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Having an Anesthetic without adverse effects : The binding sites of dexmedetomidine on adrenoceptor

具有无副作用的麻醉剂：右美托咪定对肾上腺素受体的结合位点

基本信息

批准号：
09470335
负责人：
MIZOBE Toshiki
金额：
$ 5.76万
依托单位：
Department of Anesthesiology Kyoto Prefectural University of Medicine
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 1999
项目状态：
已结题

项目摘要

Dexmedetomidine (DEX) is a subtype non-selective, alpha2 adrenergic agonist developed for anesthetic management because of its sedative/hypnotic, anestheticsparing, analgesic and sympatholytic properties. Each of these properties are thought to be mediated by activation of the alpha2A adrenoceptor subtype. We sought to determine the TMDs on the human alpha2A adrenoceptor subtype which are responsible for binding to DEX in order to design novel subtype selective alpha2A agonists. Chimeric receptor were constructed to replace defined TMDs of the wild-type human alpha2A adrenoceptor with homologous sequence from the human beta2 adrenoceptor which itself has no affinity for DEX.Chimerae of the human alpha2A and human beta2 adrenoceptors were constructed from genes encoding human alpha2A (or C10) and beta adrenoceptors. COS-7 cells were transfected with 12 new constructs as well as the wild-type human alpha2A and human beta2 adrenoceptors. Cells were harvested and membranes prepared for radiolabeled ligand binding using 3H-atipamezole and DEX as the displacing ligand.TMD#7 is pivotal for DEX binding since CRS 25 (alpha2 TMD#7) has > 10,000 for higher affinity than CRT3 (beta2 TMD#7). To a lesser extent TMD#2 is involved in DEX binding since CRS16 (alpha2 TMD#2) has >100 fold higher affinity for DEX than does beta2 adrenoceptor. TMD#6 may be involved in DEX binding since CRS 25(alpha2 TMD#6) has 10 fold higher affinity for DEX than does CR8 (beta2 TMD#6).The next generation of alpha2 adrenergic agonists needs to be alpha2A subtype selective. To better understand the amino acid residues responsible for subtype selectivity, further site-directed mutagenesis studies of the differences in amino acid residues in TMD#7 and 2 (and 6) between the three adrenoceptor subtypes need to be undertaken. This will facilitate target-based drug design of the novel alpha2A subtype selective agonists for use in anesthesia.

右美托咪定（DEX）是一种非选择性α 2肾上腺素能激动剂，因其镇静/催眠、麻醉保留、镇痛和交感神经溶解特性而被开发用于麻醉管理。这些特性都被认为是由α 2a肾上腺素受体亚型的激活介导的。我们试图确定人类α 2a肾上腺素受体亚型与DEX结合的tmd，以便设计新的亚型选择性α 2a激动剂。构建嵌合受体，用人β 2肾上腺素受体的同源序列取代野生型人α 2a肾上腺素受体的定义TMDs，而β 2肾上腺素受体本身对DEX没有亲和力。利用编码人α 2a（或C10）和β肾上腺素受体的基因构建人α 2a和β 2肾上腺素受体嵌合体。在COS-7细胞中转染了12个新构建体以及野生型人α 2a和β 2肾上腺素受体。收集细胞并制备膜，以3h -阿替帕唑和DEX作为取代配体进行放射性标记配体结合。TMD#7是DEX结合的关键，因为CRS 25 （alpha2 TMD#7）比CRT3 （bet2 TMD#7）具有更高的亲和力。在较小程度上，TMD#2参与DEX结合，因为CRS16 （alpha2 TMD#2）对DEX的亲和力比β 2肾上腺素受体高100倍。TMD#6可能参与DEX结合，因为CRS 25（alpha2 TMD#6）对DEX的亲和力比CR8 （bet2 TMD#6）高10倍。下一代α 2肾上腺素能激动剂需要具有α 2a亚型选择性。为了更好地了解负责亚型选择性的氨基酸残基，需要进行进一步的位点定向诱变研究，研究TMD#7和2（和6）中三种肾上腺素能受体亚型之间氨基酸残基的差异。这将促进用于麻醉的新型alpha2A亚型选择性激动剂的靶向药物设计。