Investigating the Effects of Block versus Statistical
Glycopolycations Containing Primary and Tertiary Amines for Plasmid
DNA Delivery
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Abstract
Polymer
composition and morphology can affect the way polymers
interact with biomolecules, cell membranes, and intracellular components.
Herein, diblock, triblock, and statistical polymers that varied in
charge center type (primary and/or tertiary amines) were synthesized
to elucidate the role of polymer composition on plasmid DNA complexation,
delivery, and cellular toxicity of the resultant polyplexes. The polymers
were synthesized via RAFT polymerization and were composed of a carbohydrate
moiety, 2-deoxy-2-methacrylamido glucopyranose (MAG), a primary amine
group, <i>N</i>-(2-aminoethyl) methacrylamide (AEMA), and/or
a tertiary amine moiety, <i><i>N,N</i></i>-(2-dimethylamino)ethyl
methacrylamide (DMAEMA). The lengths of both the carbohydrate and
cationic blocks were kept constant while the primary amine to tertiary
amine ratio was varied within the polymers. The polymers were characterized
via nuclear magnetic resonance (NMR) and size exclusion chromatography
(SEC), and the polyplex formulations with pDNA were characterized
in various media using dynamic light scattering (DLS). Polyplexes
formed with the block copolymers were found to be more colloidally
stable than statistical copolymers with similar composition, which
rapidly aggregated to micrometer sized particles. Also, polymers composed
of a higher primary amine content were more colloidally stable than
polymers consisting of the tertiary amine charge centers. Plasmid
DNA internalization, transgene expression, and toxicity were examined
with each polymer. As the amount of tertiary amine in the triblock
copolymers increased, both gene expression and toxicity were found
to increase. Moreover, it was found that increasing the content of
tertiary amines imparted higher membrane disruption/destabilization.
While both block and statistical copolymers had high transfection
efficiencies, some of the statistical systems exhibited both higher
transfection and toxicity than the analogous block polymers, potentially
due to the lack of a hydrophilic block to screen membrane interaction/disruption.
Overall, the triblock terpolymers offer an attractive composition
profile that exhibited interesting properties as pDNA delivery vehicles