Access the full text.
Sign up today, get DeepDyve free for 14 days.
Loading next page...
/lp/oxford-university-press/emerging-antibody-based-therapeutics-against-sars-cov-2-during-the-ve6PyyeHxc
- Publisher
- Oxford University Press
- Copyright
- © Published by Oxford University Press on behalf of Antibody Therapeutics 2020
- eISSN
- 2516-4236
- DOI
- 10.1093/abt/tbaa025
- Publisher site
- See Article on Publisher Site
Abstract
SARS-CoV-2 antibody therapeutics are being evaluated in clinical and preclinical stages. As of 11 October 2020, 13 human monoclonal antibodies targeting the SARS-CoV-2 spike protein have entered clinical trials with three (REGN-COV2, LY3819253/LY-CoV555, and VIR-7831/VIR-7832) in phase 3. On 9 November 2020, the US Food and Drug Administration issued an emergency use authorization for bamlanivimab (LY3819253/LY- CoV555) for the treatment of mild-to-moderate COVID-19. This review outlines the development of neutralizing antibodies against SARS-CoV-2, with a focus on discussing various antibody discovery strategies (animal immunization, phage display and B cell cloning), describing binding epitopes and comparing neutralizing activities. Broad-neutralizing antibodies targeting the spike proteins of SARS-CoV-2 and SARS-CoV might be helpful for treating COVID-19 and future infections. VIR-7831/7832 based on S309 is the only antibody in late clinical development, which can neutralize both SARS-CoV-2 and SARS-CoV although it does not directly block virus receptor binding. Thus far, the only cross-neutralizing antibody that is also a receptor binding blocker is nanobody VHH-72. The feasibility of developing nanobodies as inhaled drugs for treating COVID-19 and other respiratory diseases is an attractive idea that is worth exploring and testing. A cocktail strategy such as REGN-COV2, or engineered multivalent and multispecific molecules, combining two or more antibodies might improve the efficacy and protect against resistance due to virus escape mutants. Besides the receptor-binding domain, other viral antigens such as the S2 subunit of the spike protein and the viral attachment sites such as heparan sulfate proteoglycans that are on the host cells are worth investigating. Statement of Significance: This review summarizes ongoing efforts to develop neutralizing antibodies against SARS-CoV-2 with a focus on targets, neutralizing activities and screening strategies, including phage display, animal immunization and B cell cloning. A cocktail strategy combining two or more antibodies, including nanobodies, targeting different epitopes might protect against mutant resistance. KEYWORDS: SARS-CoV-2; spike or S protein; human antibody; cocktail therapy; single domain antibody or nanobody INTRODUCTION in an intensive care unit (1–6). As of 12 November 2020, The severe acute respiratory syndrome coronavirus 2 there are 53 001 867 confirmed cases and 1 289 231 deaths (SARS-CoV-2) first appeared in late 2019 and caused the worldwide, with 203 countries/regions affected (https://co Coronavirus Disease commonly known as COVID-19. In ronavirus.jhu.edu/map.html). Global efforts are ongoing some cases, this coronavirus results in a syndrome leading to treat COVID-19 and to flatten the pandemic curve. to a critical care condition that requires specialized care This review aims to summarize our current knowledge on To whom correspondence should be addressed: National Cancer Institute, Bethesda, MD 20892, USA. Tel: +1 (240) 760-7848; Web: https://ccr.cancer. gov/mitchell-ho; Tel: (240) 760-7848; Fax: (240) 541-4501; Email: homi@mail.nih.gov © Published by Oxford University Press on behalf of Antibody Therapeutics 2020 This work is written by US Government employees and is in the public domain in the US Antibody Therapeutics, 2020 247 antibody-based therapeutics against SARS-CoV-2 by pro- CoVs can be divided into four distinct groups based on the viding an overview of neutralizing antibody development genomic sequence alignment phylogenetically, defined as mainly targeting the spike (S) protein. α, β, γ and δ. β-Coronaviruses may further be subgrouped as lineage a, b, c, and d in classical taxonomy. Both SARS-CoV and SARS-CoV-2 belong to β-genus lineage b, CORONAVIRUS OUTBREAK HISTORY whereas MERS-CoV belongs to β-genus lineage c. HCoV- OC43 and HCoV-HKU1 are β-genus lineage a, whereas Coronaviruses (CoVs) are potentially lethal pathogens, HCoV-229E and HCoV-NL63 are α-genus (28, 29). with seven strains having emerged to infect humans in The SARS-CoV-2 viral genome of about 27–32 kb recent years. Human coronavirus-229E (HCoV-229E) and encodes for structural and non-structural proteins. The HCoV-OC43 were identified in the 1960s and reported to structural proteins include membrane (M) protein, enve- cause symptoms similar to that of a mild common cold, lope (E) protein, nucleocapsid (N) protein and spike (S) except in infants, the elderly and the immunocompromised protein. The S protein plays a role in viral entry and is cru- (7–9). Decades later, in 2002–3, outbreak of SARS-CoV cial for determining host tropism and transmission capacity infection became a global pandemic (10, 11). SARS-CoV (30–32). The S protein mainly consists of two functional is thought to be an animal virus from its natural reservoir, subunits, S1 and S2. S1 is responsible for host cell receptor perhaps bats, that spread to other animals (civet cats) as an binding, while S2 is responsible for viral and cellular intermediate host in animal-to-human transmission (12, membrane fusion (33). For many CoVs, the S protein is 13). Patients infected with SARS-CoV exhibited atypical cleaved between the S1 and S2 subunits, which can activate pneumonia that had the potential to progress to acute the protein for membrane fusion (34–38). CoVs entry into respiratory distress syndrome (14). As of 13 July 2003, susceptible cells is a complex process that requires the when the last new probable case was reported, there was process of receptor-binding and proteolytic processing of a total of 8096 probable cases and 774 deaths (case-fatality the S protein to cause the virus-cell fusion. The structure of rate: 9.56%) (15). Two more coronaviruses, HCoV-NL63 the S protein allows extensive conformational flexibility as and HCoV-HKU1, were found in 2004–5 from archived it modulates its ACE2 receptor binding and later undergoes nasopharyngeal aspirates and caused mild to serious lower dramatic conformational change to facilitate the fusion of respiratory tract infections (16–18). In 2012, almost a viral and cellular membranes (39, 40). Using cryo-electron decade after the first SARS-CoV outbreak, the Middle East microscopy and tomography, Ke et al. determined the high- respiratory syndrome coronavirus (MERS-CoV) caused resolution structure of S trimers on the virion surface (41). a total of 2494 laboratory-confirmed cases, including 858 Each virion is a spherical with a diameter of 91 ± 11 nm. associated deaths (case-fatality rate: 34.4%) globally (19). It Each individual virion contains only 24 ± 9 S trimers, lower was reported that MERS-CoV has the same receptor usage than previously estimated. Notably, the trimers do not all and cell entry as bat coronavirus HKU4, which provides protrude straight from the viral surface (41). In fact, they an insight into bat-to-human transmission of MERS-CoV can tilt by up to 90 toward the membrane, though tilts (20, 21). In December 2019, cases of mysterious pneumonia over 50 are decreasingly favored. were reported in Wuhan, Hubei Province, China, which A recent report about the molecular assembly of the were later confirmed to be caused by a new coronavirus authentic SARS-CoV-2 virus at average resolutions of 8.7– named SARS-CoV-2. Although bats are probable reservoir 11 Å largely confirms previous