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Oncolytic viruses are a promising approach for cancer treatment where viruses selectively target and kill cancer cells while also stimulating an immune response. Among viruses with this ability, bovine herpesvirus-1 (BoHV-1) has several advantages, including observations suggesting it may not require viral replication for its anti-cancer effects. We previously demonstrated that binding and penetration of enveloped virus particles are sufficient to trigger intrinsic and innate immune signaling in normal cells, while other groups have published the efficacy of non-replicating viruses as viable immunotherapies in different cancer models. In this work, we definitively show that live and UV-inactivated (UV) (non-replicating) BoHV-1-based regimens extend survival of tumor-bearing mice to similar degrees and induce infiltration of similar immune cell populations, with the exception of neutrophils. Transcriptomic analysis of tumors treated with either live or UV BoHV-1-based regimens revealed similar pathway enrichment and a subset of overlapping differentially regulated genes, suggesting live and UV BoHV-1 have similar mechanisms of activity. Last, we present a gene signature across our in vitro and in vivo models that could potentially be used to validate new BoHV-1 therapeutics. This work contributes to the growing body of literature showing that replication may not be necessary for therapeutic efficacy of viral immunotherapies.

Oncolytic virotherapies have shown excellent promise in a variety of cancers by promoting antitumor immunity. However, the effects of oncolytic virus-mediated type I interferon (IFN-I) production on antitumor immunity remain unclear. Recent reports have highlighted immunosuppressive functions of IFN-I in the context of checkpoint inhibitor and cell-based therapies. In this study, we demonstrate that oncolytic virus-induced IFN-I promotes the expression of PD-L1 in tumor cells and leukocytes in a IFN receptor (IFNAR)-dependent manner. Inhibition of IFN-I signaling using a monoclonal IFNAR antibody decreased IFN-I-induced PD-L1 expression and promoted tumor-specific T cell effector responses when combined with oncolytic virotherapy. Furthermore, IFNAR blockade improved therapeutic response to oncolytic virotherapy in a manner comparable with PD-L1 blockade. Our study highlights a critical immunosuppressive role of IFN-I on antitumor immunity and uses a combination strategy that improves the response to oncolytic virotherapy.

Summary: The angiotensin-converting enzyme 2 (ACE2) protein is a key catalytic regulator of the renin-angiotensin system (RAS), involved in fluid homeostasis and blood pressure modulation. ACE2 also serves as a cell-surface receptor for some coronaviruses such as SARS-CoV and SARS-CoV-2. Improved characterization of ACE2 regulation may help us understand the effects of pre-existing conditions on COVID-19 incidence, as well as pathogenic dysregulation following viral infection. Here, we perform bioinformatic analyses to hypothesize on ACE2 gene regulation in two different physiological contexts, identifying putative regulatory elements of ACE2 expression. We perform functional validation of our computational predictions via targeted CRISPR-Cas9 deletions of these elements in vitro, finding them responsive to immune signaling and oxidative-stress pathways. This contributes to our understanding of ACE2 gene regulation at baseline and immune challenge. Our work supports pursuit of these putative mechanisms in our understanding of infection/disease caused by current, and future, SARS-related viruses such as SARS-CoV-2.

Summary: Since the vast majority of species solely rely on innate immunity for host defense, it stands to reason that a critical evolutionary trait like immunological memory evolved in this primitive branch of our immune system. There is ample evidence that vaccines such as bacillus Calmette-Guérin (BCG) induce protective innate immune memory responses (trained immunity) against heterologous pathogens. Here we show that while BCG vaccination significantly reduces morbidity and mortality against influenza A virus (IAV), it fails to provide protection against severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). In contrast to IAV, SARS-CoV-2 infection leads to unique pulmonary vasculature damage facilitating viral dissemination to other organs, including the bone marrow (BM), a central site for BCG-mediated trained immunity. Finally, monocytes from BCG-vaccinated individuals mount an efficient cytokine response to IAV infection, while this response is minimal following SARS-CoV-2. Collectively, our data suggest that the protective capacity of BCG vaccination is contingent on viral pathogenesis and tissue tropism.

Summary: Type I interferons (IFNs) are our first line of defense against virus infection. Recent studies have suggested the ability of SARS-CoV-2 proteins to inhibit IFN responses. Emerging data also suggest that timing and extent of IFN production is associated with manifestation of COVID-19 severity. In spite of progress in understanding how SARS-CoV-2 activates antiviral responses, mechanistic studies into wild-type SARS-CoV-2-mediated induction and inhibition of human type I IFN responses are scarce. Here we demonstrate that SARS-CoV-2 infection induces a type I IFN response in vitro and in moderate cases of COVID-19. In vitro stimulation of type I IFN expression and signaling in human airway epithelial cells is associated with activation of canonical transcriptions factors, and SARS-CoV-2 is unable to inhibit exogenous induction of these responses. Furthermore, we show that physiological levels of IFNα detected in patients with moderate COVID-19 is sufficient to suppress SARS-CoV-2 replication in human airway cells.

Summary: Compared with other mammals, bats harbor more zoonotic viruses per species and do not demonstrate signs of disease on infection with these viruses. To counteract infections with viruses, bats have evolved enhanced mechanisms to limit virus replication and immunopathology. However, molecular and cellular drivers of antiviral responses in bats largely remain an enigma. In this study, we demonstrate that a serine residue in IRF3 is positively selected for in multiple bat species. IRF3 is a central regulator of innate antiviral responses in mammals. Replacing the serine residue in bat IRF3 with the human leucine residue decreased antiviral protection in bat cells, whereas the addition of this serine residue in human IRF3 significantly enhanced antiviral protection in human cells. Our study provides genetic and functional evidence for enhanced IRF3-mediated antiviral responses in bats and adds support to speculations that bats have positively selected for multiple adaptations in their antiviral immune responses. : Biological Sciences; Immunology; Evolutionary Biology Subject Areas: Biological Sciences, Immunology, Evolutionary Biology