key: cord-0857244-7vsim0zf
authors: Xu, Kun; Gao, Ping; Liu, Sheng; Lu, Shuaiyao; Lei, Wenwen; Zheng, Tianyi; Liu, Xueyuan; Xie, Yufeng; Zhao, Zhennan; Guo, Shuxin; Tang, Cong; Yang, Yun; Yu, Wenhai; Wang, Junbin; Zhou, Yanan; Huang, Qing; Liu, Chuanyu; An, Yaling; Zhang, Rong; Han, Yuxuan; Duan, Minrun; Wang, Shaofeng; Yang, Chenxi; Wu, Changwei; Liu, Xiaoya; She, Guangbiao; Liu, Yan; Zhao, Xin; Xu, Ke; Qi, Jianxun; Wu, Guizhen; Peng, Xiaozhong; Dai, Lianpan; Wang, Peiyi; Gao, George F.
title: Protective prototype-Beta and Delta-Omicron chimeric RBD-dimer vaccines against SARS-CoV-2
date: 2022-04-27
journal: Cell
DOI: 10.1016/j.cell.2022.04.029
sha: c8b6b471a450d2b2212c6874094d0196e863b808
doc_id: 857244
cord_uid: 7vsim0zf

Breakthrough infections by SARS-CoV-2 variants become the global challenge for pandemic control. Previously, we developed the protein subunit vaccine ZF2001 based on dimeric receptor-binding domain (RBD) of prototype SARS-CoV-2. Here, we developed a chimeric RBD-dimer vaccine approach to adapt SARS-CoV-2 variants. A prototype-Beta chimeric RBD-dimer was first designed to adapt the resistant Beta variant. Compared with its homotypic forms, the chimeric vaccine elicited broader sera neutralization of variants and conferred better protection in mice. The protection of the chimeric vaccine was further verified in macaques. This approach was generalized to develop Delta-Omicron chimeric RBD-dimer to adapt the currently prevalent variants. Again, the chimeric vaccine elicited broader sera neutralization of SARS-CoV-2 variants, and conferred better protection against challenge by either Delta or Omicron SARS-CoV-2 in mice. The chimeric approach is applicable for rapid updating of immunogens, and our data supported the use of variant-adapted multivalent vaccine against circulating and emerging variants.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants are 70 continually emerging and become the circulating strains in the world (www.who.int). 71

Several highly transmissible variants of concern (VOCs) showed altered pathogenicity 72 and transmission showed that prototype-Beta chimeric RBD-dimer vaccine induced the broadest and 217 most balanced neutralizing activity against SARS-CoV-2 variants, compared with 218 homotypic RBD-dimer vaccines ( Figure 2B) . 219 To further explore the protective efficacy of these RBD-dimers, the first batch of eight 220 mice in each group were evaluated for protection against SARS-CoV-2 infection. 221 BALB/c mouse is not sensitive to prototypic SARS-CoV-2 infection because of the low 222 binding affinity between mouse ACE2 and S protein, but becomes sensitive to Beta 223 variant due to the promoted affinity by N501Y mutation at S protein (Niu et al., 2021) . 224 Therefore, four mice in each group were transduced via intranasal (i.n.) route with 225 adenovirus (Ad5) expressing hACE2 and, five days later, challenged intranasally with 226 prototypic SARS-CoV-2 (hCoV-19/China/CAS-B001/2020 strain). The other four mice 227 in each group were challenged directly with Beta SARS-CoV-2 variant (GDPCC- challenged with prototype SARS-CoV-2, high levels of both viral gRNA (average:1.72 232 × 10 9 copies/g) and sgRNA (average: 1.9 × 10 8 copies/g) were detected in sham-233 immunized mice ( Figure 2C and 2D). By contrast, significantly reduced viral loads 234 (P<0.0001) were detected in vaccine-immunized mice. Averages of pulmonary viral 235 gRNA were 4.61 × 10 5 , 2.58 × 10 6 and 2.66 × 10 5 copies/g in prototype, Beta, and 236 prototype-Beta chimeric vaccine groups, respectively, with 2-4-log10 reduction 237 compared with the sham group ( Figure 2C ). In line with the trends in neutralization, 238 both prototype and prototype-Beta chimeric vaccine groups showed significantly lower 239 viral gRNA compared with the Beta vaccine group (P=0.016 and 0.0045, respectively) 240 ( Figure 2C ). All vaccine groups showed undetectable pulmonary viral sgRNA, 241

