to the non-PEGylated polymer at a ratio close to unity and for similar polymers,
P(MAPTAC-co-OEMA), with higher degree of OE substitution, 8 oligo(ethylene
glycol)methyl ether units per graft, M w ¼ 1.8 Â 10
6 g mol
À1 for 89 mol% OE, and
M w ¼ 3.5 Â 10
6 g mol
À1 for 94 mol% OE (Fig. 4b) [129]. Complete binding of
DNA by these polycations occurred at a ratio of more than 1 due to the presence of
PEG, which partially screens the charges of the polycation (the higher the PEG
content, the higher the ratio for complete binding). At a charge ratio of ~2 for these
P(MAPTAC-co-OEMA) copolymers, the zeta potential reached a plateau at a
neutral value and the hydrodynamic diameter stayed constant, meaning that the
excess polycation was not incorporated into the polyplex. On the contrary, for
the examples of the last paragraph that did not possess a steric component and for
which the same phenomenon had mainly an electrostatic explanation, the lack of
incorporation of the excess polycation into the polyplex occurred in this case
mainly because of the steric repulsion produced by the PEG corona. Moreover,
the DNA present in these polyplexes was inaccessible to DNAse I, which clearly
indicates that the PEG segments present in the outer part of the polyplexes protect
the DNA inside the polyplexes. Nevertheless, it is surprising that despite the high
content of PEG in the polycation, the DNA binding was still efficient. By contrast,
for P(MAPTAC-co-OEMA) the zeta potential was positive at a high charge ratio of
random copolymers, P(TMAEMA-co-HPMA) (Fig. 4c) [130]. Nevertheless, only
binding of DNA is not enough and a tight binding is desired. The copolymers P
(TMAEMA-co-HPMA) containing the lowest degree of ammonium (5 and 15%)
showed virtually no ability to displace EtBr and also did not protect DNA from
degradation by endonucleases, probably because their association with DNA was
too weak. The inverse tendency has been observed for PLL and PLS and their
trimethylated derivatives (PtmL and PtmLS, respectively; Fig. 4d) [131]: the
complexes with trimethylated peptides seemed to be looser (according to EtBr
complexation and exchange reaction with anions) but the compaction of DNA
occured at lower charge C/A ratio, due to their higher charge density.Interestingly,
the transfection efficiency of the trimethylated PtmLS into COS-1 cells was better
than that of PtmL and the non-quaternized derivatives, despite their similar intracellular distribution. Thus, it seems that a loose structure for the release of DNA
from the complex (at best from endosome into the cytoplasm) is necessary, as well
as the presence of functional groups such as serine residues that impart hydrophilicity and hydrogen bonding capacity.
A similar construct to PLL, based on a slightly different amide backbone, a
polyaspartamide derivative containing a quaternary ammonium, α,β-poly{(N-2hydroxyethyl)-N-carbazate[N-(3-trimethylammonium chloride) propylhydrazide]-D,Laspartamide} (PHEA-HYD-CPTA) (Fig. 4e) [132], also revealed itself to efficiently
complex DNA and reduce its rate of degradation by DNAse. Similar polymers but
with a block structure were also efficient at protecting DNA against enzymatic
degradation. Indeed, partially methylated PEG-mPDMAEMA-β and completely
methylated PEG-mPDMAEMA or butylated PEG-bPDMAEMA with PEG blocks
of various lengths (Fig. 4f) [133] formed, as previously explained, micellar-type
Polyelectrolyte Complexes of DNA and Polycations as Gene Delivery Vectors
137
P(MAPTAC-co-OEMA), with higher degree of OE substitution, 8 oligo(ethylene
glycol)methyl ether units per graft, M w ¼ 1.8 Â 10
6 g mol
À1 for 89 mol% OE, and
M w ¼ 3.5 Â 10
6 g mol
À1 for 94 mol% OE (Fig. 4b) [129]. Complete binding of
DNA by these polycations occurred at a ratio of more than 1 due to the presence of
PEG, which partially screens the charges of the polycation (the higher the PEG
content, the higher the ratio for complete binding). At a charge ratio of ~2 for these
P(MAPTAC-co-OEMA) copolymers, the zeta potential reached a plateau at a
neutral value and the hydrodynamic diameter stayed constant, meaning that the
excess polycation was not incorporated into the polyplex. On the contrary, for
the examples of the last paragraph that did not possess a steric component and for
which the same phenomenon had mainly an electrostatic explanation, the lack of
incorporation of the excess polycation into the polyplex occurred in this case
mainly because of the steric repulsion produced by the PEG corona. Moreover,
the DNA present in these polyplexes was inaccessible to DNAse I, which clearly
indicates that the PEG segments present in the outer part of the polyplexes protect
the DNA inside the polyplexes. Nevertheless, it is surprising that despite the high
content of PEG in the polycation, the DNA binding was still efficient. By contrast,
for P(MAPTAC-co-OEMA) the zeta potential was positive at a high charge ratio of
random copolymers, P(TMAEMA-co-HPMA) (Fig. 4c) [130]. Nevertheless, only
binding of DNA is not enough and a tight binding is desired. The copolymers P
(TMAEMA-co-HPMA) containing the lowest degree of ammonium (5 and 15%)
showed virtually no ability to displace EtBr and also did not protect DNA from
degradation by endonucleases, probably because their association with DNA was
too weak. The inverse tendency has been observed for PLL and PLS and their
trimethylated derivatives (PtmL and PtmLS, respectively; Fig. 4d) [131]: the
complexes with trimethylated peptides seemed to be looser (according to EtBr
complexation and exchange reaction with anions) but the compaction of DNA
occured at lower charge C/A ratio, due to their higher charge density.Interestingly,
the transfection efficiency of the trimethylated PtmLS into COS-1 cells was better
than that of PtmL and the non-quaternized derivatives, despite their similar intracellular distribution. Thus, it seems that a loose structure for the release of DNA
from the complex (at best from endosome into the cytoplasm) is necessary, as well
as the presence of functional groups such as serine residues that impart hydrophilicity and hydrogen bonding capacity.
A similar construct to PLL, based on a slightly different amide backbone, a
polyaspartamide derivative containing a quaternary ammonium, α,β-poly{(N-2hydroxyethyl)-N-carbazate[N-(3-trimethylammonium chloride) propylhydrazide]-D,Laspartamide} (PHEA-HYD-CPTA) (Fig. 4e) [132], also revealed itself to efficiently
complex DNA and reduce its rate of degradation by DNAse. Similar polymers but
with a block structure were also efficient at protecting DNA against enzymatic
degradation. Indeed, partially methylated PEG-mPDMAEMA-β and completely
methylated PEG-mPDMAEMA or butylated PEG-bPDMAEMA with PEG blocks
of various lengths (Fig. 4f) [133] formed, as previously explained, micellar-type
Polyelectrolyte Complexes of DNA and Polycations as Gene Delivery Vectors
137