Gene Delivery Using Chemical Methods - Challenges in Delivery of Therapeutic Genomics and Proteomics - page 151

after direct administration in tumors [178-181] . Following intratumoral injection of
PVP formulated with pDNA-encoding human cytokines, specifically IL-12, resulted
in noticeable levels of cytokines after 24 h. IL-12-encoding plasmid or a combina-
tion of both plasmids induces formation of CD8 T-lymphocytes, presumably CTL,
which mediate the antitumor effect. Although comparison studies using the plasmids
encoding the cytokines without PVP were not performed, the protein expression
data seem to be either comparable or higher than those obtained with cationic lip-
ids during individual studies. Formerly, researchers used PVP in multiepitope pDNA
vaccines, which demonstrated an increase in responses to the less immunogenic epit-
opes, without responses to others [182,183] . The clinical testing was performed using
HIV vaccine, EP HIV-1090 formulated with PVP, in HLA-A0201 transgenic mice.
The clinical product was formulated as 2 mg/ml EP HIV-1090 pDNA vaccine, 3.4%
PVP in 100 mM sodium phosphate (pH 7. 0) and 150 mM NaCl. Owing to the non-
toxic nature of the PVP, repeat immunizations can be done at different sites, or repeat
administrations can be given within 24 h to increase response levels. This characteris-
tic gives PVP an edge over other vectors having a less desirable toxicity profile, such
as cationic lipids.
4.3 Lipidic Vectors
Many different types of lipidic systems have been developed as vectors for gene
delivery, for example, liposomes, micelles, emulsions, and other organized structures
of lipids. Early studies done using neutral and anionic liposomes demonstrated that
pDNA is encapsulated inside the vesicles, and the structures of liposomes remain
unchanged. However, there has been a lack of progress in this direction because of the
practical difficulties in encapsulating a sufficient amount of DNA into the vesicles.
Over the last 15 years, cationic liposomes have gained much attention compared
to other nonviral vectors. The use of lipid-based vectors was first reported by Felgner
et al. in the late 1980s [184] . In 1987, the group described the use of cationic lipid as
vectors for gene delivery. The authors reported that a positively charged lipid, specifi-
cally N -[1-(2,3-dioleyloxy)propyl]- N , N , N- trimethylammonium chloride (DOTMA),
could form liposomes under physiological conditions, either alone or in combination
with neutral lipids [185] . These cationic liposome vesicles were shown to interact
spontaneously with anionic DNA to form lipid-DNA complexes known as lipoplexes.
It was observed that these cationic liposomes interact with DNA through charge inter-
action, and extensive lipid rearrangement during complex formation was noted. The
resulting complexes are substantially different from starting liposomes [186,187] .
The aim of cationic lipid-mediated gene transfer is to deliver pDNA to a desired
cell such that it becomes transcribed and translated into a desired protein/peptide.
The sequence of events involved in cationic lipid-mediated gene transfer comprises,
but is not limited to, the following steps:
1. Formation of liposome from cationic lipid offering a multivalent surface charge
2. Interaction via electrostatics with negative charges on the DNA phosphate backbone