High Potency of a Bivalent Human VH Domain in SARS-CoV-2 Animal Models.

Novel COVID-19 therapeutics are urgently needed. We generated a phage-displayed human antibody VH domain library from which we identified a high-affinity VH binder ab8. Bivalent VH, VH-Fc ab8, bound with high avidity to membrane-associated S glycoprotein and to mutants found in patients. It potently neutralized mouse-adapted SARS-CoV-2 in wild-type mice at a dose as low as 2 mg/kg and exhibited high prophylactic and therapeutic efficacy in a hamster model of SARS-CoV-2 infection, possibly enhanced by its relatively small size. Electron microscopy combined with scanning mutagenesis identified ab8 interactions with all three S protomers and showed how ab8 neutralized the virus by directly interfering with ACE2 binding. VH-Fc ab8 did not aggregate and did not bind to 5,300 human membrane-associated proteins. The potent neutralization activity of VH-Fc ab8 combined with good developability properties and cross-reactivity to SARS-CoV-2 mutants provide a strong rationale for its evaluation as a COVID-19 therapeutic.

Rapid identification of a human antibody with high prophylactic and therapeutic efficacy in three animal models of SARS-CoV-2 infection.

Effective therapies are urgently needed for the SARS-CoV-2/COVID-19 pandemic. We identified panels of fully human monoclonal antibodies (mAbs) from large phage-displayed Fab, scFv, and VH libraries by panning against the receptor binding domain (RBD) of the SARS-CoV-2 spike (S) glycoprotein. A high-affinity Fab was selected from one of the libraries and converted to a full-size antibody, IgG1 ab1, which competed with human ACE2 for binding to RBD. It potently neutralized replication-competent SARS-CoV-2 but not SARS-CoV, as measured by two different tissue culture assays, as well as a replication-competent mouse ACE2-adapted SARS-CoV-2 in BALB/c mice and native virus in hACE2-expressing transgenic mice showing activity at the lowest tested dose of 2 mg/kg. IgG1 ab1 also exhibited high prophylactic and therapeutic efficacy in a hamster model of SARS-CoV-2 infection. The mechanism of neutralization is by competition with ACE2 but could involve antibody-dependent cellular cytotoxicity (ADCC) as IgG1 ab1 had ADCC activity in vitro. The ab1 sequence has a relatively low number of somatic mutations, indicating that ab1-like antibodies could be quickly elicited during natural SARS-CoV-2 infection or by RBD-based vaccines. IgG1 ab1 did not aggregate, did not exhibit other developability liabilities, and did not bind to any of the 5,300 human membrane-associated proteins tested. These results suggest that IgG1 ab1 has potential for therapy and prophylaxis of SARS-CoV-2 infections. The rapid identification (within 6 d of availability of antigen for panning) of potent mAbs shows the value of large antibody libraries for response to public health threats from emerging microbes.

Immune Responses to MERS-CoV in Humans and Animals.

Middle East Respiratory Syndrome Coronavirus (MERS-CoV) is an emerging zoonotic coronavirus that circulates in dromedary camels and sporadically transmit into humans, subsequently resulting in community and nosocomial cases. The viral infection in humans has a range of disease severity from asymptomatic to severe pneumonia and death, whereas the infection in camels is usually asymptomatic. There is no approved antiviral therapy or vaccine for MERS-CoV infections although there have been a number of therapeutic and vaccine candidates under development, for both humans and camels. To date, there has been limited research on the immune responses and pathogenesis of MERS-CoV in both humans and camels. Here, this chapter is focused on MERS-CoV specific immunity in different species with some details regarding the various animal models.

Unique aspects of adaptive immunity in camelids and their applications.

Members of the Camelidae have unique adaptive immunological features that are not widely observed in other species. All camelids are known to have three distinct IgG isotypes - IgG1, IgG2 and IgG3. While IgG1 has a conventional antibody structure, both IgG2 and IgG3 are devoid of light chains and instead possess hypervariable regions in their heavy chain (VHH), while lacking the typical CH1 domain found in heavy chains. VHH domains are increasingly being utilized as "next generation" antibodies, as they have unique biochemical and structural properties including high pH stability as well as a lower molecular weight allowing for increased tissue penetration. These features of VHH domains offer a number of advantages for both biotechnology and clinical applications and are commonly termed "nanobodies". A second unique aspect of the camelid adaptive response is involves T cell-mediated immunity. Characterization of gamma delta (ꝩδ) T cells in camelid species has found they use somatic hypermutation in their T cell receptor gamma (TRG) and delta (TRD) loci to increase the structural stability of their ꝩδ T receptor. The use of somatic hyper mutation to increase the diversity of their T cell repertoire, is a feature that has not been observed in other mammalian species. In addition, in alpacas there is a unique subset of ꝩδ T cells called Vꝩ9Vδ2 T cells. Activation of these cells is dependent upon phosphoantigen (PAg)-mediated interaction with B7-like butyrophilin molecules (BTN-3). This makes alpacas the first species outside of primates to be identified with this unique subset and activation mechanism. Here we review some fundamentals of camelid adaptive immunity that make them distinct from other vertebrate species and their potential applications to human therapies.