indicating the complete control of viral replication ( Figure 2D ). Analysis of immune 242 correlates of protection following vaccination showed that NAb titers correlated 243 strongly with the reduction of pulmonary prototypic SARS-CoV-2 gRNA based on a 244 linear model (r = -0.8967, P<0.0001) ( Figure 2E ). 245

For mice challenged with SARS-CoV-2 Beta variant, high levels of pulmonary viral 246 gRNA (average: 4.01 × 10 8 copies/g) and sgRNA (average: 3.03 × 10 7 copies/g) were 247 detected in sham-vaccinated mice ( Figure 2F and 2G). Less than 2-log10 viral 248 reductions were observed in prototype vaccine group (average gRNA: 6.34 × 10 6 249 copies/g; average sgRNA: 4.51 × 10 5 copies/g) ( Figure 2F and 2G). In contrast, more 250 significant reduction (2-4-log10) of pulmonary viral gRNA was detected in mice 251 vaccinated with Beta (average: 5.46 × 10 5 copies/g; P=0.0079 compared to prototype 252 vaccine group) or prototype-Beta chimeric vaccine (average: 9.81 × 10 4 copies/g; To provide a further assessment of protection following vaccination, mice in each 259 group were assessed for virus-related pathology in lung at 5 DPI. Tissue sections were 260 stained with hematoxylin and eosin (H&E) to examine histopathology. Sham recipients 261 challenged with either prototype virus or Beta variant showed moderate-to-severe 262 histopathological changes in lung, including vanishment of alveolar cavities, pulmonary 263 vascular congestion, and diffuse inflammatory cell infiltration ( Figure S6A ). In contrast, 264 mice vaccinated with prototype, Beta or prototype-Beta chimeric RBD-dimer vaccine 265 exhibited relieved lung injury ( Figure S6A ). Moreover, histopathology showed that both Beta and prototype-Beta chimeric vaccines provided better protection compared with 267 prototype vaccine when mice were challenged with SARS-CoV-2 Beta variant (Figure 268 Twenty-four healthy young rhesus macaques (Table S1) were immunized 278 intramuscularly with three jabs of 25 μg vaccine (n=12), a dose used in human (Yang 279 et al., 2021), or sham (n=12), 21 days apart. Serum samples were collected before 280 (day 0) and after (day 14, 35, and 52) priming ( Figure S5B ). The average endpoint 281 titers of RBD-binding antibody raised from 512 after one dose to 46341 after two doses, 282 and further up to 73562 after three doses in the vaccine group ( Figure 3A ). The sera 283 after three doses were tested for neutralization of pseudotyped viruses expressing four 284 VOCs ( Figure 3B and S5C). Sera from vaccinated animals showed robust and 285 balanced neutralizing activities against prototype, Alpha, Beta and Delta 286 pseudoviruses (GMT ranging between 2607-3200). All vaccine-elicited sera 287 neutralized Omicron pseudovirus (GMT 496), with a 6.5-fold reduction of titer as 288 compared with prototype neutralization ( Figure 3B ). All vaccine-elicited sera 289 neutralized authentic prototype SARS-CoV-2 (GMT 837), variant Alpha (GMT 583), 290

