Poly(Propyleneimine) Transposon-Nanocarriers for Targeted Delivery of Therapeutic DNAs to Tumor Cells

治疗性核酸的递送，如编码自杀基因的dna或使用多阳离子载体大分子的shrna，如完全支链聚（丙烯亚胺）树状大分子（PPI），是治疗癌症的一种有前途的治疗选择。然而，载体复合核酸的全身递送需要大量的DNA，这通常会导致不必要的非特异性离组织和脱靶效应。这尤其如此，因为DNA的短暂表达也意味着反复的全身给药是治疗的强制性。然而，累积毒性的风险预计会增加。一种避免不必要的脱靶效应和提供DNA稳定表达的方法是引入靶向部分（配体，抗体），利用靶向传递的概念，特异性地结合靶细胞并传递治疗基因。在我们的项目中，我们计划研究一种新的靶向转座子-小环dna递送方法，使用功能化的PPIs，在同源细胞受体表面表达的靶细胞中选择性摄取和内体释放。更具体地说，功能化PPIs，结合肿瘤选择性靶向部分，如单链片段变量（scFv）和肽配体，将被用于选择性靶向胶质瘤和前列腺癌肿瘤细胞。治疗基因的稳定表达是通过共同递送睡美人转座酶来提供的。在这项概念验证性研究中，将利用报告基因（EGFP、荧光素酶）、自杀基因（铜绿假单胞菌外毒素PE38）和编码治疗性shrna的dna的基因转移，在体外研究基于选择性ppi的转座子纳米载体平台。因此，该项目的目标是建立一种生物相容性的基于ppi的转座子载体，作为将基因选择性递送到靶细胞的平台。

The delivery of therapeutic nucleic acids, such as DNAs encoding suicide genes or shRNAs using polycationic carrier macromolecules, such as perfectly-branched poly(propyleneimine) dendrimers (PPI), is a promising therapeutic option to treat cancer. However, systemic delivery of carrier-complexed nucleic acids requires huge amounts of DNA which often results in unwanted unspecific off-tissue and off-target effects. This is particularly so since the transient expression of DNA also imply that repeated systemic administration is mandatory for treatment. Yet, the risk of cumulative toxicity is expected to increase. One approach to avoid unwanted off-target effects and to provide stable expression of DNA is the introduction of targeting moieties (ligands, antibodies) that specifically bind to target cells and deliver the therapeutic gene, using the concept of targeted delivery.In our project we plan to investigate a novel targeted transposon-minicircle DNA-delivery approach using functionalized PPIs enabling selective uptake and endosomal release in target cells displaying a surface expression of the cognate cellular receptor. More specifically, functionalized PPIs, coupled to tumor-selective targeting moieties such as single chain fragment variables (scFv) and peptide ligands, will be employed to selectively target glioma and prostate carcinoma tumor cells. The stable expression of the therapeutic genes is provided by co-delivery of a sleeping beauty transposase. In this proof-of-concept study the selective PPI-based transposon-nanocarrier platform will be investigated in vitro using gene transfer of reporter genes (EGFP, Luciferase), a suicide gene (Pseudomonas aeruginosa exotoxin PE38) and DNAs encoding for therapeutic shRNAs. The objective of this project therefore is to establish a biocompatible PPI-based -transposon carriers as a platform for the selective delivery of genes into target cells.