Study: Delivery of Circular mRNA via Degradable Lipid Nanoparticles against SARS-CoV-2 Delta Variant. Image Credit: Crevis/Shutterstock

Study examines lipid nanoparticles against SARS-CoV-2

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

Study: Delivery of circular mRNA via degradable lipid nanoparticles against SARS-CoV-2 Delta Variant. Image Credit: Crevis/Shutterstock

Messenger ribonucleic acid (mRNA) vaccines have played an important role in curbing the transmission of coronavirus disease 2019 (COVID-19). However, the varying side effects reported by vaccinees, as well as the vaccine inefficiency against new SARS-CoV-2 variants, emphasize the need for the development of new mRNA vaccines.

About the study

In the current study, researchers produced a COVID-19 vaccine called circular mRNA (cmRNA)-1130 against the variant SARS-CoV-2 Delta (B.1.617.2).

Using the Michael addition reaction, the team synthesized a series of ionizable lipids with different ester bonds between the branched tails. Proton nuclear magnetic resonance (1H NMR) spectra were used to verify the structure of a lipid AX4, while estimating its vaccination potential by formulating the lipid into lipid nanoparticles (LNPs). The team then evaluated the delivery efficiency of the mRNA by using the circular firefly luciferase (Fluc) mRNA as a reporter in the LNP. The expression of the Fluc protein was later measured in mice after intramuscular (IM) administration.

Furthermore, the team investigated the impact of the tail number of the lipidoids on mRNA expression by generating AX4-2 and AX4-3 with two and three X4, respectively. To understand the effect of ester bonds in the lipidoid tails, the researchers replaced the amide bonds with some or all of the ester bonds in AX4. Then the in vivo mRNA-AX4-LNP biodistribution was determined using mRNA labeled with cyanine-5 (Cy5 mRNA).

The team also evaluated the efficacy and immunogenicity of the cmRNA-1130 vaccine encoding the receptor binding domain (RBD) at the SARS-CoV-2 peak glycoprotein of the Delta variant. This was achieved by administration of the cmRNA-1130 vaccine to BALB/c mice, followed by the collection of serum samples on days 14, 28, 42, 72, 102 and 132 along with evaluation of RBD/spike specific antibodies, SARS Cov -2 neutralizing antibodies (NAbs) and RBD-specific T cell.

Results

The study results showed that the mice receiving the Fluc mRNA-LNP had ionizable lipids with a tail containing butyloctanoic acid (X4), showed better mRNA release and higher protein expression compared to lipids containing hexyldecanoic acid (X6). The team also noted that the tertiary amines with two amine groups in the polar head allowed the introduction of up to four branched tails. These tertiary amines also showed better delivery efficiency, including AX4 with two- and AX6 with 0.2-fold higher expression of proteins than that observed in the commercially available Dlin-MC3-DMA lipid (MC3). Notably, mice treated with AX4 had consistent expression of luciferase 24 hours after dosing, with the peak expression level observed at 6 hours after dosing.

The team also found that the incorporation of ether, ester, hydroxyl and tertiary amine into the lipid head resulted in less protein expression compared to that in MC3. On the other hand, increasing the distance between the hydroxyl and the amine groups improved the delivery efficiency. Overall, the results showed that AX4 enabled the highest delivery efficiency.

The generation of AX4-2 and AX4-3 showed that increasing the number of branched X4 resulted in the improvement of protein expression levels. This was indicated by the change in protein levels by three orders and three times in AX4 compared to those in AX4-2 and AX4-3, respectively. The substantial decrease in protein expression in AX4-2 was probably due to the increase in the number of secondary amines that could reduce the rate of mRNA release due to the formation of strong electrostatic interactions.

The team noted that the addition of amide linkages to the lipidoid tails reduced the intensity of luciferase expression, which resulted from reduced lipid breakdown due to the presence of the amine linkages, leading to less mRNA release. In addition, higher protein expression was observed when the amide linkages furthest from the tertiary amine were replaced compared to those adjacent to the tertiary amide, which was attributed to the breakdown of ester linkages producing carboxyl groups, as well as shorter alkyl chains enhancing the mRNA. . Edition.

Administration of Cy5 mRNA showed significant Cy5 fluorescence in the liver, kidney and gallbladder six hours after injection, regardless of IM or intravenous (IV) administration. Interestingly, remarkable luciferase fluorescence was detected throughout the liver, indicating that mRNA-AX4-LNP had significant liver-targeted protein expression. On the other hand, the team noted that administration of mRNA-AX4-LNP IV showed significant protein expression in apolipoprotein E (ApoE) knockout mice.

The administration of the cmRNA-1130 vaccine showed a high level of NAbs after the second immunization with the vaccine. The team noted that the cmRNA-1130 vaccinated mice had high Delta . showed pseudovirus neutralization indicating that two doses of cmRNA-1130 induced a high level of SARS-CoV-2 NAbs against Delta pseudovirus.

Overall, the study results showed that the combination of AX4-LNP and circular mRNA provided safe and effective protection against SARS-CoV-2.

*Important announcement

bioRxiv publishes preliminary scientific reports that are not peer-reviewed and therefore should not be considered conclusive, should guide clinical practice/health-related behavior, or be treated as established information.

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