Extended Data Fig. 4: (matched to Fig. 5).
a. Schematic of experimental design of prophylactic antibody treatment MISTRG6-hACE2 mice received prophylactic treatment of convalescent plasma (5ml/kg) or monoclonal antibodies at 10mg/kg (clone 135 -m135 or clone 144-m144) 8 h prior to infection or left untreated (untd). Mice were euthanized 4dpi. Convalescent plasma samples from the top 30 neutralizers in a cohort of 148 individuals were pooled to create a mixture with an NT50 titer of 1597 against HIV-1 pseudotyped with SARS-CoV-2 S protein68. Sequencing the antibody genes from infected humans has revealed the expansion of closely related Receptor Binding Domain of the Spike protein (RBD)-specific B cell antibody clones in different SARS-CoV-2 infected individuals. Although most convalescent plasma samples obtained from patients who recovered from COVID-19 did not contain high levels of neutralizing activity, RBD-specific antibodies with potent antiviral activity were found in all individuals tested68. Monoclonal recombinant antibodies (mAbs) used in this study were cloned from these convalescent patients and had high neutralizing activity against SARS-CoV-2 in vitro and in vivo in mouse adapted SARS-CoV-2 infection68,72. b. Weight change in convalescent plasma treated mice at 2 and 4dpi plotted as percent change compared with original weight measured just before inoculation with SARS-CoV-2. Unpaired t-test, two-tailed comparing 4dpi values. N=4 biologically independent mice examined over 2 independent experiments for all time points. Individual values per each mouse plotted. c. Representative images (10x magnification) of H&E staining of lungs (4dpi) from mice treated prophylactically with convalescent plasma or left untreated. The histopathological findings mark transition into the organizing phase, highlighted by infiltration of the inter- and intra-alveolar space by lymphocytes macrophages, and fibroblasts. Representative of N=4 biologically independent mice examined over 2 independent experiments for all time points. Individual values for each mouse are plotted. d. Frequencies of human monocytes (CD14+ classical; CD14+CD16+ intermediate, CD16+ non classical) within human CD45+ cells in the lungs of MISTRG6-hACE2 mice which received a prophylactic treatment of monoclonal antibody clone 135 (m135) or clone 144(m144) 8 h prior to infection or left untreated (untd). Unpaired, two-tailed t-test. P-values<0.05 are plotted. Untreated N=5, treated n=6 biologically independent mice examined over 2 independent experiments for all time points. Individual values for each mouse and means are plotted. e. Frequencies of HLA-DR+CXCR3+ T cells within human CD3+CD45+ cells in lungs of MISTRG6-hACE2 mice which received a prophylactic treatment of monoclonal antibody clone 135 (m135) or clone 144(m144) 8 h prior to infection or left untreated (untd). Unpaired, two-tailed t-test. P-values<0.05 are plotted. Untreated N=5, treated n=6 N=4 biologically independent mice examined over 2 independent experiments for all time points. Individual values for each mouse and means are plotted. f. Frequencies of Tγδ cells within human CD3+CD45+ cells in lungs of MISTRG6-hACE2 mice which received prophylactic treatment of monoclonal antibody clone 135 (m135) or clone 144(m144) 8 h prior to infection or left untreated (untd). Unpaired, two-tailed t-test. N=5-6. P-values<0.05 are plotted. Untreated N=5, treated n=6 biologically independent mice examined over 2 independent experiments for all time points. Individual values for each mouse and means are plotted. g. Frequencies of human T cells within human CD45+ cells in spleens of MISTRG6-hACE2 mice received a prophylactic treatment of monoclonal antibody clone 135 (m135) or clone 144(m144) 8 h prior to infection or left untreated (untd). Unpaired, two-tailed t-test. P-values<0.05 are plotted. Untreated N=5, treated n=64 biologically independent mice examined over two independent experiments for all time points. Individual values for each mouse and means are plotted. h. Frequencies of human CD3+ T cells within human CD45+ population in the blood pre- and post-infection (2, 4, 7, 14,2 8dpi). Lines connect pre- and post-infection values for the same mouse. MISTRG6-hACE2 mice received a prophylactic treatment of monoclonal antibody clone 135 (m135) or clone 144(m144) 8 h prior to infection or left untreated (untd). Paired, two-tailed t-test. P-values<0.05 are plotted. Untreated n=5, treated n=6 biologically independent mice examined over 2 independent experiments for all time points. Individual values for each mouse are plotted. i. Schematic of experimental design of post-infection mAb treatment. MISTRG6-hACE2 mice received a mixed cocktail of monoclonal antibodies clone 135 (m135) and clone 144(m144) at 20mg/kg or left untreated (untd). Early treatment groups were treated 11 h post-infection and late treatment 35 h post-infection. Monoclonal recombinant antibodies (mAbs) used in this study were cloned from the convalescent patients (whose plasma was used prophylactically before) and had high neutralizing activity against SARS-CoV-2 in vitro and in vivo in mouse adapted SARS-CoV-2 infection68,72. j. Viral titers measured by PFU in homogenized lung tissue following early or late therapeutic mAb at 4dpi using Vero E6 (standard assay) or Vero-ACE2+TMPRSS2+ (sensitive assay) cells. Treated n=4 biologically independent mice examined over 2 independent experiments for measurement with Vero-ACE2+TMPRSS2+ (sensitive assay) cells. Individual values for each mouse are plotted. k. Frequencies of human T cells within human CD45+ cells in spleens of MISTRG6-hACE2 mice that received early, late or no treatment of monoclonal antibody mix. Unpaired, two-tailed t-test. Untreated n=7, treated n=5 biologically independent mice examined over 3 independent experiments for all time points. Pooled, infection-matched mice are presented. P-values<0.05 are plotted. Individual values representing each mouse are plotted. Individual values for each mouse are plotted. l. Number of CD16+ human monocytes in lungs of treated and untreated mice at 4dpi. Mice were either MISTRG6-hACE2 mice that received a mixed cocktail of monoclonal antibodies clone 135 (m135) and clone 144(m144) or left untreated (untd). Early treatment group was treated 11 h post-infection (hpi) and late treatment 35 hpi. Unpaired, two-tailed t-test. Untreated n=7, treated n=5 biologically independent mice examined over 3 independent experiments for all time points. Pooled, infection-matched mice are presented. P-values<0.05 are plotted. Individual values representing each mouse are plotted. m. Number of human macrophages in lungs of treated and untreated mice at 4dpi. MISTRG6-hACE2 mice that received a mixed cocktail of monoclonal antibodies clone 135 (m135) and clone 144(m144) or left untreated (untd). Early treatment group was treated 11 h post-infection and late treatment 35 h post-infection. Unpaired, two-tailed t-test. Untreated n=7, treated n=5 biologically independent mice examined over 3 independent experiments for all time points. Pooled, infection-matched mice are presented. P-values<0.05 are plotted. Individual values representing each mouse are plotted.
