Restriction lift date: 2029-09-30
Designing RNA nanoparticle formulations for oral delivery: influence of carrier chemistry, N/P ratio, and intestinal fluid components
Loading...
Files
Date
2025-12-31
Authors
O'Brien, Richard
Journal Title
Journal ISSN
Volume Title
Publisher
University College Cork
Published Version
Abstract
The development of RNA therapeutics represents a new frontier in the treatment of inflammatory bowel diseases such as Crohn’s disease, where current therapies are limited by incomplete remission rates, systemic side effects, and eventual need for surgical intervention. However, the oral delivery of RNA is severely hindered by the gastrointestinal environment, where pH gradients, enzymatic degradation, mucus entrapment, and restricted epithelial uptake impose significant barriers to stability and efficacy.
This thesis investigates the potential of nanoparticle systems-including dendrimer-based formulations and APE polymer-based nanoparticles-for the oral delivery of mRNA to the intestine. A library of amino-polyester and dendrimer polymers was synthesised and evaluated for mRNA encapsulation, particle size, stability, and transfection efficiency. Optimisation studies examined the impact of nitrogen-to-phosphate (N/P) ratios on complexation and performance in biologically relevant media, including simulated intestinal fluids and enzymatic conditions. Comparative transfections were carried out in undifferentiated and differentiated Caco-2 intestinal epithelial models to reflect differences in barrier function. Additional experiments assessed the effect of digestive enzymes such as pancreatin on nanoparticle integrity and explored inhibition strategies with EDTA.
Across these studies, both dendrimer and APE nanoparticles consistently achieved efficient RNA encapsulation with particle sizes in the 80-150 nm range and low polydispersity. Increasing N/P ratios marginally enhanced stability without compromising particle assembly. In vitro, potent gene expression was observed in Caco-2 cells. Exposure to simulated gastrointestinal fluids highlighted the destabilising effects of bile salts and enzymes, with partial protection afforded by formulation design. The impact of intestinal fluids was also shown to be highly biomaterial dependent. Collectively, these findings confirm that while nanoparticle carriers can shield mRNA payloads and mediate transfection in intestinal cells, the gastrointestinal environment remains the principal obstacle to effective oral delivery.
In conclusion, this thesis demonstrates the feasibility of nanoparticle-mediated oral RNA delivery and provides mechanistic insights into the challenges imposed by gut physiology. By identifying the critical formulation parameters that influence stability and uptake, this work contributes to the rational design of next-generation nanocarriers with the potential to advance RNA therapeutics for local treatment of Crohn’s disease and related intestinal disorders.
Description
Keywords
mRNA nanoparticles , Intestinal fluid , Polymer , Dendrimer
Citation
O’Brien, R. 2025. TDesigning RNA nanoparticle formulations for oral delivery: influence of carrier chemistry, N/P ratio, and intestinal fluid components. MRes Thesis, University College Cork.