Beta (GMT 393) and Delta (GMT 295) ( Figure 3C ). As the authentic Omicron variant 291 (BA.1 subvariant, NPRC 2.192100005 strain) was available in China CDC currently, 292 macaque sera were tested for neutralization of Omicron and showed 1.6-folds 293 reduction of activity against Omicron than against prototype SARS-CoV-2 ( Figure 3D) . 294

In addition, peripheral blood mononuclear cells (PBMCs) collected at day 6 after the 295 third dose were tested for cytokine production using enzyme-linked immunospot 296 (ELISPOT) assay. In consistent with trends of prototype RBD-dimer vaccine ZF2001 297 in rhesus macaque and human Yang et al., 2021) , prototype-Beta 298 chimeric vaccine elicited moderate but balanced TH1 (IFN-γ and IL-2) and TH2 (IL-4) 299 cytokine production ( Figure 3E ). 300

Protective efficacy of prototype-Beta chimeric RBD-dimer vaccine in rhesus 301 macaques 302

The 12 rhesus macaques receiving prototype-Beta chimeric vaccine were 303 challenged (n=4) with prototype SARS-CoV-2 (GDPCC-nCoV27 strain), Beta variant 304 (GDPCC-nCoV84 strain), and Delta (CCPM-B-V-049-2105-8), respectively, via both 305 the upper and lower respiratory tracts. The other 12 sham-immunized animals were 306 also challenged in parallel as controls (n=4) ( Figure S5B ). Body weight and 307 temperature were monitored daily until euthanasia at 7 DPI ( Figure S5D ). Swabs were 308 collected at 0, 1, 3, 5, and 7 DPI from nose, throat, and anus for viral gRNA 309 quantification. At 7 DPI, lung tissues were collected from 7 lopes for histopathology 310 examination. Lung tissues from each lope (a sample mixture of six sites) were 311 quantified for viral gRNA. we developed the Delta-Omicron vaccine with our chimeric RBD-dimer approach 338 ( Figure 1A) . 339

The Delta-Omicron chimeric RBD-dimer protein was expressed and purified with 340 high yield and purity, indicating the capacity for large-scale production ( Figure S1E ). 341

Next, the exposure of RBM and major antigenic sites of RBD-dimer vaccine were 342 verified through SPR assay using hACE2 protein and major classes of RBD-specific 343 mAbs as probes ( Figure 1C and 1D). Monomeric RBD proteins from prototype, Delta, 344 and Omicron variants were used for comparison. Delta-Omicron chimeric RBD-dimer 345 showed similar binding affinity (8.44 nM) to hACE2 in comparison to the affinities of 346 monomeric prototype (6.53 nM), Delta (5.09 nM), and Omicron RBD (6.59 nM) ( Figure  347 1D and S2B). For the mAb binding, the Delta RBD did not bind to mAb C110 and 348 exhibited decreased binding affinity to mAb CV07-270; the Omicron RBD did not bind 349 to mAb CB6 and displayed reduced binding affinities to mAbs CV07-270 and C110 350 ( Figure 1D and S2B). In contrast, Delta-Omicron chimeric RBD-dimer bound all the 351 tested representative mAbs, but like a combination of Delta and Omicron RBD with the 352 decreased affinities to mAbs CV07-270 and C110, respectively ( Figure 1D and S2B). 353

The cryo-EM structure of Delta-Omicron chimeric RBD-dimer in complexed with CB6 354

Fab was determined at resolution of 12.8 Å. It reassembles the structure of prototype-355

