Aug 2026· ACS Applied Materials and Interfaces· Vol 18, pp. 45954-45970· 0 citations· 54 references
Medicine
TL;DR
This work describes a chemical approach to deliver multi-tailed DNA-surfactant conjugates into cells using nucleic acid nanocapsules (NANs) and shows that these multi-tailed DSCs improve DNAzyme-mediated gene silencing and enhance mRNA delivery in vitro, owing to their enhanced membrane disruption capabilities reflected in both cell studies and molecular dynamics simulations.
Abstract
Enhancing the endosomal escape of nucleic acids remains a critical bottleneck in improving their therapeutic efficacy. Inspired by advancements in lipid nanoparticles, we describe a chemical approach to deliver multi-tailed DNA-surfactant conjugates (DSCs) into cells using nucleic acid nanocapsules (NANs). NANs are enzyme degradable, crosslinked DNA micelles that exhibit enhanced membrane destabilizing properties. Previously, we reported the successful delivery of therapeutic oligonucleotides to cells using single-tailed DSCs delivered by esterase-cleavable NANs. This work features the rational design of a set of asymmetrically synthesized hydrophobic micelle crosslinkers using a synthetic approach that incorporates either one or two alkyl chains and an esterase-labile group into a nonionic surfactant flanked by clickable azides. These crosslinkers are covalently incorporated into the NAN construct, and upon their enzymatic degradation within the cell, multi-tailed DSCs are released. We show that these DSCs have superior transfection capabilities over single-tailed DSCs. Additionally, we show that these multi-tailed DSCs improve DNAzyme-mediated gene silencing and enhance mRNA delivery in vitro, owing to their enhanced membrane disruption capabilities reflected in both cell studies and molecular dynamics simulations. Importantly, our chemical approach enables the formation of multi-tailed surfactants in situ, post micelle assembly. This allows an oligonucleotide nanomaterial delivery platform to act as a "pro-drug" where the active therapeutic and the potential for its cytosolic delivery are the product of the nanoformulation's degradation.
Spherical nucleic acids (SNAs) enhance antisense oligonucleotide (ASO) uptake and nuclease resistance, but their activity is often constrained by inefficient intracellular trafficking. Here, we developed a zwitterionic brush-engineered molecular SNA (EmSNA) to improve ASO delivery. EmSNA was constructed by grafting poly(carboxybetaine) side chains onto a unimolecular ASO nanostructure, generating a compact, hydrated, near-neutral corona that modulated interfacial charge, oligonucleotide crowding, and intracellular processing. This architecture preserved rapid, sequence-specific hybridization while increasing duplex thermal stability and nuclease resistance. The zwitterionic brush further enhanced cellular uptake and reduced lysosomal sequestration, thereby improving antisense activity. Using a HER2-targeted ASO as a model cargo, EmSNA achieved stronger HER2 silencing in vitro and greater tumor growth inhibition in a xenograft model after local intratumoral administration, without overt systemic toxicity under the tested conditions. These findings establish zwitterionic brush engineering as an effective strategy to enhance functional SNA-based ASO delivery.
Rui Xu, Xiuying Sun, Li Yan et al.· Nano letters (Print)· 0 citations
The efficient cytosolic delivery of nucleic acid molecular machines remains a major challenge due to the dual barriers of the cell membrane and endosomal sequestration. Here, we report a class of amphiphilic framework nucleic acids (ampFNAs) that enable direct cytosolic delivery involving energy-independent traversal of lipid membranes. Among several designed geometries, a rigid rod-like six-helix bundle functionalized with a single cholesterol moiety exhibits superior cellular binding and internalization. We find that this ampFNA can enter cells through a cholesterol-dependent, lipid raft-mediated pathway, capable of bypassing endosomal entrapment. Compared to the commercial transfection reagent Lipofectamine 3000 (Lipo3000), the ampFNA platform exhibits reduced lysosomal entrapment. When delivering small interfering RNAs (siRNAs), the ampFNA-mediated enhanced green fluorescent protein (EGFP) gene silencing was achieved with efficiency comparable to Lipo3000. By targeting the proto-oncogene Bcl-2, the ampFNA induced an apoptotic rate of 31.3% in the tumor cell population. Our work establishes ampFNAs as a programmable, efficient, and biocompatible platform for the development of next‑generation smart nucleic acid delivery machines for precision medicine.
Lixuan Lin, Kai Jiao, Biancheng Wei et al.· Angewandte Chemie· 0 citations
Lipid nanoparticles (LNPs) are the leading platform for mRNA delivery, with their in vivo performance governed by lipid composition and colloidal stability. While anionic helper lipids can bias LNP expression toward the spleen, weak RNA-lipid interactions during purification often induce nanoparticle rearrangement and reduced activity. These stability limitations effectively narrow the accessible formulation design and screening space, leaving large regions of anionic compositional space underexplored. Here, we extend our cleavable crosslinking strategy to stabilize anionic LNPs without replacing the primary lipid constituents of the parent LNP formulation. By tuning the lengths of the cholesterol-derived acid-cleavable crosslinker and PEG-diamine, we achieved balanced structural stability. The optimized crosslinked formulation exhibited a significant increase in splenic mRNA expression at 12 h compared to the uncrosslinked LNPs. Notably, 33.2% of CD45+ tdTomato+ cells in the spleen were identified as T cells. Mechanistic analyses suggest that controlled mRNA release and altered intracellular processing contribute to the improved transfection efficiency. Together, these findings define a tunable crosslinking window that expands the accessible design landscape for tissue- and cell-specific mRNA delivery.
Yunhe Su, Joseph Choy, Xiang Liu et al.· ACS Applied Materials and In...· 0 citations
Comparing two cationic reverse-hexagonal LCNs formulated from monoolein, oleic acid, poly(allylamine hydrochloride) and either poloxamer 407 (P407) or poloxamer 188 (P188) reveals a direct relationship between LCN composition, membrane affinity, and functional performance, providing mechanistic insight for the rational design of LCN-based carriers in RNA therapeutics.
Ana Vitória Pupo Silvestrini, Márcia Carvalho de Abreu Fantini, Ana Paula Ramos et al.· International Journal of Pha...· 0 citations
Lipid nanoparticles (LNPs) have enabled the clinical translation of various nucleic acid cargos, including messenger RNA. Traditional LNP formulations contain a small proportion of lipids conjugated to poly(ethylene glycol), or PEG, which provides advantages of size control, stability, and extended circulation. However, PEGylation of both LNPs and other nano‐carriers raises challenges to efficient gene delivery, including endogenous production of anti‐PEG antibodies, accelerated clearance of multiple doses of LNPs, and reduced cellular uptake and endosomal escape. Here, we design and characterize a fully PEG‐free LNP platform, stabilized alternatively through the electrostatic adsorption of charged polymeric coatings that leverage principles of layer‐by‐layer self‐assembly. A library of carboxylated polyanions confers stability to non‐PEGylated (nonPEG) LNPs under biological stresses. nonPEG layered LNPs (LLNPs) significantly improve transfection of cancer and immune cells in vitro and generate favorable transfection in vivo via multiple routes of administration. In particular, nonPEG LLNPs reduce hepatic transfection by an order of magnitude, a major challenge for targeted LNP gene delivery, while maintaining similar circulation and transfection in other organs. Ultimately, this platform offers a highly modular, tunable approach to incorporate various bioactive moieties to achieve PEG‐free extrahepatic gene delivery.
This Perspective revisits cationic polymer-lipid nanoparticles not as historical precursors to ionizable lipid LNPs, but as a macromolecular materials space that can clarify and extend current RNA delivery design.
