Optimising hybrid vesicle RNA formulations for enchanced stability and storage requirements

Nanoparticles (NP) are particles such as metals, lipids and polymers that range between 10-100nm in size and have a wide range of applications such as catalysis, magnetic materials, batteries, and drug delivery systems. There is Increasing interest into the use of NPs for the optimisation of drug delivery systems due to their ability to carry small molecules, and nucleic acids such as RNA.

Naked RNA is a negatively charged hydrophilic macromolecule that struggles to enter cells because of the electrostatic repulsion by the cellular membrane. Additionally, naked RNA can be degraded by RNAases, needing a protective carrier to facilitate internalisation. The encapsulation and delivery of these nucleic acids have gained attention due to their integral and effective use as a delivery platform that is the COVID-19 vaccine. However, the COVID-19 vaccine contains an anchored poly(ethylene glycol) (PEG) to stabilize the NP, according to recent publications, this had led to the development of PEG-specific antibodies in recipients. These antibodies can enhance the clearance of these delivered PEGylated NPs, thus reducing their efficacy and circulation time via the absorption of the mononuclear phagocyte system. These previously generated vaccines have different storage requirements indicating that different RNA formulations have varying nucleic acid affinity and nanoparticle stability. Therefore, using a hybrid vesicle delivery system that does not contain PEG, assessing the stability and storage of this hybrid vesicle would provide valuable insights and a better understanding of the nanoparticle assembly required.

The aim and goal for this project is to optimise the stability against functional performance on hybrid vesicle RNA formulations. This vesicle is a hybrid between a pure lipid and a pure co-block polymer - phosphatidylcholine/poly(ethylene oxide)-poly(butadiene) (POPC/PBd-PEO). Hybrid vesicles have been shown to provide a compromise between the biocompatibility of the lipid and the stability and robustness of polymer. This project will mainly focus on enhanced stability and storage conditions. The storage conditions assessed will be shelf life of the formulation and the ability to be stored at higher temperatures. Analysing these factors would include assessing the colloidal and chemical stability along with the particle structural stability. However, enhanced stability needs to be considered alongside maintaining a functional performance where the delivery of the encapsulated microRNA in the hybrid vesicle will be used in a model system to treat Kaposi's sarcoma-associated herpesvirus (KSHV)-infected cells, this will further assess the stability of the hybrid vesicle RNA formulation along with transfection ability.

A potential subsection of this project would be to assess the changes in physiochemical properties caused by nonspecific absorption of the nanoparticle by proteins resulting in the formation of protein corona at the interface of the hybrid vesicle. This protein corona formation changes characteristics of the nanoparticle and plays a significant role in biodistribution and endocytosis. A basic example of protein corona would give insight in physiochemical characteristic changes that may affect the function of the hybrid vesicle RNA formulation.