PRO-LEU-GLY-NH2 AMD DOPAMINE RECEPTOR MODULATION STUDY

PRO-LEU-GLY-NH2 AMD 多巴胺受体调节研究

• 批准号: 2263754
• 负责人: RODNEY L JOHNSON
• 金额: $ 19.61万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 1983
• 资助国家: 美国
• 起止时间: 1983-12-01 至 1999-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2263754
• 关键词: G protein; Parkinson's disease; X ray crystallography; chemical structure function; disease /disorder model; dopamine agonists; dopamine receptor; gene expression; haloperidol; hormone receptor; laboratory mouse; laboratory rat; lactams; neuropeptide receptor; nuclear magnetic resonance spectroscopy; nucleic acid probes; peptide chemical synthesis; peptide hormone; peptide hormone analog; piperazines; radiotracer; receptor binding; tardive dyskinesia

The long term objective of this research is to understand the structural basis by which Pro-Leu-Gly-NH2 (PLG) and its peptidomimetic analogues modulate CNS dopamine receptors and to elucidate the molecular mechanism underlying this modulation. In pursuing this objective, we plan to synthesize analogues of highly potent PLG peptidomimetics to test the hypothesis that the carbonyl groups in these molecules are important for their intrinsic activity. Analogues will also be made which are substituted with hydrophobic groups capable of mimicking the leucyl side chain found in PLG. Synthesis of analogues of the diketopiperazine lactam peptidomimetic will be made to test the hypothesis that it is the type II beta-turn conformation that is the important conformational feature of dopamine receptor modulating activity while the N-terminal C5 conformation is the structural feature responsible for the potency of the PLG peptidomimetics. Different sized ring analogues of the highly constrained spiro bicyclic lactam PLG peptidomimetic will be made so as to alter the phi2, psi2, and phi3 torsion angles and thereby determine the effect this has on dopamine receptor modulating activity. Finally, two general types of potential radioligands for the putative PLG receptor will be obtained and evaluated. One type will be a tritiated diketopiperazine lactam peptidomimetic,while in a second approach the 4- hydroxybenzyl group will be incorporated into the structures of the highly potent PLG analogues for the purpose of radiolabeling these compounds with 125I. The PLG peptidomimetics synthesized will be evaluated for their ability to modulate (3H]spiroperidol/N- propylnorapomorphine competitive binding to D2 receptors in the presence or absence of 5 prime-guanylylimidodiphosphate, a norhydrolyzable analog of GTP. They will also be evaluated for their ability to enhance the binding of dopamine receptor agonists to either the D1, D2, D3 D4 or D5 receptors using SH-SY5Y human neuroblastoma cell lines transfected with either the D1,D2, D3, D4 or D5 receptor genes. Selected PLG peptidomimetics will be tested for their activity in in vivo animal models of Parkinson's disease (6- hydroxydopamine-lesioned rat model and the MPTP mouse model) and dyskinesia disorders (haloperidol-induced D2 receptor supersensitivity model and the L-DOPA-induced dyskinesia model). Finally, studies will be conducted to further determine the functional interaction of PLG and its bioactive analogues with G-proteins in the striatum through the use of specific antisense phosphothioate oligonucleotides by examining: (1) the ability of Gs and Gi antisense oligonucleotides to modulate rotational behavior induced by dopamine agonists in 6-hydroxydopamine lesioned rats in the presence and absence of PLG analogues; (2) the G protein levels and mRNA expression for Gi alter treating rats with D2 receptor agonists in the absence and presence of PLG peptidomimetics, and (3) the effect of PLG peptidomimetics on GTPase activity of purified G-proteins uncoupled from the receptor. These studies will enhance our understanding of dopamine receptor modulation and provide groundwork for the development of new therapeutic agents for the treatment of such neurological disorders as Parkinson's disease and tardive dyskinesia.

这项研究的长期目标是了解结构性 Pro-Leu-Gly-NH2（PLG）及其肽类似类似物的基础 调节中枢神经系统多巴胺受体并阐明分子机制 该调制的基础。在追求这个目标时，我们计划 合成高度有效PLG肽仪的类似物测试 假设这些分子中的羰基对 它们的内在活性。也将制作类似物 用能够模仿叶核侧的疏水基取代 在PLG中发现的链。 Diketopiperazine lactam的类似物的合成 将提出肽分以检验是类型的假设 II是重要构象特征的Beta-Turn构象 多巴胺受体调节活性，而N端C5 构象是负责效力的结构特征 PLG肽仪。高度的不同尺寸的环类似物 将制造受约束的螺纹螺旋乳杆菌lactam plg肽瘤 要更改phi2，psi2和phi3扭转角，从而确定 这对多巴胺受体调节活性的影响。最后， 推定的PLG受体的两种一般类型的潜在辐射基因 将获得并评估。一种类型将是一种 二酮甲酰胺lactam肽类拟合，而第二次接近4- 羟基苯基将纳入 高度有效的PLG类似物，用于放射标记的目的 具有125i的化合物。 PLG肽仪合成的是 评估其调节（3H]螺旋翼醇/N-的能力 在存在的情况下，丙基诺帕丁与D2受体的竞争性结合 或缺乏5个主要限性二磷酸，一种不可氢化的类似物 GTP。他们还将评估他们增强的能力 多巴胺受体激动剂与D1，D2，D3 D4或D5的结合 使用SH-SY5Y人类神经母细胞瘤细胞系的受体转染 D1，D2，D3，D4或D5受体基因。选定的PLG 肽仪将测试其在体内动物中的活性 帕金森氏病模型（6-羟基多巴胺属性的大鼠模型和 MPTP小鼠模型）和运动障碍障碍（氟哌啶醇诱导的D2 受体超敏模型和L-DOPA诱导的运动障碍模型）。 最后，将进行研究以进一步确定功能 PLG及其生物活性类似物与G蛋白的相互作用 通过使用特定反义的纹状体 通过检查：（1）GS和 GI反义寡核苷酸，以调节由 在存在的6-羟基多巴胺病变的大鼠中，多巴胺激动剂在存在下 没有PLG类似物； （2）G蛋白水平和mRNA表达 在不存在的情况下，用D2受体激动剂治疗大鼠的胃肠道 PLG肽仪的存在，（3）PLG的作用 纯化的GTPase活性的肽仪纯化的G蛋白与未偶联 受体。这些研究将增强我们对多巴胺的理解 受体调制并为开发新 治疗此类神经系统疾病的治疗剂 帕金森氏病和迟发性运动障碍。