Gene delivery using liposomes

H. Kikuchi, N. Suzuki, K. Ebihara, H. Morita, Y. Ishii, A. Kikuchi, S. Sugaya, T. Serikawa, K. Tanaka, Gene delivery using liposome technology, J. Control. Release, 1999, 62, 269-277. 

Meyer O, et al. Cationic liposomes coated with polyethylene glycol as carriers for oligonucleotides. J Biol Chem 1998 273 15621 Shi G, et al. Efficient intracellular drug and gene delivery using folate receptor-targeted pH-sensitive liposomes composed of cationic/anionic lipid combinations. J Control Release 2002 80 309. 

Folate-linked liposomes showed efficient FR-dependent cellular uptake and transfection in vitro but have not delivered plasmid DNA successfully in vivo as part of gene therapy experiments (Hofland et al. 2002 Reddy et al. 2002). The major limitation of in vivo gene therapy using liposomes is the low transfection efficiency. A size for gene transfer vectors of less than 300 nm is required for extravasation and intratumoral diffusion, which could be limiting for targeted delivery. 

Hattori, Y., Kawakami, S., Suzuki, S., Yamashita, E, and Hashida, M. 2004. Enhancement of immune responses by Dna vaccination through targeted gene delivery using mannosylated cationic liposome formulations following intravenous administration in mice. Biochemistry and Biophysics Research Communication, 317,992-999. 

Formation of a complex between DNA and polycationic compounds appears to be the initial and quite possibly a critical parameter for nonviral gene delivery. Several synthetic vector systems, which are generally cationic in nature, including poly(lysines), cationic liposomes or various types of block copolymers and recently dendrimers, have been shown to self-assemble with plasmid DNA [13-15] [16]. Specific physicochemical properties manifested by these DNA complexes depend on the type of cationic agent used however, interesting patterns for such interactions are beginning to evolve [17, 18]. Under certain conditions, the interaction of DNA with polyvalent cations results in... 

Some investigators have used electrical current (electroporation) to improve DNA (or drug) entry into tumor cells with some preliminary success. Liposomes are attractive vehicles for gene delivery, since they can carry plasmid, antisense, or viral DNA. Compared with viral approaches, however, liposomes remain relatively inefficient at facilitating gene transfer, although their safety profile remains more desirable. Some of the attributes and limitations of the nonviral methods are listed in Table 58.1. 

Answer Aerosol delivery of the CFTR gene. Both viruses and liposome-DNA complexes are capable of successful CFTR gene transfer to the nasal and airway epithelia of patients with CF. In fact, gene transfer to the airways is one of the few areas where liposome-DNA complexes match the expression obtained using viral vectors without the viruses inflammatory side effects. Current trials are aimed at optimizing gene delivery with reduced toxicity to produce sustained correction of the epithelial transport defect. 

SW CNT functionalized with DNA showed a 10 times more effective penetration and expression of genes in vitro, in comparison with molecular DNA. Other charged macromolecules such as polypeptides and liposomes, can provide more effective transport, but they can cause destabilization of the cellular membrane exhibiting a cytotoxic effect. Whereas, using nanotubes for gene delivery has not caused any cytotoxic effects. 

Successful gene delivery by use of cationic liposomes requires the following conditions (134) (1) condensation of DNA into the complex and its protection from degradation by intracellular nucleases (2) adhesion of DNA-lipid complex onto the cellular surface (3) complex internalization (4) fusion of an internalized DNA-cationic liposome complex with the endosome membrane (5) escape of DNA from the endosome (6) entry of DNA into the nucleus followed by gene expression. 

Zuidam, N.J., Barenholz, Y. (1997). Electrostatic parameters of cationic liposomes commonly used for gene delivery as determined by 4-heptadecyl-7-hydroxycoumarin. Biochim. Biophys. Acta, 1329, 211-222. 

Lipid moieties coupled to polyethylene glycol (PEG) have been used to increase the blood circulation time of lipoplexes (Fig. 32). The PEG-lipid conjugates such as DOPE-PEG, Chol-PEG, ceramides-PEG and their derivatives are then coformulated with the cationic lipid, helper lipid, and DNA. This results in coating the surface of the lipoplexes with PEG and preventing undesired association with plasma proteins or circulating cells (stealth liposomes). Recently, a-tocopheryl PEG-succinate (TPGS) was also used in gene delivery formulations because of its ability to confer not only a stealth property but also antioxidant and absorption enhancer properties [129]. 

Several reports in the literature state that DOPE, while successfully used for in vitro gene delivery, is a poor helper lipid for in vivo applications [28-32], Instead, for reasons that are not understood, lipid mixtures for successful transfection in vivo seem to require cholesterol [33]. In fact, an equimolar mixture of cholesterol and DOTAP is widely used for in vivo experiments and clinical trials. Cholesterol has also been included in liposomes along with cationic DOTAP and fusogenic DOPE to form a potent mixture used to study the treatment of ovarian cancer by delivery of the p53 tumor suppressor gene [34, 35]. 

The major obstacle for the use of this type of liposomes in gene delivery was their intrinsically poor encapsulation efficiency of ultra-large payload compounds... 

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