Genetic and structural basis for recognition of SARS-CoV-2 spike protein by a two-antibody cocktail

TLDR: In this article, the authors determined the structures of two human monoclonal antibodies, COV2-2196 and COV2130, which formed the basis of the investigational antibody cocktail AZD7442, in complex with the receptor binding domain (RBD) of SARS-CoV-2.

Abstract: The SARS-CoV-2 pandemic has led to an urgent need to understand the molecular basis for immune recognition of SARS-CoV-2 spike (S) glycoprotein antigenic sites. To define the genetic and structural basis for SARS-CoV-2 neutralization, we determined the structures of two human monoclonal antibodies COV2-2196 and COV2-21301, which form the basis of the investigational antibody cocktail AZD7442, in complex with the receptor binding domain (RBD) of SARS-CoV-2. COV2-2196 forms an 'aromatic cage' at the heavy/light chain interface using germline-encoded residues in complementarity determining regions (CDRs) 2 and 3 of the heavy chain and CDRs 1 and 3 of the light chain. These structural features explain why highly similar antibodies (public clonotypes) have been isolated from multiple individuals1-4. The structure of COV2-2130 reveals that an unusually long LCDR1 and HCDR3 make interactions with the opposite face of the RBD from that of COV2-2196. Using deep mutational scanning and neutralization escape selection experiments, we comprehensively mapped the critical residues of both antibodies and identified positions of concern for possible viral escape. Nonetheless, both COV2-2196 and COV2130 showed strong neutralizing activity against SARS-CoV-2 strain with recent variations of concern including E484K, N501Y, and D614G substitutions. These studies reveal germline-encoded antibody features enabling recognition of the RBD and demonstrate the activity of a cocktail like AZD7442 in preventing escape from emerging variant viruses.

Figures

Table 1. Sites of strong escape from each antibody for highlighting in logo plots were 846 determined heuristically as sites whose summed mutational escape scores were at least 10 times 847 the median sitewise sum of selection, and within 10-fold of the sitewise sum of the most strongly 848 selected site. Full documentation of the computational analysis is at 849 https://github.com/jbloomlab/SARS-CoV-2-RBD_MAP_AZ_Abs. These results are also 850

Full text

1
ARTICLE 1
Genetic and structural basis for recognition of SARS-CoV-2 spike protein by 2
a two-antibody cocktail 3
4
AUTHORS: Jinhui Dong
1,*
, Seth J. Zost
1,*
, Allison J. Greaney
2,3
, Tyler N. Starr
2
,
Adam S. 5
Dingens
2
, Elaine C. Chen
4
,
Rita E. Chen
5,6
, James Brett Case
6
, Rachel E. Sutton
1
, Pavlo Gilchuk
1
, 6
Jessica Rodriguez
1
, Erica Armstrong
1
, Christopher Gainza
1
, Rachel S. Nargi
1
, Elad Binshtein
1
, 7
Xuping Xie
7
, Xianwen Zhang
7
, Pei-Yong Shi
7
, James Logue
8
, Stuart Weston
8
, Marisa E. McGrath
8
, 8
Matthew B. Frieman
8
, Tyler Brady
9
, Kevin Tuffy
9
, Helen Bright
9
, Yueh-Ming Loo
9
, Patrick 9
McTamney
9
, Mark Esser
9
, Robert H. Carnahan
1,10
, Michael S. Diamond
5,6,11
,
12
, Jesse D. Bloom
2,3,13
, 10
James E. Crowe, Jr.
1,4,10*
11
12
Affiliations: 13
1
Vanderbilt Vaccine Center, Vanderbilt University Medical Center, Nashville, TN 37232, USA 14
2
Basic Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA 15
3
Department of Genome Sciences & Medical Scientist Training Program, University of 16
Washington, Seattle, WA 98195, USA 17
4
Department of Pathology, Microbiology, and Immunology, Vanderbilt University Medical 18
Center, Nashville, TN, 37232, USA 19
5
Department of Pathology and Immunology, Washington University School of Medicine, 20
Saint Louis, MO, 63110, USA 21
6
Department of Medicine, Washington University School of Medicine, Saint Louis, MO, 22
63110, USA 23
(which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission.
The copyright holder for this preprintthis version posted March 1, 2021. ; https://doi.org/10.1101/2021.01.27.428529doi: bioRxiv preprint

2
Department of Biochemistry & Molecular Biology, The University of Texas Medical Branch 24
at Galveston, Galveston, TX, 77555, USA 25
Department of Microbiology and Immunology, The University of Maryland, College Park, 26
MD, 20742, USA 27
9
Microbial Sciences, AstraZeneca, One MedImmune Way, Gaithersburg, MD 20878, USA 28
10
Department of Pediatrics, Vanderbilt University Medical Center, Nashville, TN, 37232, USA 29
11
Department of Molecular Microbiology, Washington University School of Medicine, Saint 30
Louis, MO, 63110, USA
31
12
Andrew M. and Jane M. Bursky Center for Human Immunology and Immunotherapy 32
Programs, Washington University School of Medicine, Saint Louis, MO, 63110, USA 33
13
Howard Hughes Medical Institute, Seattle, WA, 98109, USA 34
35
*These authors contributed equally. 36
**Correspondence to: James E. Crowe, Jr., M.D., james.crowe@vumc.org
37
38
Contact information: 39
James E. Crowe, Jr., M.D. [LEAD CONTACT] 40
Departments of Pediatrics, Pathology, Microbiology, and Immunology, and the Vanderbilt 41
Vaccine Center 42
Mail: 43
Vanderbilt Vaccine Center 44
11475 Medical Research Building IV 45
2213 Garland Avenue 46
(which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission.
The copyright holder for this preprintthis version posted March 1, 2021. ; https://doi.org/10.1101/2021.01.27.428529doi: bioRxiv preprint

