Parallel Synthesis and Screening of Polymers for Non-Viral Gene Delivery

Viruses are most commonly used vectors in gene therapy. However, concerns relating to safety, immunogenicity, and degradation necessitate the discovery of new methods for delivering DNA to cells. While non-viral methods are attractive in gene delivery, they often are characterized by low efficiencies of gene transfer and/or high cytotoxicities. The use of semi-rational and combinatorial techniques can result in the rapid identification of high-efficacy transfection agents. Here we describe the use of parallel synthesis and screening for the discovery of novel, biocompitable cationic polymers as non-viral gene delivery vectors.

We synthesized a library of eighty cationic polymers using ring-opening polymerization reactions between epoxide groups of diglycidyl ethers and the amines of (poly)amines. Polymers were first screened for their DNA binding efficacies using the ethidium bromide displacement assay. Transfection efficacies of lead polymers with high DNA binding efficacies were evaluated using PC3-PSMA human prostate cancer cells and murine osteoblasts. In vitro transfections demonstrated that one candidate polymer showed significantly higher transfection efficacies and lower cytotoxicities than poly(ethyleneimine) (pEI), the current standard for polymeric transfection agents. In addition, polymers that demonstrated moderately higher and comparable transfection efficacies with respect to pEI were also identified. Factors influencing polymer-mediated transfection efficacies were investigated in closely related prostate cancer cell lines. Differences in transfection efficacies were related to differential intracellular trafficking of polymer:DNA complexes in these cells. Strategies for altering intracellular trafficking and therefore, for enhancing transfection efficacies in these cells were investigated. In summary, our studies describe the rapid identification of novel polymers for gene delivery and intracellular trafficking based approaches for enhancing transfection efficacies in cancer cells.