EGCG在纳米材料介导下增强其抗肿瘤疗效及活体中靶向治疗乳腺癌的研究

(-_Epigallocatechin-3-galate (EGCG) is a good water-soluble anticancer drug that derivates from green tea as the major components of biological activity of tea polyphenols.Comparing to other anticancer drugs, EGCG is almost no side effect to normal cells. However, it also has disadvantages such as poor liposolubility, low bioavailability and shorter half-life.In cells culture medium, EGCG could auto-oxidation, with a half-life of 30min-2h, that influnce its healing effect on cancer cells and in vivo experiments. The auto-oxidation of EGCG produce EGCG dimers, hydrogen peroxide (H2O2) and other oxidative products. Based on prior experiments，we designed a drug carrier, not only enlarged the half-life of EGCG to enhance drug effect but also trained the position of EGCG in cancer cells by self-coating fluorescent dye(FITC). The drug carrier is a multiple core-shell mesoporous SiO2 (CMS) that could be functionalized to effectively prolong half-life and enhance the anticancer ability of EGCG to inhibit HeLa cells growth and proliferation.The functionalization of the CMS surface with amino groups and pores allowed electrostatic attracting of EGCG, alleviating the interaction of free radicals for producing dimers or other polymers, to the surface and pores. Based on the results of MTT assay, cell cycle analysis and western blot, the CMS acted as a carrier to prolong the half-life and enhance the anticaner effect of EGCG. In addition, fluorchrome isothiocyanate (FITC) was entraped in the core of CMS, which endowed the CMS to contemporaneously denote the position of EGCG in cells. The load of EGCG by CMS in cancer cells was capable of increasing the anticancer ability of EGCG exceeding that of free EGCG being treated the HeLa cells in the absence of CMS. These results demonstrate that CMS plateform can effectively prolong half-life of EGCG and improve its pharmacodynamic action and it is possible to use the CMS plateform as a promoter to improve anticancer ability of the unstable chemotherapeutic agent. The surfaces of CMS were also modified cyclic nonapeptide(PEGA peptide) and cell penetrateing peptide(CPPs peptide) in shells. The design could enhance the inhibit action of EGCG to tumor cells and specificly distinguish the breast cancer from all tissues by PEGA and CPPs peptides that bestowed EGCG to target and perish breast cancer cells and treat breast cancer. The drug carrier reduces the cycle length of EGCG in vivo, decreases the degradation of EGCG and achieve the purpose of enhancing the inhibiting ability of EGCG to cancer. The accomplishment of the project will supply a new idea to enhance drug efficency of anti-cancer that have strongly academic and practice directive sences.

表没食子儿茶素没食子酸酯（EGCG）是一种很好的抗肿瘤药物并对人体的正常细胞和组织几乎没有损伤，但EGCG不稳定、代谢快，半衰期只有30分钟至两小时左右，影响了其在细胞实验及活体中的疗效。本项目根据前期实验设计了一种药物载体，既能提高EGCG的半衰期，还能通过自身包覆的荧光染料（FITC）示踪EGCG作用于肿瘤细胞的位置。该药物载体是由核内包覆FITC、壳上修饰透膜小肽（CPPs）和导引胜肽（PEGA肽）的核-壳介孔SiO2构成。借助该载体不仅可以增强EGCG对肿瘤细胞的抑制作用，而且SiO2表面修饰的小肽还能特异性识别乳腺癌，引导SiO2通过血管系统到达乳腺癌发生的部位，靶向消灭乳腺癌细胞，治疗乳腺癌。该载体的设计充分降低了EGCG在活体内的循环时间，减少了EGCG的降解，实现了增强其抑制肿瘤疗效的目的。本项目的完成，将提供一条增强抗肿瘤药物疗效的新思路，具有很强的理论和实践指导意义。

化疗是借助化学药物来抑制癌细胞的增殖、入侵和转移的癌症治疗方法，但化疗不但消灭癌细胞，对正常细胞也有很大损伤。因此，应大力研制和提取无副作用的抗癌药物。表没食子儿茶素没食子酸酯（EGCG）是绿茶的主要活性成分，在抗癌及DNA保护方面具有很好的药用价值。然而，如果直接口服的话，EGCG在体内会很快的代谢掉。因此，我们在该课题中设计了一个药物释放体系来负载EGCG从而增强EGCG在细胞及活体中的稳定性、靶向性及抗癌效果。该药物释放体系主要包括三部分：抗癌药物—EGCG；药物载体—胶体介孔二氧化硅（CMS）；靶向配体—肿瘤归巢细胞透膜肽（peptide）。根据CCK-8实验、共聚焦成像、细胞周期分析和Western blot的实验结果，我们可以得出以下结论：将EGCG负载在CMS及CMS@peptide中可增强EGCG在细胞及活体中的抗癌效果。细胞实验中，考查了三个时间点不同浓度EGCG、CMS@EGCG及CMS@peptide@EGCG对细胞的抑制率，结果显示CMS@peptide@EGCG组的抑制效果最好。EGCG是通过在细胞中产生活性氧诱导细胞凋亡的，活性氧探针的共聚焦成像结果显示CMS@peptide@EGCG处理过的细胞活性氧水平明显高于其他组。活体实验中，CMS对荷瘤小鼠没有明显的影响，单独的EGCG对肿瘤的抑制率为69.9%，但CMS@peptide@EGCG对肿瘤的抑制率高达89.66%。H＆E染色显示所有实验组的动物都没有出现组织损伤。.本研究结果证明EGCG是一种高效的无副作用的抗癌药物，CMS@peptide可以通过靶向聚集和定点释放极大的促进EGCG对乳腺癌的治疗效果，因此CMS@peptide作为药物输送体系的载体应用前景广阔。