Beta chimeric RBD-dimer in complexed with CB6 Fab as the "bilateral-lung"-like 356 structure engaging one CB6 Fab at the Delta arm ( Figure 1G , S4A-S4C and S4F). In 357 summary, the antigenic characterization and structural analysis indicated that the 358 Delta-Omicron chimeric RBD-dimer protein was correctly folded, presenting the RBM 359 and major antigenic epitopes. For mice challenged with Delta variant, the averages of pulmonary viral gRNA were 382 1.09 × 10 10 in sham group, but reduced to 1.43 × 10 8 copies/g in prototype vaccine 383 group, and additionally reduced (474-folds) to 2.37 × 10 7 copies/g in Delta-Omicron 384 chimeric vaccine group ( Figure 5C ). In line with this, the pulmonary viral sgRNA were 385 detected in all mice in the sham group with high levels (average: 1.70 × 10 8 copies/g), 386 but only detectable in three mice receiving prototype vaccine with the average titer of 387 1.07 × 10 6 copies/g, and undetectable in the all mice receiving Delta-Omicron vaccine, 388 suggesting the complete control of Delta viral replication ( Figure 5D ). 389

For mice challenged with Omicron variant, the sham group of mice developed high 390 levels of pulmonary viral gRNA (average: 1.04 × 10 9 copies/g) and sgRNA (average: 391 1.73 × 10 7 copies/g) ( Figure 5G and 5H). Mice immunized prototype vaccine showed 392 decreased pulmonary viral gRNA (average: 3.68 × 10 7 copies/g) and sg RNA (2.41 × 393 10 4 copies/g) ( Figure 5G and 5H). Encouragingly, the protection efficacy of Delta-394

Omicron vaccine was better than prototype vaccine, with no pulmonary virus 395 replication detected in all mice ( Figure 5H Twenty-four rhesus macaques (see Table S1 ) were immunized with 3 doses of 596 prototype-Beta chimeric RBD-dimer vaccine or sham (n=12), 21 days apart. Serum 597 samples were collected according to the study schedule shown in Figure S5B . 

Further information and requests for resources and reagents should be directed to 756 and will be fulfilled by the Lead Contact, George F. Gao (gaof@im.ac.cn). 757

All requests for unique/stable reagents generated in this study should be directed to 759 and will be fulfilled by the Lead Contact author with a completed Materials Transfer 760

The cryo-EM maps have been deposited in the Electron Microscopy Data Bank with 763 accession codes EMDB: EMD-33225 (SARS-CoV-2 prototype RBD-dimer bound to 764 CB6 Fab), EMDB: EMD-33234 (SARS-CoV-2 prototype-Beta chimeric RBD-dimer 765 bound to CB6 Fab) and EMDB: EMD-33235 (SARS-CoV-2 Delta-Omicron chimeric 766 RBD-dimer bound to CB6 Fab). All the other data supporting the finding of this study 767 are available within the paper and are available from the corresponding author upon 768 request. This study did not generate unique code. residues 319-537) connected as tandem repeat. Prototype-Beta chimeric SARS-CoV-835 2 RBD-dimer was one prototype RBD (S protein residues 319-537) and one Beta RBD 836 (S protein residues 319-537) connected as tandem repeat. Delta-Omicron chimeric 837 SARS-CoV-2 RBD-dimer was one Delta RBD (S protein residues 319-537) and one 838

Omicron BA.1 RBD (S protein residues 316-534) connected as tandem repeat. 839

For each construct, signal peptide sequence of MERS-CoV S protein (S protein 840 residues 1-17) was added to the protein N terminus for protein secretion, and a hexa-841

His tag was added to the C terminus to facilitate further purification processes. These Beta chimeric RBD-dimer was one prototype RBD (S protein residues R319-K537) and 855 one Beta RBD (S protein residues V320-K537) connected as tandem repeat. The R319 856 was deleted in the RBD-dimer protein to avoid the potential protease cleavage. Signal 857 peptide sequence was added to the N terminus for protein secretion. No tag sequence 858 was added at C terminus. The monomeric Omicron (BA.1) variant RBD (S protein 316-534, GISAID No. 875 EPI_ISL_6795848) was purchased from ACROBiosystems, which was expressed from 876

The hACE2 protein (residues 1-740, Genbank: NP_068576) was purchased from 878 Sino Biological Inc., China, which was expressed from HEK293T cells. 879

