key: cord-0983406-81cymdbn
authors: Gavor, Edem; Choong, Yeu Khai; Er, Shi Yin; Sivaraman, Hariharan; Sivaraman, J.
title: Structural basis of SARS-CoV-2 and SARS-CoV–antibody interactions
date: 2020-09-17
journal: Trends Immunol
DOI: 10.1016/j.it.2020.09.004
sha: 940b0678db4f06e28d74f931c28a6daae4f9e5e5
doc_id: 983406
cord_uid: 81cymdbn

The 2019 coronavirus pandemic is still a major public health concern. Neutralizing antibodies (nAbs) represent a cutting-edge antiviral strategy. Here, we focus on SARS-CoV-2 and SARS-CoV and discuss the current antibody research progress against rampant SARS-CoV-2 infections. We provide a perspective on the mechanisms of SARS-CoV-2-derived nAbs, comparing these with existing SARS-CoV-derived antibodies. We offer insight into how these antibodies cross-react and cross-neutralize by analyzing available S-glycoprotein-antibody complex structures. We also propose ways of adopting antibody-based strategies -- such as cocktail antibody therapeutics against SARS-CoV-2-- to overcome the possible resistance of currently identified mutations, and to mitigate possible antibody-dependent enhancement pathologies. Overall, this review provides a platform for the progression of antibody and vaccine design against SARS-CoV-2 and possibly, future coronavirus pandemics.

In recent decades, three highly pathogenic betacoronaviruses emerged in humans: severe acute respiratory syndrome coronavirus (SARS-CoV) [1, 2] in 2002, Middle East respiratory syndrome coronavirus (MERS-CoV) [3] in 2012, and SARS-CoV-2 in 2019, the causative agent of the rampant coronavirus disease [4, 5] . Despite appreciable progress in coronavirus research, the overwhelming number of COVID-19 deaths has warranted urgent and novel intervention [6, 7] . In terms of a strategy against SARS-CoV-2, lessons are being drawn from previous approaches: small-molecule inhibitors, of which remdesivir has received the most accolade, entering Phase2/3 clinical trials (NCT04292899) I [8] [9] . Other recent findings have included the assessment of convalescent plasma [10, 11] , polyclonal and monoclonal antibodies (mAbs) [12, 13] , as well as putative vaccines [14] . Several vaccine candidates have been rolled out for clinical trials with promising preliminary results as of August 2020, including ChAdOx1-S (Phase3: NCT04400838) II Lunar-Cov19(Phases1/2-ARCT-021:NCT04480957) III , and adenovirus-based vaccines (Ad26.nCoV:NCT04436276 ) IV [15] and (Ad5-nCoV: NCT04341389) V [16, 17] . Indeed, the current therapeutic race against SARS-CoV-2 might need to be multifaceted. New paradigms, such as T-cell based immunotherapies [18] , might be hopeful. However, of these options, we posit that neutralizing antibodies (nAbs) present timely and safe opportunities for the early intervention against viral pandemics, as noted previously for the successful control of Ebola virus [19, 20] along with many efforts towards HIV-1 treatments [21] .

NAbs hold remarkable potential for therapeutic and prophylactic applications against coronaviruses, and various avenues for antibody treatment (Tables 1, 2, 3) are currently being explored, with a surge in research findings. Previous review articles [17, [22] [23] [24] have provided a good foundation for our understanding of antibody reactivity and neutralization regarding SARS-CoV-derived antibodies. Furthermore, in recent weeks, new SARS-CoV-2-derived[25-30] and human host-specific nAbs have been reported [31] [32] [33] . Early attempts to cross-neutralize SARS-CoV-2 with SARS-CoV-derived antibodies did not yield ideal results [34] [35] [36] . However, most of the newly reported SARS-CoV-2-derived antibodies demonstrate a potent neutralizing effect in vitro and/or protection in animal models such as hamsters [27, 29] , mice [37, 38] , and rhesus macaques [39] (Tables 1, 2) . SARS-CoV-2-specific nAbs such as B38 [27] , BD-368-2 [37] , CA1/CB6-LALA[40], P2C-1F11/P2B-2F6/P2A-1A3[31], 414-1[41], ADI-55689/56046 [42] , REGN-CoV-2 (Phases1/2/3:NCT04425629 VI , NCT04426695 VII and NCT04452318 VIII ) [25, 43] , and COVI-SHIELD [17] (Tables 1 and 2), have either entered clinical trials or are in preclinical stages. Apart from these, the study of camelid antibodies (nanobodies) is also becoming an attractive research area with early positive results against COVID-19[34, [44] [45] [46] .

