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蛋白的去脂肽，用氚气体氢化得到一种新的化合物，命名为[~3H]PSB 15900。我们建立了放射性配基结合分析，发现放射性配基[~3H]PSB 15900与天然Galphaq蛋白在低纳摩尔范围内表现出高亲和力。在本项目中，这四种人类Galphaq蛋白亚型将在中国仓鼠卵巢(CHO)细胞中稳定过表达，中国仓鼠卵巢细胞被证明表达非常低水平的GQ蛋白。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、脂类和蛋白质对放射性配体结合的调节作用。放射性配基将用于在生理和病理条件下确定Galphaq在各种细胞、组织和器官中的表达水平。通过使用化合物文库的筛选方法，我们希望确定结构上的新型Galphaq抑制剂。此外，我们将优化LC MS方法，以定量从体内研究获得的生物样品中的FR900359，并测定其代谢物。

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.