SPR binding experiments were carried out using a BIAcore 8000 device (GE 881 Healthcare) at 25 °C. The buffers for all proteins used for kinetic analyses were 882 exchanged to PBST (10 mM Na2HPO4; 2 mM KH2PO4, pH 7.4; 137 mM NaCl; 2.7 mM 883 KCl; 0.005% Tween 20). Purified RBD-dimer and -monomer proteins were immobilized 884 on a CM5 chip with the standard EDC/NHS coupling method at about 1,000 response 885 units (RU). Serial dilutions of Fabs were prepared and used to flow over the chip 886 surface. Data were collected over time. After each cycle, the sensor surface was 887 regenerated via a short treatment using 10 mM NaOH. The apparent equilibrium 888 dissociation constants (apparent binding affinity, KD) for each antibody were calculated 889 using BIAcore 8000 analysis software (BIAevaluation v3.0). Each set of equilibrium 890 binding responses was fitted to the single binding site model.

For samples of prototype RBD-dimer bound to CB6 Fab and prototype-Beta chimeric 893 RBD-dimer bound to CB6 Fab, an aliquot of 3.5 μl solution (0.4 mg/ml) was applied to 894 glow-discharged Quantifiol R 1.2/1.3 holey carbon grids and blotted for 1.5 s with a 895 humidity of 95% before being plunged into liquid ethane using a Vitrobot Mark IV 896 (Thermo Fisher). The frozen grides were loaded onto a Titan Krios cryo-transmission 897 electron microscope (Thermo Fisher) that is equipped with a BioQuantum energy filter 898 (Gatan), operated at 300 kV for data collection. Automatic data collection was 899 performed using Serial-EM software. Movies were recorded with a Gatan K2 direct For the prototype-Beta chimeric RBD-dimer/CB6 Fab dataset, 302,365 particles 912 were picked from 2036 micrographs. Then the picked particles were extracted and 913 subjected to two rounds of reference-free 2D classification in Relion. A clean dataset 914 with 162,269 particles from good 2D classes were selected and the initial model was 915 generated by cryoSPARC ab initio. Then the model was used as reference in Relion 916 3D classification. After the third round of 3D classification without applying symmetry, 917 the predominant class containing a subset of 31,507 good particles. These particles 918 were subjected to 3D refinement, which yielded a reconstruction at 11.6 Å resolution

The prototype RBD-dimer/CB6 Fab dataset was processed similarly. Briefly, a total 921 of 289,269 automatically picked particles were extracted in Relion for the following 2D 922 and 3D classification. Three rounds of reference-free 2D classification were performed 923 to remove the heterogeneous particles. A clean dataset with 214,464 particles from 924 good 2D classes was selected and subjected to three rounds of 3D classification 925 without applying symmetry. A single dominant class (26,842 particles) was identified 926 and used to calculate the density map at 11.5 Å resolution by applying a 3D refinement 927 with C2 symmetry. 928

The Delta-Omicron chimeric RBD-dimer/CB6 Fab dataset was processed similarly 929 to those described above. The specific images processing and reconstruction were 930 shown in Figure S4A -S4C. 931

Due to the fierce flexibility between two RBDs, we could only obtain low resolution 932 maps as described above. However, we fitted the crystal structure of RBD/CB6 933 complex (PDB ID:7C01) or its RBD part into the two density maps using CHIMERA 934 (Pettersen et al., 2004) , which showed a high degree of matching. 935

The pseudotyped viruses displaying SARS-CoV-2 spikes express GFP in infected 937 cells. They were prepared as previously described (Zhao et al., 2021) . Mice sera were 938 2-fold serially diluted and incubated with pseudotyped virus at 37°C for 1 h. Then the 939 mixture was transferred to pre-plated Vero cell monolayers in 96-well plates. After 940 incubation for 15 h, the transducing unit numbers were calculated on a CQ1 confocal 941 image cytometer (Yokogawa). Fifty percent pseudovirus neutralization titer (pVNT50) 942 was determined by fitting nonlinear regression curves using GraphPad Prism and 943 calculating the reciprocal of the serum dilution required for 50% neutralization of 944 infection. pVNT50 below the limit of detection was determined as half the limit of 945 detection. 946