Moreover, much effort has gone into exploring host-specific antibodies. NAbs that target the host system have a unique advantage of overcoming virus mutations and might be easily repurposed for related viral outbreaks [47] [48] [49] [50] . Furthermore, decoy strategies are being explored, such as utilizing the SARS-CoV-2 binding partner --the angiotensin-converting enzyme 2 (ACE-2) receptor --fused to human immunoglobulin (IgG), and preliminary in vitro and mouse studies are encouraging [32] .

Here, we discuss the properties and mechanisms of neutralization of several antibodies. We analyze the available virus S-glycoprotein-antibody complex structures and offer insights into cross-reactivity and cross-neutralization mechanisms of SARS-CoV-2-derived and SARS-CoV-derived antibodies. We also briefly touch on the possibility of antibody-dependent enhancement (ADE) in SARS-CoV-2 and provide ways of mitigating ADE if it becomes a challenge. In addition to ADE, another emerging concern for SARS-CoV-2 therapeutic design is the emergence of more virulent escape mutants [51, 52] . We focus on residues on the S-glycoprotein recognized by antibodies and speculate on whether emerging SARS-CoV-2

The S-glycoprotein of coronaviruses is the primary determinant of viral tropism and plays a vital role in cellular receptor-binding and membrane fusion [53] [54] [55] . The S-glycoprotein assembles into a trimeric form on the virion surface in a crown-like shape [56] . Upon cleavage by host proteases, the S-glycoprotein is subdivided into two functionally distinct subunits: the S1 subunit --responsible for receptor recognition -and the S2 subunit --facilitating host-membrane fusion [53] [54] [55] (Figures 1, 2, 3) . The S-glycoprotein of both SARS-CoV-2 and SARS-CoV primarily recognize human ACE2(hACE2) [57, 58] . Given that the Sglycoprotein represents the major surface protein that interacts with host cells, it is considered a potential therapeutic target against coronaviruses [59] . The S-glycoprotein is the immunodominant target for nAbs[60-62], comprising an N-terminal domain (NTD), a receptor-binding domain (RBD/S1 B ), and an S2 subunit (Figures 1a, 3a) . The SARS-CoV RBD (amino acid, a.a: 338-506) consists of an S1 B core domain (S1 B CD) (a.a.318-424), and a receptor-binding motif (RBM) (a.a.438-498), that directly engages the human receptor hACE2 [63] (Figures 1a, 1c) . Although here, the N-terminal region of the RBD is referred to as the S1 B core domain (S1 B CD) following the literature [63] , it is important to clarify that the S1 B CD is not an independent structural unit. Antibodies can be raised against the full-length (FL) S-glycoprotein or its subdomains. Vaccination of African green monkeys with an attenuated parainfluenza virus-encoded with the FL-S-glycoprotein of the SARS-CoV Urbani strain elicited nAbs that protected monkeys from identical SARS-CoV infection [64] . On the one hand, although the FL-S-glycoprotein is highly immunogenic and induces nAb responses, it can also induce harmful immune responses in ferrets [65, 66] . On the other hand, antibodies raised against epitopes of S1 (a.a.485-625) or S2 (a.a.1,029-1,192) can neutralize virus infection by SARS-CoV strains (e.g., Tor2 and Sin2774) in Vero E6 cells [67, 68] . The RBD is also a significant neutralization determinant in the inactivated SARS-CoV vaccine; it induces potent nAbs that block SARS-CoV entry [14] . This was verified by observing reduced neutralizing activity upon depletion of RBD-specific antibodies from patient or rabbit immune sera (SARS) relative to affinity-purified anti-RBD antibodies which elicited higher potency neutralizing activity against SARS-CoV pseudovirus in 293T/ACE2 cells [69] .

