Enhancing the membrane destabilization of enzymatically released DNA Surfactant Conjugates for improved endosomal escape and gene expression

Enhancing the endosomal escape of nucleic acids remains a critical bottleneck to improve their therapeutic efficacy. Inspired by advancements in lipid nanoparticles, we describe a new chemical approach to deliver multi-tailed DNA surfactant conjugates with the potential for improved endosomal escape efficiency through enhanced membrane destabilizing properties. Previously we reported the successful delivery of therapeutic oligonucleotides to cells using single tailed DNA surfactant conjugates (DSCs) formed from enzyme degradable crosslinked micelles that we refer to as Nucleic Acid Nanocapsules (NANs). This work features the rational design of a new asymmetric set of hydrophobic crosslinkers using a synthetic approach which incorporates one or two alkyl chains and an esterase-labile group into a nonionic surfactant flanked by clickable azides. Through the covalent incorporation of these new crosslinkers into micelles and upon their enzymatic degradation within the cell, multi-tailed DNA surfactant conjugates are released which we show have superior transfection capabilities over single tailed DNA surfactant conjugates. We show that the multi-tailed DNA surfactant conjugates improve mRNA delivery and protein expression in vitro, as well as exhibit enhanced membrane disruption capabilities as seen by their dispersion in cells and through molecular dynamics simulations. Our chemical approach enables the formation of multi-tailed surfactants in situ, post micelle assembly, adding to the membrane destabilizing capacity of the delivered oligonucleotides. These studies are an exciting step towards using an oligonucleotide nanomaterial delivery platform as a “pro-drug” where the active therapeutic and potential for its cytosolic delivery is the product of the formulation’s degradation.