基于粒径程序化调控策略构建的抗肿瘤中药脂质微乳复合系统及其瘤内聚集渗透研究

Particle size of a nano-sized drug delivery system is the most important factor of accumulation and penetration in tumor tissues. Based on our previous studies, the intermediate-sized microemulsions were capable of efficient accumulation, but the smaller ones were good at deep penetration within tumor tissue. The problem we face is that a conventional vehicle could not possess two types of optimal sizes simultaneously. To overcome this issue, a lipid-microemulsion complex capable of cleavage layer by layer triggered through tumor microenvironment is being developed using programmed assembling strategy. Such coix seed oil - tripterine microemulsion-based complex system with intermediate size is coated with thermo-sensitive lipid and modified with furin-cleavable PEG shell. After accumulation in tumor sites, the PEG shells would seprate from particles under furin-rich stimulation and then the lipid shells could become unstable in the tumoral thermal environment to realize deep tumor penetration performed by small-sized coix seed oil - tripterine microemulsion. Moreover, this study will also focus on the mechanism of sequential detachment, the efficacy of accumulation and penetration in the tumor sites, and antitumor ability. It offers a new sight and technology to effectively improve the antitumor efficacy of nano-sized anticancer Traditional Chinese Medicine through resolving a contradiction of inconformity in optimal size of accumulation and penetration in the tumor sites.

粒径是决定中药纳米制剂在肿瘤组织聚集和渗透的最关键因素。基于以往对不同粒径的薏苡仁油-雷公藤红素微乳在宫颈癌组织聚集与渗透规律的研究发现，大粒径利于聚集，而小粒径利于渗透，但常规递药系统难以同时实现两种过程所需的最佳粒径，极大地限制了抗肿瘤药效的发挥。针对上述问题，本项目采用粒径程序化调控策略将两种最优粒径参数设计至同一纳米粒中，构建可被肿瘤微环境触发的“逐级蜕壳”式脂质微乳系统：该系统由热敏脂质材料包封小粒径薏苡仁油-雷公藤红素微乳，其外修饰对宫颈癌furin蛋白敏感的PEG层，组装成大粒子；该系统先在宫颈癌组织高效聚集，后由furin蛋白及微热环境刺激蜕去PEG及脂质外壳，释放出小粒径微乳继续完成组织深层渗透。本项目还将对其逐层蜕壳机制、肿瘤聚集及渗透效率、抗肿瘤药效等进行深入研究，以解决中药纳米制剂在肿瘤组织聚集与渗透尺寸矛盾的问题，为真正提高中药纳米制剂抗肿瘤疗效提供新思路与方法。

纳米粒子的粒径是决定中药纳米递药系统在肿瘤部位聚集及渗透的关键性因素。通常大粒子有利于其在肿瘤组织聚集，而小粒子利于深层渗透。但常规单一粒径纳米系统难以满足上述粒径要求，极大地限制了抗肿瘤药效的发挥。针对上述问题，本项目构建了一种中药脂质微乳复合系统（PEGcleavableTf-CTM/L），该脂质系统外部修饰宫颈癌furin蛋白敏感肽-PEG外壳，粒径为115nm，Zeta电位为-47mV，内部包裹粒径为40nm的转铁蛋白修饰的薏苡仁油-雷公藤红素微乳（Tf-CTM），雷公藤红素包封率及载药量分别为73.40%及0.3292%，稳定性良好。该系统在42℃孵育下，温敏脂质释放药物，可被HeLa细胞摄取，具有良好的抗HeLa细胞增殖和促凋亡作用。该系统凭借furin蛋白敏感肽-PEG外壳和尺寸效应可在肿瘤组织高效聚集，之后释放出的小粒径Tf-CTM可深层渗透至3D HeLa细胞瘤球及肿瘤组织。药动学研究表明PEGcleavableTf-CTM/L与裸药相比AUC提高了近2倍，显著提高了雷公藤红素体内生物利用度。体内抗肿瘤药效表明PEGcleavableTf-CTM/L-42℃组抗肿瘤药效最好，抑瘤率为80.27 ± 9.21%。肿瘤组织切片的H&E、TUNEL、Ki67染色均表明PEGcleavableTf-CTM/L-42℃对肿瘤杀伤、凋亡作用最强。并通过上调IFN-γ、IL-2和IL-12A的表达，及下调CCL2、IL-6、IL-10、TNF-α和TGF-β的表达发挥抗肿瘤作用。该系统肝、肾功能等生化指标均在正常范围以内，心、肝、脾、肾、肺等器官组织切片均未见明显异常，制剂安全性良好。本项目的研究意义在于通过智能调控中药纳米制剂粒径，为同时实现中药多组分纳米制剂高效肿瘤组织聚集与深层渗透、精准定位，真正提高中药多组分纳米递药系统抗肿瘤疗效提供新思路和新方法。