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Adding further complexity to antibody design is the high propensity of viruses to adapt and develop mutations that dodge therapeutics. Indeed, viral mutations facilitate virus adaptation, potentially increasing transmissibility, worsening disease symptoms, and sometimes creating resistant mutants to therapeutics [51] .

Two key mutations have been identified in the SARS-CoV-2 S-glycoprotein: Asp614Gly and Ser943Pro.

The Gly614 mutation seems to cause enhanced infectivity of pseudotype single-cycle vesicular stomatitis virus (VSV) displaying the SARS-CoV-2 S-glycoprotein, relative to the Asp614 reference strain in 293T/ACE2, 293T/ACE2-TMPRSS2, and Vero cells [51, 52] . These residues might be considered in antibody therapeutics development, as they might render mutant strains resistant to existing nAbs as shown in 293T/hACE2, and Vero cells [52] , and also perhaps cause ADE-related pathologies [3, 70] . Nevertheless, the impact of the Asp614Gly mutation is currently a topic of debate [71] . One argument is the fact that said mutation is not located in the RBD of the S-glycoprotein [71] . Besides, antibodies raised with either Asp614 or Gly614 harbor cross-neutralizing activity [71] . It is debatable whether there is enough scientific evidence to confirm that this variant will worsen COVID-19 [71] . However, new evidence suggests that the Asp614Gly strain does increase infectivity; and, other sporadic mutations such as Asn234Gln, Leu452Arg, Ala475Val, and Val483Ala (of which most are found in the RBD) do present marked resistance to some nAbs [52] . Specifically, Ala475Val reduced the sensitivity to mAbs 157, 247, CB6, P2C-1F11, B38, and CA1, whereas Phe490Leu reduced the sensitivity to mAbs X593, 261-262, H4, and P2B2F6 in 293T/hACE2, and Vero cells [52] . Moreover, Val483Ala became resistant to mAbs X593 and P2B-2F6, and Leu452Arg to mAbs X593 and P2B-2F6 [52] . Also, Tyr508His/Asp614Gly+Ala435Ser, Asn439Lys, Ala831Val, Asp614Gly+Ile472Val reduced the sensitivity to mAbs H014, H00S022, B38, and X593, respectively by more than 4 times in 293T/hACE2, and Vero cells [52] . These results demand immediate improvement of antibody and vaccine design strategies against COVID-19. Antibody combination/cocktail therapies may be an excellent place to start since the REGN-COV cocktail antibody, and other upcoming cocktails have been demonstrated to neutralize SARS-CoV-2 pseudovirus particle mutant escape variants in The fight against SARS-CoV-2 using antibody therapies has been extensive, with many antibody-virus subunit complex structures determined at high resolutions (Protein Database (PDB):

7BYR/7BZ5/7BWJ) [27, 31, 37, 72] . Therefore, here, we aim to dissect, synthesize, and link the knowledge gap of many independent research findings of antibody-mediated neutralization against SARS-CoV-2.

Antibodies against coronaviruses are predominantly designed to target the S-glycoprotein because of the Sglycoprotein's ability to induce specific immune responses that are crucial against viral infections[60-62].

The S glycoprotein's conformational state and the domain targeted by an antibody seems to determine antibody cross-reactivity and cross-neutralization between SARS-CoV-2 and SARS-CoV [38, 73] . Despite the similarity in the SARS-CoV-2-RBD and SARS-CoV-RBD structures, the domains have different electrostatic surface potential maps [35, 74] , perhaps accounting for the differential S-glycoprotein immunogenicity observed so far [31, 35, 36, 75] .

Most of the nAbs bind directly to the RBM [25, 27, 38, 43, 76] , which results in the direct blocking of virusreceptor interactions ( Figure. 2a) . Although nAbs that target the RBM of SARS-CoV-2 and SARS-CoV tend to have higher potency than non-RBM-targeting antibodies [77] [78] [79] , the RBM-targeting nAbs are often virus-specific and generally bind poorly or show limited cross-neutralization [80] . This might be due to the poorly conserved nature of the RBM of the two viruses' S-glycoprotein (Figure 1a, 1c) . S-glycoproteins sequence comparison of SARS-CoV-2 and SARS-CoV shows that S1 B CD has a higher sequence identity than the RBM (85% versus ~50%; Figure 1c ) [81] .

