The pandemic severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the cause of Coronavirus Disease 2019 (COVID-19). Worldwide efforts are being made to develop vaccines to mitigate this pandemic. We engineered two recombinant Newcastle disease virus (NDV) vectors expressing either the full-length SARS-CoV-2 spike protein (NDV-FLS) or a version with a 19 amino acid deletion at the carboxy terminus (NDV-Δ19S). Hamsters receiving two doses (prime-boost) of NDV-FLS developed a robust SARS-CoV-2-neutralizing antibody response, with elimination of infectious virus in the lungs and minimal lung pathology at five days post-challenge. Single-dose vaccination with NDV-FLS significantly reduced SARS-CoV-2 replication in the lungs, but only mildly decreased lung inflammation. NDV-Δ19S-treated hamsters had a moderate decrease in SARS-CoV-2 titers in lungs and presented with severe microscopic lesions, suggesting that truncation of the spike protein was a less effective strategy. In summary, NDV-vectored vaccines represent a viable option for protection against COVID-19.

Adeno-associated virus (AAV) vector mediated expression of therapeutic monoclonal antibodies is an alternative strategy to traditional vaccination to generate immunity in immunosuppressed or immunosenescent individuals. In this study, we vectorized a human monoclonal antibody (31C2) directed against the spike protein of SARS-CoV-2 and determined the safety profile of this AAV vector in mice and sheep as a large animal model. In both studies, plasma biochemical parameters and hematology were comparable to untreated controls. Except for mild myositis at the site of injection, none of the major organs revealed any signs of toxicity. AAV-mediated human IgG expression increased steadily throughout the 28-day study in sheep, resulting in peak concentrations of 21.4–46.7 µg/ mL, demonstrating practical scale up from rodent to large animal models. This alternative approach to immunity is worth further exploration after this demonstration of safety, tolerability, and scalability in a large animal model.

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Epithelial ovarian cancer is the deadliest gynecological malignancy. The lack of effective treatments highlights the need for novel therapeutic interventions. The aim of this study was to investigate whether sustained adeno-associated virus (AAV) vector-mediated expression of vascular normalizing agents 3TSR and Fc3TSR and the antiangiogenic monoclonal antibody, Bevacizumab, with or without oncolytic virus treatment would improve survival in an orthotopic syngeneic mouse model of epithelial ovarian carcinoma. AAV vectors were administered 40 days post-tumor implantation and combined with oncolytic avian orthoavulavirus-1 (AOaV-1) 20 days later, at the peak of AAV-transgene expression, to ascertain whether survival could be extended. Flow cytometry conducted on blood samples, taken at an acute time point post-AOaV-1 administration (36 h), revealed a significant increase in activated NK cells in the blood of all mice that received AOaV-1. T cell analysis revealed a significant increase in CD8+ tumor specific T cells in the blood of AAV-Bevacizumab+AOaV-1 treated mice compared to control mice 10 days post AOaV-1 administration. Immunohistochemical staining of primary tumors harvested from a subset of mice euthanized 90 days post tumor implantation, when mice typically have large primary tumors, secondary peritoneal lesions, and extensive ascites fluid production, revealed that AAV-3TSR, AAV-Fc3TSR+AOaV-1, or AAV-Bevacizumab+AOaV-1 treated mice had significantly more tumor-infiltrating CD8+ T cells than PBS controls. Despite AAV-mediated transgene expression waning faster in tumor-bearing mice than in non-tumor bearing mice, all three of the AAV therapies significantly extended survival compared to control mice; with AAV-Bevacizumab performing the best in this model. However, combining AAV therapies with a single dose of AOaV-1 did not lead to significant extensions in survival compared to AAV therapies on their own, suggesting that additional doses of AOaV-1 may be required to improve efficacy in this model. These results suggest that vectorizing anti-angiogenic and vascular normalizing agents is a viable therapeutic option that warrants further investigation, including optimizing combination therapies.

Novel immunotherapies continue to be developed and tested for application against a plethora of diseases. The clinical translation of immunotherapies requires an understanding of their mechanisms. The contributions of antibodies in driving long-term responses following immunotherapies continue to be revealed given their diverse effector functions. Developing an in-depth understanding of the role of antibodies in treatment efficacy is required to optimize immunotherapies and improve the chance of successfully translating them into the clinic. However, analyses of antibody responses can be challenging in the context of antigen-agnostic immunotherapies, particularly in the context of cancers that lack pre-defined target antigens. As such, robust methods are needed to evaluate the capacity of a given immunotherapy to induce beneficial antibody responses, and to identify any therapy-limiting antibodies. We previously developed a comprehensive method for detecting antibody responses induced by antigen-agnostic immunotherapies for application in pre-clinical models of vaccinology and cancer therapy. Here, we extend this method to a high-throughput, flow cytometry-based assay able to identify and quantify isotype-specific virus- and tumor-associated antibody responses induced by immunotherapies using small sample volumes with rapid speed and high sensitivity. This method provides a valuable and flexible protocol for investigating antibody responses induced by immunotherapies, which researchers can use to expand their analyses and optimize their own treatment regimens.