rAAV expressing recombinant antibody for emergency prevention and long-term prophylaxis of COVID-19.

Introduction: Numerous agents for prophylaxis of SARS-CoV-2-induced diseases are currently registered for the clinical use. Formation of the immunity happens within several weeks following vaccine administration which is their key disadvantage. In contrast, drugs based on monoclonal antibodies, enable rapid passive immunization and therefore can be used for emergency pre- and post-exposure prophylaxis of COVID-19. However rapid elimination of antibody-based drugs from the circulation limits their usage for prolonged pre-exposure prophylaxis. Methods: In current work we developed a recombinant adeno-associated viral vector (rAAV), expressing a SARS-CoV-2 spike receptor-binding domain (RBD)-specific antibody P2C5 fused with a human IgG1 Fc fragment (P2C5-Fc) using methods of molecular biotechnology and bioprocessing. Results and discussions: A P2C5-Fc antibody expressed by a proposed rAAV (rAAV-P2C5-Fc) was shown to circulate within more than 300 days in blood of transduced mice and protect animals from lethal SARS-CoV-2 virus (B.1.1.1 and Omicron BA.5 variants) lethal dose of 105 TCID50. In addition, rAAV-P2C5-Fc demonstrated 100% protective activity as emergency prevention and long-term prophylaxis, respectively. It was also demonstrated that high titers of neutralizing antibodies to the SARS-CoV-2 virus were detected in the blood serum of animals that received rAAV-P2C5-Fc for more than 10 months from the moment of administration.Our data therefore indicate applicability of an rAAV for passive immunization and induction of a rapid long-term protection against various SARS-CoV-2 variants.

Single-domain antibody delivery using an mRNA platform protects against lethal doses of botulinum neurotoxin A.

Single-domain antibodies (sdAbs, VHHs, or nanobodies) are a promising tool for the treatment of both infectious and somatic diseases. Their small size greatly simplifies any genetic engineering manipulations. Such antibodies have the ability to bind hard-to-reach antigenic epitopes through long parts of the variable chains, the third complementarity-determining regions (CDR3s). VHH fusion with the canonical immunoglobulin Fc fragment allows the Fc-fusion single-domain antibodies (VHH-Fc) to significantly increase their neutralizing activity and serum half-life. Previously we have developed and characterized VHH-Fc specific to botulinum neurotoxin A (BoNT/A), that showed a 1000-fold higher protective activity than monomeric form when challenged with five times the lethal dose (5 LD50) of BoNT/A. During the COVID-19 pandemic, mRNA vaccines based on lipid nanoparticles (LNP) as a delivery system have become an important translational technology that has significantly accelerated the clinical introduction of mRNA platforms. We have developed an mRNA platform that provides long-term expression after both intramuscular and intravenous application. The platform has been extensively characterized using firefly luciferase (Fluc) as a reporter. An intramuscular administration of LNP-mRNA encoding VHH-Fc antibody made it possible to achieve its rapid expression in mice and resulted in 100% protection when challenged with up to 100 LD50 of BoNT/A. The presented approach for the delivery of sdAbs using mRNA technology greatly simplifies drug development for antibody therapy and can be used for emergency prophylaxis.

Single-Domain Antibodies Efficiently Neutralize SARS-CoV-2 Variants of Concern.

Virus-neutralizing antibodies are one of the few treatment options for COVID-19. The evolution of SARS-CoV-2 virus has led to the emergence of virus variants with reduced sensitivity to some antibody-based therapies. The development of potent antibodies with a broad spectrum of neutralizing activity is urgently needed. Here we isolated a panel of single-domain antibodies that specifically bind to the receptor-binding domain of SARS-CoV-2 S glycoprotein. Three of the selected antibodies exhibiting most robust neutralization potency were used to generate dimeric molecules. We observed that these modifications resulted in up to a 200-fold increase in neutralizing activity. The most potent heterodimeric molecule efficiently neutralized each of SARS-CoV-2 variant of concern, including Alpha, Beta, Gamma, Delta and Omicron variants. This heterodimeric molecule could be a promising drug candidate for a treatment for COVID-19 caused by virus variants of concern.