Neutralizing antibody activities induced by vaccines in rhesus macaques against 948 prototype, Alpha, Beta and Delta SARS-CoV-2 were titrated on basis of inhibition of cytopathogenic effect (CPE) in IMBCAMS. Briefly, equal volume of serially diluted 950 serum and 100 TCID50 SARS-CoV-2 was mixed and incubated for one hour at 37°C. 951

The mixture of serum and virus was added to Vero E6 cells, followed by incubation at 952 37°C for 3 days. CPE was recorded for determination of antibody neutralizing titer. CoV-2 was mixed and incubated for one hour at 37°C. The mixture of serum and virus 961 was added to Vero cells, followed by incubation at 37°C for 4 days. CPE was recorded 962 for determination of antibody neutralizing titer. Prototype SARS-CoV-2 (IVDC-QD-11-963 2P2), Beta (GDPCC-nCoV84, NPRC 2.062100001) and Omicron (BA.1, NPRC 964 2.192100005) variants were used in this study. 965

Binding properties of sera to SARS-CoV-2 prototype-Beta chimeric RBD-dimer 967 protein were determined by ELISA. 96-well plates (3590; Corning, USA) were coated 968 over-night with 3 μg/ml of prototype-Beta chimeric RBD-dimer protein in 0.05 M 969 carbonate-bicarbonate buffer, pH 9.6, and blocked in 5% skim milk in PBS. Serum 970 samples from macaques were serially diluted and added to each well. The plates were 971 incubated for 2 hours and then washed. The plates were incubated with goat anti-972 monkey IgG-HRP antibody (Abcam, ab112767), incubated for 1.5 hours and then 973 washed. The plates subsequently developed with 3,3',5,5'-tetramethylbenzidine (TMB) 974 substrate. Reactions were stopped with 2 M hydrochloric acid, and the absorbance 975 was measured at 450 nm using a microplate reader (PerkinElmer, USA). The endpoint 976 titers were defined as the highest reciprocal dilution of serum to give an absorbance 977 greater than 2.5-fold of the background values. Antibody titer below the limit of detection was determined as half the limit of detection. 979

To evaluate the protection efficacy of vaccine candidates against prototype SARS-981

CoV-2, BALB/c mice model transduced intranasally with a recombinant adenovirus 982 recombinant adenovirus 5 expressing human ACE2 (Ad5-hACE2) was used. 983

Immunized BALB/c mice were i.n infected with 8 × 10 9 vp of Ad5-hACE2. Five days 984 later, the transduced mice were challenged with 5 × 10 5 TCID50 of SARS-CoV-2 (hCoV- 

To perform the histopathology analysis, mice lung tissues were fixed in 4% 1004 paraformaldehyde, dehydrated, embedded in paraffin, and then sectioned. Tissue 1005 sections (4 μm) were deparaffinized in xylene and stained with haematoxylin and eosin 1006

New SARS-CoV-2 variants -clinical, 1108 public health, and vaccine implications

Effectiveness of the BNT162b2 Covid-19 vaccine against the

B.1.1.7 and B.1.351 variants

Adjuvanting a subunit COVID-19 1114 vaccine to induce protective immunity

A tandem-repeat dimeric RBD protein-based COVID-19 vaccine ZF2001 1117 protects mice and nonhuman primates

SARS-CoV-1120 2 neutralizing antibody structures inform therapeutic strategies

Broadly neutralizing 1123 antibodies overcome SARS-CoV-2 Omicron antigenic shift

Omicron escapes the majority of existing SARS-CoV-2 1126 neutralizing antibodies

Omicron extensively but 1129 incompletely escapes Pfizer BNT162b2 neutralization

SARS-CoV-2 infection 1132 protects against rechallenge in rhesus macaques

COVID-19 vaccine effectiveness against the B.1.1.7 and B.1.351 variants and severe 1136 COVID-19 disease in Qatar

