RESUMO
Emerging variants of concern (VOCs) are threatening to limit the effectiveness of SARS-CoV-2 monoclonal antibodies and vaccines currently used in clinical practice; broadly neutralizing antibodies and strategies for their identification are therefore urgently required. Here we demonstrate that broadly neutralizing antibodies can be isolated from peripheral blood mononuclear cells (PBMCs) of convalescent patients using SARS-CoV-2 receptor binding domains (RBDs) carrying epitope-specific mutations. This is exemplified by two human antibodies, GAR05, binding to epitope class 1, and GAR12, binding to a new epitope class 6 (located between class 3 and class 5). Both antibodies broadly neutralize VOCs, exceeding the potency of the clinical monoclonal sotrovimab (mAb S309) by orders of magnitude. They also provide potent prophylactic and therapeutic in vivo protection of hACE2 mice against viral challenge. Our results indicate that exposure to Wuhan SARS-CoV-2 induces antibodies that maintain potent and broad neutralization against emerging VOCs using two unique strategies: either by targeting the divergent class 1 epitope in a manner resistant to VOCs (ACE2 mimicry, as illustrated by GAR05 and mAbs P2C-1F11/S2K14); or alternatively, by targeting rare and highly conserved epitopes, such as the new class 6 epitope identified here (as illustrated by GAR12). Our results provide guidance for next generation monoclonal antibody development and vaccine design.
RESUMO
Phagocytic responses by effector cells to antibody or complement-opsonised viruses have been recognized to play a key role in anti-viral immunity. These include antibody dependent cellular phagocytosis mediated via Fc-receptors, phagocytosis mediated by classically activated complement-fixing IgM or IgG1 antibodies and antibody independent phagocytosis mediated via direct opsonisation of viruses by complement products activated via the mannose-binding lectin pathway. Limited data suggest these phagocytic responses by effector cells may contribute to the immunological and inflammatory responses in SARS-CoV-2 infection, however, their development and clinical significance remain to be fully elucidated. In this cohort of 62 patients, acutely ill individuals were shown to mount phagocytic responses to autologous plasma-opsonised SARS-CoV-2 Spike protein-coated microbeads as early as 10 days post symptom onset. Heat inactivation of the plasma prior to use as an opsonin caused 77-95% abrogation of the phagocytic response, and pre-blocking of Fc-receptors on the effector cells showed only 18-60% inhibition. These results suggest that SARS-CoV-2 can provoke early phagocytosis, which is primarily driven by heat labile components, likely activated complements, with variable contribution from anti-Spike antibodies. During convalescence, phagocytic responses correlated significantly with anti-Spike IgG titers. Older patients and patients with severe disease had significantly higher phagocytosis and neutralisation functions when compared to younger patients or patients with asymptomatic, mild, or moderate disease. A longitudinal study of a subset of these patients over 12 months showed preservation of phagocytic and neutralisation functions in all patients, despite a drop in the endpoint antibody titers by more than 90%. Interestingly, surface plasmon resonance showed a significant increase in the affinity of the anti-Spike antibodies over time correlating with the maintenance of both the phagocytic and neutralisation functions suggesting that improvement in the antibody quality over the 12 months contributed to the retention of effector functions. Author SummaryLimited data suggest antibody dependent effector functions including phagocytosis may contribute to the immunological and inflammatory responses in SARS CoV-2 infection, however, their development, maintenance, and clinical significance remain unknown. In this study we show: O_LIPatients with acute SARS CoV-2 infection can mount phagocytic responses as early as 10 days post symptom onset and these responses were primarily driven by heat labile components of the autologous plasma. These results indicate that the current approach of studying phagocytosis using purified or monoclonal antibodies does not recapitulate contribution by all components in the plasma. C_LIO_LIIn convalescent patients, high phagocytic responses significantly correlated with increasing age, increasing disease severity, high neutralisation functions and high anti-Spike antibody titers, particularly IgG1. C_LIO_LILongitudinal study of convalescent patients over a 12-month period showed maintenance of phagocytic and neutralisation functions, despite a drop in the anti-Spike endpoint antibody titers by more than 90%. However, we found significant increase in the affinity of the anti-Spike antibodies over the 12-month period and these correlated with the maintenance of functions suggesting that improvement in the antibody quality over time contributed to the retention of effector functions. Clinically, measuring antibody titers in sera but not the quality of antibodies is considered a gold standard indicator of immune protection following SARS-CoV 2 infection or vaccination. Our results challenge this notion and recommends change in the current clinical practice. C_LI
