Neutralizing activity of Sputnik V vaccine sera against SARS-CoV-2 variants

The novel pandemic betacoronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has infected at least 120 million people since its identification as the cause of a December 2019 viral pneumonia outbreak in Wuhan, China1,2. Despite the unprecedented pace of vaccine development, with six vaccines already in use worldwide, the emergence of SARS-CoV-2 ‘variants of concern’ (VOC) across diverse geographic locales have prompted re-evaluation of strategies to achieve universal vaccination3. All three officially designated VOC carry Spike (S) polymorphisms thought to enable escape from neutralizing antibodies elicited during initial waves of the pandemic4–8. Here, we characterize the biological consequences of the ensemble of S mutations present in VOC lineages B.1.1.7 (501Y.V1) and B.1.351 (501Y.V2). Using a replication-competent EGFP-reporter vesicular stomatitis virus (VSV) system, rcVSV-CoV2-S, which encodes S from SARS coronavirus 2 in place of VSV-G, and coupled with a clonal HEK-293T ACE2 TMPRSS2 cell line optimized for highly efficient S-mediated infection, we determined that only 1 out of 12 serum samples from a cohort of recipients of the Gamaleya Sputnik V Ad26 / Ad5 vaccine showed effective neutralization (IC90) of rcVSV-CoV2-S: B.1.351 at full serum strength. The same set of sera efficiently neutralized S from B.1.1.7 and showed only moderately reduced activity against S carrying the E484K substitution alone. Taken together, our data suggest that control of some emergent SARS-CoV-2 variants may benefit from updated vaccines.

INTRODUCTION. 45 In the 15 months since its emergence in late 2019 1 , SARS-CoV-2 has caused over 131 46 million confirmed COVID-19 cases worldwide, leading to at least 2.85 million deaths 2 .
SARS-CoV-2 is closely related to two other zoonotic betacoronaviruses, MERS-CoV 48 and SARS-CoV, that also cause life-threatening respiratory infections 3 . 49 This global health emergency has spurred the development of COVID-19 preventive 50 vaccines at an unprecedented pace. Six are already authorized for human use across 51 the globe [4][5][6][7][8][9] . These vaccines focus on the SARS-CoV-2 spike protein (S), due to its 52 critical roles in cell entry. Indeed, the presence of serum neutralizing antibodies 53 directed at S correlate strongly with protection against COVID-19 10,11 . Although these 54 six vaccines are efficacious, the recent emergence of novel SARS-CoV-2 variants has 55 reignited concerns that the pandemic may not be so easily brought under control. 56 In December 2020, the United Kingdom reported the sudden emergence of a novel 57 SARS-CoV-2 lineage, termed B.1.1.7 (501Y.V1, VOC 202012/01), which was 58 designated as the first SARS-CoV-2 variant of concern (VOC) . The lineage had rapidly 59 increased in prevalence since first being detected in November 2020 12 . Its genome 60 showed an unusually high number of non-synonymous substitutions and deletions, 61 including eight in the S gene, suggesting a substantial degree of host adaptation that 62 may have occurred during prolonged infection of an immunocompromised person 13 . 63 The B.1.1.7 lineage has now been shown to exhibit enhanced transmissibility 14 as well 64 as an increased case fatality rate 15,16 .

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A key public health concern related to emergent SARS-CoV-2 variants is that by 228 incrementally accruing mutations that escape neutralizing antibodies, they will 229 penetrate herd immunity and spread to reach unvaccinated individuals, some of whom 230 will be susceptible to severe or fatal disease.  Table 2 summarizes peer-reviewed studies that have tested post-vaccination 268 sera from the major vaccines against the VOC/mutant spikes used in this study. Our 269 study shows a similar mean reduction in GMT (reciprocal IC50) against E484K and  While the E484K substitution appears to be a common route of escape from many 285 RBD-targeting monoclonal antibodies, it is somewhat surprising that a single mutation 286 can confer a significant degree of neutralization resistance from polyclonal responses.

