Gene Delivery Using Chemical Methods - Challenges in Delivery of Therapeutic Genomics and Proteomics - page 175

opinion as to whether L-lysine dendrimers should be classified under the original
term dendrimer , described by Tomalia et al. in 1985 as consisting of asymmetrical
L-lysine residues [286] . Also, dendritic molecules, consisting of amino acids, are
expected to facilitate the construction of gene vectors. Block copolymers consisting
of dendritic PLLs and poly(ethylene glycol) were synthesized for the gene vector
[325,326] .
Okuda et al. analyzed PLL complex with atomic force microscopy to understand
the mechanism of complex formation of sixth-generation dendritic PLL (G6). The
transfection activity observed was related to the size of the dendrimer-DNA com-
plex. Larger complexes (greater than1 m) exhibited the highest transfection. The
complex uptake was by endocytosis pathway. Further proton sponge effect mediates
endosomal escape of complex to reach the nucleus of the cell [327] .
Ohsaki et al. prepared first- to sixth-generation (G1-G6) monodispersed dendritic
PLL having a hexamethylenediamine core, to investigate DNA-binding abilities
and transfection efficiency. It was observed that G3 and higher generations formed
complexes efficiently with pDNA. Moreover, degree of compaction of the DNA was
increased with increasing generational number. G5 and G6 demonstrated higher
transfection efficiency comparable to Superfect and Lipofectin TM , in vitro , without
significant toxicity [328] . The sixth-generation structure also exhibited substantial
serum stability. The same group has performed an in vivo study of PLL dendrimers
that prolonged circulation time and significant tumor accumulation. However, gene
expression was not observed at all, possibly due to much stronger dendrimer-DNA
binding, which prevents release of DNA postendocytosis [329] .
Although cellular uptake of PLL dendriplexes is fourfold lesser than linear PLL,
PLL dendriplexes-mediated gene expression is 100-fold higher. The reason behind the
dramatic increase in transfer activity in spite of reduced cellular uptake was assumed
to be improved endosomal release and higher availability to RNA polymerase inside
the nucleus due to less compactness [330] . Koyama et al. performed biodistribution of
pDNA in normal and tumor-bearing mice after IV injection of DNA complexes with
PLL dendrimers. It was observed that PLL dendrimers possess stealth properties that
maintain their presence in blood circulation for a longer duration. The stealth ability
of PLL dendrimers would also enhance their permeability and retention (EPR) effect
in the tumor [331] .
Various modifications to PLL dendrimer structure have been investigated to
improve transfection efficiency. The effect of substituting terminal lysine groups with
either arginine or histidine was observed on gene delivery into cells. Arginine deriva-
tive bound to the pDNA as strongly as PLL dendrimer, whereas histidine derivative
showed decreased binding ability. The transfection efficiency of arginine derivative
was 3 to 12 folds higher than unmodified PLL dendrimer, whereas histidine deriva-
tive does not demonstrate any gene transfer ability. However, when it was mixed with
DNA under acidic conditions (pH 5.0), DNA complexes were formed that showed high
transfection efficiency, comparable to that obtained with arginine derivative-mediated
transfection. This result highlighted pH-dependent in vitro and in vivo gene transfec-
tion systems [332] . In another investigation, PLL dendrimers with a cubic octa(3-ami
nopropyl)silsesquoxane core were synthesized. These PLL dendrimers displayed supe-