A Potent SARS-CoV-2 Neutralizing Human Monoclonal Antibody That Reduces Viral Burden and Disease Severity in Syrian Hamsters

Abstract

The emergence of COVID-19 has led to a pandemic that has caused millions of cases of disease, variable morbidity and hundreds of thousands of deaths. Currently, only remdesivir and dexamethasone have demonstrated limited efficacy, only slightly reducing disease burden, thus novel approaches for clinical management of COVID-19 are needed. We identified a panel of human monoclonal antibody clones from a yeast display library with specificity to the SARS-CoV-2 spike protein receptor binding domain that neutralized the virus in vitro. Administration of the lead antibody clone to Syrian hamsters challenged with SARS-CoV-2 significantly reduced viral load and histopathology score in the lungs. Moreover, the antibody interrupted monocyte infiltration into the lungs, which may have contributed to the reduction of disease severity by limiting immunopathological exacerbation. The use of this antibody could provide an important therapy for treatment of COVID-19 patients.

Keywords: COVID; SARS-CoV-2; coronavirus; monoclonal Ab; therapeutic antibodies.

Figure 1

Properties of clone AvGn-B. (A) Binding curve of biotinylated RBD to the virus neutralizing AvGn-B clone by ELISA. (B) IC₅₀ curve of the AvGn-B for competing binding of SARS-CoV-2 RBD to immobilized ACE2-Fc receptor. (C) Binding kinetics of RBD to AvGn-B measured by BLI, using an Octet system. (D) Binding curve of clone AvGn-B compared to ACE2-Fc or isotype control IgG to 293 cells expressing full-length SARS-CoV-2 spike protein as a GFP-fusion protein. (E) EC₅₀ curve for the ability of clone AvGn-B to neutralize viral-induced cell death in the CPE assay. Curve fitting in (A, B, D, E) was performed using Prism software.

Figure 2

Clinical presentation and viral load in the lungs of Syrian hamsters dosed with AvGn-B following SARS-CoV-2 infection. (A) Hamster body weights were recorded daily (0–5 dpi), and weight loss was defined as percentage loss from 0dpi. Hamsters were separated by treatment group and weight loss was analyzed using multiple t-tests in Prism GraphPad for each day (p > 0.05). (B) Viral load (gene copy number/reaction) in infected hamster lung was compared between treatment groups (analyzed by Mann-Whitney U Test in Prism GraphPad) (*p < 0.05).

Figure 3

Manual annotation and comparison of histological scores of bronchointerstitial pneumonia in control and treated hamsters. (A) Lungs from an uninfected hamster were visualized and scored for lack of significant hypercellularity. (B) Affected parenchyma of infected hamsters (Untreated) characterized by intense hypercellularity is manually annotated (blue) excluding unaffected lung parenchyma, main stem bronchi and hilar adipose tissue. (C) Lung section from a control antibody treated hamster (Ab Control) is showing marked reduction in total cellularity or inflammation. (D) Lung section from a low-dose AvGn-B antibody-treated hamster (AvGn-B Low) showing very significant reduction of inflammation reducing the total area of affected pulmonary parenchyma. (E) After manual annotation depicted in (B) and based on the generated montage including training set and threshold, the automated classifier to discriminate lesional from non-lesional parenchyma was generated in grey and then in blue. (F) After manual annotation of affected lungs, all of the defined classes in preset ROI, i.e., inflammatory foci characterized by infiltration of inflammatory cells into alveolar spaces were identified to generate total number of nuclei (hematoxylin and DAPI-stained nuclei) that were accurately recorded.

Figure 4

Monocytosis and histiocytic arteritis in SARS-CoV-2-infected and untreated hamsters. (A) Major branches of pulmonary artery contain numerous monocytes adhered to endothelial cells. (B) The luminal monocytes in pulmonary artery lumen (PAL) include multinucleate cells attached to the luminal surface of endothelium (red arrow) and the vessels is surrounded by many macrophages. (C) Close-up of pulmonary artery tunica intima showing endothelial-adherent monocytes some of which are subintimal (blue arrows) or migrating through the wall of the vessels, leukocyoclastic vasculitis. (D) Luminal monocytes and perivascular macrophages showing immunoreactivity against SARCoV-2 nucleocapsid. (E) Staining the vessels intima with endothelial marker, factor VIII show discontinuity of the tunical intima by migrating monocytes. (F) Most of the cells crowding tunica intima are showing strong immunoreactivity for the macrophage marker, CD204.

Figure 5

Reduction of SARS-CoV-2 and macrophage infiltration by AvGn-B treatment. (A–E) Overall hypercellularity within lung tissue, resulting in pathological bronchointerstitial pneumonia, was determined using ROI delineations on hematoxylin and eosin stained sections for all groups. (N) Quantification of the total area of the lung tissue affected with bronchointerstitial pneumonia was conducted using automated focal point determination within ROIs following manual thresholding. (O) The total IBA-1+ monocyte lineage cellular intensity was quantified in the lung tissue (for each experimental group, as depicted in the high resolution (white bar in each high resolution image represents 100 µm) and 10× immunofluorescent montage images (F–M). (P) Co-localization of SARS-CoV2 (red) and IBA-1 (cyan) was also quantified within the SARS-CoV2, SARS-CoV2+AvGn-B 1mg, SARS-CoV2+AvGn-B 2.5 mg groups (*p < 0.05, **p < 0.01, ****p < 0.0001; N = 5 hamsters/group) and in the uninfected+AvGn-B 2.5 mg and SARS-CoV-2+IgG groups (*p < 0.05, **p < 0.01, ****p < 0.0001; N = 2 hamsters/group).