Department of Chemical and Structural Biology, Weizmann Institute of Science, 7610001 Rehovot, Israel.
Department of Chemistry and Chemical Biology, TU Dortmund University, Otto-Hahn-Strasse 4a, 44227 Dortmund, Germany.
J Med Chem. 2022 Aug 11;65(15):10341-10356. doi: 10.1021/acs.jmedchem.1c02206. Epub 2022 Jul 30.
High-throughput nanomole-scale synthesis allows for late-stage functionalization (LSF) of compounds in an efficient and economical manner. Here, we demonstrated that copper-catalyzed azide-alkyne cycloaddition could be used for the LSF of covalent kinase inhibitors at the nanoscale, enabling the synthesis of hundreds of compounds that did not require purification for biological assay screening, thus reducing experimental time drastically. We generated crude libraries of inhibitors for the kinase MKK7, derived from two different parental precursors, and analyzed them the high-throughput In-Cell Western assay. Select inhibitors were resynthesized, validated conventional biological and biochemical methods such as western blots and liquid chromatography-mass spectrometry (LC-MS) labeling, and successfully co-crystallized. Two of these compounds showed over 20-fold increased inhibitory activity compared to the parental compound. This study demonstrates that high-throughput LSF of covalent inhibitors at the nanomole-scale level can be an auspicious approach in improving the properties of lead chemical matter.
高通量毫摩尔尺度合成能够高效经济地对化合物进行后期功能化（LSF）。在这里，我们证明了铜催化的叠氮-炔环加成反应可用于在毫摩尔尺度对共价激酶抑制剂进行 LSF，从而能够合成数百种无需纯化即可用于生物测定筛选的化合物，从而大大缩短了实验时间。我们生成了两种不同母体前体衍生的激酶 MKK7 的抑制剂粗文库，并通过高通量细胞内 Western 分析进行了分析。选择的抑制剂通过传统的生物学和生化方法（如 Western blot 和液相色谱-质谱（LC-MS）标记）进行重新合成和验证，并成功进行了共结晶。其中两种化合物的抑制活性比母体化合物提高了 20 多倍。这项研究表明，毫摩尔尺度的共价抑制剂的高通量 LSF 可能是改善先导化学物质性质的一种有前途的方法。
