Abstract
BackgroundGene therapy is most effective when delivery is both efficient and safe. However, it has often proven difficult to find a balance between efficiency and safety in case of viral or polymeric vectors for gene therapy. Peptide based delivery systems may be attractive alternatives but their relative instability to proteolysis is a major concern in realizing their potential application in biomedical sciences. In this work we report gene delivery potential of nanoparticles (Nps) synthesized from cationic dipeptides containing a non-protein amino acid α, β-dehydrophenylalanine (∆Phe) residue.MethodsDipeptides were synthesized using solution phase peptide synthesis method. Nps were formed using self-assembly. Nps were characterized using light scattering, electron microscopy. Transfection efficiency was tested in hepatocellular carcinoma (HuH 7) cells.ResultsThe cationic dipeptides condensed plasmid DNA into discrete vesicular nanostructures. Dipeptide Nps are non-cytotoxic, protected the condensed DNAs from enzymatic degradation and ferried them successfully inside different types of cells. GFP encoding plasmid DNA loaded dipeptide Nps showed positive transfection and gene expression in HuH 7 cells.ConclusionsThe cationic dipeptide Nps can successfully deliver DNA without exerting any cytotoxic effect. Owing to their simple dipeptide origin, ease of synthesis, enhanced enzymatic stability as well unmatched biocompatibility, these could be successfully developed as vehicles for effective gene therapy.
Highlights
Gene therapy is most effective when delivery is both efficient and safe
Nanoparticles (Nps) loaded with plasmid DNA may serve as efficient sustained release gene delivery systems due to their rapid escape from the degradative endo-lysosomal compartment to the cytoplasmic compartment [7]
Materials Materials used in this study are: THF, N-methyl morpholine (NMM), DMF, piperidine, DIPCDI, Isobutyl chloroformate (IBCF), TIPS, Trifluoroacetic acid (TFA), HFIP, phenol, DL-threophenylserine, MTT (Sigma-Aldrich, Munich, Germany) Boc-Arg(Mtr)-OH, Boc-Lys(Boc)-OH (Novabiochem, Merck, Darmstadt, Germany) sodium acetate, ethyl acetate, acetonitrile (Spectrochem Pvt Ltd, Mumbai, India), anhydrous sodium sulfate, citric acid (Merck, Munich, Germany), human ovarian cancer (HeLa), hepatocellular carcinoma (HuH-7) and human dermal fibroblast (L929) cells were obtained from American Type Culture
Summary
Gene therapy is most effective when delivery is both efficient and safe. it has often proven difficult to find a balance between efficiency and safety in case of viral or polymeric vectors for gene therapy. Most early applications of gene therapy were concentrated around viral vectors for efficient gene delivery. Many of these vectors raised undesirable immune responses, hindering their therapeutic effect [6]. Polycationic polymer based Nps as non-viral gene delivery vectors, have been developed and polyethylenimines is currently the most popular polymer used to deliver genes into various cell types, including neurons. Polyethylenimines is able to condense genes into small Nps and protect the DNA from degradation by nucleases Polymeric complexes such as PLGA– polyethylenimines Nps have been demonstrated as new delivery systems to carry genes to the lung epithelium [16]. Cationic bovine serum albumin conjugated with poly(ethyleneglycol)–poly(lactide) nanoparticle, was developed as a promising brain drug delivery carrier with low toxicity [17]
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