Laboratorium für Organische Chemie, Eidgenössische Technische Hochschule Zürich, Vladimir-Prelog-Weg 3, 8093 Zürich, Switzerland.
Roche Pharma Research & Early Development, Roche Innovation Center Basel, F. Hoffmann-La Roche Ltd., 4070 Basel, Switzerland.
J Am Chem Soc. 2020 Oct 7;142(40):16953-16964. doi: 10.1021/jacs.0c05587. Epub 2020 Sep 29.
Pharmacological modulation of cannabinoid type 2 receptor (CBR) holds promise for the treatment of numerous conditions, including inflammatory diseases, autoimmune disorders, pain, and cancer. Despite the significance of this receptor, researchers lack reliable tools to address questions concerning the expression and complex mechanism of CBR signaling, especially in cell-type and tissue-dependent contexts. Herein, we report for the first time a versatile ligand platform for the modular design of a collection of highly specific CBR fluorescent probes, used successfully across applications, species, and cell types. These include flow cytometry of endogenously expressing cells, real-time confocal microscopy of mouse splenocytes and human macrophages, as well as FRET-based kinetic and equilibrium binding assays. High CBR specificity was demonstrated by competition experiments in living cells expressing CBR at native levels. The probes were effectively applied to FACS analysis of microglial cells derived from a mouse model relevant to Alzheimer's disease.
大麻素受体 2（CBR）的药理学调节有望用于治疗多种疾病，包括炎症性疾病、自身免疫性疾病、疼痛和癌症。尽管该受体具有重要意义，但研究人员缺乏可靠的工具来解决有关 CBR 信号表达和复杂机制的问题，特别是在细胞类型和组织依赖的情况下。在此，我们首次报道了一种用于模块化设计一系列高特异性 CBR 荧光探针的多功能配体平台，该探针可成功应用于不同物种和细胞类型的应用。这些探针包括对表达内源性 CBR 的细胞进行流式细胞术分析、实时共聚焦显微镜观察小鼠脾细胞和人巨噬细胞，以及基于荧光共振能量转移（FRET）的动力学和平衡结合测定。通过在表达天然水平 CBR 的活细胞中的竞争实验证明了这些探针具有高 CBR 特异性。这些探针有效地应用于源自与阿尔茨海默病相关的小鼠模型的小神经胶质细胞的 FACS 分析。
