Single-cell-resolved interspecies comparison identifies a shared inflammatory axis and a dominant neutrophil-endothelial program in severe COVID-19 (preprint)

Key issues for research of COVID-19 pathogenesis are the lack of biopsies from patients and of samples at the onset of infection. To overcome these hurdles, hamsters were shown to be useful models for studying this disease. Here, we further leveraged the model to molecularly survey the disease progression from time-resolved single-cell RNA-sequencing data collected from healthy and SARS-CoV-2-infected Syrian and Roborovski hamster lungs. We compared our data to human COVID-19 studies, including BALF, nasal swab, and post-mortem lung tissue, and identified a shared axis of inflammation dominated by macrophages, neutrophils, and endothelial cells, which we show to be transient in Syrian and terminal in Roborovski hamsters. Our data suggest that, following SARS-CoV-2 infection, commitment to a type 1 or type 3-biased immunity determines moderate versus severe COVID-19 outcomes, respectively.

Tissue protective role of Ganetespib in SARS-CoV-2-infected Syrian golden hamsters (preprint)

The emergence of new SARS-CoV-2 variants, capable of escaping the humoral immunity acquired by the available vaccines, together with waning immunity and vaccine hesitancy, challenges the efficacy of the vaccination strategy in fighting COVID-19. Improved therapeutic strategies are therefore urgently needed to better intervene particularly in severe cases of the disease. They should aim at controlling the hyper-inflammatory state generated upon infection, at reducing lung tissue pathology and endothelial damages, along with viral replication. Previous research has pointed a possible role for the chaperone HSP90 in SARS-CoV-2 replication and COVID-19 pathogenesis. Pharmacological intervention through HSP90 inhibitors was shown to be beneficial in the treatment of inflammatory diseases, infections and reducing replication of diverse viruses. In this study, we analyzed the effects of the potent HSP90 inhibitor Ganetespib in vitro on alveolar epithelial cells and alveolar macrophages to characterize its effects on cell activation and viral replication. Additionally, to evaluate its efficacy in controlling systemic inflammation and the viral burden after infection in vivo, a Syrian hamster model was used. In vitro, Ganetespib reduced viral replication on AECs in a dose-dependent manner and lowered significantly the expression of pro-inflammatory genes, in both AECs and alveolar macrophages. In vivo, administration of Ganetespib led to an overall improvement of the clinical condition of infected animals, with decreased systemic inflammation, reduced edema formation and lung tissue pathology. Altogether, we show that Ganetespib could be a potential medicine to treat moderate and severe cases of COVID-19.

A live attenuated vaccine confers superior mucosal and systemic immunity to SARS-CoV-2 variants (preprint)

Vaccines are a cornerstone in COVID-19 pandemic management. Here, we compare immune responses to and preclinical efficacy of the mRNA vaccine BNT162b2, an adenovirus-vectored spike vaccine, and the live-attenuated-virus vaccine candidate sCPD9 after single and double vaccination in Syrian hamsters. All regimens containing sCPD9 showed superior efficacy. The robust immunity elicited by sCPD9 was evident in a wide range of immune parameters after challenge with heterologous SARS-CoV-2 including rapid viral clearance, reduced tissue damage, fast differentiation of pre-plasmablasts, strong systemic and mucosal humoral responses, and rapid recall of memory T cells from lung tissue. Our results demonstrate that use of live-attenuated vaccines may offer advantages over available COVID-19 vaccines, specifically when applied as booster, and may provide a solution for containment of the COVID-19 pandemic.

Insights into standards of care: dexamethasone and antibodies against COVID-19 in hamster models (preprint)

Rationale: In face of the ongoing SARS-CoV-2 pandemic, effective and well-understood treatment options are still scarce. While vaccines have proven instrumental in fighting SARS-CoV-2, their efficacy is challenged by vaccine hesitancy, novel variants and short-lasting immunity. Therefore, understanding and optimization of therapeutic options remains essential. Objectives: We aimed at generating a deeper understanding on how currently used drugs, specifically dexamethasone and anti-SARS-CoV-2 antibodies, affect SARS-CoV-2 infection and host responses. Possible synergistic effects of both substances are investigated to evaluate combinatorial treatments. Methods: By using two COVID-19 hamster models, pulmonary immune responses were analyzed to characterize effects of treatment with either dexamethasone, anti-SARS-CoV-2 spike monoclonal antibody or a combination of both. scRNA sequencing was employed to reveal transcriptional response to treatment on a single cell level. Measurements and main results: Dexamethasone treatment resulted in similar or increased viral loads compared to controls. Anti-SARS-CoV-2 antibody treatment alone or combined with dexamethasone successfully reduced pulmonary viral burden. Dexamethasone exhibited strong anti-inflammatory effects and prevented fulminant disease in a severe COVID-19-like disease model. Combination therapy showed additive benefits with both anti-viral and anti-inflammatory potency. Bulk and single-cell transcriptomic analyses confirmed dampened inflammatory cell recruitment into lungs upon dexamethasone treatment and identified a candidate subpopulation of neutrophils specifically responsive to dexamethasone. Conclusions: Our analyses i) confirm the anti-inflammatory properties and indicate possible modes of action for dexamethasone, ii) validate anti-viral effects of anti-SARS-CoV-2 antibody treatment, and iii) reveal synergistic effects of a combination therapy and can thus inform more effective COVID-19 therapies.

