Human-Immune-System (HIS) humanized mouse model (DRAGA: HLA-A2.HLA-DR4.Rag1KO.IL-2RγcKO.NOD) for COVID-19.

We report a Human Immune System (HIS)-humanized mouse model ("DRAGA": HLA-A2.HLA-DR4.Rag1KO.IL-2 RγcKO.NOD) for COVID-19 research. DRAGA mice express transgenically HLA-class I and class-II molecules in the mouse thymus to promote human T cell development and human B cell Ig-class switching. When infused with human hematopoietic stem cells from cord blood reconstitute a functional human immune system, as well as human epi/endothelial cells in lung and upper respiratory airways expressing the human ACE2 receptor for SARS-CoV-2. The DRAGA mice were able to sustain SARS-CoV-2 infection for at least 25 days. Infected mice showed replicating virus in the lungs, deteriorating clinical condition, and human-like lung immunopathology including human lymphocyte infiltrates, microthrombi and pulmonary sequelae. Among the intra-alveolar and peri-bronchiolar lymphocyte infiltrates, human lung-resident (CD103+) CD8+ and CD4+ T cells were sequestered in epithelial (CD326+) lung niches and secreted granzyme B and perforin, suggesting anti-viral cytotoxic activity. Infected mice also mounted human IgG antibody responses to SARS-CoV-2 viral proteins. Hence, HIS-DRAGA mice showed unique advantages as a surrogate in vivo human model for studying SARS-CoV-2 immunopathological mechanisms and testing the safety and efficacy of candidate vaccines and therapeutics.

Preclinical evaluation of a candidate naked plasmid DNA vaccine against SARS-CoV-2.

New generation plasmid DNA vaccines may be a safe, fast and simple emergency vaccine platform for preparedness against emerging viral pathogens. Applying platform optimization strategies, we tested the pre-clinical immunogenicity and protective effect of a candidate DNA plasmid vaccine specific for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The DNA vaccine induced spike-specific binding IgG and neutralizing antibodies in mice, rabbits, and rhesus macaques together with robust Th1 dominant cellular responses in small animals. Intradermal and intramuscular needle-free administration of the DNA vaccine yielded comparable immune responses. In a vaccination-challenge study of rhesus macaques, the vaccine demonstrated protection from viral replication in the lungs following intranasal and intratracheal inoculation with SARS-CoV-2. In conclusion, the candidate plasmid DNA vaccine encoding the SARS-CoV-2 spike protein is immunogenic in different models and confers protection against lung infection in nonhuman primates. Further evaluation of this DNA vaccine candidate in clinical trials is warranted.

A Human-Immune-System (HIS) humanized mouse model (DRAGA: HLA-A2. HLA-DR4. Rag1 KO.IL-2Rγc KO. NOD) for COVID-19.

We report the first Human Immune System (HIS)-humanized mouse model ("DRAGA": HLA-A2.HLA-DR4.Rag1KO.IL-2RγcKO.NOD) for COVID-19 research. This mouse is reconstituted with human cord blood-derived, HLA-matched hematopoietic stem cells. It engrafts human epi/endothelial cells expressing the human ACE2 receptor for SARS-CoV-2 and TMPRSS2 serine protease co-localized on lung epithelia. HIS-DRAGA mice sustained SARS-CoV-2 infection, showing deteriorated clinical condition, replicating virus in the lungs, and human-like lung immunopathology including T-cell infiltrates, microthrombi and pulmonary sequelae. Among T-cell infiltrates, lung-resident (CD103+) CD8+ T cells were sequestered in epithelial (CD326+) lung niches and secreted granzyme B and perforin, indicating cytotoxic potential. Infected mice also developed antibodies against the SARS-CoV-2 viral proteins. Hence, HIS-DRAGA mice showed unique advantages as a surrogate in vivo human model for studying SARS-CoV-2 immunopathology and for testing the safety and efficacy of candidate vaccines and therapeutics.

Opsono-Adherence Assay to Evaluate Functional Antibodies in Vaccine Development Against Bacillus anthracis and Other Encapsulated Pathogens.

The opsono-adherence assay is a functional assay that enumerates the attachment of bacterial pathogens to professional phagocytes. Because adherence is requisite to phagocytosis and killing, the assay is an alternative method to opsono-phagocytic killing assays. An advantage of the opsono-adherence assay is the option of using inactivated pathogens and mammalian cell lines, which allows standardization across multiple experiments. The use of an inactivated pathogen in the assay also facilitates work with biosafety level 3 infectious agents and other virulent pathogens. In our work, the opsono-adherence assay was used to assess the functional ability of antibodies, from sera of animals immunized with an anthrax capsule-based vaccine, to induce adherence of fixed Bacillus anthracis to a mouse macrophage cell line, RAW 264.7. Automated fluorescence microscopy was used to capture images of bacilli adhering to macrophages. Increased adherence was correlated with the presence of anti-capsule antibodies in the serum. Non-human primates that exhibited high serum anti-capsule antibody concentrations were protected from anthrax challenge. Thus, the opsono-adherence assay can be used to elucidate the biological functions of antigen specific antibodies in sera, to evaluate the efficacy of vaccine candidates and other therapeutics, and to serve as a possible correlate of immunity.