C.S. Yang1, D. Vanover2, J.A. Assumpção2, J. Joo2, P. Kumar E. K.2, H.E. Peck2, P.J. Santangelo2, J. Lyons1 1Emory University Hospital, Department Of Surgery, Atlanta, GA, USA 2Emory University and Georgia Instititute of Technology, Wallace H. Coulter Department Of Biomedical Engineering, Atlanta, GA, USA
Introduction:
Messenger RNA (mRNA) therapeutics are highly versatile and can combat a wide range of diseases. Exogenous mRNA sequences are encapsulated in nanoparticles and delivered to cells, where they are translated to functional protein by host cell machinery. Several delivery methods have been examined for either systemic or organ-specific protein expression, including aerosolized delivery to lung tissue. However, this technique has largely been untested in the face of infection or active inflammation. Cellular stress responses repress mRNA translation during infection, potentially limiting the clinical relevance of this therapeutic approach. In this study, we tested the efficacy of polymer-encapsulated mRNA delivery and protein expression during Pseudomonas aeruginosa pneumonia.
Methods:
C57Bl/6j Mice were anesthetized and received a 40 uL intratracheal injection of Pseudomonas aeruginosa (3×10^8 CFU/mL) or saline. 24 hours after the procedure, mice received 300 ug of aerosolized luciferase-encoding mRNA encapsulated in either p76 or p81 polymer. Lungs were harvested 24 hours after mRNA delivery. To measure expression of luciferase protein, extracted whole lungs were incubated in luciferase substrate and radiance measurements were taken with an in-vivo imaging system (IVIS).
Results:
There was no significant difference in luciferase activity between sham and infected groups in both p76 (p=0.86, n=3/group) and p81 (p=0.77, n=1-3/group) conditions (Figure 1). Several mice succumbed to their infections in the initial study, so a subsequent experiment examining p81 delivery and translation with a larger sample size was conducted. Similarly, there was no significant change in radiance between the sham and infected groups (p=0.44, n=4-8/group). Furthermore, luciferase activity was 10-fold higher in the p81 group than in the p76 group (p=0.03, n=3/group) (Figure 1).
Conclusion:
Aerosolized mRNA delivery can achieve similar levels of protein expression in both healthy and infected lung tissue. P81, a novel polymer, yields greater protein levels than previously published polymers. It is unclear whether this polymer improves delivery of mRNA cargo or increases mRNA translation. These findings illustrate that inhalable mRNA is a promising therapeutic strategy even in the setting of active lung inflammation.