一种肝细胞肝癌特异性靶分子的鉴定及其靶向纳米药物载体的设计

Hepatocellular carcinoma (HCC) has a high morbidity and is difficult to treat due to high incidence of recurrence and metastasis, lacking specific therapy target and severe side effects from chemotherapy. Targeted drug delivery systems that can selectively deliver high dose therapeutic drugs into HCC tumors as well as metastases nodules will undoubtedly take the spotlight in HCC treatment. In our previous work, we biomimetically synthesized a theranostic protein cage nanoprobe with intrinsic tumor targeting property (Nat Nanotech, 2012; PNAS, 2014). More importantly, we recently identified a specific tumor target molecule GRP78 in HCC and found that the expression of GRP78 corelated to the invasion and metastasis of HCC. Based on these findings, this application will design protein cage nanoprobes for HCC diagnosis and treatment, whose protein shell possessing HCC tumor targeting property, while interior cavity being capable of loading of chemotherapeutic drugs and diagnostic probes. Employing HCC subcutaneous tumor and lung metastasis mice models, we will systemically study the molecular mechanisms of GRP78 in specific diagnosis and treatment of HCC, clarify the functions of GRP78 in invasion and metastasis of HCC, and develop the HCC specific targeting theranostic nanocarriers. This study will provide a novel tumor target and new targeting nanocarrier for HCC therapy.

肝细胞肝癌（HCC）因其高复发、高转移的特点，致死率居高不下，目前仍缺乏有效的治疗手段。HCC治疗的难点在于缺乏特异性的肿瘤标志物。申请人在前期工作中，根据仿生原理，制备了具有肿瘤靶向性的笼状蛋白纳米诊疗探针（Nat Nanotech, 2012; PNAS, 2014）。更重要的是，申请人最新发现并鉴定了HCC的一个特异性肿瘤靶分子GRP78，并证实其与HCC的侵袭和转移有直接相关性。据此，本项目将以新鉴定的GRP78靶分子为基础，设计特异性靶向HCC的笼状蛋白纳米探针，其外壳靶向HCC肿瘤细胞，内腔可装载化疗药物或诊断试剂；利用HCC皮下瘤和肺转移模型，系统研究GRP78在HCC靶向诊断和治疗中的分子机制和影响因素，阐明GRP78在肝细胞肝癌的侵袭和转移中发挥的作用；评估特异性靶向HCC的笼状蛋白纳米药物载体的有效性和安全性，为HCC的治疗提供肿瘤新靶点和靶向性纳米药物新载体。

抗肿瘤药物的有效靶向递送是改善肿瘤治疗效率的关键。到目前为止，肝细胞肝癌（HCC）的靶向药物递送仍缺乏有效的手段，鉴定HCC特异性靶点并发展对应的靶向药物递送系统是解决这一问题的关键。本项目基于对HCC特异性靶向肽（SP94）膜受体的分离鉴定技术，通过质谱分析及分子生物学验证，鉴定出膜蛋白GRP78是HCC的特异性肿瘤标志物，并发现其与HCC的侵袭和转移有相关性。基于此，本项目利用铁蛋白独特的笼状结构及自组装特性，将SP94融合展示在铁蛋白表面，发展了一种靶向GRP78的HCC靶向性笼状铁蛋白药物载体（HccFn）。同时，本项目基于铁蛋白的结构特征，开发了一种高效的药物装载新技术，实现了单个铁蛋白装载400个阿霉素分子（Dox）的高装载率。装载药物的HccFn-Dox通过与受体GRP78相互作用，特异性结合到HCC肿瘤细胞上，进而高效内化进入溶酶体，实现药物Dox的高浓度富集、释放及肿瘤杀伤，克服了HCC靶向药物有效递送的瓶颈问题。通过对HCC皮下瘤模型和肺转移癌模型的药效评估，证实了HccFn-Dox是一种有效治疗HCC原位及转移癌的靶向性纳米药物载体。本项目基于以上研究工作，开发了基于HCC靶向的铁蛋白药物新载体，发展了铁蛋白药物装载新方法，建立了HCC靶向铁蛋白载体的标准化制备体系，在国际上发表SCI论文22篇，包括 Nano Today, Nano Letters, Theranostics等，获得授权发明专利1项，全面完成计划书规定的内容和指标。铁蛋白药物载体用于肿瘤靶向治疗是由中国人发现、国际认可并实现成果转化的系统性研究工作，本项目进一步拓展了铁蛋白药物载体的靶向适应症，并应用于肝细胞肝癌治疗，期冀尽快为国民健康做出实际贡献。