Viral targets for vaccines against COVID-19

A universal design of betacoronavirus vaccines against COVID-19

SARS-CoV-2 variants escape neutralization by vaccine-induced humoral immunity

SARS-CoV-2 variants

1.351 and P.1 escape from neutralizing antibodies

The Omicron variant 1155 is highly resistant against antibody-mediated neutralization: Implications for control of 1156 the COVID-19 pandemic

Serum 1158 sample neutralisation of BBIBP-CorV and ZF2001 vaccines to SARS-CoV-2 501Y.V2

Omicron SARS-CoV-2 variant: a new chapter 1161 in the COVID-19 pandemic

Neutralizing 1164 antibody levels are highly predictive of immune protection from symptomatic SARS

CoV-2 infection

SARS-CoV-2 variants and 1168 vaccines

A therapeutic non-self-reactive

SARS-CoV-2 antibody protects from lung pathology in a COVID-19 hamster model

Omicron-specific mRNA vaccine induced 1175 potent neutralizing antibody against Omicron but not other SARS-CoV-2 variants

Viral loads of Delta-variant SARS-CoV-1179 et al. (2020). The impact of mutations in SARS-CoV-2 spike on viral infectivity and 1183 antigenicity

Reduced neutralization of SARS

CoV-2 B.1.617 by vaccine and convalescent serum

Striking antibody evasion manifested by the Omicron 1189 variant of SARS-CoV-2

Neutralizing activity of BNT162b2-elicited 1192 serum

SARS-CoV-2 variants on mRNA vaccine-induced immunity

Efficacy of the ChAdOx1 1198 nCoV-19 Covid-19 vaccine against the B.1.351 variant

A case of new variant COVID-19 first emerging in South Africa 1201 detected in a security guard at the isolation point

UCSF Chimera--a visualization system for exploratory 1205 research and analysis

Cross-neutralization of SARS-CoV-1208 2 by a human monoclonal SARS-CoV antibody

Considerable 1211 escape of SARS-CoV-2 Omicron to antibody neutralization

cryoSPARC: 1213 algorithms for rapid unsupervised cryo-EM structure determination

China's Zhifei says unit's COVID shot shows 81.76% efficacy in late-1216 stage trial

Neutralizing antibody vaccine for 1222 pandemic and pre-emergent coronaviruses

Neutralization of SARS-CoV-2 variants B.1.429 and B.1

A human neutralizing antibody targets the receptor-binding site of 1228 SARS-CoV-2

Efficacy of NVX-CoV2373

Covid-19 vaccine against the B.1.351 variant

Community transmission and viral load kinetics of the SARS-CoV-2 delta (B.1.617.2) 1235 variant in vaccinated and unvaccinated individuals in the UK: a prospective, 1236 longitudinal, cohort study

Prospective mapping of viral mutations that escape 1239 antibodies used to treat COVID-19

BNT162b2 1242 and mRNA-1273 COVID-19 vaccine effectiveness against the SARS-CoV-2 Delta 1243 variant in Qatar

Detection of a SARS-CoV-1246 2 variant of concern in South Africa

An infectious SARS

CoV-2 B.1.1.529 Omicron virus escapes neutralization by therapeutic monoclonal 1250 antibodies

Rapid epidemic expansion of 1253 the SARS-CoV-2 Omicron variant in southern Africa

Susceptibility of circulating SARS-CoV-2 1260 variants to neutralization

Structural and functional basis of SARS-CoV-2 entry by using 1263 human ACE2

Analysis of SARS-CoV-2 variant mutations reveals neutralization 1266 escape mechanisms and the ability to use ACE2 receptors from additional species

Description of the first strain of 2019-nCoV, C-Tan-nCoV Wuhan strain -1270 National Pathogen Resource Center

