Differential Polymer Structure Tunes Mechanism of
Cellular Uptake and Transfection Routes of Poly(β-amino ester)
Polyplexes in Human Breast Cancer Cells
posted on 2014-01-15, 00:00authored byJayoung Kim, Joel C. Sunshine, Jordan J. Green
Successful gene delivery with nonviral
particles has several barriers,
including cellular uptake, endosomal escape, and nuclear transport.
Understanding the mechanisms behind these steps is critical to enhancing
the effectiveness of gene delivery. Polyplexes formed with poly(β-amino
ester)s (PBAEs) have been shown to effectively transfer DNA to various
cell types, but the mechanism of their cellular uptake has not been
identified. This is the first study to evaluate the uptake mechanism
of PBAE polyplexes and the dependence of cellular uptake on the end
group and molecular weight of the polymer. We synthesized three different
analogues of PBAEs with the same base polymer poly(1,4-butanediol
diacrylate-co-4-amino-1-butanol) (B4S4) but with
small changes in the end group or molecular weight. We quantified
the uptake and transfection efficiencies of the pDNA polyplexes formulated
from these polymers in hard-to-transfect triple negative human breast
cancer cells (MDA-MB 231). All polymers formed positively charged
(10–17 mV) nanoparticles of ∼200 nm in size. Cellular
internalization of all three formulations was inhibited the most (60–90%
decrease in cellular uptake) by blocking caveolae-mediated endocytosis.
Greater inhibition was shown with polymers that had a 1-(3-aminopropyl)-4-methylpiperazine
end group (E7) than the others with a 2-(3-aminopropylamino)-ethanol
end group (E6) or higher molecular weight. However, caveolae-mediated
endocytosis was generally not as efficient as clathrin-mediated endocytosis
in leading to transfection. These findings indicate that PBAE polyplexes
can be used to transfect triple negative human breast cancer cells
and that small changes to the same base polymer can modulate their
cellular uptake and transfection routes.