Nonviral Gene Therapy and its Delivery Systems

Abstract

Nonviral gene therapy has significant clinical potential, yet its therapeutic utility has been hindered by low transfection efficiency due to a combination of extracellular and intracellular barriers. Recent developments in formulation and delivery methodology have allowed a number of advances toward high efficiency gene delivery to various cell types and organs. In particular, the extracellular and intracellular pharmacokinetics of plasmid DNA trafficking are better understood in a number of cell systems. Using cationic lipid or polymers (often with receptor targeting), more than 10(5) plasmids can be delivered to a single cell. Endosomolytic agents promote endosome disruption, and include: weak bases, proton-sponge polymers, fusogenic peptides, viral particles, and photosensitizing compounds. Both classical and nonclassical nuclear localization signal (NLS) peptides have also been tested for enhancement of the probability of nuclear import events, a major rate-limiting step in DNA delivery to nondividing cells. For example, the M9 sequence from heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1) protein, a nonclassical NLS, has been found to increase gene expression level by more than 10 to 150-fold in a variety of cell types. This review will concentrate on the current understandings of the basic mechanisms of nonviral gene delivery and new approaches in the field.

DNA vaccines: are they still just a powerful tool for the future?

“…DNA carriers tested for DNA vaccination are various molecules which complex with DNA by 1) electrostatic forces between negatively charged DNA molecules and a positively charged carrier (or cationic ions on a negatively charged carrier), 2) analogous to the natural DNA-protein interaction, or 3) artificial covalent linkage between DNA and a carrier [81]. The DNA/carrier complexes protect DNA from serum DNases, increase transmission of DNA through the cytoplasmic membrane of target cells, allow targeting to specific tissues, and some of them induce the escape of DNA entrapped in endosomes by promoting endosomal disruption (weak bases such as chloroquine, the proton-sponge effect of many polymers) [69]. Non-viral DNA vaccine delivery systems are based on 1) electrostatic complexation of DNA with cationic polymers (poly-L-lysine, protamine sulfate, polyethyleneimine, chitosan, polyethylene glycol, poly--(D,L-lactide-co-glycolide)), complexes commonly termed DNA/polyplexes, 2) electrostatic complexation and condensation of DNA with artificial cationic lipids or lipopolyamines (DC-chol, DOTMA, DOTAP, DOSPA, DOGS) which are mixed together with zwitterionic helper lipids responsible for the fusion of complexes with the target cell membrane (Chol, DOPE, DPPC), complexes commonly termed DNA/lipoplexes or DNA/lipopolyplexes, 3) complexation of DNA with artificial anionic lipids (DMPG) through electrostatic interaction mediated by Na + and K + ions which are supplemented with zwitterionic helper lipids (DPPC), complexes commonly termed fluid DNA/liposomes, 4) association of DNA with proteins or peptides (histones, peptide tyrosine-lysine-alanine-(lysine) 8 -tryptophan-lysine, Fab fragments of anti-DNA antibodies, cationic viral proteins µ and Vp1) which in combination with cationic polymers or lipids facilitate nuclear targeting, and 5) complexation of DNA with dendrimers with a very low degree of polydispersity (PAMAM), complexes commonly termed DNA/dendrimers (Table 1) [8,68,69,78,84,86,104].…”

“…Liposomes are often used for systemic (intravenous, i.m., i.d.) or topical (nasal, oral) DNA administration [31,43,50,69,74,84]. The final DNA/lipopolyplex structures, DNA concentration, ratio of cationic moiety to DNA, and supplements such as condensing agents, endosmolytic agents, or nuclear targeting molecules are the most critical factors in the transfection efficacy of each formulation [36,50,86].…”

Model system to study classical nuclear export signals

“…In native proteins, NLSs are peptide sequences encoded by DNA that direct the protein to the nucleus of the cell. In the case of gene delivery, NLS peptide sequences are fused to pieces of DNA to be delivered to the nucleus, 2 or fused to cationic lipids or polymers that act as targeting agents or carriers for DNA delivery 3 to enhance delivery to the nucleus. Less well studied by the pharmaceutical field are nuclear export signals (NESs).…”

Tissue specific expression of suicide genes delivered by nanoparticles inhibits gastric carcinoma growth

“…Recently, nonviral vectors, such as nanoparticles, have emerged as one of the most effective vectors for gene transfer. 22,23 Several clinical trials for gene therapy using nanoparticles are ongoing for human genetic disease and cancer. 24,25 Nanoparticles are non-immunogenic, non-inflammatory, and quite applicable in a variety of gene therapy protocols.…”