Mohamad‐Gabriel Alameh

Recent years have witnessed incredible growth in RNA therapeutics, which has benefited significantly from decades of research on lipid nanoparticles, specifically its key component—the ionizable lipid. This comment discusses the major ionizable lipid types, and provides perspectives for future development. RNA therapeutics have benefited significantly from decades of research on lipid nanoparticles, specifically its key component—the ionizable lipid. This comment discusses the major ionizable lipid types, and provides perspectives for future development.

The recent success of mRNA vaccines in SARS-CoV-2 clinical trials is in part due to the development of lipid nanoparticle delivery systems that not only efficiently express the mRNA-encoded immunogen after intramuscular injection, but also play roles as adjuvants and in vaccine reactogenicity. We present an overview of mRNA delivery systems and then focus on the lipid nanoparticles used in the current SARS-CoV-2 vaccine clinical trials. The review concludes with an analysis of the determinants of the performance of lipid nanoparticles in mRNA vaccines.

Lipid Nanoparticles (LNPs) are used to deliver siRNA and COVID-19 mRNA vaccines. The main factor known to determine their delivery efficiency is the pKa of the LNP containing an ionizable lipid. Herein, we report a method that can predict the LNP pKa from the structure of the ionizable lipid. We used theoretical, NMR, fluorescent-dye binding, and electrophoretic mobility methods to comprehensively measure protonation of both the ionizable lipid and the formulated LNP. The pKa of the ionizable lipid was 2-3 units higher than the pKa of the LNP primarily due to proton solvation energy differences between the LNP and aqueous medium. We exploited these results to explain a wide range of delivery efficiencies in vitro and in vivo for intramuscular (IM) and intravascular (IV) administration of different ionizable lipids at escalating ionizable lipid-to-mRNA ratios in the LNP. In addition, we determined that more negatively charged LNPs exhibit higher off-target systemic expression of mRNA in the liver following IM administration. This undesirable systemic off-target expression of mRNA-LNP vaccines could be minimized through appropriate design of the ionizable lipid and LNP.

SARS-CoV-2 mRNA vaccination induces robust humoral and cellular immunity in the circulation; however, it is currently unknown whether it elicits effective immune responses in the respiratory tract, particularly against variants of concern (VOCs), including Omicron. We compared the SARS-CoV-2 S-specific total and neutralizing antibody responses, and B and T cell immunity, in the bronchoalveolar lavage fluid (BAL) and blood of COVID-19-vaccinated individuals and hospitalized patients. Vaccinated individuals had significantly lower levels of neutralizing antibody against D614G, Delta (B.1.617.2), and Omicron BA.1.1 in the BAL compared with COVID-19 convalescents despite robust S-specific antibody responses in the blood. Furthermore, mRNA vaccination induced circulating S-specific B and T cell immunity, but in contrast to COVID-19 convalescents, these responses were absent in the BAL of vaccinated individuals. Using a mouse immunization model, we demonstrated that systemic mRNA vaccination alone induced weak respiratory mucosal neutralizing antibody responses, especially against SARS-CoV-2 Omicron BA.1.1 in mice; however, a combination of systemic mRNA vaccination plus mucosal adenovirus-S immunization induced strong neutralizing antibody responses not only against the ancestral virus but also the Omicron BA.1.1 variant. Together, our study supports the contention that the current COVID-19 vaccines are highly effective against severe disease development, likely through recruiting circulating B and T cell responses during reinfection, but offer limited protection against breakthrough infection, especially by the Omicron sublineage. Hence, mucosal booster vaccination is needed to establish robust sterilizing immunity in the respiratory tract against SARS-CoV-2, including infection by the Omicron sublineage and future VOCs.