Preliminary results were reported on PHEMA substituted with amino acids such
as glycine [P(HEMA-Gly), Fig. 11c-R 1 ], alanine [P(HEMA-Ala), Fig. 11c-R 2 ],
valine [P(HEMA-Val), Fig. 11c-R 3 ], phenylalanine [P(HEMA-Phe), Fig. 11c-R 4 ],
and lysine [P(HEMA-Lys, Fig. 11c-R 5 ] [179]. These polymers were able to condense DNA, while having very little toxicity (up to 250 μg mL
À1 tested on COS-7
and SPCA-1 cell lines); unfortunately, only preliminary results were reported and
there was no comparison between these amino acid-substituted PHEMAs, despite
their interesting structures.
Several poly(methacrylate)- and poly(methacrylamide)-based homopolymers
with various side chains bearing primary, tertiary, and quaternary ammonio groups
were designed by the group of Seymour in order to study the influence of (1) length
of side chains bearing cationic residues; (2) the nature of the amine, i.e., primary,
secondary, or tertiary amines and quaternary ammonio groups; (3) charge spacing
along the polymer backbone; and (4) molecular weight or degree of polymerization
[180]. These polymers are presented Fig. 11d–g: PVA.HCl (Fig. 11d), PAA.HCl
(Fig. 11e), PMAEDA.HCl (Fig. 11f-R 1 ), PMAGEDA.HCl (Fig. 11f-R 2 ),
PMADGHDA.HCl (Fig. 11f-R 3 ), PDMAEMAm (Fig. 11f-R 4 ), PTMAEM.Cl
(Fig. 11f-R 5 ), P(HPMA-co-TMAEM.Cl (Fig. 11g), and PBTMAIPM.I 2
(Fig. 11f-R 6 ). At an N:P ratio of 2, considering first the influence of side-chain
length, polymers with long side chains such as PMADGHDA.HCl were incapable
of efficient complex formation with DNA (considerable residual EtBr fluorescence)
but did show considerable transfection activity (positively charged complexes, thus
their uptake by cells may be facilitated), despite their poor gene expression via
direct intranuclear injection. At the other extreme, complexes formed using
polymers with very short side chains, such as PVA.HCl and PAA.HCl, were
efficient at complex formation (little residual EtBr fluorescence) and were remarkably stable to polyanion-mediated disruption. Efficient charge neutralization may
explain their lack of surface charge, which probably underlies their tendency to
aggregate. Their low transfection activity could be a consequence of their low
surface charge (not uptaken by cells to a great extent), but their ability to undergo
efficient intranuclear transcription was surprising in light of their stability to
polyanions, as mentioned by the authors. The influence of cationic charge strength,
using primary or tertiary amines or ammonio groups, on the properties of
complexes formed with DNA was investigated. Polymers containing ammonio
groups such as PTMAEM.Cl complexed DNA relatively efficiently, reflecting a
stronger bond than with primary amino groups. The transfection activity of their
complexes was 10- to 100-fold less than complexes based on most other cationic
polymers. This poor transfection activity may be the result of poor access to the
cytoplasm/nucleus, due to either cytotoxicity or poor endosomal release due to the
absence of pH responsiveness (no proton sponge effect). By contrast, PDMAEMam
containing tertiary amines was less effective at DNA condensation but showed good
transfection activity in HEK 293 cells in vitro, probably because of the pK a of its
amines in the endosomal range. In the next step, the influence of the density
of positive charge along the cationic polymer on the properties of complexes formed
with DNA was investigated. Comparing PTMAEM.Cl with IBTMAIPM.I 2 , which
Polyelectrolyte Complexes of DNA and Polycations as Gene Delivery Vectors
153
as glycine [P(HEMA-Gly), Fig. 11c-R 1 ], alanine [P(HEMA-Ala), Fig. 11c-R 2 ],
valine [P(HEMA-Val), Fig. 11c-R 3 ], phenylalanine [P(HEMA-Phe), Fig. 11c-R 4 ],
and lysine [P(HEMA-Lys, Fig. 11c-R 5 ] [179]. These polymers were able to condense DNA, while having very little toxicity (up to 250 μg mL
À1 tested on COS-7
and SPCA-1 cell lines); unfortunately, only preliminary results were reported and
there was no comparison between these amino acid-substituted PHEMAs, despite
their interesting structures.
Several poly(methacrylate)- and poly(methacrylamide)-based homopolymers
with various side chains bearing primary, tertiary, and quaternary ammonio groups
were designed by the group of Seymour in order to study the influence of (1) length
of side chains bearing cationic residues; (2) the nature of the amine, i.e., primary,
secondary, or tertiary amines and quaternary ammonio groups; (3) charge spacing
along the polymer backbone; and (4) molecular weight or degree of polymerization
[180]. These polymers are presented Fig. 11d–g: PVA.HCl (Fig. 11d), PAA.HCl
(Fig. 11e), PMAEDA.HCl (Fig. 11f-R 1 ), PMAGEDA.HCl (Fig. 11f-R 2 ),
PMADGHDA.HCl (Fig. 11f-R 3 ), PDMAEMAm (Fig. 11f-R 4 ), PTMAEM.Cl
(Fig. 11f-R 5 ), P(HPMA-co-TMAEM.Cl (Fig. 11g), and PBTMAIPM.I 2
(Fig. 11f-R 6 ). At an N:P ratio of 2, considering first the influence of side-chain
length, polymers with long side chains such as PMADGHDA.HCl were incapable
of efficient complex formation with DNA (considerable residual EtBr fluorescence)
but did show considerable transfection activity (positively charged complexes, thus
their uptake by cells may be facilitated), despite their poor gene expression via
direct intranuclear injection. At the other extreme, complexes formed using
polymers with very short side chains, such as PVA.HCl and PAA.HCl, were
efficient at complex formation (little residual EtBr fluorescence) and were remarkably stable to polyanion-mediated disruption. Efficient charge neutralization may
explain their lack of surface charge, which probably underlies their tendency to
aggregate. Their low transfection activity could be a consequence of their low
surface charge (not uptaken by cells to a great extent), but their ability to undergo
efficient intranuclear transcription was surprising in light of their stability to
polyanions, as mentioned by the authors. The influence of cationic charge strength,
using primary or tertiary amines or ammonio groups, on the properties of
complexes formed with DNA was investigated. Polymers containing ammonio
groups such as PTMAEM.Cl complexed DNA relatively efficiently, reflecting a
stronger bond than with primary amino groups. The transfection activity of their
complexes was 10- to 100-fold less than complexes based on most other cationic
polymers. This poor transfection activity may be the result of poor access to the
cytoplasm/nucleus, due to either cytotoxicity or poor endosomal release due to the
absence of pH responsiveness (no proton sponge effect). By contrast, PDMAEMam
containing tertiary amines was less effective at DNA condensation but showed good
transfection activity in HEK 293 cells in vitro, probably because of the pK a of its
amines in the endosomal range. In the next step, the influence of the density
of positive charge along the cationic polymer on the properties of complexes formed
with DNA was investigated. Comparing PTMAEM.Cl with IBTMAIPM.I 2 , which
Polyelectrolyte Complexes of DNA and Polycations as Gene Delivery Vectors
153