copolymers with hydrophilic groups were capable of producing complexes with
DNA that were more soluble than the PLL alone; as expected, the most efficient
was the PEG-b-PLL with the longest PEG segment (12 kDa) at the highest grafting
ratio (10%) and it was also the least toxic. PLL-b-dextran showed increased toxicity
compared to low molecular weight PLL.
Polyoxazolines are thought to be an alternative to PEG as biocompatible blocks.
In this frame, PLL-b-PMOx (Fig. 17e) and the polyplexes made thereof were
synthesized and studied by the group of Lu ¨hmann [200]. As for PEG or other
block copolymers comprising a cationic block and some hydrophilic modification,
the complexation of DNA by PLL-b-PMOx was compromised at high grafting rates
and/or long PMOx chains (as shown by gel electrophoresis). Condensates made of
polymers with less than 7% grafting density showed aggregate formation at ratios
supposedly above the neutral point (zeta potential not reported) and were over
500 nm in diameter. Above 7% grafting density, independent of the length of
PMOx, the diameters of the polyplexes were less than 200 nm. These polymers
were able to protect DNA from DNAse I digestion, but only PLL-g-PMOx with low
molecular weight PMOx and low grafting densities of 7–14% showed significant
gene expression in COS-7 cells (good transfection efficiency for this polymer was
found at N:P ratio of 3.125), which correlated with their good cellular uptake.
Other Amino Acid-Based Polymers
PEG-co-(PLL-g-His) (Fig. 18a), a copolymer of PEG (1.45 kDa) and PLL modified
with histidine at various grafting rates (5, 9, 16, and 22%) showed relatively poor
EtBr displacement capacity [201]. EtBr displacement was only achieved at N:P
ratio of 5 for the polymers with the two lowest grafting rates, and at ratios of 10 for
the two others, which can be explained by a looser complexation due to the
presence of PEG or the bulky imidazole groups or by the copolymer nature or by
less cationic residues due to the grafting. Consistent with these high N:P ratios were
the diameters of most of the complexes that remained between 150 and 200 nm for
N:P ratios of 10 and above, whereas the complexes based on PLL were much
smaller. The complexes of DNA based on PEG-co-(PLL-g-His 16% ) showed the
highest transfection efficiency in the A7r5 cell line compared to the other polymers
and the efficiency increased with the N:P ratio. Compared to polymers with lower
degree of modification, this could be explained by the presence of more histidine
residues, which are known to have endosomal buffering capacity (the buffering
capacities increased with increasing His content), facilitating the escape of the
polyplex into the cytoplasm. In addition, the decreased efficiency of PEG-co(PLL-g-His 22% ) indicated the importance of having enough complexing units in
the polymer. Probably because of the ester bonds, the polymer was totally degraded
into its constituent PEG and PLL blocks after 24 h, which is an interesting approach
to obtaining biodegradable polyplexes.
PEG has also been coupled to polyamino acids other than PLL in order to
increase solubility (critical in the case of polyhistidine for instance) and design a
polymer with great DNA complexation and transfection properties.
164
A. Bertin
DNA that were more soluble than the PLL alone; as expected, the most efficient
was the PEG-b-PLL with the longest PEG segment (12 kDa) at the highest grafting
ratio (10%) and it was also the least toxic. PLL-b-dextran showed increased toxicity
compared to low molecular weight PLL.
Polyoxazolines are thought to be an alternative to PEG as biocompatible blocks.
In this frame, PLL-b-PMOx (Fig. 17e) and the polyplexes made thereof were
synthesized and studied by the group of Lu ¨hmann [200]. As for PEG or other
block copolymers comprising a cationic block and some hydrophilic modification,
the complexation of DNA by PLL-b-PMOx was compromised at high grafting rates
and/or long PMOx chains (as shown by gel electrophoresis). Condensates made of
polymers with less than 7% grafting density showed aggregate formation at ratios
supposedly above the neutral point (zeta potential not reported) and were over
500 nm in diameter. Above 7% grafting density, independent of the length of
PMOx, the diameters of the polyplexes were less than 200 nm. These polymers
were able to protect DNA from DNAse I digestion, but only PLL-g-PMOx with low
molecular weight PMOx and low grafting densities of 7–14% showed significant
gene expression in COS-7 cells (good transfection efficiency for this polymer was
found at N:P ratio of 3.125), which correlated with their good cellular uptake.
Other Amino Acid-Based Polymers
PEG-co-(PLL-g-His) (Fig. 18a), a copolymer of PEG (1.45 kDa) and PLL modified
with histidine at various grafting rates (5, 9, 16, and 22%) showed relatively poor
EtBr displacement capacity [201]. EtBr displacement was only achieved at N:P
ratio of 5 for the polymers with the two lowest grafting rates, and at ratios of 10 for
the two others, which can be explained by a looser complexation due to the
presence of PEG or the bulky imidazole groups or by the copolymer nature or by
less cationic residues due to the grafting. Consistent with these high N:P ratios were
the diameters of most of the complexes that remained between 150 and 200 nm for
N:P ratios of 10 and above, whereas the complexes based on PLL were much
smaller. The complexes of DNA based on PEG-co-(PLL-g-His 16% ) showed the
highest transfection efficiency in the A7r5 cell line compared to the other polymers
and the efficiency increased with the N:P ratio. Compared to polymers with lower
degree of modification, this could be explained by the presence of more histidine
residues, which are known to have endosomal buffering capacity (the buffering
capacities increased with increasing His content), facilitating the escape of the
polyplex into the cytoplasm. In addition, the decreased efficiency of PEG-co(PLL-g-His 22% ) indicated the importance of having enough complexing units in
the polymer. Probably because of the ester bonds, the polymer was totally degraded
into its constituent PEG and PLL blocks after 24 h, which is an interesting approach
to obtaining biodegradable polyplexes.
PEG has also been coupled to polyamino acids other than PLL in order to
increase solubility (critical in the case of polyhistidine for instance) and design a
polymer with great DNA complexation and transfection properties.
164
A. Bertin