The study investigates lipid nanoparticles for SARS-CoV-2

In a recent study posted on bioRxiv* Preprint server, researchers evaluated the efficiency of lipid nanoparticles against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).

Messenger ribonucleic acid (mRNA) vaccines have played an important role in controlling coronavirus disease 2019 (COVID-19) infection. However, the various side effects reported by vaccinated individuals and the inefficiency of the vaccine against novel SARS-CoV-2 mutants underscore the need for the development of new mRNA vaccines.

About research

In this study, the researchers produced a COVID-19 vaccine named Circular mRNA (cmRNA) -1130 against the SARS-CoV-2 delta (B.1.617.2) variant.

Using the Michael addition reaction, the team synthesized a series of ionizable lipids with several ester bonds between the branched tails. Proton nuclear magnetic resonance (¹The structure of lipid AX4 was verified using 1 H NMR) spectrum, and the possibility of vaccination was estimated by incorporating the lipid into lipid nanoparticles (LNP). The team then used firefly luciferase (Fluc) cyclic mRNA as a reporter for LNP to assess the delivery efficiency of the mRNA. Fluc protein expression was later measured in mice after intramuscular (IM) administration.

In addition, the team investigated the effect of lipidoid tail numbers on mRNA expression by producing AX4-2 and AX4-3, which have two and three X4s, respectively. To understand the effects of ester bonds present in the lipidoid tail, researchers replaced amide bonds with some or all of the ester bonds found in AX4. afterwards, In vivo The biodistribution of mRNA-AX4-LNP was determined using mRNA labeled with cyanine-5 (Cy5-mRNA).

The team also evaluated the efficacy and immunogenicity of the cmRNA-1130 vaccine, which encodes the receptor-binding domain (RBD) of SARS-CoV-2 spikes. Glycoprotein Of the delta variant. Serum samples were collected on days 14, 28, 42, 72, 102, and 132 days after administration of the cmRNA-1130 vaccine to BALB / c mice to the RBD / spike-specific antibody SARS-Cov. Achieved by evaluation. -2 Neutralizing antibody (NAbs) and RBD-specific T cells.

result

The results of the study showed that mice treated with Fluc mRNA-LNP had an ionizable lipid with a tail containing butyloctanoic acid (X4) and were more of mRNA compared to lipids with hexyldecanoic acid (X6). It was shown to show good delivery and higher expression of the protein. The team also observed that a tertiary amine with two amine groups on the polar head allows the introduction of up to four branched tails. These tertiary amines also showed superior delivery efficiency, including AX4 and AX6, with 0.2-fold higher protein expression than those observed with the commercially available Dlin-MC3-DMA lipid (MC3). In particular, mice treated with AX4 showed consistent expression of luciferase 24 hours after dosing and peak expression levels were observed 6 hours after dosing.

The team also found that incorporating ethers, esters, hydroxyls, and tertiary amines into the lipid head reduced protein expression compared to MC3. On the other hand, increasing the distance between the hydroxyl and amine groups improved delivery efficiency. Overall, the results show that AX4 enables the highest delivery efficiency.

Production of AX4-2 and AX4-3 showed that increasing the number of branched X4s increased protein expression levels. This was indicated by changes in protein levels of 3rd and 3x, respectively, in AX4 compared to AX4-2 and AX4-3. The significant reduction in protein expression in AX4-2 may be due to an increase in the number of secondary amines that may reduce the degree of mRNA release due to the formation of strong electrostatic interactions.

The team noted that the addition of amide bonds to the lipidoid tail reduced the intensity of luciferase expression. This is a result of reduced lipid degradation due to the presence of amine bonds, resulting in reduced mRNA release. In addition, higher protein expression was observed when the amide bond farthest from the tertiary amine was replaced compared to that adjacent to the tertiary amine. This is due to the breakdown of the ester bonds that form the carboxyl groups and the short alkyl chains that improve the mRNA. release.

Cy5-mRNA administration showed significant Cy5 fluorescence in the liver, kidney, and gallbladder 6 hours after injection, regardless of IM or intravenous (IV) administration. Interestingly, significant luciferase fluorescence was detected throughout the liver and mRNA-AX4-LNP showed substantial liver target protein expression. Meanwhile, the team noted that mRNA-AX4-LNP IV administration showed significant protein expression in apolipoprotein E (ApoE) knockout mice.

Administration of the cmRNA-1130 vaccine showed high levels of NAb after the second immunization with the vaccine.The team noted that mice vaccinated with cmRNA-1130 showed high deltas. Pseudo virus Neutralization showed that two doses of cmRNA-1130 induced high levels of SARS-CoV-2NAb against delta pseudovirus.

Overall, research results have shown that the combination of AX4-LNP and circular mRNA provides safe and effective protection against SARS-CoV-2.

*Important Notices

bioRxiv publishes unpeer-reviewed preliminary scientific reports and should not be considered definitive, guide clinical / health-related behaviors, or be treated as established information.