Interim Statement on COVID-19 vaccines in the context of the circulation 1272 of the Omicron SARS-CoV-2 Variant from the WHO Technical Advisory Group on

COVID-19 Vaccine Composition

SARS-CoV-2 501Y.V2 escapes neutralization by South African COVID-19 donor 1280 plasma

Virological assessment 1283 of hospitalized patients with COVID-2019

Serum Neutralizing

Activity Elicited by mRNA-1273 Vaccine

Humoral and cellular immunity and the safety of

COVID-19 vaccines: a summary of data published by 21

Safety and immunogenicity of a recombinant tandem-repeat dimeric

Boosting with Omicron-matched or 1297 historical mRNA vaccines increases neutralizing antibody responses and protection 1298 against B.1.1.529 infection in mice

A highly conserved cryptic epitope in the receptor binding domains of SARS

Effects of a prolonged booster interval on neutralization of 1304 Omicron variant

Neutralisation of ZF2001-elicited antisera to SARS-CoV-2 variants

MotionCor2: anisotropic correction of beam-induced motion for improved cryo-1310 electron microscopy

New tools for automated high-resolution cryo-EM structure 1313 determination in RELION-3

primarily lymphocytes, surrounding a bronchiole), perivasculitis (inflammatory cells, 1008 primarily lymphocytes, surrounding a blood vessel), interstitial pneumonitis (increased 1009 thickness of alveolar walls associated with inflammatory cells, primarily neutrophils), 1010 and alveolitis (inflammatory cells, primarily neutrophils and macrophages, within 1011 alveolar spaces). 1012

To evaluate the protection efficacy of vaccine candidates against Delta and Omicron 1014 variants, the immunized mice were challenged with 6 × 10 5 TCID50 of Delta variant 1015 

Twenty-four rhesus monkeys (n = 12) were immunized with clinical-grade prototype-1030 Beta chimeric vaccine (25 μg antigen + Aluminum hydroxide adjuvant) and placebo 1031 (Aluminum hydroxide adjuvant), respectively. Three doses were administrated at days 1032 0, 21 and 42. These monkeys were bled for humoral immunogenicity evaluation before 1033 immunization and 14, 35 and 52 days after receiving the first dose.To evaluate the cellular immune responses elicited by prototype-Beta chimeric 1035 vaccine, PBMCs were collected at day 48 and stimulated with peptide pool consisting 1036 of 15-18-mers (overlapping by 11 amino acids) and spanning the RBD of prototype 1037 SARS-CoV-2. INF-γ, IL-2 and IL-4 ELISpot assays were performed with ELISpot kits 1038 according to the manufacturer's protocols (MabTech, cat#3421M-4APW-10, 1039 cat#3445M-4APW-10 and cat#3410-2APW-10). 1040At the day 56, 57 and 58, animals of prototype-Beta chimeric vaccine group (2 1041 female and 2 male) and sham group (2 female and 2 male) were intranasally and 1042 intratracheally challenged with total 1×10 6 TCID50 prototype SARS-CoV-2 (GDPCC- Anal temperature was measured via electronic thermometer. 1047Before challenge and at the days 1, 3, 5, 7 post infection, nasal, throat, and anal 1048 swabs were collected from anesthetized animals into clean tube and lysed with Trizol. 1049 RNA was eluted with RNase/DNase-free distill water and stored at -80°C. Trizol. RNase/DNase-free distilled water was used to elute RNA, which was stored at 1057 -80°C. 1058One-step qRT-PCR was performed to measure viral genomic RNA (gRNA) with 1059 primers and probe paired with viral N gene that is recommended by WHO and China 1060 CDC. The sequences of primers and probe were same as those used in the prototype 1061 and preparation of the standard for N gene were performed according to SOP or the 1063 manufacture's protocols. 1064

Lung tissue samples of macaques were collected from seven lobes as described