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) 是催化生物合成的最后一步的酶 肾上腺素(Epi)。肾上腺皮质激素约占中枢神经系统的5% (中枢神经系统)儿茶酚胺含量，它与许多 大脑中的神经调节过程。它被证明(我们的实验室 和其他)，PNMT的抑制剂可以降低血压 然而，目前所有的抑制剂都是自发性高血压大鼠 对α2肾上腺素能受体有很高的亲和力，这可能 对观察到的药理作用有很大贡献。一个数字 用于确定PNMT和For的活性部位结合要求的探针 我们实验室已经合成了A2-肾上腺素能受体，并将其用于 开发计算机图形模型(比较分子场分析； CoMFA)。这些模型已被用于设计新的配体。 根据初步研究，合成具有潜在的 表现出对PNMT活性部位的所需水平的选择性 A2肾上腺素受体。对这些配体的评估结果将 用来改进我们的计算机模型并提高其帮助 高效、选择性PNMT抑制剂的设计 亲脂性能穿过血脑屏障。人脑PNMT(HPNMT) 最近被克隆和表达(与M.McLeish合作) 在三种PNMT存在下，hPNMT有三种不同的结晶 已经生长出具有不同结合特性的抑制剂(在 与J.Martin合作)；晶体结构正在等待解决。 计算机建模、蛋白质结晶学(基于结构 配体设计)、定点突变和同源模建 HPNMT的活性部位将使我们能够开发出更有选择性和更有效的 抑制剂。一种用于PNMT的高通量屏幕(基于与 将与R.Borchardt合作开发和 用于筛选结构多样的化合物的独特文库 斯米斯曼和默特斯在堪萨斯大学收集样本。线索将是 在适当的情况下采用并行合成技术进行优化。 毛细管电泳微透析实验(合作 将允许测量对中枢神经系统EPI水平的影响 清醒大鼠体内新的PNMT抑制剂。结果来自所有人 子项目将协同行动，以导致合成一个高度 有效的和选择性的PNMT抑制剂，这将是有用的 探索EPI I在中枢神经系统中的作用的药理工具(S)，以及，在 特别是，潜在地确定了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.