observations using recom- hosts for the new coronavirus (22), any intermediate binant S proteins (42). The biological explanation for the host that may facilitate transfer to humans has not been tilted S trimer on the virion is unclear. It might be possi- identified. While the researchers have isolated a coronavirus ble that they represent different prefusion stages of the S from a Malayan pangolin, the S protein receptor-binding protein. Based on the S protein sequence alignment, the domain (RBD) of pangolin-CoV is similar to that of overall similarities between SARS-CoV-2 S and SARS- SARS-CoV-2 (23, 24), indicating that pangolin could CoV S (isolated from human, civet or bat) are ∼76–78% be a potential intermediate host. It has been speculated for the whole protein and 73–76% for the receptor binding that SARS-CoV-2 might be the result of a recombination domain (RBD) (22, 39, 43). The sequence similarity may between bat (RaTG13) and pangolin coronaviruses based partly explain why SARS-CoV-2 and SARS-CoV share on the analysis of the S protein sequences (25). The SARS- the receptor ACE2 on host cells. Additionally, this shared CoV-2 S protein contains a few residues (e.g., F486 and characteristic may provide the rationale or possibility to N501) for stronger contacts with human angiotensin develop cross-neutralizing antibodies to both of CoVs (27, converting enzyme 2 (ACE2) (26). These residues are also 39, 44, 45). found in the sequence of pangolin coronavirus (27). CORONAVIRUS SPIKE PROTEIN IS IMPORTANT ANTIBODY THERAPEUTICS FOR COVID-19 FOR VIRUS ENTRY The US Food and Drug Administration (FDA) has CoVs are enveloped viruses containing single-stranded approved the repurposing of some drugs as emergency positive-sense RNA that belongs to the Coronaviridae treatment for severe COVID-19 patients (46). However, family of the Orthocoronavirinae subfamily, which can major ongoing preclinical and clinical studies have focused cause illness in animals and humans. CoVs are a large on identifying anti-SARS-CoV-2 antibodies targeting its family that is genotypically and phenotypically diverse. spike protein, thereby blocking virus entry effectively (27, 248 Antibody Therapeutics, 2020 47). Three antibody drugs specific for the S protein of protein (53), since the VH library panning on RBD protein SARS-CoV-2 are being evaluated in phase 3 clinical trials: was unable to get neutralizing antibodies as mentioned REGN-COV2 (REGN10933 + REGN10987; Regeneron/- in the study. The two human VH antibodies, n3130 and NIAID), LY3819253 (LY-CoV555; AbCellera/Eli Lilly/NI- n3088, were identified to bind to the cryptic epitope located AID) and VIR-7831/VIR-7832 (Vir biotechnology/GSK). in the spike trimeric interface. The study reported that Meanwhile, antibodies targeting other antigens are also both antibodies had neutralizing ability against SARS- under investigation and will be discussed later in this review. CoV-2 with an IC of ∼2.6 µg/mL (17.3 nM). Overall, the antibodies isolated from phage libraries have relatively low neutralizing activities against SARS-CoV-2 without Antibodies targeting the S protein affinity maturation. Further improvement on the library Neutralizing monoclonal antibodies against the S protein size and screening strategies might be necessary to isolate may block virus entry. The RBD located in the S protein potent neutralizing antibodies by phage display technology. is responsible for host cell receptor binding, making it a Nevertheless, phage display might have an advantage primary target of neutralizing antibody development (27). over other screening strategies to isolate cross-reactive There are two conformations, prefusion and postfusion, antibodies against multiple SARS-related coronaviruses for the S trimer structure (40). It has been experimen- or multiple variants/mutants of SARS-CoV-2. Further tally shown that ∼97% of S trimers are in the prefusion affinity maturation using phage display (51, 54), yeast form, and only 3% in the postfusion form (41). A previ- display (55, 56) or mammalian cell display (57) might be ously reported SARS-CoV monoclonal antibody, CR3022 needed to improve their neutralizing activities. (48, 49), was demonstrated for the first time to also bind Animal immunization has also been used to isolate potently with SARS-CoV-2 RBD at nanomolar affinity antibodies targeting SARS-CoV-2 S protein. Wang et al. (50); however, it does not show cross-neutralizing ability identified SARS-CoV-2 reactive antibodies from S pro- with SARS-CoV-2. The most promising preclinical studies tein immunized transgenic mice (H2L2) that encode of antibodies targeting the spike protein are summarized in chimeric immunoglobulins with human antibody variable Table 1. regions and rat antibody constant regions. Of all the Different screening strategies such as phage display, hybridoma supernatant, one antibody (47D11) exhibited animal immunization or single B cell cloning, were used cross-neutralizing activity of SARS-CoV and SARS- to isolate neutralizing antibodies in these studies. By CoV-2 pseudotyped virus infection. The chimeric 47D11 phage library panning, Wrapp et al. isolated single-domain antibody was humanized by cloning of the human variable camelid antibodies, V Hs including VHH-72, from a regions into a human IgG1 framework (58). Taken llama immunized with prefusion-stabilized coronavirus together, animal immunization with S protein from multiple spikes (45). These V Hs could neutralize MERS-CoV or CoVs is an efficient way to identify cross-neutralizing SARS-CoV pseudoviruses. After V H engineered into a antibodies. bivalent format with human IgG1 Fc-fusion (VHH-72- As the most popular strategy so far, single B-cell Fc), it obtained the cross-neutralization ability with IC cloning allows for the rapid generation of antigen-specific of 0.2 µg/mL (2.7 nM) on pseudotyped SARS-CoV, as well monoclonal antibodies in a matter of several weeks, as SARS-CoV-2, suggesting a strategy using a nanobody to which is highly efficient for antibody development against engineer cross-neutralizing antibodies for future study. Its emerging infectious virus (59). Pinto et al. reported activities on live virus are unknown. Additionally, Huo human monoclonal antibodies targeting SARS-CoV-2 S et al. has isolated H11-D4 from a naïve llama single- protein isolated from memory B cells of an individual domain antibody library using the RBD of SARS-CoV- who was infected with SARS-CoV in 2003. One of these 2 as an antigen for phage panning (51). They improved antibodies, named S309 (the antibody used as the basis the affinity maturation of H11-D4 via affinity maturation for developing VIR-7831/7832), neutralizes both SARS- by phage display and obtained the high affinity mutant CoV-2 and SARS-CoV pseudoviruses, as well as authentic H11-H4. These two V H nanobodies, H11-D4 and H11- SARS-CoV-2 by binding the RBD (60). Interestingly, S309 H4, were capable of binding the RBD with KD of 39 recognizes an epitope containing the N343 glycan (N330 and 12 nM, respectively, and blocked the attachment in SARS-CoV S glycoprotein) conserved within SARS- of S protein to ACE2 in vitro. After fused to Fc, both related coronavirus spike proteins without competing nanobodies could neutralize SARS-CoV-2 live virus, with with ACE2 binding. Like 47D11 (58), S309 is an ACE2 H11-H4-Fc showing a particularly high potency (IC : non-blocker although both human antibodies 47D11 and 4–6 nM) after affinity maturation. Hanke et al. reported S309 are SARS-CoV-2 and SARS-CoV cross-neutralizing the isolation and characterization of an alpaca-derived antibodies (27). Up to date, the only cross-neutralizing single domain antibody Ty1 by immunizing one alpaca antibody that is also an ACE2 blocker is nanobody with SARS-CoV-2 S1-Fc