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Fig. 7.2 Alkyl-acrylate, alkyl-acrylamide, and amino molecules were used to synthesize a combi-
natorial library of lipidoids (reproduced with permission from [ 18 ] )
paramount importance because endocytosis is the main process by which cells
uptake nanoparticles, and nucleic acids are membrane impermeable [ 22, 23 ] . Brie fl y,
amines possessing the appropriate p K a can accept protons that are pumped into the
endosome as it acidifies to become a catabolic lysosome [ 24 ] , preventing degrada-
tion of the compartment's contents. The net positive charge on the quarternary
amines creates an electrochemical gradient, leading to the influx of negatively
charged chloride ions. The high salt concentration inside the endosome produces a
strong osmotic gradient, leading to the inflow of water molecules, which eventually
rupture the endosomal membrane and release the drug cargo into the cytosol.
7.4
Structure-Activity Relationships
The ability of the lipidoids to deliver siRNA to mammalian cells was first investi-
gated in vitro [ 18 ]. A HeLa cell line was established to express stably both firely
( Photinus pyralis ) and Renilla ( Renilla reniformis ) luciferases. These biolumines-
cent enzymes have unique substrates and can, thus, be differentiated by performing
sequential assays using DualGlo™(Promega). Cells were plated into 96-well plates
and allowed to attach overnight in growth medium. Lipidoids were dissolved in
25 mM sodium acetate (pH 5) and mixed with siRNA targeting firefly luciferase at
multiple lipidoid-to-siRNA weight-to-weight ratios. These mixtures were incubated
for 20 min to allow for complex formation prior to addition to the cells. After 24 h,
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