成对应于 潜在的活跃站点。这些多肽将被检测其 在体外阻断或模拟多种NGF生物活性的能力。 通过系统地改变每一个中包含哪些NGF残基 通过制造不同长度的多肽，一个人可以开始 映射活动站点。然后，活性多肽将被 形成第一代NGF类似物的常规技术。这 合成肽策略已被多次用于定义活性 并产生许多蛋白质的活性类似物，其中大多数是 比NGF复杂得多。事实上，我们的初步研究与 NGF多肽提示，这种方法将在NGF AS中获得成功 井。 建议授权期内的具体目标为：1) 神经生长因子多肽及其类似物的合成和纯化 它们都是活跃的。2)测试这些多肽及其类似物的 能够模拟或抑制NGF的三种特性良好的活性 在试管中。3)测试最有效的多肽的能力 与NGF在其受体上的结合形成竞争。

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.