Potent Neutralizing Antibodies Directed to Multiple Epitopes on SARS-CoV-2 Spike

Abstract

23The SARS-CoV-2 pandemic rages on with devasting consequences on human lives and the 24 global economy. The discovery and development of virus-neutralizing monoclonal 25 antibodies could be one approach to treat or prevent infection by this novel coronavirus. 26Here we report the isolation of 61 SARS-CoV-2-neutralizing monoclonal antibodies from 5 27 infected patients hospitalized with severe disease. Among these are 19 antibodies that 28 potently neutralized the authentic SARS-CoV-2 in vitro, 9 of which exhibited exquisite 29 potency, with 50% virus-inhibitory concentrations of 1 to 9 ng/mL. Epitope mapping showed 30 this collection of 19 antibodies to be about equally divided between those directed to the 31 receptor-binding domain (RBD) and those to the N-terminal domain (NTD), indicating that 32 both of these regions at the top of the viral spike are quite immunogenic. In addition, two 33 other powerful neutralizing antibodies recognized quaternary epitopes that are overlapping 34 with the domains at the top of the spike. Cyro-electron microscopy structures of one 35 antibody targeting RBD, a second targeting NTD, and a third bridging RBD and NTD 36 revealed recognition of the closed, "all RBD-down" conformation of the spike. Several of 37 these monoclonal antibodies are promising candidates for clinical development as potential 38 therapeutic and/or prophylactic agents against SARS-CoV-2. 39 40 Background 41A novel coronavirus, now termed SARS-CoV-2 1,2 , has caused nearly 8 million confirmed 42 infections globally, leading to about 450,000 deaths. This pandemic has also put much of the 43 world on pause, with unprecedented disruption of lives and unparalleled damage to the economy. 44A return to some semblance of normalcy will depend on science to deliver an effective solution, 45 and the scientific community has responded admirably. Drug development is well underway, and 46 vaccine candidates are entering clinical trials. Another promising approach is the isolation of 47 SARS-CoV-2-neutralizing monoclonal antibodies (mAbs) that could be used as therapeutic or 48 prophylactic agents. The primary target for such antibodies is the viral spike, a trimeric protein 3,4 49 that is responsible for binding to the ACE2 receptor on the host cell 1,3,5,6 . The spike protein is 50 comprised of two subunits. The S1 subunit has two major structural elements: RBD and NTD; the 51 S2 subunit mediates virus-cell membrane fusion after the RBD engages ACE2. Reports of 52 discovery of neutralizing mAbs that target the RBD have been published recently 7-11 . We now 53 describe our efforts in isolating and characterizing a collection of mAbs that not only target 54 multiple epitopes on the viral spike but also show exquisite potency in neutralizing SARS-CoV-2. 55 56 Patient Selection 57Forty patients with PCR-confirmed SARS-CoV-2 infection were enrolled in an observational 58 cohort study on virus-neutralizing antibodies. Plasma samples from all subjects were first tested 59 for neutralizing activity against SARS-CoV-2 pseudovir...

An ultra-potent synthetic nanobody neutralizes SARS-CoV-2 by locking Spike into an inactive conformation

“…Most recent strategies to prevent SARS-CoV-2 entry into the host cell aim at blocking the ACE2-RBD interaction. High-affinity monoclonal antibodies, many identified from convalescent patients, are leading the way as potential therapeutics (22)(23)(24)(25)(26)(27)(28)(29). While highly effective in vitro, these agents are expensive to produce by mammalian cell expression and need to be intravenously administered by healthcare professionals (30).…”

“…For one Class II nanobody, Nb3, we identified a likely binding site in the Spike S1 domain external to the RBDs. Previously discovered neutralizing antibodies from convalescent patients bind an epitope in a similar region of Spike (24,26,27). Binding of Nb3 to this epitope may allosterically stabilize RBDs in the down-state, thereby decreasing ACE2 binding.…”

A glycan cluster on the SARS-CoV-2 spike ectodomain is recognized by Fab-dimerized glycan-reactive antibodies

“…to HIV-1 Env versus is binding to SARS-CoV-2 S is that on the HIV-1 Env all the binding glycans come from a single gp120 monomer in the HIV-1 Env trimer, whereas in the SARS-CoV-2 S protein the binding site is quaternary and the 2G12-binding glycans at each site are contributed by two of the three protomers, making this one of the first examples of a quaternary epitope spanning multiple protomers on the SARS-CoV-2 S protein (28), and the first in the S2 domain.…”

Escape from neutralizing antibodies by SARS-CoV-2 spike protein variants

“…Recent studies have shown that related, potently neutralizing monoclonal antibodies that recognize the SARS-CoV-2 receptor binding domain (RBD) are often elicited in SARS-CoV-2 infection (3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17). These antibodies have great potential to be clinically impactful in the treatment and prevention of SARS-CoV-2 infection.…”