J o u r n a l P r e -p r o o f

Human anti-SARS-CoV-2 mAbs: 311mab-31B5 and 311mab-32D4 [82] (Table 1 ) specifically bind to the RBD, most likely at the hACE2-receptor-binding interface [82] . Both mAbs can neutralize SARS-CoV-2 pseudovirus[82] but have not been tested against SARS-CoV. Thus, structure information is required to elucidate the neutralizing mechanism of these two antibodies.

In addition, six SARS-CoV-2-specific mAbs-P2C-1F11, P2C-1A3, P2B-2F6, P2A-1A10, P2A-1B3, and P2C-1C10 (Table 1) BD-368-2 [37] , and CB6(LALA) [40] . Respectively, these antibodies have been reported to protect hamster (4dpi), mouse(5dpi), and rhesus macaque (4dpi) animals against SARS-CoV-2 infections[29, 37, 40] . Despite its potent neutralizing effect, mAb2-15 has not neutralized three mutants: Lys455Arg, Ala475Arg, and Gly502Arg, suggesting that mAb2-15 might function better if combined with other antibodies. The CB6(LALA) antibody is an Fc-domain engineered form of mAb CB6 in an attempt to minimize Fcmediated cytotoxicity [17, 40] . Nanobodies [44] lacking the Fc domain can neutralize viruses and mitigate ADE [85] , currently of concern for vaccine and antibody design against SARS-CoV-2 [17, 86] . This has been evidenced by the potent neutralizing activities of the typical nanobody (ALX-0171) against respiratory syncytial virus (RSV) in respiratory epithelial cells, currently being tested in phase 2 clinical trials [87] .

Although not yet observed with SARS-CoV-2 [86] , perhaps as a way of caution, researchers might consider routinely adopting strategies to mitigate possible ADE, perhaps by considering cocktail therapy where, for example, two nAbs are applied simultaneously or by ensuring adequate optimization of antibody concentrations to ensure a complete neutralization of the virus [25, 27, 39, 43] . Fcγ receptor (FcγR) engineering is another technique that might help mitigate possible ADE because in vitro modeling of ADE has attributed increased pathogenesis to FcγR-mediated viral entry [85] .

Altogether, the newly isolated SARS-CoV-2 S-glycoprotein-derived mAbs that directly target the RBM might offer hopes of protection against COVID-19 and might potentially perform best when used as a cocktail, although cross-reactivity and neutralization with SARS-CoV might be less effective by virtue of the poor RBM sequence identity. However, these possibilities still warrant further robust investigation.

Many of the highly potent SARS-CoV nAbs that target the hACE2 binding site on the RBD, such as S230.15, cross-react with and cross-neutralize other CoV strains, including human (Urbani, GZ02, CUHK-W1), raccoon dog (A031G), and palm civet (HC/SZ/61/03) strains in in vitro assays and/or in BALB/c mice (2dpi) [77] . Most of these antibodies recognize an epitope in the RBM (Leu443, Thr487), validated with J o u r n a l P r e -p r o o f Leu443Arg and Thr487Ser virus escape mutants [88] as supported by plaque reduction assays in Vero E6 cells (Table 2 ) [77] . The same is true for mAb S230 (Tyr408, Tyr442, Leu443, Tyr475) (PDB: 6NB7), also substantiated with the Leu443Arg escape mutant in Vero E6 cells [73] . Likewise, the human mAb m396 (Table 2) replaces Tyr442 in SARS-CoV, and Leu443 --a key residue for virus neutralization --is replaced by Phe456; these findings are also consistent for mAb S230.15 [77] . Accordingly, for mAb m396, extensive mapping of SARS-CoV-Fab fragment complexes [78, 79] Overall, the structural and functional characterization of SARS-CoV Abs suggests that relying on existing SARS-CoV antibodies might not provide ideal therapeutic results, and by contrast, targeting the SARS-CoV-2 RBD might represent a more promising therapeutic strategy.

