A novel Drug Delivery System Using HBV Envelop Protein Particles

使用 HBV 包膜蛋白颗粒的新型药物递送系统

基本信息

批准号：
13470243
负责人：
UEDA Masakazu
金额：
$ 3.07万
依托单位：
Keio University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2001
资助国家：
日本
起止时间：
2001 至 2002
项目状态：
已结题

项目摘要

There are many hepatopathies that may be candidates for gene therapy. Gene therapy needs a vector that is carried together with its genes into a target cell or organ. Desirable vectors should be extremely safe, have good transfection efficiency, and demonstrate a high specificity for their target. In 1992, Kuroda et al. reported on the large amount of HBsAg envelope L particles that could be synthesized in yeast cells. HBV envelope L particles overproduced by yeast are hollow nanoparticles(average diameter 80nm) displaying the pre-S1 peptide indispensable for human liver-specific infection. Either a GFP(green fluorescence protein)-expression plasmid or a fluorescent dye(calcein) was incorporated into L particles by electroporation, and directly used for the transfection of various cells in vitro. Either GFP- or calcein-derived fluorescence was observed only in human liver-derived cells such as NuE and HepG2 not but WiDr and A431. We transplanted human hepatocellular carcinoma cells and human colon cancer cells subcutaneously in nude rats or nude mice. L particles with. GFP-expression plasmide or carcein were infected into the tumor-bearing nude rats or mice through tail vein. After 7 days, fluorescence was observed specifically in the subcutaneous heptocellular carcinoma, but not was observed in the subcutaneous colon cancer, brain, heart, lung, liver, spleen, kidney, adrenal gland, intestine, or muscle of rats or mice. When the human clotting factor IX gene, a therapeutic gene for hemophilia B, was transferred into the xenograft model by the L particles, therapeutic factor IX levels were obtained in the plasma for at least 1 month. In conclusion, HBsAg L particles showed a specific recognition ability for human-derived liver tissue and demonstrated a transgenie ability in an in vivo system.

有许多肝病可能适合基因治疗。基因治疗需要一种载体，将其与其基因一起携带到靶细胞或器官中。理想的载体应该非常安全，具有良好的转染效率，并对其靶点表现出高度的特异性。 1992 年，黑田等人。据报道，酵母细胞中可以合成大量 HBsAg 包膜 L 颗粒。酵母过量产生的HBV包膜L颗粒是中空纳米颗粒（平均直径80nm），显示出人类肝脏特异性感染所必需的前S1肽。通过电穿孔将GFP（绿色荧光蛋白）表达质粒或荧光染料（钙黄绿素）掺入L颗粒中，直接用于体外转染各种细胞。 GFP 或钙黄绿素衍生的荧光仅在人肝源性细胞（如 NuE 和 HepG2）中观察到，但在 WiDr 和 A431 中观察不到。我们将人肝癌细胞和人结肠癌细胞皮下移植到裸鼠或裸鼠体内。 L颗粒带。将表达GFP的质粒或癌素通过尾静脉感染荷瘤裸鼠或小鼠。 7天后，在皮下肝细胞癌中特异性观察到荧光，但在大鼠或小鼠的皮下结肠癌、脑、心脏、肺、肝、脾、肾、肾上腺、肠或肌肉中未观察到荧光。当人类凝血因子IX基因（血友病B的治疗基因）通过L颗粒转移到异种移植模型中时，在血浆中获得治疗因子IX水平至少1个月。总之，HBsAg L颗粒表现出对人源性肝组织的特异性识别能力，并在体内系统中表现出转基因能力。