Antitumor Compounds from Cordyceps Militaris

We established a large-scale separation and purification platform to obtain kilogram amounts of natural compounds from the extraction of the fruiting bodies of <i>C. militaris. </i>Seven monomeric compounds, N6-(2-hydroxyethyl) adenosine (HEA), ergosterol (E), ergosta-7,22-diene-3,5,6-triol (EI), 5α,8α-epidioxy-(22<i>E</i>,24<i>R</i>)-ergosta-6,22-dien-3β-ol (ED), ergosta-7,22-dien-3β,5α-dihydroxy-6-one(EO),(20<i>S</i>,22<i>E</i>,24<i>R</i>)-Eegosta-7,22-dien-3β,5α,6β,9α-tetraol22<i>E</i>,24<i>R</i>-ergosta-7,22-diene-3β,5α,6β,9α-tetraol (ET), and (24<i>S</i>)-5,22-stigmastadien-3β-ol (SE), were harvested using different solvents, and the structure of each compound was identified. The activities and functions of the isolated compounds were tested by label-free, real-time cell analysis methods at the cellular level, and their antitumor effects were verified using mouse models of Lewis and H22 tumors. The anti-insomnia effect of HEAadenosine was tested in an anti-insomnia mouse model. The interactions between E and 8 A549 cell proteins were determined. The biosynthetic pathways of HEA and E, which possess pharmacologically active monomers, were determined. This platform can provide a theoretical basis for the further development and discovery of novel natural medicines.

本研究构建了一套大规模分离纯化平台，可从蛹虫草（*C. militaris*）子实体的提取物中获取千克级天然化合物。本平台通过不同溶剂体系分离得到7种单体化合物，分别为：N⁶-(2-羟乙基)腺苷（HEA）、麦角固醇（E）、麦角甾-7,22-二烯-3,5,6-三醇（EI）、5α,8α-过氧桥-(22*E*,24*R*)-麦角甾-6,22-二烯-3β-醇（ED）、麦角甾-7,22-二烯-3β,5α-二羟基-6-酮（EO）、(20*S*,22*E*,24*R*)-麦角甾-7,22-二烯-3β,5α,6β,9α-四醇（ET）、(24*S*)-5,22-豆甾二烯-3β-醇（SE），并对所有化合物的结构完成了鉴定。本研究采用无标记实时细胞分析技术，在细胞水平上对上述分离得到的化合物的活性与功能进行了测试；通过Lewis肿瘤小鼠模型与H22肿瘤小鼠模型验证了其抗肿瘤效果。针对N⁶-(2-羟乙基)腺苷（HEA）的抗失眠活性，本研究通过失眠小鼠模型开展了验证实验。此外，本研究还解析了麦角固醇（E）与A549细胞中8种蛋白质的相互作用，并阐明了具有药理学活性的单体化合物HEA与麦角固醇（E）的生物合成途径。该大规模分离纯化平台可为新型天然药物的研发与发现提供理论基础。