Single cell RNA-seq uncovers the nuclear decoy lincRNA PIRAT as a regulator of systemic monocyte immunity during COVID-19 (preprint)

The systemic immune response to viral infection is shaped by master transcription factors such as NF{kappa}B or PU.1. Although long non-coding RNAs (lncRNAs) have been suggested as important regulators of transcription factor activity, their contributions to the systemic immunopathologies observed during SARS-CoV-2 infection have remained unknown. Here, we employed a targeted single-cell RNA-seq approach to reveal lncRNAs differentially expressed in blood leukocytes during severe COVID-19. Our results uncover the lncRNA PIRAT as a major PU.1 feedback-regulator in monocytes, governing the production of the alarmins S100A8/A9 - key drivers of COVID-19 pathogenesis. Knockout and transgene expression, combined with chromatin-occupancy profiling characterized PIRAT as a nuclear decoy RNA, diverting the PU.1 transcription factor from alarmin promoters to dead-end pseudogenes in naive monocytes. NF{kappa}B-dependent PIRAT down-regulation during COVID-19 consequently releases a transcriptional brake, fueling alarmin production. Our results suggest a major role of nuclear noncoding RNA circuits in systemic antiviral responses to SARS-CoV-2 in humans.

A SARS-CoV-2 neutralizing antibody protects from lung pathology in a COVID-19 hamster model (preprint)

The emergence of SARS-CoV-2 led to pandemic spread of coronavirus disease 2019 (COVID-19), manifesting with respiratory symptoms and multi-organ dysfunction. Detailed characterization of virus-neutralizing antibodies and target epitopes is needed to understand COVID-19 pathophysiology and guide immunization strategies. Among 598 human monoclonal antibodies (mAbs) from ten COVID-19 patients, we identified 40 strongly neutralizing mAbs. The most potent mAb CV07-209 neutralized authentic SARS-CoV-2 with IC50 of 3.1 ng/ml. Crystal structures of two mAbs in complex with the SARS-CoV-2 receptor-binding domain at 2.55 and 2.70 A revealed a direct block of ACE2 attachment. Interestingly, some of the near-germline SARS-CoV-2 neutralizing mAbs reacted with mammalian self-antigens. Prophylactic and therapeutic application of CV07-209 protected hamsters from SARS-CoV-2 infection, weight loss and lung pathology. Our results show that non-self-reactive virus-neutralizing mAbs elicited during SARS-CoV-2 infection are a promising therapeutic strategy.

Age-dependent progression of SARS-CoV-2 infection in Syrian hamsters (preprint)

In late 2019, an outbreak of a severe respiratory disease caused by an emerging coronavirus, SARS-CoV-2, resulted in high morbidity and mortality in infected humans1. Complete understanding of COVID-19, the multi-faceted disease caused by SARS-CoV-2, requires suitable small animal models, as does the development and evaluation of vaccines and antivirals2. Because age-dependent differences of COVID-19 were identified in humans3, we compared the course of SARS-CoV-2 infection in young and aged Syrian hamsters. We show that virus replication in the upper and lower respiratory tract was independent of the age of the animals. However, older hamsters exhibited more pronounced and consistent weight loss. In situ hybridization in the lungs identified viral RNA in bronchial epithelium, alveolar epithelial cells type I and II, and macrophages. Histopathology revealed clear age-dependent differences, with young hamsters launching earlier and stronger immune cell influx than aged hamsters. The latter developed conspicuous alveolar and perivascular edema, indicating vascular leakage. In contrast, we observed rapid lung recovery at day 14 after infection only in young hamsters. We propose that comparative assessment in young versus aged hamsters of SARS-CoV-2 vaccines and treatments may yield valuable information as this small-animal model appears to mirror age-dependent differences in human patients.