DESIGN OF INHIBITORS OF EPINEPHRINE BIOSYNTHESIS

描述：苯乙醇胺N-甲基转移酶（PNMT，E. C. 2.1.1.28请参阅 是催化生物合成的最后一步的酶， 肾上腺素（Epi）。 Epi占中枢神经系统的5% (CNS)儿茶酚胺含量，它与许多疾病有关 大脑中的神经调节过程。 我们的实验室（Our Laboratory） 和其他人），PNMT抑制剂可以降低血压， 自发性高血压大鼠然而，目前所有的抑制剂 可用的对α 2-肾上腺素能受体具有高亲和力， 对观察到的药理学作用有显著贡献。 一些 的探针，以确定PNMT的活性位点结合要求， α 2-肾上腺素受体已经在我们实验室合成， 开发计算机图形模型（比较分子场分析; CoMFA）的两个网站。 这些模型已被用于设计新的配体 在初步研究的基础上， 表现出对PNMT活性位点的所需选择性水平， α 2肾上腺素受体。 这些配体的评价结果将 用于改进我们的计算机模型，并提高其辅助 设计一种有效和选择性的PNMT抑制剂， 通过血脑屏障的能力。 人脑PNMT（hPNMT） 最近被克隆和表达（与M. McLeish） 在三种PNMT存在下， 具有不同结合特性的抑制剂已经生长（在 与J. Martin合作）;晶体结构有待解决。 结合计算机建模，蛋白质晶体学（基于结构 配体设计），定点诱变和同源性建模， hPNMT的活性位点将使我们能够开发更具选择性和有效性的 抑制剂的 PNMT的高通量筛选（基于与 与R. Borchardt）将被开发， 用于筛选结构多样的化合物的独特库， Smissman和Mertes在美国的样本收集。来自堪萨斯。 电极导线 在适当的情况下用平行合成技术进行优化。 毛细管电泳微透析实验（合作 链球菌Lunte）将允许测量以下物质对CNS Epi水平的影响： 新的PNMT抑制剂在清醒大鼠体内。 所有结果 次级项目将协同作用， PNMT的有效的和选择性的抑制剂，这将是有用的， 药理学工具，以探测Epi在CNS中发挥的作用， 特别是，可能确定一种新的药物治疗机制， 高血压

DESCRIPTION: Phenylethanolamine N-methyltransferase (PNMT, E. C. 2.1.1.28) is the enzyme that catalyzes the terminal step in the biosynthesis of epinephrine (Epi). Epi comprises about 5 percent of central nervous system (CNS) catecholamine content and it has been implicated in a number of neuroregulator processes in the brain. It was demonstrated (our laboratory and others) that inhibitors of PNMT can lower blood pressure in spontaneously hypertensive rats However, all of the inhibitors presently available have high affinity for a2-adrenergic receptors, which could contribute significantly to the observed pharmacological effects. A number of probes to determine the active site binding requirements for PNMT and for the a2-adrenoceptor have been synthesize in our laboratory and used to develop computer graphics models (comparative molecular field analysis; CoMFA) of the two sites. These models have been used to design new ligands for synthesis that, based on preliminary studies, have the potential of exhibiting the desired level of selectivity for the PNMT active site over the a2-adrenoceptor. The results from the evaluation of these ligands will be used to refine our computer model and improve its ability to aid in the design of a potent and selective inhibitor of PNMT with sufficient lipophilicity to cross the blood brain barrier. Human brain PNMT (hPNMT) has recently been cloned and expressed (in collaboration with M. McLeish) and thre different crystals of hPNMT in the presence of three PNMT inhibitors with different binding characteristics have been grown (in collaboration with J. Martin); the crystal structures are awaiting solution. A combination of computer modeling, protein crystallography (structure-based ligand design), site-directed mutagenesis and homology modeling of the active site of hPNMT will allow us to develop more selective and potent inhibitors. A high throughput screen for PNMT (based on coupling with AdoHcy hydrolase, in collaboration with R. Borchardt) will be developed and used to screen unique libraries of structurally diverse compounds in the Smissman and Mertes sample collections at the U. of Kansas. Leads will be optimized with parallel synthesis techniques where appropriate. Microdialysis experiments with capillary electrophoresis (in collaboration with S. Lunte) will allow measurement of the effects on CNS Epi levels of new PNMT inhibitors in vivo in conscious rats. Results from all sub-projects will act synergistically to lead to the synthesis of a highly potent and selective inhibitor of PNMT, which would be useful as a pharmacological tool to probe the role(s) played by Epi i the CNS, and, in particular, potentially identify a new mechanism for drug treatment of hypertension.