Gene delivery mediated by synthetic oligopeptide

已知的是，质粒DNA与多种阳离子多肽和脂类形成络合物，已被探索作为DNA在哺乳动物细胞中的可能载体。我们合成了由赖氨酸(K)、色氨酸(W)和半胱氨酸(C)等9个氨基酸残基组成的寡肽及其以二硫键为载体的对称性二聚体。PDNA(PGL3)/寡肽复合体对HeLa S3的转染率较低，但对HeLa S3的细胞毒性较小。由胆固醇衍生物(DMB-Chol)和二油基磷脂酰乙醇胺(DOPE)组成的DNA/寡肽/脂质体三元体系的有效基因表达(10 8~109水平，RLU/min/mg蛋白)是相应的PDNA/寡肽复合体的10 4~10 5倍。在10%的血清存在下，三元络合物维持在10^7水平。溴化乙锭排斥研究表明，三元络合物与PDNA的亲和力远大于相应的PDNA/寡肽络合物。质粒对DNaseI降解的敏感性表明，三元络合物能够很好地保护其免受消化。人工合成的寡肽是阳离子脂质体介导的药物转染剂的潜在促进剂。这些发现对体内成功的转基因具有一定的指导意义。

Plasmid DNA is known to form complexes with a variety of cationic peptides and lipids, which have been explored as possible carriers for DNA transfection in mammalian cells. We synthesized, oligopeptides consisting of nine amino acid residues including lysine (K) tryptophan (W), and cysteine (C), and also their symmetrical dimmers with a disulfide bond as possible carriers. The pDNA (pGL3)/oligopeptide complexes generally showed poor transfection efficiencies but little cytotoxicity for HeLa S3. The ternary system of DNA/oligopeptide/liposome containing cationic liposome formulated from the cholesterol derivative (DMB-Chol) and dioleylphosphatidylethanolamine (DOPE) showed 10^4-10^5 fold greater effective gene expression (10^8-10^9 level, RLU/min/mg protein) than those of the corresponding pDNA/oligopeptide complexes. In the presence of 10% serum, the ternary complexes were maintained at 10^7 level. The ethidium bromide exclusion studies showed the ternary complexes have much greater affinity to pDNA than the corresponding pDNA/oligopeptide complexes. Plasmid sensitivity against DnaseI degradation showed that the ternary complexes were well protected from the digestion. Synthetic oligopeptides are active as potential enhancers for DOPE-containing cationic liposome-mediated transfection. These findings have implications for successful in vivo transfection.