Expanding lipid nanoparticle platforms for effective mRNA therapeutics
Authors:   , Estapé Sentí Mariona 
Published: September 2024

Small molecule drugs have been the cornerstone of drug development for decades, offering treatments for various conditions and significantly improving countless lives. However, traditional drug development faces challenges as many diseases remain untreatable. RNA therapeutics have recently emerged as a promising alternative, offering potential treatments for cancer, infections, and genetic disorders. Their key advantage is targeting genetic instructions within cells, treating diseases in ways small molecule drugs cannot.

A major challenge for RNA therapeutics is delivering these fragile molecules to specific locations in the body. RNA degrades easily outside cells, so researchers developed protective systems. Lipid nanoparticles (LNPs) have shown the most promise, protecting RNA and ensuring it reaches its target. LNPs are typically composed of four lipids: ionizable lipids, non-cationic phospholipids, cholesterol, and PEG-lipids. Together, these components help RNA enter cells, evade immune detection, and remain stable during delivery.

However, challenges remain in optimizing LNPs for effective delivery. A key hurdle is ensuring RNA reaches the right location without accumulating in the liver. Another focus is the LNP coating, particularly PEG-lipids, which help evade the immune system but may also hinder interaction with target cells. Over time, the immune system can recognize PEG-lipids, reducing future treatment effectiveness. Ongoing research aims to find the right balance for efficient delivery while minimizing immune responses.

Overall, LNPs play a pivotal role on RNA therapeutics and have enabled their therapeutic success but still require extensive improvement to ensure the efficacy and safety of RNA therapeutics. The work described in this thesis is aimed at addressing this need, with a main aim of contributing into the development of the revolutionary LNP-based RNA therapeutics. In this thesis, we reviewed the basics of mRNA cancer vaccines and discussed how mRNA triggers the immune system and the ways scientists modify it to enhance its stability and reduce its immunogenicity. We also extensively reviewed the different drug delivery systems to deliver mRNA cancer vaccines. Additionally, we provided a thorough examination of LNPs, highlighting the function and challenges associated with each component. We went on to investigate the mechanism and consequences of complement activation triggered by anti-PEG antibodies, which arise from the immune system's response to the PEG-lipid, on PEGylated lipid nanoparticles. In chapter 4, we applied a Design of Experiments approach to refine LNP formulations for delivering mRNA directly into tumors. This method enables us to systematically evaluate multiple factors simultaneously, maximizing efficiency in identifying the optimal formulation. We also show experimentally how using a pair of complimentary peptides can improve the efficiency of LNP-mediated mRNA delivery to the heart when injected directly into this tissue. In Chapter 6, we investigated the effectiveness of delivering mRNA to the heart using LNPs and compared their performance to that of naked RNA, the current standard for direct heart injection. Additionally, we examined which cell types are transfected following intramyocardial LNP injection. Finally, we summarized the key findings from the thesis, reflecting on the results and discussing the current challenges and future prospects for lipid-based delivery systems in therapy.