and RBD (52). Ty1 showed the VHH-72 (45). Nevertheless, VIR-7831/7832 based on neutralization on SARS-CoV-2 pseudotyped viruses at an S309 is currently being tested in phase 3 clinical trials IC of 0.77 µg/mL (64 nM). A cryo-electron microscopy and it is the only antibody in late clinical development structure demonstrated that Ty1 binds to an epitope on the that can neutralize both SARS-CoV-2 and SARS-CoV RBD accessible in both the ‘up’ and ‘down’ conformations, viruses. It would be interesting to examine the potential sterically blocking RBD-ACE2 binding. In another study, clinical benefits of this novel cross-neutralizing antibody Wu et al. isolated two human VH single domain antibodies for treating current COVID-19 patients and potential from an engineered VH library by panning on S1 subunit SARS-related CoV infections in the future. Antibody Therapeutics, 2020 249 Table 1. Preclinical studies of antibodies targeting the spike protein of SARS-CoV-2 VHH-72-Fc Llama V H, fused to hIgG1 Fc SARS-CoV RBD was used for phage panning RBD Pseudovirus SARS-CoV, SARS-CoV-2 Daniel Wrapp et al., by an immunized llama library IC 0.2 µg/mL (2.7 nM) Cell, 2020 (45) H11-H4-Fc Llama V H, fused to hIgG1 Fc SARS-CoV-2 RBD was used for phage panning RBD Live SARS-CoV-2 IC 4–6 nM Jiangdong Huo et al., H 50 by a naïve llama library Nature Structural & Molecular Biology, 2020 (48) Ty1 Alpaca V H Immunized one alpaca with SARS-CoV-2 S1-Fc RBD Pseudoviruse SARS-CoV-2 and RBD on a 60-day immunization schedule IC of 0.77 µg/mL (64 nM) Leo Hanke et al., Nature Communications, (52) n3130, n3088 Human VH SARS-CoV-2 S1 was used for phage panning by cryptic epitope located an engineered human VH library in the spike trimeric interface Live SARS-CoV-2 Yanling Wu et al., Cell Host&Microbe, 2020 (53) IC ∼ 2.6 µg/mL (17.3 nM) 47D11 Reformat to human IgG1 Immunized transgenic H2L2 mice with a conserved epitope in Live SARS-CoV IC 0.19 µg/mL (1.2 nM) SARS-CoV-1 S protein RBD Live SARS-CoV-2 IC Chunyan Wang et al., Nature 0.57 µg/mL (3.8 nM) Communications, 2020 (58) P2C-1F11, P2B-2F6, P2C-1A3 Human IgG1 Single B cell antibody isolation of 8 RBD live SARS-CoV-2 IC ∼ 0.1 µg/mL Bin Ju et al., Nature, SARS-CoV-2 infected individuals (0.7 nM) 2020 (68) CB6 Human IgG1 Utilized SARS-CoV-2 RBD as the bait to sort Overlapping with Live virus IC 36 ng/mL (0.24 nM) specific memory B cells PBMCs of a ACE2-binding sites in convalescent COVID-19 patient SARS-CoV-2 RBD CB6 (50 mg/kg) inhibited Rui Shi et al., Nature, 2020 (69) SARS-CoV-2 infection in rhesus monkeys at both prophylactic and treatment settings C121, C144, C135 Human IgG1 Single B cell antibody isolation from 6 Different binding Live SARS-CoV-2 IC 1.64, 2.55 and Davide F. Robbiani convalescent individuals epitope from CR3022 2.98 ng/mL (10.9 pM, 17 pM, 19.8 pM) et al., Nature, 2020 (70) CC12.1 Human IgG1 Single B cell antibody isolation from 3 RBD Live SARS-CoV-2 IC 22 ng/mL Thomas F. Rogers et al., convalescent individuals (0.14 nM) CC12.1 (4 mg/kg) inhibited Science, 2020 (71) SARS-CoV-2 infection in Syrian hamsters in prophylaxis setting COVA1–18 COVA2–15 Human IgG1 Single B cell antibody isolation from 3 Competition with ACE2 Live SARS-CoV-2 IC of 7 and 9 ng/mL Philip J. M. Brouwer SARS-CoV-2 infected individuals binding site to RBD (46 and 60 pM) et al., Science, 2020 (72) 4A8 Human IgG1 Single B cells antibody isolation of 10 COVID-19 NTD Live SARS-CoV-2 IC 0.6ug/mL (4 nM) Xiangyang Chi et al., recovered patients with different ages and Science, 2020 (73) different infection phase 250 Antibody Therapeutics, 2020 Jones et al. recently reported the isolation of LY3819253/ single B cells of eight SARS-CoV-2 infected individuals LY-CoV555 (bamlanivimab) to the RBD of the SARS- (68). The most potent antibodies, P2C-1F11, P2B-2F6, CoV-2 spike protein using two single B cell screening and P2C-1A3, neutralize live SARS-CoV-2 with an IC s methods, multiplexed bead-based assay and live cell-based of 0.03, 0.41, and 0.28 µg/mL (200 pM, 2.7 nM, 1.8 nM) assay, from a patient hospitalized with COVID-19 in respectively. These antibodies are most competitive with mid-February 2020 (61). Next-generation sequencing of ACE2, indicating that blocking the RBD and ACE2 inter- antibody genes from selected single B-cells shows that of the action is a useful surrogate for neutralization. However, 440 unique antibodies identified, only 4% are cross-reactive none of the anti-SARS-CoV-2 antibodies cross-react with to both full-length SARS-CoV-2 and SARS-CoV spike SARS-CoV RBD. Similarly, Shi et al. reported a human proteins. Notably, the neutralization potency of Ab169 monoclonal antibody CB6 utilizing SARS-CoV-2 RBD (later called LY3819253/LY-CoV555 or bamlanivimab), as the bait to sort specific memory B cells PBMCs of a an RBD binder and ACE2 blocker, exhibits the greatest convalescent COVID-19 patient (69). CB6 exhibits strong activity with the IC value of 100 pM in live virus assay neutralizing activity against live SARS-CoV-2 infection among all the antibodies. In a rhesus macaque challenge of Vero E6 cells, with an observed IC of 0.036 µg/mL model, prophylaxis doses as low as 2.5 mg/kg reduce (240 pM). In addition, CB6 inhibits SARS-CoV-2 infection viral replication in the upper and lower respiratory tract. in rhesus monkeys in both prophylactic and treatment Mechanistically, LY-CoV555 binds the spike protein RBD settings. At present, CB6 is in phase 1 clinical trials in China in both up and down conformations such as mAb114 that and the USA. However, CB6 is not a cross-neutralizing binds the Ebola virus glycoprotein RBD in both the pre- antibody and cannot cross-bind to SARS-CoV S either. activation and activated states for treating Ebola infection Robbiani et al. reported antibody isolation on 149 COVID- (62, 63). LY-CoV555 (bamlanivimab) is being evaluated 19 convalescent individuals (70). Plasma samples binding in phase 3 clinical trials and has been recently approved to the SARS-CoV-2 RBD and trimeric spike proteins were as an emergency use authorization for the treatment of collected, followed by neutralization activity testing on mild-to-moderate COVID-19. SARS-CoV-2 pseudovirus. Lastly, 534 paired IgG heavy In order to overcome virus escape mutation, Regen- and light chain sequences were obtained by reverse tran- eron has described parallel efforts utilizing both animal scription PCR from individual RBD-binding B cells from immunization (genetically humanized mice) and B cell six convalescent individuals. Potent neutralizing antibodies, cloning from convalescent humans to generate a large C121, C144, and C135 with an IC s of 1.64, 2.55, and collection of highly potent human neutralizing antibodies 2.98 ng/mL (10.9 pM, 17 pM, 19.8 pM), against authentic targeting the RBD of the spike protein of SARS-CoV-2 SARS-CoV-2 were identified. The bilayer interferometry (64). Genetically humanized mice were immunized with result has shown that these three antibodies can bind with a DNA plasmid that expresses SARS-CoV-2 S protein different epitopes from CR3022. Negative stain electron and boosted with a recombinant RBD protein. The most microscopy imaging has confirmed the different binding potent antibodies with IC values of low pM (e.g., 37 epitope. Using similar methodology, Rogers et al. reported pM for REGN10933, 42 pM for REGN10987) might be a rapid screening platform to generate over 2045 antibodies isolated from humanized mice, suggesting that animal from a cohort of SARS-CoV-2 recovered participants in immunization induced high affinity antibodies to the 2 weeks (71). CC12.1 isolated by single B cell cloning virus spike protein. The antibody cocktail to SARS-CoV- from recovery patient donors was able to show the 100% 2, REGN-COV2 (REGN10933 + REGN10987), could neutralization