3
Nashville, TN 37232-0417, USA 47
Telephone (615) 343-8064 48
Email james.crowe@vumc.org
49
50
51
Keywords: Coronavirus; SARS-CoV-2; SARS-CoV; COVID-19; Antibodies, Monoclonal; 52
Human; Adaptive Immunity. 53
(which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission.
The copyright holder for this preprintthis version posted March 1, 2021. ; https://doi.org/10.1101/2021.01.27.428529doi: bioRxiv preprint

4
The SARS-CoV-2 pandemic has led to an urgent need to understand the molecular basis 54
for immune recognition of SARS-CoV-2 spike (S) glycoprotein antigenic sites. To define the 55
genetic and structural basis for SARS-CoV-2 neutralization, we determined the structures 56
of two human monoclonal antibodies COV2-2196 and COV2-2130
1
, which form the basis of 57
the investigational antibody cocktail AZD7442, in complex with the receptor binding 58
domain (RBD) of SARS-CoV-2. COV2-2196 forms an “aromatic cage” at the heavy/light 59
chain interface using germline-encoded residues in complementarity determining regions 60
(CDRs) 2 and 3 of the heavy chain and CDRs 1 and 3 of the light chain. These structural 61
features explain why highly similar antibodies (public clonotypes) have been isolated from 62
multiple individuals
1-4
. The structure of COV2-2130 reveals that an unusually long LCDR1 63
and HCDR3 make interactions with the opposite face of the RBD from that of COV2-2196. 64
Using deep mutational scanning and neutralization escape selection experiments, we 65
comprehensively mapped the critical residues of both antibodies and identified positions of 66
concern for possible viral escape. Nonetheless, both COV2-2196 and COV2-2130 showed 67
strong neutralizing activity against SARS-CoV-2 strain with recent variations of concern 68
including E484K, N501Y, and D614G substitutions. These studies reveal germline-encoded 69
antibody features enabling recognition of the RBD and demonstrate the activity of a 70
cocktail like AZD7442 in preventing escape from emerging variant viruses. 71
72
The current coronavirus disease 2019 (COVID-19) pandemic is caused by SARS-CoV-2, a clade 73
B betacoronavirus (Sarbecovirus subgenus) with 96.2% or 79.6% genome sequence identity to 74
the bat coronavirus RaTG13 or SARS-CoV respectively
5,6
. The S glycoprotein mediates viral 75
attachment via binding to the host receptor angiotensin converting enzyme 2 (ACE2) and 76
(which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission.
The copyright holder for this preprintthis version posted March 1, 2021. ; https://doi.org/10.1101/2021.01.27.428529doi: bioRxiv preprint

5
possibly other host factors, and subsequent entry into cells after priming by the host 77
transmembrane protease serine 2 (TMPRSS2)
7-9
. The trimeric S protein consists of two subunits, 78
designated S1 and S2. The S1 subunit binds to ACE2 with its receptor binding domain (RBD), 79
while the central trimeric S2 subunits function as a fusion apparatus after S protein sheds the S1 80
subunits
10
. The human humoral immune response to SARS-CoV-2 has been well documented
11-
81
13
, and numerous groups have isolated monoclonal antibodies (mAbs) that react to SARS-CoV-2 82
S protein from the B cells of patients previously infected with the virus. A subset of the human 83
mAbs neutralize virus in vitro and protect against disease in animal models
1,2,13-21
. Studies of the 84
human B cell response to the virus have been focused mostly on S protein so far, due to its 85
critical functions in attachment and entry into host cells
1,2,13-21
. For these S-protein-targeting 86
antibodies, the RBD of S protein is the dominant target of human neutralizing antibody 87
responses
1,2,13-21
. This high frequency of molecular recognition may be related to the accessibility 88
of the RBD to B cell receptors, stemming from a low number of obscuring glycosylation sites 89
(only 2 sites on the RBD versus 8 or 9 sites on the N-terminal domain [NTD] or S2 subunit, 90
respectively)
13
. The RBD also occupies an apical position and exhibits exposure due to the 91
“open-closed” dynamics of the S trimer observed in S protein cryo-EM structures
22-24
. Potently 92
neutralizing mAbs predominantly target the RBD, since this region is directly involved in 93
receptor binding. 94
95
In previous studies, we isolated a large panel of SARS-CoV-2 S-protein-reactive human mAbs 96
from the B cells of patients previously infected with the virus. that bind to the SARS-CoV-2 S 97
protein
25
. A subset of these mAbs was shown to bind to recombinant RBD and S protein 98
ectodomain and exhibit neutralization activity against SARS-CoV-2 by blocking S-protein-99
(which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission.
The copyright holder for this preprintthis version posted March 1, 2021. ; https://doi.org/10.1101/2021.01.27.428529doi: bioRxiv preprint