Medicinal chemistry of Gq proteins and their inhibitors: synthesis, computational calculations and (bio)analytical studies

天然产物FR900359是一种选择性抑制Galphaq蛋白的沉积肽，使用氚气氢化，得到一个名为[3H]PSB 15900的新化合物。我们建立了放射性配体结合实验，发现放射性配体[3H]PSB 15900在低纳摩尔范围内对天然Galphaq蛋白具有高亲和力。在本项目中，四种人类Galphaq蛋白亚型将在中国仓鼠卵巢（CHO）细胞中稳定过表达，而CHO细胞的Gq蛋白表达水平非常低。这些细胞系将用于研究放射性配体和其他新分离和合成的化合物的选择性。将研究离子、核苷酸、脂质和蛋白质对放射性配体结合的调节。该放射配体将用于测定生理和病理条件下各种细胞、组织和器官中Galphaq的表达水平。通过利用化合物文库的筛选方法，我们希望鉴定结构新颖的Galphaq抑制剂。此外，我们将优化LCMS方法来定量体内研究获得的生物样品中的FR900359，并确定其代谢产物。

The natural product FR900359, a depsipeptide which selectively inhibits Galphaq proteins, was hydrogenated using tritium gas to provide a new compound designated [3H]PSB 15900. We established a radioligand binding assay, and found that the radioligand [3H]PSB 15900 displays high affinity in the low nanomolar range for native Galphaq proteins. In the present project the four human Galphaq protein subtypes will be stably overexpressed in Chinese hamster ovary (CHO) cells which were shown to express very low levels of Gq proteins. These cell lines will be used for investigating the selectivity of the radioligand and of other, newly isolated and synthesized compounds. Modulation of radioligand binding by ions, nucleotides, lipids and proteins will be studied. The radioligand will be used to determine Galphaq expression levels in various cells, tissues and organs under physiological and pathological conditions. With a screening approach utilizing compound libraries we want to identify structurally novel Galphaq inhibitors. Moreover we will optimize an LCMS method to quantify FR900359 in biological samples obtained from in vivo studies, and determine its metabolites.