of live SARS-CoV-2 at a concentration of prevent rapid mutational escape of virus variants that 22 ng/mL (146 pM). Most importantly, CC12.1 at a dose of have arisen in the human population (65). Genomics 500 µg/animal (on average 4 mg/kg) could protect against analysis of SARS-CoV-2 from the same individual with re- weight loss and lung viral replication in Syrian hamsters infection shows genetically significant differences between challenged intranasally with 1 × 10 PFU of SARS-CoV- the variants associated with early infection and re-infection 2. Another study, Brouwer et al. isolated 19 neutralizing (66). The second infection is symptomatically even more antibodies from single B cell derived from three SARS- severe than the first one. REGN-COV2 and other cocktail CoV-2 infected individuals (72). Two of them, COVA1–18 or multispecific therapeutics might be useful to overcome and COVA2–15, showed picomolar neutralizing activities potential epitope escape variants in re-infection. In against authentic SARS-CoV-2 with an IC of 7 and addition, REGN-COV2 appears highly potent therapeutic 9 ng/mL (46 and 60 pM), respectively. Through large-scale antibodies against SARS-CoV-2 S protein with low pM SPR-based competition assay and electron microscopy activities on live virus. REGN-COV2 cocktail therapy is studies, antibodies with different binding epitopes to the being evaluated in phase 3 clinical trial. spike protein were demonstrated, including RBD and non- Researchers from Astrazeneca have isolated 389 SARS- RBD epitopes. However, these antibodies targeting non- CoV-2 S-protein-reactive human monoclonal antibodies RBD epitope are not able to neutralize SARS-CoV-2. The from the B cells of two convalescent individuals who had above two most potent antibodies can compete with ACE2 been infected with SARS-CoV-2 in Wuhan, China. Among binding site to RBD. these human antibodies, COV2–2196 and COV2–2130 Currently, most of the antibodies developed are targeting bound simultaneously to the S protein and neutralized RBD in the spike protein. However, Chi et al. isolated wild-type SARS-CoV-2 virus in a synergistic manner (67). monoclonal antibodies derived from 10 patients that have Ju et al. reported the isolation and characterization of recovered from SARS-CoV-2 viral infection, the patient’s 206 RBD-specific monoclonal antibodies derived from age ranging from 25 to 53 years, and memory B cells were Antibody Therapeutics, 2020 251 collected from different infection phase. 4A8 is a human COVID-19 (27). Recently, Clausen et al. showed that monoclonal antibody that targets the N-terminal domain SARS-CoV-2 S protein interacted with cell surface heparan (NTD) of the SARS-CoV-2 S protein and exhibits high sulfate and ACE2 through its RBD. Interestingly, the S neutralization potency against SARS-CoV-2 although it protein binding to heparan sulfate and ACE2 on the cell does not directly inhibit the interaction between RBD and surface may occur co-dependently. Heparin and purified ACE2 (73). Liu et al. reported the isolation of 19 antibod- heparan sulfate can block S protein binding and infection ies from five patients infected with SARS-CoV-2, which by SARS-CoV-2 virus, suggesting using heparin as bait could neutralize SARS-CoV-2 in vitro. Epitope mapping to attract the virus away from human cells. It would be showed that this collection of 19 human antibodies was interesting to further validate whether heparin, an approved about evenly divided against the RBD and NTD, indicating medication to treat blood clots, might be repurposed to that these two regions at the top of the viral spike are reduce SARS-CoV-2 infection. In another study, Zhang immunogenic (74). et al. also showed that heparan sulfate facilitated spike- Interestingly, Ma et al. reported a strategy using cell- dependent viral entry and screened approved drugs to based chimeric antigen receptor (CAR) technology. They identify inhibitors targeting the HS-dependent cell entry have developed a novel approach for the generation (88). Altogether, these studies indicate heparan sulfate as of CAR-NK cells using the scFv fragment of CR3022 a co-receptor for viral entry and support the rationale (henceforth, CR3022-CAR-NK) for targeting SARS-CoV- for developing therapeutics that target heparan sulfate 2, which showed specifically killing to pseudo-SARS-CoV- for inhibiting SARS-CoV-2 and other virus infections. 2 infected target cells in vitro (75). While it could be a Biochemical analysis for identification of specific binding complimentary strategy worth exploring, many questions motifs of HS (e.g., 2-O, 3-O, or 6-O sulfation (87) and should be addressed in more biologically relevant assays, N-sulfation (82)) for SARS-CoV-2 attachment would be including animal testing. In particular, how biologically useful for designing specific anti-viral inhibitors. and therapeutically this cell-based therapy could stop SARS-CoV-2 virus proliferation and spread is unclear. The potential side-effects induced by CAR-based cell therapies ONGOING CLINICAL TRIALS OF ANTIBODIES need to be carefully evaluated in proof-of-concept animal TARGETING THE SPIKE PROTEIN studies before they can be used in humans. There are 13 clinical trials ongoing related to human mon- oclonal antibodies targeting SARS-CoV-2 spike protein Antibodies targeting the host derived proteins described in Table 2. According to the COVID-19 Anti- Some studies have investigated the changes of several body Therapeutics Tracker (https://chineseantibody.org/ cytokines in serum of the COVID-19 patients that generates covid-19-track/) (46), three antibody drugs have entered a series of immune responses, and the cytokine storm into phase 3 clinical trials. Among them, REGN10933 and syndrome was proportional to the severity of disease (3, REGN10987 represent a non-competing pair of antibodies 4, 76). The pro-inflammatory cytokine IL-6 may have that can simultaneously bind to RBD and thus can be a prominent role, leading to the inflammatory cascade, partners for a therapeutic antibody cocktail aimed at which may result in increased alveolar-capillary blood-gas decreasing the potential for mutant viral strain escaping. exchange dysfunction (77, 78). Antibodies targeting IL-6, Regeneron in collaboration with the National Institute of such as Olokizumab and Siltuximab, are in phase 3 trial Allergy and Infectious Diseases (NIAID) at the National at present. Clinically, Stoclin et al. reported the case of Institutes of Health (NIH) initiated a phase 3 clinical trial a patient with a respiratory failure linked to COVID-19 evaluating REGN-COV2 (REGN10933 + REGN10987) who had a rapid favorable outcome after two infusions for the treatment and prevention of COVID-19 in late June of Tocilizumab, an anti-IL-6 receptor antibody (79). 