Aerosolizable Lipid Nanoparticles for Pulmonary Delivery of mRNA through Design of Experiments.

Hairui Zhang,Melissa R Soto,Debadyuti Ghosh,Jasmim Leal,Hugh D C Smyth

doi:10.3390/pharmaceutics12111042

Hairui Zhang, Melissa R Soto + Show 3 more

Open Access

https://doi.org/10.3390/pharmaceutics12111042

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Journal: Pharmaceutics	Publication Date: Oct 30, 2020
Citations: 84	License type: CC BY 4.0

Affiliation: The University of Texas at Austin

Abstract

Messenger RNA is a class of promising nucleic acid therapeutics to treat a variety of diseases, including genetic diseases. The development of a stable and efficacious mRNA pulmonary delivery system would enable high therapeutic concentrations locally in the lungs to improve efficacy and limit potential toxicities. In this study, we employed a Design of Experiments (DOE) strategy to screen a library of lipid nanoparticle compositions to identify formulations possessing high potency both before and after aerosolization. Lipid nanoparticles (LNPs) showed stable physicochemical properties for at least 14 days of storage at 4 °C, and most formulations exhibited high encapsulation efficiencies greater than 80%. Generally, upon nebulization, LNP formulations showed increased particle size and decreased encapsulation efficiencies. An increasing molar ratio of poly-(ethylene) glycol (PEG)-lipid significantly decreased size but also intracellular protein expression of mRNA. We identified four formulations possessing higher intracellular protein expression ability in vitro even after aerosolization which were then assessed in in vivo studies. It was found that luciferase protein was predominately expressed in the mouse lung for the four lead formulations before and after nebulization. This study demonstrated that LNPs hold promise to be applied for aerosolization-mediated pulmonary mRNA delivery.

Highlights

Successful delivery of nucleic acids for gene replacement therapy and editing offers the promise of long-term correction and a cure for genetic diseases affecting the lung, such as cystic fibrosis (CF) and alpha-1 antitrypsin deficiency
To establish a baseline and initially identify the optimal N/P molar ratio desired for intracellular protein expression, six lipid nanoparticles (LNPs) formulations encapsulating Enhanced Green Fluorescent Protein (EGFP) messenger RNA (mRNA) were prepared by varying the N/P ratio between 6:1 to 200:1, using as reference a commercial formulation as previously published [28,33] for siRNA delivery in vivo
This work highlights a Design of Experiments (DOE) approach to discover LNP-mRNA formulations that can be aerosolized for pulmonary delivery

Summary

Introduction

Successful delivery of nucleic acids for gene replacement therapy and editing offers the promise of long-term correction and a cure for genetic diseases affecting the lung, such as cystic fibrosis (CF) and alpha-1 antitrypsin deficiency. Translate Bio, a company focusing on mRNA therapeutics, developed LNPs that carry cystic fibrosis transmembrane conductance regulator (CFTR) mRNA (MRT5005), and is the first company to execute a clinical trial investigating nebulized delivery of mRNA treatment [8]. While these studies reflect the ability of LNPs to be delivered to the lungs, there remain no reports using Design of Experiments (DOE) methodologies to investigate the stability and delivery post nebulization of different compositions of LNP libraries in order to achieve efficient mRNA expression in the lungs

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