槐糖脂的结构修饰及其联合依托泊苷对食管癌细胞增殖的影响和机制研究

Sophorolipids (SLs) are the most promising biosurfactant to substitute the conventional chemical surfactants. Preliminary works showed that SLs performed significant surface activity and antitumor activity, and different SLs structure have different antitumor effects or physicochemical properties. Etoposide (ETO) is an antineoplastic drug that has been extensively used to couple DNA damage to apoptosis in a variety of cell types. The major bottlenecks in its clinical application are drug tolerance, normal tissue toxicity and produced adverse reactions. To some extent, combination drug therapy can solve the above problems. Therefore, the project intends to select the SLs produced from pretreated rice straw by W. domercqiae as study object. SL is modified by chemical and biological methods to obtain different SL derivatives firstly. Then, experiments are carried out to screen SL derivatives with excellent anticancer effect or physicochemical property rapidly . The corresponding relations of structure and property are also illustrated. After esophageal cancer animal model is established, effects and mechanisms of drug therapy of SL derivatives combination with ETO would be investigated. Meanwhile, substitution effects of SL derivatives in place of Tween-80 in ESE are studied. The mentioned research results could explore new application field of SLs and provide important evidiences for cancer treatment.

槐糖脂（SLs）是一类重要的生物表面活性剂，前期研究发现其具有优秀的表面活性和抗肿瘤活性，且不同结构SL的理化/生物活性不同。依托泊苷（ETO）是国际公认的广谱抗肿瘤药物，长期用药后的耐受性、对正常组织的毒副作用及易产生不良反应是其在临床治疗上难以克服的瓶颈。联合用药可一定程度解决上述问题。本项目拟以拟威克酵母变种（W. domercqiae）所产SLs为研究对象，首先利用生物酶法和化学法对SL修饰改性获得不同类组衍生物；然后快速筛选出抗肿瘤/理化活性优秀的SL衍生物并探讨结构与抗肿瘤/理化活性的对应关系；随后建立食管癌移植瘤模型，考察抗肿瘤活性优良的SL衍生物与ETO联合用药的效果和作用机制；考察理化活性优良的SL衍生物在ETO亚微乳（ESE）中的Tween-80替代效果来降低不良反应的发生率；通过上述研究开拓SLs应用新领域并探索肿瘤治疗新策略。

槐糖脂作为一类表面活性/乳化能力/抗肿瘤活性优良的生物表面活性剂,其在新型抗肿瘤药物和药用辅料的探索开发中存在巨大开发潜力和应用前景,值得科研工作者关注。本项目主要研究内容和成果概述如下：.1. 廉价底物产槐糖脂研究：采用SO3微热爆的预处理方法处理稻草秸秆制备糖化液；考察用预处理后糖化液替代葡萄糖，用单细胞油替代油酸降低槐糖脂生产成本的可行性；在实验室小试基础上，采用50L发酵罐对槐糖脂发酵生产进行了中试放大；对大规模（300吨/年）槐糖脂的发酵生产与分离工艺流程进行了设计，上述工作为今后开展以木质纤维素为底物大规模生产槐糖脂提供理论基础和技术手段。.2. C18:1 DLSL及其衍生物的制备及活性筛选：成功利用沉降-醇沉法结合柱层析大量制备获得C18:1 DLSL；利用酶法联合化学法成功制备了4种乙酰基化程度不同及4种不同氨基酸酯化的槐糖脂衍生物，并对其进行了结构鉴定。以吐温-80为参比，MTT法体外检测结果提示双乙酰基内酯型槐糖脂抑制食管癌细胞增殖效果最好，双乙酰基酸型槐糖脂降低表面张力的能力最强。.3. 槐糖脂联合依托泊苷抗食管癌增殖作用及机制研究：MTT法体外实验结果显示槐糖脂联合依托泊苷可以显著抑制食管癌细胞增殖；与两药单独使用相比，联合用药抑制食管癌细胞增殖的作用明显增强；动物体内实验提示联合用药对食管癌小鼠移植瘤有更好的抑制作用，低、中、高剂量联合给药组的抑瘤率均较单独给药组高，且对小鼠的毒性更小；联合用药主要通过促进食管癌细胞的凋亡达到提高药物的体内外抗肿瘤效果的目的。.4. 依托泊苷-槐糖脂亚微乳的制备及评价：采取高压均质法成功制备不同依托泊苷-槐糖脂亚微乳；单因素实验考察了槐糖脂结构与用量、油相组成和用量、豆磷脂用量、体系pH值等对亚微乳制剂的物理稳定性、化学稳定性及载药率的影响；结果显示使用槐糖脂替代吐温-80制备新型亚微乳是可行的，且所得亚微乳理化性质稳定，毒副作用低，载药量高；槐糖脂具有被开发成药用辅料的潜力。