2020. LY3819253 (LY-CoV555) developed by AbCeller- However, Stone et al. reported a randomized, double-blind, a/Eli Lilly in collaboration with the NIAID/NIH also placebo-controlled phase 3 trial involving 243 patients entered phase 3 clinical trial. Notably, on 9 November 2020, with confirmed SARS-CoV-2 infection and found that the US FDA issued an emergency use authorization for Tocilizumab was not effective for preventing intubation bamlanivimab (LY3819253/LY-CoV555) for the treatment or death in moderately ill hospitalized patients with of mild-to-moderate COVID-19 in adult and pediatric COVID-19 (80). patients (https://www.fda.gov/news-events/press-announce Heparan sulfate proteoglycans (HSPGs) provide the ments/coronavirus-covid-19-update-fda-authorizes-mono attachment sites for virus such as polyomaviruses, papil- clonal-antibody-treatment-covid-19). Bamlanivimab has lomavirus, and hepatitis C virus, to make primary contact been shown in two randomized, double-blind, placebo- with the host cell surface (81–83). Treatment of the cells controlled clinical trial in 465 non-hospitalized adults with heparinase or heparin prevents the binding of the with mild-to-moderate COVID-19 symptoms to reduce S protein to host cells and inhibits SARS pseudovirus COVID-19-related hospitalization or emergency room vis- infection (84). Based on the findings in previous studies its. However, a clinical benefit of bamlanivimab treatment including ours using the HS20 human monoclonal anti- has not been established in hospitalized patients due to body targeting heparan sulfate to inhibit viral infection COVID-19. (81, 84–87), we speculated that in addition to ACE2, Recently, GlaxoSmithKline (GSK) and Vir Biotech- HSPGs might be another potential target on human cells nology declared that they had launched the phase 2/3 that can be blocked by therapeutic antibodies for treating study of VIR-7831, which also has the development name 252 Antibody Therapeutics, 2020 Table 2. Ongoing clinical trials of antibodies targeting the spike protein of SARS-CoV-2 REGN-COV2 Animal immunization RBD Phase 3 Regeneron Live SARS-CoV-2 Johanna Hansen (REGN10933 + using genetically- IC of 40 pM et al., Science, REGN10987) umanized mice 2020 (62, 63); NCT04452318 LY3819253 (LY-CoV555; B cell cloning from RBD Phase 3; emergency AbCellera/Eli Lilly Live SARS-CoV-2 Bryan E. Jones, bamlanivimab) convalescent patients use authorization and Company IC of 100pM et al, bioRxiv, for treating 2020 (61); mild-to-moderate NCT04497987 COVID-19 patients VIR-7831/7832 (S309) Single B cells antibody glycan epitope Phase 3 Vir Biotechnolo- Live SARS-CoV-2 Dora Pinto et al., isolation of an individual contains position gy/GlaxoSmithK- IC of 500 pM Nature, 2020 (60); who was infected with N343 line NCT04545060 SARS-CoV in 2003 DXP-593 High-throughput single B N/A Phase 2 Beigene/Singlomics N/A NCT04551898 cell sequencing from over Biopharmaceuti- 60 convalescent patients cals/Peking University JS016 Utilized SARS-CoV-2 Overlapping with Phase 1 Junshi Live SARS-CoV-2 Rui Shi et al., RBD as the bait to sort ACE2-binding Biosciences/Eli IC of 240 pM Nature, 2020 (59); specific memory B cells sites in Lilly and Company NCT04441918 PBMCs of a convalescent SARS-CoV-2 RBD COVID-19 patient TY027 N/A N/A Phase 1 Tychan Pte. Ltd. N/A NCT04429529 CT-P59 N/A N/A Phase 1 Celltrion N/A NCT04525079 BRII-196 N/A N/A Phase 1 Brii Biosciences N/A NCT04479631 BRII-198 N/A N/A Phase 1 Brii Biosciences N/A NCT04479644 SCTA01 N/A N/A Phase 1 Sinocelltech Ltd. N/A NCT04483375 AZD7442 B cell cloning of two Overlapping with Phase 1 AstraZeneca Live SARS-CoV-2 NCT04507256 (AZD8895 + AZD1061) convalescing individuals ACE2-binding IC of 100 pM who had been infected sites in with SARS-CoV-2 in SARS-CoV-2 RBD Wuhan, China MW33 N/A N/A Phase 1 Mabwell N/A NCT04533048 (Shanghai) Bioscience Co., Ltd STI-1499/COVI-SHIELD Screening antibodies in its Overlapping with Phase 1 Sorrento N/A NCT04454398 proprietary G-MAB™ ACE2-binding Therapeutics, Inc. fully human antibody sites in library SARS-CoV-2 RBD N/A: not available. Antibody Therapeutics, 2020 253 GSK4182136. This study, named COMET-ICE, will enroll that inhibit SARS-CoV-2 viral infection at sub-ng/ml 1300 patients worldwide to test VIR-7831/GSK4182136 in concentration (98). Furthermore, multivalent nanobody early treatment of patients infected with SARS-CoV-2 who constructs can achieve high neutralization potency (IC s are at high risk of hospitalization. as low as 0.058 ng/ml). For respiratory infection such as DXP-593 was identified from peripheral blood mononu- COVID-19, nanobodies are particularly attractive because clear cells collected from convalescent patients by high- they might be administered as an inhaler directly to the throughput single-cell sequencing by a joint research site of infection (99). Further studies are necessary to team from the Beijing Advanced Innovation Center for evaluate the feasibility of developing nanobody-based Genomics at Peking University. It has been demonstrated therapeutics as inhaled drugs for treating COVID-19 and to show highly potent neutralizing antibodies against other respiratory infections. SARS-CoV-2; the phase 2 study is currently ongoing. Around 85% of antibodies are developed targeting the S JS016, which is discussed above as antibody CB6, is protein, primarily focused on targeting RBD in an effort the first SARS-CoV-2 neutralizing antibody to enter clin- to block the viral entry at the initial step of binding to the ical trials in China. These trials are led by Junshi and host cell receptor. In the perspective of cross-neutralizing Eli Lilly in China and the rest of the world, respectively. antibody development for SARS related CoVs, it seems JS016 has been tested in rhesus monkeys in prophylactic that antibody, which is derived from SARS-CoV patient and treatment settings. TY027, developed by Tychan in or SARS-CoV immunization animals, could have the cross- Singapore, is also in phase 1 clinical trial. CT-P59, devel- neutralizing activities on SARS-CoV-2. However, antibod- oped by Celltrion, has entered a phase 1 clinical trial in ies derived from SARS-CoV-2 patients usually do not show mild COVID-19 patients. Celltrion previously showed its cross-neutralizing ability on SARS-CoV. Although both S antiviral activities in neutralizing the mutated G-variant proteins from SARS-CoV-2 and SARS-CoV bind human strain (D614G variant) which might be associated with the ACE2 on the cells for viral entry and both S proteins have increased viral transmission of COVID-19. Brii Biosciences highly similar structures, hypothetically, an ACE2 blocker (Brii Bio) company, in collaboration with Tsinghua Univer- that can neutralize both viruses is feasible. However, it sity and 3rd People’s Hospital of Shenzhen, has launched seems rare to identify such a shared epitope using existing the phase 1 clinical trial of BRII-196, BRII-198 for assess- SARS-CoV-2 antibodies. This apparent discrepancy will ing safety, tolerability, and pharmacokinetics in healthy require further investigation on the structure and function adult volunteers. Sinocelltech Ltd. has developed an anti- of the S protein of these two viruses. Cross-neutralizing SARS-CoV-2 monoclonal antibody, SCTA01, and started antibodies are being explored by using various strategies, phase 1 clinical trial in healthy subjects in China. Most including single domain antibodies. recently, AstraZeneca launched the phase 1 clinical trial of Next-generation sequencing of neutralizing antibodies AZD7442 (AZD8895 + AZD1061) in UK as a potential against SARS-CoV-2 has been conducted in several studies. combination therapy for the prevention and treatment of In Regeneron’s study, over 200 antibodies that were isolated COVID-19. Mabwell (Shanghai) Bioscience Co., Ltd., ini- from humanized mice show predominant lineage of anti- tiated a phase I clinical trial with the MW33 antibody. Sor- bodies, which utilize VH3–53 paired with VK1–9, VK1–33, rento Therapeutics, Inc., started a phase 1 clinical trial of or VK1–39, while antibodies isolated from infected humans STI-1499 (COVI-GUARD™) for hospitalized COVID-19 utilize VH3–66 paired with VK1–33 or VH2–70 paired patients. with VK1–39. Interestingly, VH3–53 usage (e.g., VH3– 53/VK1–9 pair (100)) has been found in other human- derived neutralizing antibody against SARS-CoV-2 spike CONCLUSION AND PERSPECTIVE protein (71, 100). In a recent study conducted by Eli Lilly, Most of the therapeutic antibody development against sequencing of about 400 antibodies shows that the VH3 SARS-CoV-2 are currently in the preclinical stage, with germline gene family (e.g., VH3–53, VH3–66) representing about 35% of antibodies in various stages of clinical trials. 