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Nonetheless, our data show that resistance conferred by E484K mutation be overcome      We cloned VSV-eGFP sequence into pEMC vector (pEMC-VSV-eGFP), which includes 337 an optimized T7 promoter and hammerhead ribozyme just before the 5' end of the viral 338 . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

(which was not certified by peer review)
The copyright holder for this preprint this version posted May 29, 2021. genome. The original VSV-eGFP sequence was from pVSV-eGFP, a generous gift of 339 Dr. John Rose 62 .

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We generated pEMC-VSV-eGFP-CoV2-S (Genbank Accession: MW816496) as follows:  Table 2).   We gratefully acknowledge all submitting authors and collecting authors on whose 445 work this research is based, and to all researchers, clinicians, and public health 446 . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

(which was not certified by peer review)
The copyright holder for this preprint this version posted May 29, 2021. ; https://doi.org/10.1101/2021.03.31.21254660 doi: medRxiv preprint authorities who make SARS-CoV-2 sequence data available in a timely manner via the (-) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

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The copyright holder for this preprint this version posted May 29, 2021. ; https://doi.org/10.1101/2021.03.31.21254660 doi: medRxiv preprint Table 2. Summary of post-vaccine sera evaluated for neutralization potency against the indicated SARS-CoV-2 variants of concern (VOC). . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

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The copyright holder for this preprint this version posted May 29, 2021 Acknowledgement of B.1.1.7 and B.1.351 viruses used for selection of S variants evaluated in this study.
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The copyright holder for this preprint this version posted May 29, 2021. ; https://doi.org/10.1101/2021.03.31.21254660 doi: medRxiv preprint between cell lines at any given dilution. Adjusted p values from Tukey's multiple comparisons test are given (ns; not significant, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001). (C) rcVSV-CoV-2-S containing the prevailing WT (D614G) and VOC (B.1.1.7 and B.1.351) spikes were inoculated into one 6-well each of F8-2 cells (MOI 0.1) and subsequently overlaid with methylcellulose-DMEM to monitor syncytia formation. Representative images of syncytial plaques at 48 hpi are shown. White bar equals 1 millimeter. (D) shows the growth of GFP positive area / infectious unit (IU) in the 6 well plate. GFP positive areas were imaged and measured by the Celigo imaging cytometer. IU was checked at 10 hpi in the same well. Bar shows the average of 3 independent experiments with error bar indicating standard deviation. No statistically significant differences were detected between WT and VOC spikes in the size of GFP+ syncytia at any given time point (two-way ANOVA as above, 'ns' not indicated in graph).
. CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.   . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review)  where Y= 100/(1+10^((LogIC50-X)*HillSlope))). Log IC50 and Hill slope values were obtained for each curve generated in Fig. 3 . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review)   CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted May 29, 2021. ; https://doi.org/10.1101/2021.03.31.21254660 doi: medRxiv preprint Extended Data Figure S1. Robust and efficient generation of an EGFP-reporter replicationcompetent VSV bearing SARS-CoV-2 spike (rcVSV-CoV2-S). (A) Schematic of the rcVSV-CoV2-S genomic coding construct and the virus rescue procedure. The maximal T7 promoter (T7prom) followed by a hammer-head ribozyme (HhRbz) and the HDV ribozyme (HDVRbz) plus T7 terminator (T7term) are positioned at the 3' and 5' ends of the viral cDNA, respectively. An EGFP(E) transcriptional unit is placed at the 3' terminus to allow for high level transcription. SARS-CoV-2-S is cloned in place of VSV-G using the indicated restriction sites designed to facilitate easy exchange of spike variant or mutants. (B) For virus rescue, highly permissive 293T cells stably expressing human ACE2 and TMPRSS2 (293T-[ACE2+TMPRSS2], F8-2 clone) cells were transfected with the genome coding plasmid, helper plasmids encoding CMV-driven N, P, M, and L genes, and pCAGS encoding codon-optimized T7-RNA polymerase(T7opt). 48-72 hpi, transfected cells turn EGFP+ and start forming syncytia. Supernatant containing rcVSV-CoV2-S are then amplified in Vero-TMPRSS2 cells at the scale shown. The blue arrows at the bottom indicate the timeline for production of each sequence verified stock.
. CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted May 29, 2021. ; https://doi.org/10.1101/2021.03.31.21254660 doi: medRxiv preprint