Department of Pharmacognosy, Kyoto Pharmaceutical University, Kyoto 607-8412, Japan.
NPR Medical Resource Laboratory, Kyoto 604-0924, Japan.
Toxins (Basel). 2022 Mar 16;14(3):212. doi: 10.3390/toxins14030212.
We examined a two-step target protein binding strategy that uses cofilin as the target protein to analyze the active constituents in . In the first step, we prepared the target protein, and used it to analyze the compounds binding to it in the second step. We used the methanolic extract of as a library of possible active compounds. We conducted LC-MS analysis using information from our previous study. The peaks in the HPLC profile were identified as cucurbitacin D, isocucurbitacin D, and cucurbitacin I. As far as we know, there is no known study of the activity of isocucurbitacin D in this research field. Therefore, we examined the effects of isocucurbitacin D on cell proliferation and cofilin protein in human fibrosarcoma cell line HT1080 to confirm the effectiveness of this strategy. The cytotoxicity assay, the fibrous/globular actin ratio assay, and the immunoblotting analysis revealed that isocucurbitacin D showed a cytotoxic effect with disruption of target protein cofilin. The target protein binding strategy is a direct and straightforward method for finding new drug seeds from crude sources, such as natural plant extracts.
我们研究了一种两步靶蛋白结合策略，该策略使用趋化因子作为靶蛋白，以分析 中的活性成分。在第一步中，我们制备了靶蛋白，并在第二步中用它来分析与它结合的化合物。我们使用 的甲醇提取物作为可能的活性化合物库。我们使用来自先前研究的信息进行了 LC-MS 分析。HPLC 图谱中的峰被鉴定为葫芦素 D、异葫芦素 D 和葫芦素 I。据我们所知，在这个研究领域中，没有关于异葫芦素 D 活性的已知研究。因此，我们研究了异葫芦素 D 对人纤维肉瘤细胞系 HT1080 中细胞增殖和趋化因子蛋白的影响，以确认该策略的有效性。细胞毒性测定、纤维/球状肌动蛋白比测定和免疫印迹分析表明，异葫芦素 D 表现出细胞毒性作用，并破坏了靶蛋白趋化因子。靶蛋白结合策略是一种直接而简单的方法，可从天然植物提取物等粗提物中寻找新的药物种子。