57% of total diversity. Among them, VH3–30 usage is the Nearly 82% of current antibodies are human monoclonal most common (38%). However, the selected Ab169 (LY- antibodies with 3, REGN-COV2, LY3819253/LY-CoV555, CoV555) has VH1–69 and VK1–39 germline framework and VIR-7831/VIR-7832, in phase 3 clinical trials, and the sequences. Further sequence analysis of neutralizing anti- second largest group is single-domain antibody (also com- bodies against SARS-CoV-2 RBD might provide valuable monly called nanobody), indicating that emerging single insights of the usage of germline sequences for potent domain antibody development after FDA approved the anti-viral neutralizing activities. Such information could be first nanobody caplacizumab in 2019 (89). Single-domain useful for not only therapeutic antibody development or antibodies can bind novel epitopes including buried cavities optimization but also vaccine or adjuvant design. inaccessible by conventional antibodies (90, 91). Naturally Antibody targeting S2 subunit may also worth studying occurring nanobodies derived from camels (45, 92–94) and since it shows more overall sequence similarity not only sharks (90, 95–97) are stable and relatively easy to express between SARS-CoV and SARS-CoV-2 but also among and fold in various conditions; therefore, they can be other human coronaviruses. Up to date, antibodies target- effective building blocks for the construction of multivalent ing S2 with potent neutralization activities against SARS- and multispecific molecules to effectively neutralize virus. CoV-2 have not been reported. It appears challenging to When revising this review article, another paper reported isolate antibodies to S2 probably due to the dynamic struc- nanobodies (e.g., Nab20, Nab21) isolated from an RBD- ture of the S2 subunit as part of the virus spike trimer and immunized llama with picomolar to femtomolar affinities elusive conformational change at the fusion stage. Further 254 Antibody Therapeutics, 2020 biochemical studies of the S2 subunit and establishment of ds/2020/10/MemoFromThePresdentsPhysician-3.png). On a suitable screening assay might be crucial for generating 2 October 2020, the President received a single 8-gram neutralizing antibodies targeting S2. dose of Regeneron’s REGN-COV2 as a “compassionate Antibodies specific for host targets including the viral use” request from the President’s physicians. On 5 October attachment sites on human cells such as heparan sulfate 2020, President Trump returned to the White House after should be explored as well. A cocktail combination therapy being discharged from the Walter Reed National Military targeting two or more distinct sites or pathways on the viral Medical Center in Bethesda, Maryland. On November 21, surface or host attachment sites might be one of the most 2020, REGN-COV2 (casirivimab and imdevimab) received effective approaches to eliminate virus in the host. Besides Emergency Use Authorization from the U.S. FDA for the cocktail combination therapies of two or more monoclonal treatment of mild to moderate COVID-19 in adults, as well antibodies targeting multiple epitopes on the virus, engi- as in pediatric patients at least 12 years of age and weighing neered multivalent or multispecific molecules using var- at least 40 kg (https://www.fda.gov/news-events/press-a ious antibody binding sites such as single chain Fvs or nnouncements/coronavirus-covid-19-update-fda-authori single domain antibodies might emerge as a new class of zes-monoclonal-antibodies-treatment-covid-19). promising antibody therapeutics for anti-viral therapy. The mechanisms of neutralizing antibodies in terms of protection against SARS-CoV-2 viral infection as well as ACKNOWLEDGEMENTS additional immune functions that may have both protective The content of this publication does not necessarily reflect and pathological consequences are being studied (101). the views or policies of the Department of Health and Besides neutralization, antibodies may have additional Human Services, nor does mention of trade names, com- anti-viral activities mediated by Fc, including antibody- mercial products or organizations imply endorsement by dependent cellular phagocytosis (ADCP), complement- the U.S. Government. dependent cytotoxicity, and antibody-dependent cellular cytotoxicity (ADCC). Using primary natural killer (NK) effector cells and SARS-CoV-2 S-glycoprotein-expressing FUNDING expiCHO as target cells, S309 shows the Fc-mediated This work was supported by the NIH Intramural Tar- ADCC of SARS-CoV-2 S-glycoprotein-transfected cells geted Anti-COVID-19 (ITAC) Program (ZIA BC 011943), (102). In addition, using NK cells or macrophage from Intramural Research Program of NIH, Center for Cancer healthy donors, REGN10987 shows strong ADCC and Research (CCR), National Cancer Institute (NCI) (Z01 ADCP activities against human Jurkat cells expressing BC010891 and ZIA BC010891) and NCI CCR Antibody SARS-CoV-2 spike protein (103). However, antibody Engineering Program (ZIC BC 011891). responses can also cause pathological damages (101). Although sub-neutralizing antibody titers from sec- ond infections have been related to antibody-dependent CONFLICT OF INTEREST STATEMENT enhancement (ADE) in patients with dengue. Evidence of M.H. is the Editor-in-Chief of the journal and is blinded ADE in SARS-CoV-2 patients has not been established so from reviewing or making decisions on the manuscript. far (104). Further understanding how antibodies may play protective and potential pathogenic roles is important for REFERENCES drug design and clinical development. Besides ADE, anti- 1. Zhu, N, Zhang, D, Wang, W et al. A novel coronavirus from drug antibody (ADA) response might be worth evaluating patients with pneumonia in China, 2019. N Engl J Med 2020; 382: as well. Since most current antibodies in the clinical 727–33. trials are human antibodies, immunogenicity and ADA 2. Chan, JF, Yuan, S, Kok, KH et al. A familial cluster of pneumonia effect have not been reported so far. Future analysis of associated with the 2019 novel coronavirus indicating ADA, in particular animal-derived antibodies (e.g., camelid person-to-person transmission: a study of a family cluster. Lancet 2020; 395: 514–23. nanobodies), would be useful for better understanding of 3. Chen, N, Zhou, M, Dong, X et al. Epidemiological and clinical potential ADA in patients. Humanization of nanobodies characteristics of 99 cases of 2019 novel coronavirus pneumonia in might be necessary to reduce immunogenicity in humans. Wuhan, China: a descriptive study. Lancet 2020; 395: 507–13. The review is focused on discussing screening strategies 4. Huang, C, Wang, Y, Li, X et al. Clinical features of patients infected with 2019 novel coronavirus in Wuhan, China. Lancet for isolating neutralizing antibodies against SARS-CoV- 2020; 395: 497–506. 2 and their functional properties. We are also aware that 5. The, L. Emerging understandings of 2019-nCoV. Lancet 2020; 395: such antibodies with high affinity and specificity, including nanobodies, can also be utilized as detectors for diagnostics 6. Bastola, A, Sah, R, Rodriguez-Morales, AJ et al. The first 2019 novel coronavirus case in Nepal. Lancet Infect Dis 2020; 20: 279–80. (105). A microfluidic device or magnetic beads using anti- 7. Tyrrell, DA, Bynoe, ML. Cultivation of a novel type of bodies highly specific for viral proteins can be developed to common-cold virus in organ cultures. Br Med J 1965; 1: 1467–70. capture virus for detection (106). 8. Hamre, D, Procknow, JJ. A new virus isolated from the human respiratory tract. Proc Soc Exp Biol Med 1966; 121: 190–3. 