化疗与抗体联合治疗的双靶向重组蛋白大分子药物的构建与评价

In view of the maldistribution and less prominent of targets cancer cells due to tumor heterogeneity, the low therapeutic index, insufficient targeting of cancer drugs, the high drug accumulation in healthy tissues especially liver and spleen for the present nano-drug delivery system, the high preparation cost, hard modification and drug resistance of conventional antibodies, we are proposed to adopt gene engineering technology to recombine the targeting nanobodies and the targeting short peptides with the elastin-like peptides to form EGFR and αv integrins dual-targeting recombinant protein. With further introduction of hydrazone bond into the recombinant protein structure and conjugation with the chemotherapeutic drug doxorubicin (DOX), it will result a dual-targeting recombinant protein-based macromolecule prodrug platform which combines chemotherapy with antibody. Targeting groups carrying chemotherapeutic drugs at tumor site-specific enrichment The dual-targeting ability of the macromolecular prodrug and the related influencing factors were studied in vitro and in mice subcutaneous tumor models. And further biosafety and the anti-cancer efficacy of the dual-targeting recombinant protein-based macromolecular prodrug. The dual-targeting vector designed in this project achieve the specific active targeting of the drug to the tumor, increase drug accumulation in the tumor site and meanwhile reduce the drug content in normal tissue especially in liver and spleen and therefore improve the therapeutic index of the drug. This study will provide a basis for the development of safe, effective new drug carriers and improvement of the drug targeting ability.

针对肿瘤异质性导致的靶点分布不均及不突出，肿瘤治疗中药物治疗指数较低、靶向性不够，现存的纳米载药系统在正常组织中特别是肝脾中的含量较多，传统抗体制备成本较高，不易改造以及耐药性的问题，我们拟采用基因工程技术，将靶向纳米抗体和靶向短肽与类弹性蛋白重组形成EGFR和αv整合素双靶向的重组蛋白，并进一步在重组蛋白结构中引入腙键，连接上化疗小分子药物阿霉素（DOX），得到化疗与抗体联合的双靶向重组蛋白大分子药物平台。通过体外实验以及小鼠皮下肿瘤模型研究大分子药物的双靶向性及其相关的影响因素，并进一步研究双靶向重组蛋白大分子药物的生物安全性以及抗肿瘤能力。本项目利用双靶向载体的设计实现药物DOX对肿瘤的特异性主动靶向，提高药物在肿瘤部位的累积量，同时降低药物在正常组织特别是肝脾中的含量，提高药物的治疗指数。本研究将为开发安全有效的新型药物载体，提高药物的靶向性提供依据。

尽管化疗以及靶向疗法在肿瘤领域应用广泛，但是1）肿瘤的发生发展是一个不断异质化的过程，肿瘤细胞表面的受体表达不均匀，单一靶向难以实现有效靶向；2）肿瘤复杂的微环境导致单一的化疗难以实现良好的抗肿瘤效果。针对这些问题，本项目开展了化疗与抗体联合治疗的双靶向重组蛋白大分子药物的构建与评价工作。利用基因工程技术和分子生物学手段，以纳米抗体和配体代替传统抗体分子，构建了基于类弹性蛋白的双靶向重组蛋白，并将基于抗体的重组蛋白与化疗药物偶联，合成了化疗与抗体联合治疗的双靶向重组蛋白大分子药物。双靶向重组蛋白大分子药物改善了药物的体内分布和代谢行为，双靶向的结构设计提高了药物对肿瘤细胞的靶向性，通过生物学环境的调节改善了肿瘤的力学微环境，打破了致密的细胞外基质，促进了所携带的化疗药物在肿瘤中的累积与分布，显著提高了药物的抗肿瘤治疗效果。本项目研究工作的开展为开发双/多靶向的基于蛋白的大分子药物提供了新的思路，具有良好的应用前景。项目执行以来，我们在国内外学术期刊上发表SCI相关论文4篇，包括Advanced Materials 1篇，获得授权专利1项，申请专利3项。