J o u r n a l P r e -p r o o f Nabs in this group bind epitopes in the RBD region, distal from the RBM, and block the S-glycoprotein to the host receptor( Figure 3c) [47, 76, 91] . Thus, they induce neutralization via a mechanism that is dependent on inhibiting hACE2-RBD binding and a pre-to-post-fusion conformational change to the Sglycoprotein [89] . These properties provide broad reactivity and neutralization, making these nAbs promising candidates for achieving synergistic effects when combined with other nAbs [25, 42, 43, 75] . This unique phenomenon has been demonstrated by the SARS-VHH-72[34, 91, 92] and ADI-56046 [42] nAbs (Table 2) . The above analysis suggests that the S1 B CD antigenic surface exhibits extensive conservation among SARSlike coronaviruses, as revealed by sequence alignment (Figure 1c) . Thus, it is tempting to speculate that S1 B CD-targeting antibodies might provide broad and potent neutralizing activity against coronaviruses. We argue that these should be tested alongside RBM or NTD-targeting antibodies in cocktail combinations.

The SARS-CoV S-glycoprotein-derived human mAb 47D11, can neutralize SARS-CoV-2 authentic and pseudoviruses in Vero E6 and HEK293T cells [63] . Others have also revealed that mAb S309, another human mAb designed against SARS-CoV S-glycoprotein, can potently neutralize both SARS-CoV and SARS-CoV-2 authentic, and pseudoviruses in Vero E6 cells, by recognizing the RBD [12] . Both 47D11 and S309 bind to the S1 B CD of the two viruses with similar affinities but do not compete with hACE2 binding to the RBD [12, 63] . This property is also exhibited by the highly potent SARS-CoV-2 derived mAb EY6A There is currently no structural information for mAb 47D11. However, the cryo-EM reconstruction of SARS-CoV-2-S-S309-Fab [12] complex (PDB: 6WPS/6WPT/6WS6) and the crystal structure of SARS-CoV-2-S-EY6A-Fab complex[93] suggest one or multiple mechanisms of neutralization, including S-trimer cross-linking or spike prefusion destabilization [36, 75] . These neutralizing mechanism(s) might also apply to 47D11, although this remains to be tested [12, 63] (Table 2) . On a positive note, S309 Fc variants with increased half-life and effector functions have been fast-tracked for clinical trials against COVID-19 [12] .

Unlike 47D11 and S309, other antibodies have shown discrepancies in neutralization results. Specifically, the SARS-CoV-specific human mAb, CR3022 (Table 2 )--used as a control antibody in many studies [38, 42, 93, 95] --targets the RBD (PDB: 6W41) and neutralizes SARS-CoV potently [76, 96] but shows no neutralizing activity against SARS-CoV-2, even though it binds tightly to the RBD [36, 76] . However, others have reported that CR3022 neutralizes SARS-CoV-2 [96] by destabilizing and destroying the prefusion S-glycoprotein trimer [96] . The latter study suggested that the microneutralization assay mode of J o u r n a l P r e -p r o o f assessment might have contributed to the observed non-neutralizing effect of CR3022 on SARS-CoV-2 [76] , compared with a plaque reduction neutralization test which showed a positive neutralization of SARS-CoV-2 by CR3022 [96] . Hence, to avoid similar discrepancies researchers should consider using different virus neutralization methods to validate their results.

The crystal structure of SARS-CoV RBD with CR3022 (PDB: 6W41), superimposed with that of SARS-CoV-2 RBD with hACE2 (PDB: 6LZG) shows that CR3022 occupies the S1 B CD [36, 76] . The crystal structure shows that, of the 28 residues in the CR3022 epitope, 24 are conserved between SARS-CoV-2 and SARS-CoV[97]. The high sequence conservation of the S1 B CD might explain the cross-reactivity and perhaps neutralization potential of CR3022 and remains to be investigated [96, 97] . According to one [12] . We posit that this conservation of residues likely accounts for the observed cross-reactivity and neutralization, and certainly merits further testing. Of note, a cocktail of S309 with weaker nAbs (e.g., S315 or S304) has significantly enhanced the neutralization of SARS-CoV-2, lending further support to the notion that Ab cocktail therapy might be therapeutically effective against COVID-19.

Collectively, these groups of antibodies utilize unknown mechanisms to neutralize either SARS-CoV-2 or SARS-CoV without necessarily blocking virus-receptor engagement. However, these mechanisms might result in lower neutralization efficiency [30] . Consequently, these groups of antibodies might require cobinding for effective neutralization [63, 76] , and might be tested as part of antibody cocktails.

