新型Ⅰ型大麻素受体拮抗剂LMJ07的体外生物活性

发布时间：2018-03-17 05:17

本文选题：型大麻素受体　切入点：选择性拮抗剂　出处：《国际药学研究杂志》2015年01期 　论文类型：期刊论文

【摘要】：目的 LMJ07为靶向大麻素Ⅰ型受体(CB1R)设计合成的选择性拮抗剂,本文从分子水平受体结合、细胞水平CB1R受体内在化、CB1R诱导的细胞骨架和信号分子的变化等方面对其CB1R拮抗活性进行了评价。方法以已知的CB1R选择性拮抗剂利莫那班(SR141716A)为对照,采用放射性配体竞争结合和增强型绿色荧光蛋白(EGFP)示踪大麻素受体(CBR)内在化的高内涵分析实验分析了LMJ07对2种CBR亚型(CB1R和CB2R)的结合及G蛋白不依赖的受体激活的拮抗活性及选择性;分别采用CELLKEY无标记检测系统和均相时间分辨荧光(HTRF)测定了LMJ07对CBR激活诱导的CHO-CB1细胞骨架和胞内环腺苷酸(c AMP)含量的影响;最后采用连续荧光检测技术使用表达CB1R的原代海马细胞检测LMJ07对胞内Ca2+的影响。结果 LMJ07高亲和、高选择地与CB1R结合,选择性地拮抗CB1R的激活导致的受体内吞,且选择性和拮抗活性与利莫那班相当;在CHO-CB1R细胞上,LMJ07(0.01~10μmol/L)剂量依赖地逆转CBR激动剂Win55212-2诱导的细胞骨架变化相关的细胞电阻改变、剂量依赖地逆转Win55212-2降低胞内c AMP含量效应;在原代培养的海马细胞上,LMJ07(10 nmol/L~1μmol/L)可拮抗Win55212-2诱导的胞内Ca2+增加效应。在上述实验中,LMJ07拮抗CB1R通路的信号分子和功能的活性与利莫那班相当。结论 LMJ07是一个与利莫那班活性相当的新结构类型CB1R选择性拮抗剂。
[Abstract]:The purpose of LMJ07 targeting cannabinoid receptors (CB1R) selective antagonist synthesis, this paper from the molecular level, cellular level of CB1R receptor binding, receptor internalization, cytoskeleton and signaling molecules in CB1R induced changes in other aspects of the evaluation of the antagonistic activity of CB1R. Methods CB1R selective antagonist rimonabant known (SR141716A) as control, using radioligand binding and enhanced green fluorescent protein (EGFP) tracer cannabinoid receptor (CBR) analysis of high content of internalized LMJ07 experimental analysis of 2 kinds of CBR subtypes (CB1R and CB2R) antagonistic activity and selective activation of G protein binding and receptor independent respectively; using CELLKEY marker detection system and homogeneous time-resolved fluorescence (HTRF) induced activation of LMJ07 cells and CHO-CB1 cells of the inner skeleton of adenylate CBR was determined (C AMP) content influence; finally, with the continuous Fluorescence detection technology using expression detection of primary hippocampal cell LMJ07 CB1R on intracellular Ca2+. Results LMJ07 high affinity, high selective binding to CB1R, the selective antagonist of CB1R receptor activation results in endocytosis, and selective antagonist activity and rimonabant and equivalent; in CHO-CB1R cells, LMJ07 (0.01~10 mol/L) cell resistance dose dependently reversed CBR agonist agent Win55212-2 induced changes of cytoskeleton related changes, dose dependently reversed Win55212-2 effect of intracellular AMP content decreased C; in primary cultured hippocampal cells, LMJ07 (10 nmol/L~1 mol/L) can antagonize Win55212-2 induced intracellular Ca2+ increase effect. In the experiment, activity and rimonabant LMJ07 signaling molecules of the CB1R pathway and antagonistic function. Conclusion LMJ07 is a new type of structure with a CB1R selective antagonist rimonabant activity equivalent.

【作者单位】：军事医学科学院毒物药物研究所;沈阳药科大学;
【基金】：国家“重大新药创制”科技重大专项资助项目(2012ZX09301003-001,2012ZX09301003-003) 国家自然科学基金资助项目(81102308)
【分类号】：R96

【共引文献】