Laboratory of Chemical Biology, Department of Biomedical Engineering and Institute for Complex Molecular Systems, Technische Universiteit Eindhoven, Den Dolech 2, Eindhoven, 5612 AZ, The Netherlands.
ACS Chem Biol. 2021 Mar 19;16(3):510-519. doi: 10.1021/acschembio.0c00941. Epub 2021 Feb 17.
The RORγt nuclear receptor (NR) is of critical importance for the differentiation and proliferation of T helper 17 (Th17) cells and their production of the pro-inflammatory cytokine IL-17a. Dysregulation of RORγt has been linked to various autoimmune diseases, and small molecule inhibition of RORγt is therefore an attractive strategy to treat these diseases. RORγt is a unique NR in that it contains both a canonical, orthosteric and a second, allosteric ligand binding site in its ligand binding domain (LBD). Hence, dual targeting of both binding pockets constitutes an attractive alternative molecular entry for pharmacological modulation. Here, we report a chemical biology approach to develop a bitopic ligand for the RORγt NR, enabling concomitant engagement of both binding pockets. Three candidate bitopic ligands, , , and , comprising an orthosteric and allosteric RORγt pharmacophore linked via a polyethylene glycol (PEG) linker, were designed, synthesized, and evaluated to examine the influence of linker length on the RORγt binding mode. and show convincing evidence of concomitant engagement of both RORγt binding pockets, while the shorter does not show this evidence, as was anticipated during the ligand design. As the most potent bitopic RORγt ligand, , antagonizes RORγt function in a potent manner in both a biochemical and cellular context. Furthermore, displays an increased selectivity for RORγt over RORα and PPARγ compared to the purely orthosteric and allosteric parent compounds. Combined, these results highlight potential advantages of bitopic NR modulation over monovalent targeting strategies.
RORγt 核受体（NR）对于辅助性 T 细胞 17（Th17）细胞的分化和增殖及其促炎细胞因子 IL-17a 的产生至关重要。RORγt 的失调与各种自身免疫性疾病有关，因此，小分子抑制 RORγt 是治疗这些疾病的一种有吸引力的策略。RORγt 是一种独特的 NR，因为它在其配体结合域（LBD）中既包含一个经典的、正构的配体结合位点，也包含第二个变构的配体结合位点。因此，双重靶向两个结合口袋构成了一种有吸引力的替代分子进入途径，用于药理学调节。在这里，我们报告了一种化学生物学方法来开发 RORγt NR 的双位配体，从而能够同时结合两个结合口袋。设计、合成并评估了三种候选双位配体、、和，它们包含一个正构和变构的 RORγt 药效基团，通过聚乙二醇（PEG）接头连接，以研究接头长度对 RORγt 结合模式的影响。和显示出同时结合两个 RORγt 结合口袋的令人信服的证据，而较短的则没有显示出这种证据，这在配体设计时就已经预料到了。作为最有效的双位 RORγt 配体，对 RORγt 功能具有很强的拮抗作用，无论是在生化还是细胞环境中都是如此。此外，与纯正构和变构母体化合物相比，对 RORγt 的选择性高于 RORα 和 PPARγ。综上所述，这些结果突出了双位 NR 调节相对于单价靶向策略的潜在优势。