9. McIntosh, K, Dees, JH, Becker, WB et al. Recovery in tracheal Addendum organ cultures of novel viruses from patients with respiratory disease. Proc Natl Acad Sci U S A 1967; 57: 933–40. When preparing this review, US President Trump and the 10. Ksiazek, TG, Erdman, D, Goldsmith, CS et al. A novel coronavirus First Lady were diagnosed with COVID-19 on 1 Octo- associated with severe acute respiratory syndrome. N Engl J Med ber 2020 (https://www.whitehouse.gov/wp-content/uploa 2003; 348: 1953–66. Antibody Therapeutics, 2020 255 11. Peiris, JS, Guan, Y, Yuen, KY. Severe acute respiratory syndrome. 38. Kirchdoerfer, RN, Cottrell, CA, Wang, N et al. Pre-fusion structure Nat Med 2004; 10: S88–97. of a human coronavirus spike protein. Nature 2016; 531: 118–21. 12. Li, W, Shi, Z, Yu, M et al. Bats are natural reservoirs of SARS-like 39. Walls, AC, Park, YJ, Tortorici, MA et al. Structure, function, and coronaviruses. Science 2005; 310: 676–9. antigenicity of the SARS-CoV-2 spike glycoprotein. Cell 2020; 181: 13. Song, HD, Tu, CC, Zhang, GW et al. Cross-host evolution of 281. e286–92. severe acute respiratory syndrome coronavirus in palm civet and 40. Wrapp, D, Wang, N, Corbett, KS et al. Cryo-EM structure of the human. Proc Natl Acad Sci U S A 2005; 102: 2430–5. 2019-nCoV spike in the prefusion conformation. Science 2020; 367: 14. Peiris, JS, Lai, ST, Poon, LL et al. Coronavirus as a possible cause 1260–3. of severe acute respiratory syndrome. Lancet 2003; 361: 1319–25. 41. Ke, Z, Oton, J, Qu, K et al. Structures and distributions of 15. Cherry, JD. The chronology of the 2002-2003 SARS mini SARS-CoV-2 spike proteins on intact virions. Nature 2020. https:// pandemic. Paediatr Respir Rev 2004; 5: 262–9. doi.org/10.1038/s41586-020-2665-2. 16. van der Hoek, L, Pyrc, K, Jebbink, MF et al. Identification of a 42. Yao, H, Song, Y, Chen, Y et al. Molecular architecture of the new human coronavirus. Nat Med 2004; 10: 368–73. SARS-CoV-2 virus. Cell 2020; 183: 730, e713–8. 17. Fouchier, RA, Hartwig, NG, Bestebroer, TM et al. A previously 43. Wan, Y, Shang, J, Graham, R et al. Receptor recognition by the undescribed coronavirus associated with respiratory disease in novel coronavirus from Wuhan: an analysis based on decade-Long humans. Proc Natl Acad Sci U S A 2004; 101: 6212–6. structural studies of SARS coronavirus. J Virol 2020; 94: 18. Woo, PC, Lau, SK, Chu, CM et al. Characterization and complete e00127–20. genome sequence of a novel coronavirus, coronavirus HKU1, from 44. Ge, XY, Li, JL, Yang, XL et al. Isolation and characterization of a patients with pneumonia. J Virol 2005; 79: 884–95. bat SARS-like coronavirus that uses the ACE2 receptor. Nature 19. Coronavirus., M.E.R.S. (2020), Vol. 2020. 2013; 503: 535–8. 20. Yang, Y, Du, L, Liu, C et al. Receptor usage and cell entry of bat 45. Wrapp, D, De Vlieger, D, Corbett, KS et al. Structural basis for coronavirus HKU4 provide insight into bat-to-human transmission potent neutralization of betacoronaviruses by single-domain of MERS coronavirus. Proc Natl Acad Sci U S A 2014; 111: camelid antibodies. Cell 2020; 181: 1436–41. 12516–21. 46. Yang, L, Liu, W, Yu, X et al. COVID-19 antibody therapeutics 21. Wang, Q, Qi, J, Yuan, Y et al. Bat origins of MERS-CoV tracker: a global online database of antibody therapeutics for the supported by bat coronavirus HKU4 usage of human receptor prevention and treatment of COVID-19. Antibody Therapeutics CD26. Cell Host Microbe 2014; 16: 328–37. 2020; 3: 204–11. 22. Zhou, P, Yang, XL, Wang, XG et al. A pneumonia outbreak 47. An, Z, Zhang, N, Salazar, GTA et al. Antibody therapies for the associated with a new coronavirus of probable bat origin. Nature treatment of COVID-19. Antibody Therapeutics 2020; 3: 101–8. 2020; 579: 270–3. 48. ter Meulen, J, van den Brink, EN, Poon, LL et al. Human 23. Lam, TT, Jia, N, Zhang, YW et al. Identifying monoclonal antibody combination against SARS coronavirus: SARS-CoV-2-related coronaviruses in Malayan pangolins. Nature synergy and coverage of escape mutants. PLoS Med 2006; 3: 2020; 583: 282–5. e237. 24. Xiao, K, Zhai, J, Feng, Y et al. Isolation of SARS-CoV-2-related 49. Huo, J, Zhao, Y, Ren, J et al. Neutralization of SARS-CoV-2 by coronavirus from Malayan pangolins. Nature 2020; 583: destruction of the prefusion spike. Cell Host Microbe 2020; 28: 497. 286–9. 50. Tian, X, Li, C, Huang, A et al. Potent binding of 2019 novel 25. Andersen, KG, Rambaut, A, Lipkin, WI et al. The proximal origin coronavirus spike protein by a SARS coronavirus-specific human of SARS-CoV-2. Nat Med 2020; 26: 450–2. monoclonal antibody. Emerg Microbes Infect 2020; 9: 382–5. 26. Yan, R, Zhang, Y, Li, Y et al. Structural basis for the recognition 51. Huo, J, Le Bas, A, Ruza, RR et al. Neutralizing nanobodies bind of SARS-CoV-2 by full-length human ACE2. Science 2020; 367: SARS-CoV-2 spike RBD and block interaction with ACE2. Nat 1444–8. Struct Mol Biol 2020; 27: 846–54. 27. Ho, M. Perspectives on the development of neutralizing antibodies 52. Hanke, L, Vidakovics Perez, L, Sheward, DJ et al. An alpaca against SARS-CoV-2. Antibody Therapeutics 2020; 3: 109–14. nanobody neutralizes SARS-CoV-2 by blocking receptor 28. Perlman, S, Netland, J. Coronaviruses post-SARS: update on interaction. Nat Commun 2020; 11: 4420. replication and pathogenesis. Nat Rev Microbiol 2009; 7: 53. Wu, Y, Li, C, Xia, S et al. Identification of human single-domain 439–50. antibodies against SARS-CoV-2. Cell Host Microbe 2020; 27: 891, 29. Graham, RL, Donaldson, EF, Baric, RS. A decade after SARS: e895–8. strategies for controlling emerging coronaviruses. Nat Rev 54. Ho, M, Kreitman, RJ, Onda, M et al. In vitro antibody evolution Microbiol 2013; 11: 836–48. targeting germline hot spots to increase activity of an anti-CD22 30. Masters, PS. The molecular biology of coronaviruses. Adv Virus immunotoxin. J Biol Chem 2005; 280: 607–17. Res 2006; 66: 193–292. 55. Boder, ET, Midelfort, KS, Wittrup, KD. Directed evolution of 31. Lu, G, Wang, Q, Gao, GF. Bat-to-human: spike features antibody fragments with monovalent femtomolar antigen-binding determining ’host jump’ of coronaviruses SARS-CoV, MERS-CoV, affinity. Proc Natl Acad Sci U S A 2000; 97: 10701–5. and beyond. Trends Microbiol 2015; 23: 468–78. 56. Boder, ET, Wittrup, KD. Yeast surface display for screening 32. Li, F. Structure, function, and evolution of coronavirus spike combinatorial polypeptide libraries. Nat Biotechnol 1997; 15: proteins. Annu Rev Virol 2016; 3: 237–61. 553–7. 33. He, Y, Zhou, Y, Liu, S et al. Receptor-binding domain of 57. Ho, M, Nagata, S, Pastan, I. Isolation of anti-CD22 Fv with high SARS-CoV spike protein induces highly potent neutralizing affinity by Fv display on human cells. Proc Natl Acad Sci U S A antibodies: implication for developing subunit vaccine. Biochem 2006; 103: 9637–42. Biophys Res Commun 2004; 324: 773–81. 58. Wang, C, Li, W, Drabek, D et al. A human monoclonal antibody 34. Belouzard, S, Chu, VC, Whittaker, GR. Activation of the SARS blocking SARS-CoV-2 infection. Nat Commun 2020; 11: 2251. coronavirus spike protein via sequential proteolytic cleavage at two 59. Guthmiller, JJ, Dugan, HL, Neu, KE et al. An efficient method to distinct sites. Proc Natl Acad Sci U S A 2009; 106: 5871–6. generate monoclonal antibodies from human B cells. Methods Mol 35. Bosch, BJ, van der Zee, R, de Haan, CA et al. The coronavirus Biol 2019; 1904: 109–45. spike protein is a class I virus fusion protein: structural and 60. Pinto, D, Park, YJ, Beltramello, M et al. Cross-neutralization of functional characterization of the fusion core complex. J Virol SARS-CoV-2 by a human monoclonal SARS-CoV antibody. 2003; 77: 8801–11. Nature 2020; 583: 290–5. 36. Burkard, C, Verheije, MH, Wicht, O et al. Coronavirus cell entry 61. Jones, BE, Brown-Augsburger, PL, Corbett, KS et al. LY-CoV555, occurs through the endo−/lysosomal pathway in a a rapidly isolated potent neutralizing antibody, provides protection proteolysis-dependent manner. PLoS Pathog 2014; 10: e1004502. in a non-human primate model of SARS-CoV-2 infection. 