In addition to the most common antibody groups identified as the RBD-targeting antibodies, recent studies have demonstrated that NTD and S2-targeting antibodies are also elicited in COVID-19 patients and might be effective nAbs. Specifically, nAbs 4A8(PDB: 7C2L) [95] , COV57[28], 2-17, 5-24, and 4-8[29] target the NTD of SARS-CoV-2 S protein (Figure 1b) while mAbs 9A1 and 0304-3h3 [95] target the S2 region ( Figure   1a ) [63] . All these antibodies have neutralized the authentic SARS-CoV-2 virus except 9A1, which is nonneutralizing [95] . Of note, COV57 is a SARS-CoV-2-derived antibody, that almost for the first time, has recognized the MERS-S protein in ELISA assays --a special property observed so far[28].

Overall, non-RBD binding antibodies might also neutralize SARS-CoV-2 without interrupting virusreceptor engagement, although the neutralizing effect of these antibodies might be less effective than RBDbinding antibodies and therefore, might require co-binding antibodies in a cocktail to exert a potent therapeutic effect. Further future testing is thus eagerly awaited.

NAbs might be an alternative source of treatment against COVID-19. Unfortunately, the poor cross- and S309 with other antibodies in a cocktail is particularly attractive because these mAbs demonstrate a potent synergistic neutralizing effect with many antibodies tested [12, 41] . Moreover, mAb CR3022 might be combined with mAbs COV21, C105, or B38 in a cocktail since CR3022 seems to non-compete with these three antibodies in terms of binding to the SARS-CoV-2 S-glycoprotein and, therefore, might offer synergistic neutralizing effects [26, 27] . Similarly, the potent NTD-binding nAb 4A8 might also be considered in a cocktail with RBD-binding antibodies because the 4A8-binding to the NTD leaves the RBD region of the S-glycoprotein free for co-binding antibodies which might offer additive neutralizing effect. Of note, in addition to cocktail antibody therapies, a cocktail with other antiviral drugs such as remdesivir might be therapeutically explored against COVID-19. Moving forward, because ADE of the disease cannot be reliably predicted after vaccination or antibody treatment, careful analysis of safety will have to be conducted in humans for COVID -19 (outstanding questions box) .

Overall, the alarming number of COVID-19 deaths is disheartening and calls for immediate public health interventions. Nevertheless, the above discussion suggests that there is hope against COVID-19, considering the many vaccines, antibodies targeting the S-glycoprotein, and small-molecule compound candidates are currently being tested in clinical trials and preclinical stages. In view of this, we expect that vaccines or antibody therapeutics might become available sooner rather than later, although we need to be vigilant of emerging mutations in the S-glycoprotein that might thwart current therapeutics efforts in the future. Secondary structure (α for helix, and β for strand) of NTD and RBD were labeled above the sequence based on PDB 5X4S (SARS-CoV S NTD), PDB 5WRG (SARS-CoV S) and PDB 6VXX (SARS-CoV-2 S). For illustrative purposes, the structure-based sequence alignment was carried out using DALI, and subsequently Clustal W and ESPript. The interacting residues of S-glycoprotein to mAbs 4A8 and B38 were highlighted in green (interacting with NTD in panel b) and white (interacting with RBD in panel c) respectively. The residues of human SARS-CoV and human SARS-CoV-2 interacting with human ACE2 receptor were bolded red in panel c. Sequence alignment of RBD highlighted with orange and cyan are S1 B CD and RBM respectively. Percent identity matrix (PIM) of NTD, RBD (S1 B CD and RBM), S1BCD, and RBM were indicated in the table. Note: These calculations were done for illustration purposes of this review.

