key: cord-0429255-6mv86am7
authors: Carreno, J. M.; Alshammary, H.; Singh, G.; Raskin, A.; Amanat, F.; Amoako, A.; Gonzalez-Reiche, A. S.; van de Guchte, A.; PARIS study group,; Srivastava, K.; Sordillo, E. M.; Sather, D. N.; van Bakel, H.; Krammer, F.; Simon, V.
title: Reduced neutralizing activity of post-SARS-CoV-2 vaccination serum against variants B.1.617.2, B.1.351, B.1.1.7+E484K and a sub-variant of C.37
date: 2021-07-23
journal: nan
DOI: 10.1101/2021.07.21.21260961
sha: 0c4622bbd8e0a5efafa0462fc58dbcf50ca330f0
doc_id: 429255
cord_uid: 6mv86am7

Highly efficacious vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have been developed. However, the emergence of viral variants that are more infectious than the earlier SARS-CoV-2 strains is concerning. Several of these viral variants have the potential to partially escape neutralizing antibody responses warranting continued immune-monitoring. Here, we tested a number of currently circulating viral variants of concern/interest, including B.1.526 (Iota), B.1.1.7+E484K (Alpha), B.1.351 (Beta), B.1.617.2 (Delta) and C.37 (Lambda) in neutralization assays using a panel of post-mRNA vaccination sera. The assays were performed with authentic SARS-CoV-2 clinical isolates in an assay that mimics physiological conditions. We found only small decreases in neutralization against B.1.526 and an intermediate phenotype for B.617.2. The reduction was stronger against a sub-variant of C.37, followed by B.1.351 and B.1.1.7+E484K. C.37 is currently circulating in parts of Latin America and was detected in Germany, the US and Israel. Of note, reduction in a binding assay that also included P.1, B.1.617.1 (Kappa) and A.23.1 was negligible. Taken together, these findings suggest that mRNA SARS-CoV-2 vaccines may remain effective against these viral variants of concern/interest and that spike binding antibody tests likely retain specificity in the face of evolving SARS-CoV-2 diversity.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged in late 2019 in China 48

and has since then caused the coronavirus disease 2019 (COVID-19) pandemic. Vaccines were developed 49 and distributed in record speed with emergency use authorizations as early as December 2020 (in the 50 US). Coronaviruses are an exception among RNA viruses because their replication machinery possesses 51 proofreading activity. 4 It was, therefore, expected that they would evolve slower than other RNA 52

viruses. The emergence of variant viruses, first in Europe in the summer of 2020 5-7 -including in minks in 53

Denmark -and then in late 2020 in the UK 8 , South Africa 9 and Brazil 10 was a surprising event. Since the 54 detection of these first variants, several other variants of interest (VoIs) and variants of concern (VoCs) 55 have spread locally as well as globally. These include B. changes in the N-terminal domain (NTD) and/or the receptor binding domain (RBD) of spike, both of 62 which harbor neutralizing epitopes. In addition, several of these variants also contain additional 63 mutations in their spike gene that may enhance affinity to human angiotensin converting enzyme 2 64 (ACE2). 16 These changes may impact negatively on binding and neutralization by vaccine-induced 65

antibodies, which are discussed as mechanistic correlates of protection. 17, 18 It is, therefore, important to 66 monitor neutralizing and binding activity of sera from vaccinees against these variants. 67

Here, we tested the neutralizing activity of a panel of 30 post mRNA SARS-CoV-2 vaccination 68 sera against the three B.1.526 sublineages, B.1.1.7+E484K, B.1.617.2, B.1.351 and a C.37 sub-variant 69 using a microneutralization assay. This assay was designed to allow for multicycle replication with 70 sera/antibodies being present in the overlay at all times to better mimic physiological conditions. 19 In 71 addition, we performed binding assays against recombinant RBD and full-length spike proteins of several 72

VoIs and VoCs. 73 . 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 July 23, 2021. (Figure 2A and B) Figure 2C and D) . 106

Monitoring the effectiveness of vaccination in the context of evolving viral variants is of great 108 public health concern for control of the COVID-19 pandemic. Indeed, neutralizing antibodies play a 109 major role in protection from COVID-19, as shown both in animal models as well as in humans. 17, 18, [21] [22] [23] is, therefore, important to continuously monitor emerging variants and their impact on neutralizing 111

activity of post-vaccination serum. Our data show a spectrum of neutralizing activities against a panel of 112

VoIs and VoCs. The least negative impact on neutralization was seen for B.1.526 variant, which is 113 reasonable since this isolate has very few changes in the spike protein (T95I, D253G and D614G) 114 compared to the wild type WA1 isolate. The impact was higher for the B.1.526 isolates that carry, in 115 . 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.

