The effect of pH on PAMAM dendrimer–siRNA complexation — Endosomal considerations as determined by molecular dynamics simulation

Abstract

Intracellular degradation of genes, most notably within the endo-lysosomal compartment is considered a significant barrier to (non-viral) gene delivery in vivo. Previous reports based on in vitro studies claim that carriers possessing a mixture of 1º, 2º & 3º amines are able to buffer the acidic environment within the endosome, allowing for timely release of their contents, leading to higher transfection rates. In this report, we adopt an atomistic molecular dynamics (MD) simulation approach, comparing the complexation of 21-bp siRNA with lowgeneration polyamidoamine (PAMAM) dendrimers (G0 and G1) at both neutral and acidic pHs, the latter of which mimics the degradative environment within maturing 'lateendosomes'. Our simulations reveal that the time taken for the dendrimer-gene complex (dendriplex) to reach equilibrium is appreciably longer at low pH and this is accompanied by more compact packaging of the dendriplex, as compared to simulations performed at neutral pH. We also note higher calculated binding free energies of the dendriplex at low pH, indicating a higher dendrimer-gene affinity in comparison with neutral pH. These novel simulations provide a more detailed understanding of low molecular-weight polymer-siRNA behaviour, mimicking the endosomal environment and provide input of direct relevance to the "proton sponge theory", thereby advancing the rational design of non-viral gene delivery systems.

Multiscale Modeling for Host-Guest Chemistry of Dendrimers in Solution

“…The types of guest molecules considered in this review will include low molecular weight organic molecules [24,29,50,51], and nucleic acids [38,[69][70][71][72][73]. The review of interactions of dendrimers with lipid membrane has been clearly explained in Reference [37].…”

Systematic coarse-grained modeling of complexation between small interfering RNA and polycations

“…[53][54][55][56][57] For binding with lower-generation dendrimers as well as certain polycations smaller in size than siRNA, such as lowmolecular-weight linear or branched poly(L-lysine), Ouyang et al have reported that polycations with lower total charge preferably bind within the major groove of siRNA, whereas those with higher total charge appear to bind less specifically on different regions of the siRNA. 53 Considering that linear PEI shows a better safety profile than branched PEI, 58,59 here we explore how linear PEI2.2k-g-PEG0.9k chains, which are longer than siRNA and far more flexible, bind to siRNA and how this condensation affects the siRNA structure.…”

Comparison of the cellular transport mechanism of cationic, star-shaped polymers and liposomes in HaCat cells

“…The "proton sponge effect" was proposed as one of the potential mechanisms for the endosomal release of the polymers. 13 The ATPase proton pump inhibitor bafilomycin A1 and the lysosomotropic base chloroquine were thus applied in our study, both of which would disturb endosomal acidification and increase the endosomal pH. We measured the effect of the endosomal acidification inhibitors on transfection efficiency by assaying the MMP-9 mRNA transcription level at 48 h after transfection to explore the endosomal release mechanisms ( Figure 7B).…”

“…We concluded that both polyplexes and liposomes were dependent on the proton sponge mechanism for endosomal escape. Ouyang et al, described the mode of siRNA release from cationic polymers as the proton sponge mechanism, 13 which implies that polymers sequester the protons transported into the endosomal compartment, thus preventing endosomal acidification. Subsequently, the increased osmolarity in this compartment causes the influx of water, which eventually leads to endosome lysis and the release of its contents into the cytoplasm.…”