Strikingly, the chimeric VHH1

Strikingly, the chimeric VHH1.1, both VHH-IgG1.1 and VHH-IgA1.1, showed over 3-folds increased binding activity against RBD antigen of SARS-CoV-2 (VHH-IgG1.1 EC50 = 0.737nM and VHH-IgA1.1 EC50 = 0.774nM) when compared to the monomeric VHH1.1 (EC50 = 2.359nM) ( Figure?1B). variant and its sub lineages BA.1.1, BA.2 and BA.2.12.1. VHH-IgA1.1 is also much more potent against Omicron variants as compared to an IgG Fc fusion construct, demonstrating the importance of IgA mediated mucosal protection for Omicron infection. Intranasal administration of VHH-IgA1.1 prior to or after challenge conferred significant protection from severe respiratory disease in K18-ACE2 transgenic mice infected with SARS-CoV-2 VOC. More importantly, for cost-effective production, VHH-IgA1.1 produced in had comparable potency to mammalian produced antibodies. Our study demonstrates that intranasal administration of affordably produced VHH-IgA fusion protein provides effective mucosal immunity against infection of SARS-CoV-2 including emerging variants. Keywords: biological sciences, microbiology, SARS-CoV-2, VOC, nanobody, IgA, neutralization, antiviral prophylaxis and therapeutics Introduction SARS-CoV-2 is a coronavirus that has led to a global pandemic and causes a severe respiratory disease known as COVID-19. The rapid spread of SARS-CoV-2 Rabbit Polyclonal to OR5AP2 globally has resulted in hundreds of millions of infections and over 6.1 million deaths as of March 2022 (1). Despite the rollout of first-generation vaccines and monoclonal antibody therapeutics, additional preventive modalities are still required for breakthrough infections and unvaccinated individuals. Newly emerged SARS-CoV-2 variants of concern (VOC) and interest (VOI) are continuing to evolve globally, including some in which the effectiveness of monoclonal antibodies and vaccines is diminished (2C6). The latest and heavily mutated Omicron and its sub-variants, also exhibit increased transmissibility and risks of infection (7). Thus, new broad variant-resistant treatments and non-invasive delivery strategies remain a high priority (8C10). Clinical trials have demonstrated that SARS-CoV-2 receptor-binding domain (RBD) targeted neutralizing IgG monoclonal antibodies (MAbs) are safe and effective against COVID-19. Pre- or post-exposure treatment with neutralizing IgG antibodies provide immediate immunity against SARS-CoV-2 in vulnerable patient populations (11, 12). A number of IgG antibodies have received emergency authorization for clinical use (13). However, emerging SARS-CoV-2 VOC continue to diminish the effectiveness of these antibodies (8C10). Intravenous infusion of IgG is not only invasive but also costly with the traditional CHO cell bio-manufacturing platform. Recently, our group demonstrated that compared to IgG, a human IgA monoclonal antibody, MAb362, is more potent at neutralizing SARS-CoV-2 in immunoglobulins natural mucosal form: secretory IgA (14). This study raised the possibility of using BIBR-1048 (Dabigatran etexilate) mucosal IgA as prophylactic therapy against SARS-CoV-2 directly at the infection sites of the respiratory tract. The relevance of this to SARS-CoV-2 infection has been the demonstration that breakthrough infections in vaccinated individuals were seen more frequently in those with lower serum IgA responses to RBD (15). Furthermore, intranasal vaccine boost elicited significantly stronger mucosal IgA responses and provided complete protection of mice from infection (16). Given that SARS-CoV-2 is a respiratory infection, the mucosal response may be more contributory to protection than what is measured in the serum. The camelid heavy-chain-only antibodies (known as nanobodies or VHHs) are a specific alternative class of monoclonal antibodies, which are single-domain antigen binding fragments derived from Alpaca and Llama. These antigen-binding variable domains are relatively small (~15 kDa), soluble, and highly stable with no associated light chains. Like conventional monoclonal antibodies, nanobodies have emerged as very promising antibody-based therapeutic treatments, diagnostic tools or delivery systems for many diseases, including cancer, infectious disease, neurodegenerative BIBR-1048 (Dabigatran etexilate) disorders, immune diseases and rare BIBR-1048 (Dabigatran etexilate) blood diseases (17C27). Compared to monoclonal antibodies, nanobodies are unique biologics that often recognize conserved epitopes on hypervariable pathogens. Due to their smaller paratope diameters and longer complementarity-determining region 3 (CDR3), nanobodies can access structurally distinct, spatially restricted epitopes, such as highly conserved epitopes in recessed BIBR-1048 (Dabigatran etexilate) regions of viral glycoproteins (28, 29). These unique biophysical advantages have led to the evaluation of mucosal delivery of nanobodies for preventing/treating respiratory pathogens, including respiratory syncytial virus (RSV), whereby nebulized nanobodies greatly reduced RSV infection in newborn lambs (30, 31). To date, there is no mucosal delivered therapeutic nanobody that has been approved by the FDA. In 2019 Caplacizumab (Sanofi), the first nanobody based drug, was approved by FDA to treat acquired thrombotic thrombocytopenic purpura (aTTP) intravenous and subcutaneous injection (20, 32). Considering more than a dozen nanobodies are currently at different stages of clinical development (18), nanobodies are progressively BIBR-1048 (Dabigatran etexilate) being demonstrated as a valid clinical alternative to monoclonal antibody therapy to treat.