Synthesis and characterization of chemically condensed oligoethylenimine containing beta-aminopropionamide linkages for siRNA delivery.

Polyethylenimines (PEI) are often inefficient in gene knockdown experiments with small interfering RNA (siRNA), presumably due to the strong complexing properties. A more efficient and potentially degradable oligoethylenimine-based carrier was synthesized by the condensation of 800 molecular weight PEI oligomers with hexanedioldiacrylate. Reaction conditions were chosen such that Michael reaction occurs followed by complete N-acylation of all residual ester bonds resulting in beta-aminopropionamide linkage sites and an average molecular weight of 30,000.

Cellular dynamics of EGF receptor-targeted synthetic viruses.

The epidermal growth factor receptor (EGFR) is overexpressed on a high percentage of human carcinomas. EGFR is an attractive therapeutic target for tissue-specific targeting by non-viral vectors in cancer gene therapy. In this study we analyzed and compared the effects of EGFR-targeted and untargeted polyplexes in respect to internalization into EGFR overexpressing HuH7 cells.

Interaction of polyamine gene vectors with RNA leads to the dissociation of plasmid DNA-carrier complexes.

Background: Plasmid DNA (pDNA) dissociation from polyamine gene vectors after cellular uptake has not been well characterized. A more detailed understanding of this process could lead to more efficient gene transfer agents. Since RNA is present in the cytoplasm at high concentrations and due to its structural similarity to DNA, we were interested in its conceivable interaction with polyamine gene vectors.

The internalization route resulting in successful gene expression depends on both cell line and polyethylenimine polyplex type.

Understanding cellular uptake and intracellular processing of nonviral gene delivery systems is a key aspect in developing more efficient vectors. In this study, the impact of clathrin- and caveolae/lipid-raft-dependent endocytosis on cell entry and overall transfection efficiency of polyethylenimine (PEI) polyplexes was evaluated. Most remarkably, the internalization pathway mediating successful transfection depended on both cell type and polyplex type applied.

Cryoconserved shielded and EGF receptor targeted DNA polyplexes: cellular mechanisms.

Recently, cryoconservable polyethylene glycol (PEG)-shielded and epidermal growth factor receptor (EGFR)-targeted polyplexes (EGF+ polyplexes) were engineered in our laboratory for tumor-directed transfer and expression of DNA. Here, we further analyzed specificity and kinetics of EGFR-mediated cellular uptake of these polyplexes. Similar to our previous results, EGF+ polyplexes significantly enhanced the transfection efficiency as compared to polyplexes without EGF (EGF- polyplexes) in HUH-7 hepatoma cells and Renca-EGFR renal carcinoma cells.

Purification of polyethylenimine polyplexes highlights the role of free polycations in gene transfer.

Background: Nonviral vectors based on polyethylenimine (PEI) usually contain an excess of PEI that is not complexed to DNA. Since unbound PEI contributes to cellular and systemic toxicity, purification of polyplexes from unbound PEI is desirable.

Methods: Size exclusion chromatography (SEC) was used to purify PEI polyplexes of free PEI.