The cannabinoid receptor 1 (CB₁R) regulates synaptic transmission in the central nervous system, but also has important roles in the peripheral organs controlling cellular metabolism. While earlier generations of brain penetrant CB₁R antagonists advanced to the clinic for their effective treatment of obesity, such molecules were ultimately shown to exhibit negative effects on central reward pathways that thwarted their further therapeutic development. The peripherally restricted CB₁R inverse agonists MRI-1867 and MRI-1891 represent a new generation of compounds that retain the metabolic benefits of CB₁R inhibitors while sparing the negative psychiatric effects. To understand the mechanism of binding and inhibition of CB₁R by peripherally restricted antagonists, we developed a nanobody/fusion protein strategy for high-resolution cryo-EM structure determination of the GPCR inactive state, and used this method to determine structures of CB₁R bound to either MRI-1867 or MRI-1891. These structures reveal how these compounds retain high affinity and specificity for CB₁R’s hydrophobic orthosteric site despite incorporating polar functionalities that lead to peripheral restriction. Further, the structure of the MRI-1891 complex along with accompanying molecular dynamics simulations shows how differential engagement with transmembrane helices and the proximal N-terminus can propagate through the receptor to contribute to biased inhibition of β-arrestin signaling.

大麻素受体 1 （CB₁R） 调节中枢神经系统的突触传递，但在控制细胞代谢的外周器官中也起着重要作用。虽然前几代脑渗透性 CB₁R 拮抗剂因其有效治疗肥胖而进入临床，但这些分子最终被证明对中枢奖励通路表现出负面影响，阻碍了它们的进一步治疗发展。外周限制性 CB₁R 反向激动剂 MRI-1867 和 MRI-1891 代表了新一代化合物，它们保留了 CB₁R 抑制剂的代谢益处，同时避免了负面的精神影响。为了了解外周限制性拮抗剂结合和抑制 CB₁R 的机制，我们开发了一种纳米抗体/融合蛋白策略，用于高分辨率冷冻电镜结构测定 GPCR 非活性状态，并使用该方法确定 CB₁R 与 MRI-1867 或 MRI-1891 结合的结构。这些结构揭示了这些化合物如何保持对 CB₁R 的疏水性正构位点的高亲和力和特异性，尽管它们结合了导致外周限制的极性官能团。此外，MRI-1891 复合物的结构以及伴随的分子动力学模拟显示了与跨膜螺旋和近端 N 末端的差异结合如何通过受体传播，从而导致 β-arrestin 信号转导的偏向抑制。