J o u r n a l P r e -p r o o f (a) Antibodies bind to the RBM of the Spike protein, compete hACE2 binding, and block hACE2-RBD interactions. Antibodies bind to (b) NTD or S2 and (c) RBD (excluding RBM), but do not compete with hACE2 binding. These antibodies exhibit neutralization activities against the virus with unknown or multiple neutralizing mechanisms. (d) Neutralizing antibodies binding to RBD, compete with hACE2 binding, restricting conformational change or causing steric hindrance, preventing the virus to interact with the receptor. (e) Single antibody or antibodies cocktails binding to multiple epitopes could also mediate a neutralizing mechanism by restricting the conformational change of the S protein. (f) Antibodies binding to the S protein, allow S-trimer cross-linking, and could lead to virus particle aggregation and neutralization. Abs, antibodies; CoVs, coronaviruses; RBM, receptor binding motif; hACE2, human angiotensin-converting enzyme 2; mAbs, monoclonal antibodies; pAbs, polyclonal antibodies; NTD, N-terminal domain; S2, subunit 2; RBD, receptor binding domain; S1 B CD, S1 B core domain.

J o u r n a l P r e -p r o o f J o u r n a l P r e -p r o o f [12] Abbreviations: SARS, SARS-CoV; SARS-2, SARS-CoV-2; SARS-S, SARS-CoV-Spike; SARS-2-S, SARS-CoV-2-Spike; mAb, monoclonal antibody; pAb, polyclonal antibody; scFv, single chain variable fragment; HCAb, heavy chain antibody; RBD, Receptor Binding Domain; S1 B CD, S1 B Core Domain; RBM, Receptor Binding Motif; hACE2, Human Angiotensin-converting enzyme 2; NA, Not Applicable.

J o u r n a l P r e -p r o o f  What is the half-life of current therapeutic antibodies? Will patients treated or individuals given antibodies as prophylactics require multiple administrations? How often should subsequent administrations be given to ensure adequate protection? What is the antibody dosage? It will take extensive retrospective studies to contribute to understanding these parameters.

J o u r n a l P r e -p r o o f  Will positive in vitro neutralization assays and animal models mirror human trials?

 Finally, as and when they become available, will vaccines be applicable for every individual? This is relevant as older and/or immunocompromised persons might respond poorly or present adverse reactions to vaccine immunization.

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Structural basis of neutralization by a human anti-severe acute respiratory syndrome spike protein antibody, 80R

Structure of severe acute respiratory syndrome coronavirus receptorbinding domain complexed with neutralizing antibody

Cross-Neutralization of Human and Palm Civet Severe Acute Respiratory Syndrome Coronaviruses by Antibodies Targeting the Receptor-Binding Domain of Spike Protein

Genomic characterisation and epidemiology of 2019 novel coronavirus: implications for virus origins and receptor binding

Human monoclonal antibodies block the binding of SARS-CoV-2 spike protein to angiotensin converting enzyme 2 receptor

A human monoclonal 1 antibody blocking SARS-CoV-2 infection

Analysis of a SARS-CoV-2-Infected Individual Reveals Development of Potent Neutralizing Antibodies with Limited Somatic Mutation

The role of IgG Fc receptors in antibody-dependent enhancement

A perspective on potential antibody-dependent enhancement of SARS-CoV-2

New therapies for acute RSV infections: where are we?

Structural Basis for Potent Cross-Neutralizing Human Monoclonal Antibody Protection against Lethal Human and Zoonotic Severe Acute Respiratory Syndrome Coronavirus Challenge

Cryo-EM structure of the 2019-nCoV spike in the prefusion conformation

Structure, Function, and Antigenicity of the SARS-CoV-2 Spike Glycoprotein

Molecular basis for the preferential cleft recognition by dromedary heavychain antibodies

Chimeric camel/human heavy-chain antibodies protect against MERS-CoV infection

Structural basis for the neutralization of SARS-CoV-2 by an antibody from a convalescent patient

Structure-based design of prefusion-stabilized SARS-CoV-2 spikes

A neutralizing human antibody binds to the N-terminal domain of the Spike protein of SARS-CoV-2. Science (80-. )

Neutralization of SARS-CoV-2 by Destruction of the Prefusion Abbreviations: SARS

SARS-2, SARS-CoV-2

CoV-2-Spike; mAb, monoclonal antibody; Ab: antibody; RBD, Receptor-Binding Domain

Receptor Binding Motif

J.S. acknowledges partial support from Ministry of Education Singapore grants R154-000-A72114 (Tier 1), R-154-000-B03-112 (Tier 2), and R-154-000-697-112 (Tier 3).