The copyright holder for this preprint this version posted July 23, 2021. ; https://doi.org/10.1101/2021.07.21.21260961 doi: medRxiv preprint addition, E484K or S477N substitutions in the RBD. Interestingly, S477N exerted a stronger impact than 116 E484K, which was surprising given the strong reduction caused by introduction of E484K into the WA1 117 wild type backbone. 24 However, S477N has also been described as escape mutation. 25 The reduction in 118 neutralization against B.1.617.2 was moderate, which aligns well with the preserved vaccine 119 effectiveness against this variant. 26,27 The combination of B.1.1.7 (which already harbors two deletions in 120 the NTD) with E484K led to a substantial reduction in neutralizing activity. However, although 121 B.1.1.7+E484K emerged several times independently, it does not seem to spread well -potentially due 122

to the detrimental effect that the E484K mutation can have on binding affinity to human ACE2. 16 As 123 expected and already shown by several publications, 28-33 neutralizing activity to B.1.351 was strongly 124 reduced likely due to the three changes in RBD and extensive changes in the NTD. Interestingly, 125

neutralizing activity was even more reduced for the local isolate of C.37 used in the current study which 126 has more extensive changes in the NTD (Δ63-75 , Δ246-252) and the RBD (L452Q, E471Q, F490S) 127 compared to the predominant C.37 variant (NTD: G75V, T76I, Δ246-252; RBD: L452Q, F490S). 3 While 128

there was a strong reduction in neutralizing activity of post-vaccination sera against this variant, all 30 129 sera retained at least partial neutralization activity against this C37 variant virus which may indicate that 130 -similar to B.1.351 34 -mRNA vaccines will remain effective against these VoIs. Of note, the additional 131 changes in the local C.37 isolate likely contribute to the immune evasion phenotype shown here and 132

should be seen as worst case scenario for the C.37 variant. 133

Another interesting finding was that the reduction in binding activity to variant RBD and spike proteins 134 was negligible. However, while neutralization activity is mostly dependent on only a few epitopes in the 135 RBD and NTD, many more binding epitopes exist on the spike, 16 which may explain our findings. 136

Interestingly, a recent study found even better correlation between binding activity and vaccine 137 effectiveness than for neutralizing activity and vaccine effectiveness. 17 This may indicate that binding 138

antibodies -even if they lack neutralizing activity -are involved in protection. It is well known that non-139 neutralizing antibodies raised upon infection or vaccination by ebolavirus or influenza viruses, can be 140 protective via Fc-mediated effector functions. [35] [36] [37] [38] Future studies are needed to establish if this is also 141 the case for SARS-CoV-2. However, the finding that the drop in binding activity is negligible even against 142 spike proteins from variants with extensive changes like B. . 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.

The copyright holder for this preprint this version posted July 23, 2021. was removed and initial dilutions of heat-inactivated sera (in PBS-T 1%-milk powder) were added to the 176 plates, followed by 2-fold serial dilutions. After 2-hour incubation, plates were washed three times with 177 PBS-T and 50 μl/well of the pre-diluted secondary antibody anti-human IgG (Fab-specific) horseradish 178 peroxidase antibody (produced in goat; Sigma, A0293) diluted 1:3,000 in PBS-T containing 1% milk 179 powder were added. After 1-hour incubation at RT, plates were washed three times with PBS-T and 180

SigmaFast o-phenylenediamine dihydrochloride (Sigma) was added (100 μl/well) for 10min, followed by 181 addition of 50 μl/well of 3 M hydrochloric acid (Thermo Fisher) to stop the reaction. Optical density was 182 measured at a wavelength of 490 nm using a plate reader (BioTek were incubated with 1,000 tissue culture infectious dose 50 (TCID50) of wt USA-WA1/2020 SARS- or the corresponding variant virus for 1 hour at RT, followed by transfer of 120μl of virus-sera mix to 199 . 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.

The copyright holder for this preprint this version posted July 23, 2021. The Icahn School of Medicine at Mount Sinai has filed patent applications relating to SARS-CoV-2 239 serological assays and NDV-based SARS-CoV-2 vaccines which list Florian Krammer as co-inventor. 240

Viviana Simon and Fatima Amanat are also listed on the serological assay patent application as co-241 . 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.

The copyright holder for this preprint this version posted July 23, 2021. ; https://doi.org/10.1101 https://doi.org/10. /2021 inventors. Mount Sinai has spun out a company, Kantaro, to market serological tests for SARS- Florian Krammer has consulted for Merck and Pfizer (before 2020), and is currently consulting for Pfizer, 243

Seqirus and Avimex. The Krammer laboratory is also collaborating with Pfizer on animal models of SARS-244

CoV-2. 245 246 . 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.

The copyright holder for this preprint this version posted July 23, 2021. . 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.

The copyright holder for this preprint this version posted July 23, 2021. 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.

The copyright holder for this preprint this version posted July 23, 2021. ; https://doi.org/10.1101/2021.07.21.21260961 doi: medRxiv preprint 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 July 23, 2021. ; https://doi.org/10.1101/2021.07.21.21260961 doi: medRxiv preprint 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 July 23, 2021. . 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 July 23, 2021. ; https://doi.org/10.1101/2021.07.21.21260961 doi: medRxiv preprint

Supplementary Figure 1: Phylogenetic tree including all patient isolates used in this study