The world has recently seen immense activity in vaccine development due to the coronavirus disease 2019 (COVID-19) pandemic, as the causative virus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV- 2), continues to emerge in new variants with increased transmissibility, immune evasion capacity, and virulence attributes.
A new paper describes a novel pathway for prevention using messenger ribonucleic acid (mRNA) molecules that code for protective antibodies in the recipient.
The use of mRNA to encode vaccines as the SARS-CoV-2 spike protein was pioneered by Pfizer/BioNTech and Moderna vaccines. These were among many vaccine technologies pressed to be used to counter the relentless spread and rising death toll of the COVID-19 pandemic.
Additionally, monoclonal antibodies have also been isolated for their neutralizing activity against the virus, both before and after exposure. Emergency use authorization (EUA) has been obtained for four mAb protocols: the therapeutic mAbs casirivimab + imdevimab, bamlanivimab + etesevimab, and sotrovimab and the prophylactic mAbs tixagevimab + cilgavimab. Of these, the first two are no longer effective, and their EUA has been withdrawn.
These antibodies must be administered intravenously, requiring a medical environment. They must be delivered in relatively large doses of 10 to 100 mg kg−1 to compensate for the fact that only a small fraction reaches the site of interest. This drives up the cost of treatments, limiting their availability, especially in low- and middle-income (LMIC) countries.
Alternative methods of mAb production should be explored. The current article, published in Advanced sciences, discusses one such alternative, where a nebulization-ready formulation was designed to allow the introduction of mRNA encoding an antibody into the lungs to neutralize the disease. Both in vitro and live the results support the use of this new technology to fight not only COVID-19, but also other viral respiratory infections.
The use of mRNA is safe in that it does not enter the nucleus, unlike DNA or DNA-bearing viral vectors, which rely on their effect on nuclear entry and integration with the host DNA genome. Second, the circulating mRNA half-life is relatively short, avoiding long-term consequences. By reducing the required dose to one hundredth of the original amount, when administered directly into the airways rather than systemically, the overall cost of treatment is significantly reduced.
Previous research has shown this approach to be feasible, using intravenous mRNAs encapsulated in lipid nanoparticles (LNPs) encoding a neutralizing antibody against the chikungunya virus, which would be expressed in the liver.
Previous research by the same authors showed the ability to direct mRNA to the lungs to produce mAbs there. By avoiding having to introduce the recombinant spike protein, the researchers encoded a membrane anchor in the heavy chain of the immunoglobulin G (IgG) antibody molecule. This allowed the tissue to retain the antibody for several weeks.
The current study went further by switching from prior intratracheal administration to nebulization, which can be performed by the individual outside of a medical setting. This would allow mAbs to be expressed in high concentrations at the mucosal surface, the main entry site for respiratory viruses, while avoiding the need for large systemic doses.
What did the study show?
Study results show ability of nebulized mRNA-encoded neutralizing antibody (nAb) to counter SARS-CoV-2 infection in hamster lung, reducing virus count and alleviating signs lung disease and infection-related weight loss in hamsters.
The membrane anchor molecule glycosylphosphatidylinositol (GPI) allowed antibodies to remain bound to the cell membrane. The antibodies tested here included six mAbs isolated from B cells taken from individuals infected with SARS-CoV-2.
Direct stochastic optical reconstruction microscopy (dSTORM) has improved the ability to visualize antibodies once expressed in cell culture. One formed long ropes on the surface of the cell and was therefore disqualified for further testing live.
All bound and anchored mAbs retained their neutralizing capacity, as shown by their cytopathic effect (CPE) in monolayer cell culture. Although most of them were able to neutralize the original variant and the B.1.1.7 variant, one failed to compensate for the latter. The protective ability was linked to the specific antibodies encoded by the mRNA and not to the GPI anchor, as shown by a control antibody with a GPI attached.
All mAbs could neutralize the virus at low concentrations, with a low nanomolar half-maximal inhibitory concentration (IC50). Additional tests were performed using two selected mAbs, COV2-2832 and DH1041.
Following these promises in vitro results, further testing was performed on Syrian golden hamsters, which provide a robust animal model for human COVID-19 infection.
Longer retention in the lungs
This demonstrated that the antibodies were, as expected, anchored to the cell membrane in the lung by the GPI molecule. Compared to the secreted or unanchored form, it was found to remain in the lung tissue. At the same time, the latter caused an increase in serum concentrations, the difference in post-transfection serum levels being 27-fold in favor of the secreted form. This occurred despite efficient translation of both types in lung tissue after nebulization, peaking at 24-48 hours.
The Coded Anchor improved lung retention from just over a day to seven days, which could signify a single-dose approach for post-exposure treatment of COVID-19.
This approach would not be expensive compared to other nebulizer-based treatments, which usually consist of several doses per day or a single dose for up to 22 hours..”
The nebulized formulation reached all parts of the lung evenly and richly, both the alveolar space and the airway epithelium.
Since the virus is primarily found in the alveolar space, this finding indicates the potential of transfected antibodies to prevent COVID-19 and serious disease. The effective dose delivered to the lungs can be increased by increasing the concentration of the formulation.
The ratio of lung-delivered mRNA to total delivered mRNA is about 12% in hamsters, but is likely to be much higher, closer to 30-50%, in large mammals. This would mean that a much smaller amount of the nebulized formulation would be needed to achieve the required dosage.
These data indicate that a low dose of mRNA can achieve high expression of long-lasting mAb constructs in a large part of the alveolar and epithelial compartments of the hamster lung airways, with minimal pulmonary toxicity..”
Additionally, hamsters treated with the mRNA-encoded mAbs before being inoculated two days later with the virus showed a 1% weight gain on day 5, starting on the second day of treatment. At the same time, untreated exposed controls lost 5% of their body weight on the fifth day after inoculation.
Examination of lung tissue from treated and control hamsters showed more than 80% lower viral titers in the former, compared to controls, with an approximately 80% decrease in viral RNA load as measured by quantitative polymerase chain reaction (qPCR). Lung pathology was also largely attenuated in treated hamsters.
The study also suggested that weight loss was the most reliable correlate of animal health, while lung pathology reflects the strength of the immune response rather than the intensity of viral replication. The mRNA delivery did not cause lung inflammation or lung tissue damage.
What are the implications?
Despite the use of human mAbs in a hamster model, which shows suboptimal Fc-mediated immune functions, the present study demonstrated the ability of these antibodies, when expressed by nebulized mRNA in the lung tissue, to induce an encouraging degree of protection against the virus. This appears to be a successful option for passive immunization that reduces the time required for the host to neutralize the virus after infection while overcoming any deficiencies in the host’s own immune system.
The nebulization approach allows for self-administration, ensuring wider distribution in low-resource settings.
mRNA-expressed COV2-2832 and DH1041 provide a clear complementary prophylactic strategy to currently used therapies.”