The Thermodynamics of Endosomal Escape and DNA Release from Lipoplexes

TL;DR

This paper models the thermodynamic forces behind endosomal escape and DNA release from lipoplexes, revealing how lipid interactions and composition influence transfection efficiency.

Contribution

It introduces a thermodynamic model explaining how lipoplex destabilization and DNA release are driven by lipid interactions and composition, advancing understanding of gene delivery mechanisms.

Findings

Lipoplex becomes thermodynamically unstable after endocytosis.

Free energy gain during lipoplex breakdown is linearly related to cationic lipid fraction.

Transfection efficiency exponentially increases with cationic lipid content.

Abstract

Complexes of cationic and neutral lipids and DNA (lipoplexes) are emerging as promising vectors for gene therapy applications. Their appeal stems from their non pathogenic nature and the fact that they self-assemble under conditions of thermal equilibrium. Lipoplex adhesion to the cell plasma membrane initiates a three-stage process termed transfection, consisting of (i) endocytosis, (ii) lipoplex breakdown, and (iii) DNA release followed by gene expression. As successful transfection requires lipoplex degradation, it tends to be hindered by the lipoplex thermodynamic stability; nevertheless, it is known that the transfection process may proceed spontaneously. Here, we use a simple model to study the thermodynamic driving forces governing transfection. We demonstrate that after endocytosis [stage (i)], the lipoplex becomes inherently unstable. This instability, which is triggered by interactions between the cationic lipids of the lipoplex and the anionic lipids of the enveloping plasma membrane, is entropically controlled involving both remixing of the lipids and counterions release. Our detailed calculation shows that the free energy gain during stage (ii) is approximately linear in $\Phi_+$, the mole fraction of cationic lipids in the lipoplex. This free energy gain, $\Delta F$, reduces the barrier for fusion between the enveloping and the lipoplex bilayers, which produces a hole allowing for DNA release [stage (iii)]. The linear relationship between $\Delta F$ and the fraction of cationic lipids explains the experimentally observed exponential increase of transfection efficiency with $\Phi_+$ in lamellar lipoplexes.