The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) presents a global health emergency that is in urgent need of intervention 1-3. The entry of SARS-CoV-2 into its target cells depends on binding between the receptor-binding domain (RBD) of the viral spike protein and its cellular receptor, angiotensin-converting enzyme 2 (ACE2) 2,4-6. Here we report the isolation and characterization of 206 RBD-specific monoclonal antibodies derived from single B cells from 8 individuals infected with SARS-CoV-2. We identified antibodies that potently neutralize SARS-CoV-2; this activity correlates with competition with ACE2 for binding to RBD. Unexpectedly, the anti-SARS-CoV-2 antibodies and the infected plasma did not cross-react with the RBDs of SARS-CoV or Middle East respiratory syndrome-related coronavirus (MERS-CoV), although there was substantial plasma cross-reactivity to their trimeric spike proteins. Analysis of the crystal structure of RBD-bound antibody revealed that steric hindrance inhibits viral engagement with ACE2, thereby blocking viral entry. These findings suggest that anti-RBD antibodies are largely viral-species-specific inhibitors. The antibodies identified here may be candidates for development of clinical interventions against SARS-CoV-2. The rapid international transmission of SARS-CoV-2 poses a serious global health emergency with no available treatments or vaccine 1-3. SARS-CoV-2 shares substantial genetic and functional similarity with other human betacoronaviruses, including SARS-CoV and MERS-CoV 2,4-8. SARS-CoV-2 uses an envelope homotrimeric spike glycoprotein to interact with the cellular receptor ACE2 2,5,6,8. Binding with ACE2 triggers a cell membrane fusion cascade that results in viral entry. This suggests that disruption of the RBD-ACE2 interaction would block SARS-CoV-2 cell entry. The high-resolution structure of SARS-CoV-2 RBD bound to the N-terminal peptidase domain of ACE2 has recently been determined 6-8. The ACE2-binding mechanism is nearly identical between SARS-CoV-2 and SARS-CoV RBDs 7-10. Animal studies on RBD-based vaccines against SARS-CoV and MERS-CoV have shown strong polyclonal antibody responses that inhibit viral entry 11,12. These findings suggest that anti-RBD antibodies should effectively block SARS-CoV-2 entry. In this study, we report on RBD-specific monoclonal antibodies obtained from individuals infected with SARS-CoV-2. Plasma antibody response against SARS-CoV-2 We collected cross-sectional and longitudinal blood samples from eight patients infected with SARS-CoV-2, who were infected during the early outbreak in Shenzhen (Supplementary Table 1). Samples were named according to patient ID and A, B, or C depending on when they were collected. Six patients (P1 to P4, P8 and P16) had recently travelled to Wuhan and the others (P5 and P22) had direct contact with people who had recently been in Wuhan. P1 to P5 comprise a family cluster, including the first documented case of human-to-human transmission...

The recently identified Middle East respiratory syndrome coronavirus (MERS-CoV) causes severe and fatal acute respiratory illness in humans. However, no prophylactic and therapeutic agents specifically against MERS-CoV are currently available. Entry of MERS-CoV into target cells depends on binding of the receptor binding domain (RBD) of the viral envelope spike glycoprotein to the cellular receptor dipeptidyl peptidase 4 (DPP4). We report the isolation and characterization of two potent human RBD-specific neutralizing monoclonal antibodies (MERS-4 and MERS-27) derived from single-chain variable region fragments of a nonimmune human antibody library. MERS-4 and MERS-27 inhibited infection of both pseudotyped and live MERS-CoV with IC50 (half-maximal inhibitory concentration) at nanomolar concentrations. MERS-4 also showed inhibitory activity against syncytia formation mediated by interaction between MERS-CoV spike glycoprotein and DPP4. Combination of MERS-4 and MERS-27 demonstrated a synergistic effect in neutralization against pseudotyped MERS-CoV. Biochemical analysis indicated that MERS-4 and MERS-27 blocked RBD interaction with DPP4 on the cell surface. MERS-4, in particular, bound soluble RBD with an about 45-fold higher affinity than DPP4. Mutagenesis analysis suggested that MERS-4 and MERS-27 recognized distinct regions in RBD. These results suggest that MERS-4 and MERS-27 are RBD-specific potent inhibitors and could serve as promising candidates for prophylactic and therapeutic interventions against MERS-CoV infection.

SARS-CoV-2 variants continue to emerge during the global pandemic and may facilitate escape from current antibody therapies and vaccine protection. Here, we showed that the South African variant B.1.351 was the most resistant to current monoclonal antibodies and convalescent plasma from COVID-19-infected individuals, followed by the Brazilian variant P.1 and the UK variant B.1.1.7. This resistance hierarchy corresponded with Y144del and 242-244del mutations in the N-terminal domain and K417N/T, E484K and N501Y mutations in the receptor binding domain (RBD) of SARS-CoV-2. Crystal structural analysis of B.1.351 triple mutant (417N-484K-501Y) RBD complexed with monoclonal antibody P2C-1F11 revealed the molecular basis for antibody neutralization and escape. B.1.351 and P.1 also acquired the ability to use mouse and mink ACE2 receptor for entry. Our results demonstrate major antigenic shifts and potential broadening of the host range for B.1.351 and P.1 variants, which pose serious challenges to our current antibody therapies and vaccine protection.

The pandemic caused by emerging coronavirus SARS-CoV-2 presents a serious global public health emergency in urgent need of prophylactic and therapeutic interventions. SARS-CoV-2 cellular entry depends on binding between the viral Spike protein receptor-binding domain (RBD) and the angiotensin converting enzyme 2 (ACE2) target cell receptor. Here, we report on the isolation and characterization of 206 RBD-specific monoclonal antibodies (mAbs) derived from single B cells of eight SARS-CoV-2 infected individuals. These mAbs come from diverse families of antibody heavy and light chains without apparent enrichment for particular families in the repertoire. In samples from one patient selected for further analyses, we found coexistence of germline and germline divergent clones. Both clone types demonstrated impressive binding and neutralizing activity against pseudovirus and live SARS-CoV-2. However, the antibody neutralizing potency is determined by competition with ACE2 receptor for RBD binding. Surprisingly, none of the SARS-CoV-2 antibodies nor the infected plasma cross-reacted with RBDs from either SARS-CoV or MERS-CoV although substantial plasma cross-reactivity to the trimeric Spike proteins from SARS-CoV and MERS-CoV was found. These results suggest that antibody response to RBDs is viral species-specific while that cross-recognition target regions outside the RBD. The specificity and neutralizing characteristics of this plasma cross-reactivity requires further investigation. Nevertheless, the diverse and potent neutralizing antibodies identified here are promising candidates for prophylactic and therapeutic SARS-CoV-2 interventions.