小分子化合物石蒜碱靶向EGFR抑制神经胶质瘤的分子机制研究

Gliomas are the most common primary brain malignancies of high incidence and fatality rate, which are seriously harmful to human health. Epidermal growth factor receptor (EGFR) has been reported as one of the most important drug target for glioma. Small molecular lycorine, possessing fewer side effect, higher specificity and better effectiveness, can inhibit the growth of a wide range of cancers, and could readily traverse blood-brain barrier. However, lycorine inhibiting glioma has not yet been reported. Here, previous study of the project shows that lycorine has been discovered suppressing glioma cell proliferation, and down-regulating the expression of EGFR. Molecular docking was used for the study of lycorine and intracellular EGFR tyrosine kinase binding. Lycorine has been found binding to tyrosine kinase via hydrogen bond to Asn842, Lys745 and Thr854 in the docking structure. Nevertheless, whether lycorine competitively inhibits ATP to block tyrosine kinase autophosphorylation in the signaling pathway downstream, as well as molecular mechanism of lycorine resisting glioma cell proliferation are still unknown. Structural biology and molecular docking technological means are applied in the project to explore binding sites of lycorine and tyrosine kinase. Molecular mechanism could be explicitly clarified via EGFR mutagenesis cell proliferation assay, ITC and other tests. This will provide theoretical basis for clinical application of lycorine curing glioma targeting EGFR, and plays vital role in novel drug discovery and development for glioma.

神经胶质瘤，一种常见的原发性恶性脑肿瘤，因其高发病率和高死亡率严重危害人类的健康。表皮生长因子受体EGFR是抗神经胶质瘤的重要靶点。小分子化合物石蒜碱具有副作用小、特异性高且药效高等特点，对多种肿瘤具有较好的抑制作用，且能够透过血脑屏障。然而，目前关于石蒜碱抗神经胶质瘤作用尚没有文献报道。本项目前期研究发现，石蒜碱显著抑制神经胶质瘤的生长并通过下调EGFR起作用。分子对接结果显示，石蒜碱与EGFR胞内酪氨酸激酶Asn842、Lys745和Thr854通过氢键结合。但石蒜碱是否竞争性抑制ATP结合酪氨酸激酶阻断其自磷酸化从而抑制神经胶质瘤作用的分子机制尚不清楚。本项目拟采用结构生物学手段、通过细胞增殖实验、免疫组化、western blot 以及ITC等温滴定等实验，阐明其具体的分子机制。这将为石蒜碱靶向EGFR治疗神经胶质瘤的临床应用提供理论依据，对神经胶质瘤新药研发具有重要临床意义。

神经胶质瘤是一种原发性恶性脑肿瘤，并且其复发性和耐药性也是长期存在的问题，因此寻找替代性药物是解决耐药性问题的方法之一。小分子化合物具有特异性高、副作用小且能透过血脑屏障等特点对多种肿瘤具有较好的抑制作用。本项目拟通过计算机模拟进行筛选，并进行实验验证，筛选到小分子化合物石蒜碱、小檗碱以及异鼠李素能够有效抑制神经胶质瘤的生长，并研究其分子机制。（1）石蒜碱依赖于EGFR来抑制神经胶质瘤的生长。并通过长链非编码RNA测序分析研究其分子机制。（2）小檗碱能够有效抑制神经胶质瘤C6细胞的生长和迁移，并通过细胞周期信号通路以及细胞凋亡信号通路起作用。（3）异鼠李素能够抑制神经胶质瘤C6细胞的生长，并且对肿瘤细胞迁移具有较好的抑制作用。本研究首次提出石蒜碱可以抑制神经胶质瘤的生长，并找到其分子机制，这为石蒜碱后期临床试验提供了坚实的理论依据。并且，我们发现小檗碱和异鼠李素对神经胶质瘤具有潜在治疗价值。这为治疗神经胶质瘤成药提供一定的理论依据，也为神经胶质瘤治疗的耐药性提供一些选择方案。