Liposome/polycation/DNA

Liposome-Polycation-DNA A Nonviral Gene Vector Turned into a Potent Vaccine Carrier... 

Liposome-polycation-DNA (LPD) nanoparticles (1) are formed by spontaneous rearrangement of a lipid shell around a polycation-condensed bacterial plasmid DNA core to form a virus-like structure (2). The LPD complexes consist of liposomes that are either made of cationic (LPDI) or anionic (LPDII) lipids and are sometimes referred to as lipopolyplexes, a broader category that also includes other lipid-based vectors (2). 

LPDI nanoparticles are homogenous, self-forming spheres between 100 and 200 nm in diameter that are formed from the spontaneous rearrangement of a lipid bilayer around a polycation condensed DNA core. The LPDI particles (lipopolyplexes) have benefits over lipoplexes, which are composed of liposomes and DNA. Homogenous particles are formed during preparation and thus allow a more consistent production of particles, as required by the FDA for clinical use. The LPDI particles also have a lower toxicity associated with them as opposed to lipoplexes, which can generate severe systemic inflammatory responses, most likely to the increased DNA content on the surface of the particles. The internalization of DNA inside the LPDI also has a benefit of DNA protection. The DNA is not nearly as accessible to nuclease attack and mechanical stress. Therefore, a lower quantity of DNA is used because it is protected inside of the LPDI for delivery. 

The weakly immunogenic protamine sulfate USP (1) condenses DNA to form a toroid structure of super-coiled DNA about 50 nm in diameter (2). The DNA in this form or in the preformed LPDI complex cannot be displaced from the protamine by polycations such as spermidine and histones or by other nucleic acids like genomic DNA (2). DNA in this toroid structure is transcriptionally inactive and this conformation allows for protection of DNA from enzymatic degradation by nucleases and other environmental assaults such as mechanical stress (1,2). After the liposome surrounds the toroid, the resulting homogenous LPDI nanoparticles are slightly less than... 

Initial vaccination studies with LPDI nanoparticles were completed using liposomes prepared with both 1,2-dioleyltriammonium propane (DOTAP) and cholesterol. After it was determined that cholesterol played only a small structural role and was not necessary for activity, the liposomes were then prepared using only DOTAP to become an LPDI type of formulation. Regardless of the lipid used, the ratio of cationic lipid, polycation, and DNA must be maintained to have all properties associated with LPDI particles (2). 

Physical methods of gene transfer. Genes can often be transferred without the use of a cloning vehicle. This is especially important for certain plant cells, such as those of cereal grains, for which transfer of genes via the Ti plasmid has been difficult.167 If DNA, which may be in a plasmid, is coprecipitated with calcium phosphate, it can often be taken up directly either by animal cells or by plant protoplasts.168 169 Polycations also facilitate DNA uptake cationic liposomes seem to be especially effective.170 In the widely used electroporation technique a short electrical pulse of a few hundred volts / cm is applied to create transient pores in the plasma membrane through which the DNA can enter a cell.111 8,171 175 Chromosomes can be transferred by cell fusion and either... 

The use of polycationic moieties in drug delivery has been a subject of considerable interest for the past several decades. One of the earliest demonstrations of the potential value of polycations in chug delivery was a study by Tan in 1977, which demonstrated increased uptake of the SV40 virus when infections were carried out in the presence of polycations.1 Other early efforts involved direct cationization of proteins to enhance cellular uptake.2,3 Simultaneously, extensive investigations of the ability of a variety of polycationic vehicles to enhance the uptake of DNA were made.4 These vehicles include cationic liposomes,5 6 poly-L-lysine,7 polyethe-leneimine,8 amino-dendrimers,9 and a variety of other natural and synthetic polycations. Only in recent years, however, has the surprisingly diverse potential of polycation-based delivery systems been realized. This revelation has come primarily from the discovery and utilization of nonclassical transport-based pathways. 

Polycation-Condensed DNA Entrapped in Cationic Liposome LPD-I Formulation... 

Liposomes and lipoplexes are usually self-assembling, easy to prepare and biodegradable. They allow increased uptake of naked DNA and DNA NPs. They can also be combined with polycations to form lipid-DNA NPs. Caracciola et al. [8] observed that lipid-protamine-DNA (LPD) NPs were more efficient than lipoplexes for gene delivery in CHO (Chinese hamster ovary cells), HEK293 (human embryonic kidney cells), NIH 3T3 (mouse embryraial cells) and A17 (murine cancer cells) cells. Unfortunately, cationic liposomes exhibit significant variability in gene delivery efficiency and are often toxic to cells. 

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