Issa Naiem T, Shen Tingzhen, Vizurraga Alexander, Pronin Alexey, Henry Taylor, Wang Qiang, Kwarcinski Frank E, Schürer Stephan, Badiavas Evangelos, Tall Gregory G, Slepak Vladlen Z
Department of Dermatology, University of Miami Miller School of Medicine, Miami, Florida.
Department of Pharmacology, University of Michigan Medical School, Ann Arbor, Michigan.
Mol Pharmacol. 2025 Aug;107(8):100059. doi: 10.1016/j.molpha.2025.100059. Epub 2025 Jul 5.
The cycle of GTP binding and hydrolysis controls heterotrimeric G proteins, and mutations reducing GTPase activity result in constitutive G protein signaling. In Gαq (gene: GNAQ) such mutations cause uveal melanoma and Sturge-Weber syndrome. Finding pharmacological agents that inhibit Gαq will be beneficial for research with therapeutic potential. Previously discovered bacterial depsipeptides (FR900359 and YM-254890) bind directly to Gαq and stabilize its inactive complex with GDP, but suffer from limitations of distribution and bioavailability. We used the established Gαq-YM-254890 complex structure to dock small-molecule drugs into the depsipeptide binding site of Gαq. Our in silico screen of 5000 Food Drug and Administration-approved, experimental, and withdrawn drugs predicted that thiazolidinediones are potential ligands of Gαq. Analysis of G protein coupled receptor-stimulated G protein GTPγS binding demonstrated that troglitazone (441 Da) inhibited Gq nucleotide exchange with the IC of ∼31.7 μM. The thiazolidinedione analogs, rosiglitazone and pioglitazone, had no effect. High concentrations of troglitazone modestly inhibited Gi and Gs, but not G13. In G protein thermal stability assays, troglitazone and FR900359 stabilized purified Gαq-GDP, indicating direct binding. Consistent with its negative effect on Gq signaling, in MIN6 mouse insulinoma cells, troglitazone inhibited Ca mobilization, extracellular regulated protein kinase phosphorylation, and insulin secretion stimulated by the Gq-coupled M3 muscarinic cholinergic receptor. Troglitazone and FR900359 inhibited proliferation of MEL92.1 uveal melanoma cells driven by a GNAQ-Q209L driver mutation, but not of SK-MEL-28 cells driven by BRAF-V600E. Together, our study shows that troglitazone may be a promising new lead for the development of a <500 Da small-molecule therapeutic Gαq inhibitor. SIGNIFICANCE STATEMENT: Troglitazone, unlike other thiazolidinediones, directly binds and inhibits activity of heterotrimeric G protein Gq, with a weaker effect on Gi. Troglitazone may find usage as a repurposed drug scaffold to build novel small-molecule Gαq inhibitors with better bioavailability than depsipeptide Gαq inhibitors.
GTP结合与水解循环控制着异源三聚体G蛋白，降低GTPase活性的突变会导致G蛋白组成型信号传导。在Gαq（基因：GNAQ）中，此类突变会引发葡萄膜黑色素瘤和斯特奇-韦伯综合征。寻找抑制Gαq的药物制剂将有助于具有治疗潜力的研究。先前发现的细菌环缩肽（FR900359和YM-254890）直接与Gαq结合，并使其与GDP的无活性复合物稳定，但存在分布和生物利用度方面的局限性。我们利用已确定的Gαq-YM-254890复合物结构，将小分子药物对接至Gαq的环缩肽结合位点。我们对5000种美国食品药品监督管理局批准的、实验性的和已撤市的药物进行的计算机模拟筛选预测，噻唑烷二酮类药物是Gαq的潜在配体。对G蛋白偶联受体刺激的G蛋白GTPγS结合的分析表明，曲格列酮（441 Da）以约31.7 μM的IC抑制Gq核苷酸交换。噻唑烷二酮类似物罗格列酮和吡格列酮则无此作用。高浓度的曲格列酮适度抑制Gi和Gs，但不抑制G13。在G蛋白热稳定性测定中，曲格列酮和FR900359使纯化的Gαq-GDP稳定，表明存在直接结合。与其对Gq信号传导的负面影响一致，在MIN6小鼠胰岛素瘤细胞中，曲格列酮抑制由Gq偶联的M3毒蕈碱胆碱能受体刺激引起的Ca动员、细胞外调节蛋白激酶磷酸化和胰岛素分泌。曲格列酮和FR900359抑制由GNAQ-Q209L驱动突变驱动的MEL92.1葡萄膜黑色素瘤细胞的增殖，但不抑制由BRAF-V600E驱动的SK-MEL-28细胞的增殖。总之，我们的研究表明，曲格列酮可能是开发分子量小于500 Da的小分子治疗性Gαq抑制剂的一个有前景的新先导物。意义声明：与其他噻唑烷二酮类药物不同，曲格列酮直接结合并抑制异源三聚体G蛋白Gq的活性，对Gi的作用较弱。曲格列酮可能作为一种重新利用的药物支架，用于构建比环缩肽Gαq抑制剂具有更好生物利用度的新型小分子Gαq抑制剂。