2020; 37. Millet, JK, Whittaker, GR. Host cell entry of Middle East 2020.2009.2030.318972. respiratory syndrome coronavirus after two-step, furin-mediated 62. Mulangu, S, Dodd, LE, Davey, RT Jr et al. A randomized, activation of the spike protein. Proc Natl Acad Sci U S A 2014; 111: controlled trial of Ebola virus disease therapeutics. N Engl J Med 15214–9. 2019; 381: 2293–303. 256 Antibody Therapeutics, 2020 63. Misasi, J, Gilman, MSA, Kanekiyo, M et al. Structural and glypican-3 for liver cancer therapy. Hepatology 2014; molecular basis for Ebola virus neutralization by protective human 60: 576–87. antibodies. 2016; 351: 1343–6. 86. Kim, H, Ho, M. Isolation of antibodies to heparan sulfate on 64. Hansen, J, Baum, A, Pascal, KE et al. Studies in humanized mice glypicans by phage display. Curr Protoc Protein Sci 2018; 94: and convalescent humans yield a SARS-CoV-2 antibody cocktail. e66. Science 2020; 369: 1010–4. 87. Gao, W, Xu, Y, Liu, J et al. Epitope mapping by a Wnt-blocking 65. Baum, A, Fulton, BO, Wloga, E et al. Antibody cocktail to antibody: evidence of the Wnt binding domain in heparan sulfate. SARS-CoV-2 spike protein prevents rapid mutational escape seen Sci Rep 2016; 6: 26245. with individual antibodies. Science 2020; 369: 1014–8. 88. Zhang, Q, Chen, CZ, Swaroop, M et al. Heparan sulfate assists 66. Tillett, RL, Sevinsky, JR, Hartley, PD et al. Genomic evidence for SARS-CoV-2 in cell entry and can be targeted by approved drugs in reinfection with SARS-CoV-2: a case study. Lancet Infect Dis 2020. vitro. 2020; 2020.2007.2014.202549. 67. Zost, SJ, Gilchuk, P, Case, JB et al. Potently neutralizing and 89. Scully, M, Cataland, SR, Peyvandi, F et al. Caplacizumab protective human antibodies against SARS-CoV-2. Nature 2020; treatment for acquired thrombotic thrombocytopenic purpura. N 584: 443–9. Engl J Med 2019; 380: 335–46. 68. Ju, B, Zhang, Q, Ge, J et al. Human neutralizing antibodies elicited 90. Stanfield, RL, Dooley, H, Flajnik, MF et al. Crystal structure of a by SARS-CoV-2 infection. Nature 2020; 584: 115–9. shark single-domain antibody V region in complex with lysozyme. 69. Shi, R, Shan, C, Duan, X et al. A human neutralizing antibody Science 2004; 305: 1770–3. targets the receptor binding site of SARS-CoV-2. Nature 2020; 584: 91. Ho, M. Inaugural editorial: searching for magic bullets. Antibody 120–4. Therapeutics 2018; 1: 1–5. 70. Robbiani, DF, Gaebler, C, Muecksch, F et al. Convergent antibody 92. Hamers-Casterman, C, Atarhouch, T, Muyldermans, S et al. responses to SARS-CoV-2 in convalescent individuals. Nature 2020; Naturally occurring antibodies devoid of light chains. Nature 1993; 584: 437–42. 363: 446–8. 71. Rogers, TF, Zhao, F, Huang, D et al. Isolation of potent 93. Nguyen, VK, Hamers, R, Wyns, L et al. Camel heavy-chain SARS-CoV-2 neutralizing antibodies and protection from disease antibodies: diverse germline VHH and specific mechanisms enlarge in a small animal model. Science 2020; 369: 956–63. the antigen-binding repertoire. EMBO J 2000; 19: 921–30. 72. Brouwer, PJM, Caniels, TG, van der Straten, K et al. Potent 94. De Genst, E, Silence, K, Decanniere, K et al. Molecular basis for neutralizing antibodies from COVID-19 patients define multiple the preferential cleft recognition by dromedary heavy-chain targets of vulnerability. Science 2020; 369: 643–50. antibodies. Proc Natl Acad Sci U S A 2006; 103: 4586–91. 73. Chi, X, Yan, R, Zhang, J et al. A neutralizing human antibody 95. Feng, M, Bian, H, Wu, X et al. Construction and next-generation binds to the N-terminal domain of the spike protein of sequencing analysis of a large phage-displayed VNAR SARS-CoV-2. Science 2020; 369: 650–5. single-domain antibody library from six naive nurse sharks. 74. Liu, L, Wang, P, Nair, MS et al. Potent neutralizing antibodies Antibody Therapeutics 2019; 2: 1–11. against multiple epitopes on SARS-CoV-2 spike. Nature 2020; 584: 96. English, H, Hong, J, Ho, M. Ancient species offers contemporary 450–6. therapeutics: an update on shark VNAR single domain antibody 75. Ma, M, Badeti, S, Geng, K et al. Efficacy of targeting SARS-CoV-2 sequences, phage libraries and potential clinical applications. by CAR-NK cells. bioRxiv 2020. Antibody Therapeutics 2020; 3: 1–9. 76. Channappanavar, R, Perlman, S. Pathogenic human coronavirus 97. Flajnik, MF. A cold-blooded view of adaptive immunity. Nat Rev infections: causes and consequences of cytokine storm and Immunol 2018; 18: 438–53. immunopathology. Semin Immunopathol 2017; 39: 529–39. 98. Xiang, Y, Nambulli, S, Xiao, Z et al. Versatile and multivalent 77. Conti, P, Ronconi, G, Caraffa, A et al. Induction of nanobodies efficiently neutralize SARS-CoV-2. eabe4747 2020. pro-inflammatory cytokines (IL-1 and IL-6) and lung inflammation 99. Van Heeke, G, Allosery, K, De Brabandere, V et al. Nanobodies by Coronavirus-19 (COVI-19 or SARS-CoV-2): anti-inflammatory as inhaled biotherapeutics for lung diseases. Pharmacol Ther 2017; strategies. J Biol Regul Homeost Agents 2020; 34: 327–31. 169: 47–56. 78. Xu, Z, Shi, L, Wang, Y et al. Pathological findings of COVID-19 100. Cao, Y, Su, B, Guo, X et al. Potent neutralizing antibodies against associated with acute respiratory distress syndrome. Lancet Respir SARS-CoV-2 identified by high-throughput single-cell sequencing Med 2020; 8: 420–2. of convalescent patients’ B cells. Cell 2020; 182: 73, 79. Michot, JM, Albiges, L, Chaput, N et al. Tocilizumab, an anti-IL6 e16–84. receptor antibody, to treat Covid-19-related respiratory failure: a 101. Zohar, T, Alter, G. Dissecting antibody-mediated protection case report. Ann Oncol 2020; 31: 961–4. against SARS-CoV-2. Nat Rev Immunol 2020; 20: 392–4. 80. Stone, JH, Frigault, MJ, Serling-Boyd, NJ et al. Efficacy of 102. Pinto, D, Park, Y-J, Beltramello, M et al. Structural and functional Tocilizumab in patients hospitalized with Covid-19. N Engl J Med analysis of a potent sarbecovirus neutralizing antibody. 2020; 2020. doi: 10.1056/NEJMoa2028836. 2020.2004.2007.023903. 81. Geoghegan, EM, Pastrana, DV, Schowalter, RM et al. Infectious 103. Hansen, J, Baum, A, Pascal, KE et al. Studies in humanized mice entry and neutralization of pathogenic JC polyomaviruses. Cell Rep and convalescent humans yield a SARS-CoV-2 antibody cocktail. 2017; 21: 1169–79. 2020; eabd0827. 82. Kines, RC, Cerio, RJ, Roberts, JN et al. Human papillomavirus 104. Long, Q-X, Liu, B-Z, Deng, H-J et al. Antibody responses to capsids preferentially bind and infect tumor cells. 2016; 138: 901–11. SARS-CoV-2 in patients with COVID-19. Nat Med 2020; 26: 83. Zhang, F, Sodroski, C, Cha, H et al. Infection of hepatocytes with 845–8. HCV increases cell surface levels of heparan sulfate proteoglycans, 105. Berkenbrock, JA, Grecco-Machado, R, Achenbach, S. Microfluidic uptake of cholesterol and lipoprotein, and virus entry by devices for the detection of viruses: aspects of emergency up-regulating SMAD6 and SMAD7. Gastroenterology 2017; 152: fabrication during the COVID-19 pandemic and other outbreaks. 257, e257–70. 2020; 476: 20200398. 84. Lang, J, Yang, N, Deng, J et al. Inhibition of SARS pseudovirus 106. Wang, S, Ai, Z, Zhang, Z et al. Simultaneous and automated cell entry by lactoferrin binding to heparan sulfate proteoglycans. detection of influenza a virus hemagglutinin H7 and H9 based on PLoS One 2011; 6: e23710. magnetism and size mediated microfluidic chip. Sensors Actuators 85. Gao, W, Kim, H, Feng, M et al. Inactivation of Wnt signaling by a B Chem 2020; 308: 127675. human antibody that recognizes the heparan sulfate chains of
Journal
Antibody Therapeutics – Oxford University Press
Published: Nov 24, 2020
Keywords: SARS-CoV-2; spike or S protein; human antibody; cocktail therapy; single domain antibody or nanobody