Comparison of Mucosal and Intramuscular Immunization against SARS-CoV-2 with Replication-Defective and Replicating Single-cycle Adenovirus Vaccines (preprint)

SARS-CoV-2 enters the body at mucosal surfaces, such as the nose and lungs. These events involve a small number of virions at these mucosal barriers and are therefore a strategic point to stop a COVID-19 infection before it starts. Despite this, most vaccines against COVID-19 are being injected into the muscle where they will not generate the highest levels of mucosal protection. The vaccines that are approved for use in humans are all replication-defective (RD) mRNA, DNA, or adenovirus (Ad) vaccines that do not amplify antigen transgenes. We developed single cycle adenovirus (SC-Ad) vectors that replicate antigen genes up to 10,000-fold in human cells, but that are disabled from producing infectious Ad particles. We show here that SC-Ad expressing the full-length SARS-CoV-2 spike protein produces 100-fold more spike protein than a matched RD-Ad-Spike vector. When Ad-permissive hamsters were immunized with these vaccines by intranasal (IN) or intramuscular (IM) routes, SC-Ad produced significantly stronger antibody responses as compared to RD-Ad against the spike protein that rose over 14 weeks after one immunization. Single IN or IM immunizations generated significant antibody responses in serum and in bronchoalveolar lavages (BALs). IN priming, but not IM priming, generated HLA-restricted CD8 T cell responses in BALs. SC-Ad-Spike generated antibodies that retain binding to spike receptor binding domains (RBDs) with mutations from new viral variants. These data suggest empowering the genomes of gene-based vaccines with the ability to amplify antigen genes can increase potency. This may be particularly advantageous when applying mucosal vaccines to combat mucosal pathogens like SARS-CoV-2. One Sentence SummaryArming adenovirus vaccines with the ability to replicate vaccine antigen genes may increase potency for systemic, or more importantly, mucosal immunization against mucosal pathogens.

The Fc-mediated effector functions of a potent SARS-CoV-2 neutralizing antibody, SC31, isolated from an early convalescent COVID-19 patient, are essential for the optimal therapeutic efficacy of the antibody (preprint)

SARS-CoV-2-neutralizing antibodies are promising therapeutics for COVID-19. However, little is known about the mechanisms of action of these antibodies or their effective dosing windows. We report the discovery and development of SC31, a potent SARS-CoV-2 neutralizing IgG1 antibody, originally isolated from a convalescent patient at day 27 after the onset of symptoms. Neutralization occurs via a binding epitope that maps within the ACE2 interface of the SARS-CoV-2 Spike protein, conserved across all common circulating SARS-CoV-2 mutants. In SARS-CoV-2 infected K18-human ACE2 transgenic mice, SC31 demonstrated potent survival benefit by dramatically reducing viral load concomitant with attenuated pro-inflammatory responses linked to severe systemic disease, such as IL-6. Comparison with a Fc-null LALA variant of SC31 demonstrated that optimal therapeutic efficacy of SC31 requires intact Fc-mediated effector functions that can further induce an IFN{gamma}-driven anti-viral immune response. Dose-dependent efficacy for SC31 was observed down to 5mg/kg when dosed before the activation of lung inflammatory responses. Importantly, despite Fc{gamma}R binding, no evidence of antibody dependent enhancement was observed with the Fc-competent SC31 even at sub-therapeutic doses. Therapeutic efficacy was confirmed in SARS-CoV-2-infected hamsters, where SC31 again significantly reduced viral load, decreased lung lesions and inhibited progression to severe disease manifestations. This study underlines the potential for significant COVID-19 patient benefit for the SC31 antibody that justifies rapid advancement to the clinic, as well as highlighting the importance of appropriate mechanistic and functional studies during development. One Sentence SummaryAnti-SARS-CoV-2 IgG1 antibody SC31 controls infection in vivo by blocking SP:ACE2 binding and triggering a Fc-mediated anti-viral response.

anti-IL-6 versus anti-IL-6R Blocking Antibodies to Treat Acute Ebola Infection in BALB/c Mice with Potential Implications for Treating Patients Presenting with COVID-19 (preprint)

Cytokine release syndrome (CRS) is known to be a factor in morbidity and mortality associated with acute viral infections including those caused by filoviruses and coronaviruses. IL-6 has been implicated as a cytokine negatively associated with survival after filovirus and coronavirus infection. However, IL-6 has also been shown to be an important mediator of innate immunity and important for the host response to an acute viral infection. Clinical studies are now being conducted by various researchers to evaluate the possible role of IL-6 blockers to improve outcomes in critically ill patients with CRS. Most of these studies involve the use of anti-IL-6R monoclonal antibodies (-IL6R mAbs). We present data showing that direct neutralization of IL-6 with an -IL-6 mAb in a BALB/c Ebolavirus (EBOV) challenge model produced a statistically significant improvement in outcome compared with controls when administered within the first 24 hours of challenge and repeated every 72 hours. A similar effect was seen in mice treated with the same dose of -IL-6R mAb when the treatment was delayed 48 hrs post-challenge. These data suggest that direct neutralization of IL-6, early during the course of infection, may provide additional clinical benefits to IL-6 receptor blockade alone during treatment of patients with virus-induced CRS.

An Effective CTL Peptide Vaccine for Ebola Zaire Based on Survivors' CD8+ Targeting of a Particular Nucleocapsid Protein Epitope with Potential Implications for COVID-19 Vaccine Design (preprint)

The 2013-2016 West Africa EBOV epidemic was the biggest EBOV outbreak to date. An analysis of virus-specific CD8+ T-cell immunity in 30 survivors showed that 26 of those individuals had a CD8+ response to at least one EBOV protein. The dominant response (25/26 subjects) was specific to the EBOV nucleocapsid protein (NP). It has been suggested that epitopes on the EBOV NP could form an important part of an effective T-cell vaccine for Ebola Zaire. We show that a 9-amino-acid peptide NP44-52 (YQVNNLEEI) located in a conserved region of EBOV NP provides protection against morbidity and mortality after mouse adapted EBOV challenge. A single vaccination in a C57BL/6 mouse using an adjuvanted microsphere peptide vaccine formulation containing NP44-52 is enough to confer immunity in mice. Our work suggests that a peptide vaccine based on CD8+ T-cell immunity in EBOV survivors is conceptually sound and feasible. Nucleocapsid proteins within SARS-CoV-2 contain multiple class I epitopes with predicted HLA restrictions consistent with broad population coverage. A similar approach to a CTL vaccine design may be possible for that virus.