ABSTRACT
Antibodies (Abs) are essential for the host immune response against SARS-CoV-2, and all the vaccines developed so far have been designed to induce Abs targeting the SARS-CoV-2 spike. Many studies have examined Ab responses in the blood from vaccinated and infected individuals. However, since SARS-CoV-2 is a respiratory virus, it is also critical to understand the mucosal Ab responses at the sites of initial virus exposure. Here, we examined plasma versus saliva Ab responses in vaccinated and convalescent patients. Although saliva levels were significantly lower, a strong correlation was observed between plasma and saliva total Ig levels against all SARS-CoV-2 antigens tested. Virus-specific IgG1 responses predominated in both saliva and plasma, while a lower prevalence of IgM and IgA1 Abs was observed in saliva. Antiviral activities of plasma Abs were also studied. Neutralization titers against the initial WA1 (D614G), B.1.1.7 (alpha) and B.1.617.2 (delta) strains were similar but lower against the B.1.351 (beta) strain. Spike-specific antibody-dependent cellular phagocytosis (ADCP) activities were also detected and the levels correlated with spike-binding Ig titers. Interestingly, while neutralization and ADCP potencies of vaccinated and convalescent groups were comparable, enhanced complement deposition to spike-specific Abs was noted in vaccinated versus convalescent groups and corresponded with higher levels of IgG1 plus IgG3 among the vaccinated individuals. Altogether, this study demonstrates the detection of Ab responses after vaccination or infection in plasma and saliva that correlate significantly, although Ig isotypic differences were noted. The induced plasma Abs displayed Fab-mediated and Fc-dependent functions with comparable neutralization and ADCP potencies, but a greater capacity to activate complement was elicited upon vaccination.
ABSTRACT
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, China. 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 suggests herd immunity may fail to eliminate the virus. All three officially designated VOC carry Spike (S) polymorphisms thought to enable escape from neutralizing antibodies elicited during initial waves of the pandemic. 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 8 out of 12 (75%) of serum samples from 12 recipients of the Russian Sputnik V Ad26 / Ad5 vaccine showed dose response curve slopes indicative of failure to neutralize rcVSV-CoV2-S: B.1.351. 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 emergent SARS-CoV-2 variants may benefit from updated vaccines.
ABSTRACT
Entry of SARS-CoV-2 is facilitated by endogenous and exogenous proteases. These proteases proteolytically activate the SARS-CoV-2 spike glycoprotein and are key modulators of virus tropism. We show that SARS-CoV-2 naive serum exhibits significant inhibition of SARS-CoV-2 entry. We identify alpha-1-antitrypsin (AAT) as the major serum protease inhibitor that potently restrict protease-mediated entry of SARS-CoV-2. AAT inhibition of protease-mediated SARS-CoV-2 entry in vitro occurs at concentrations far below what is present in serum and bronchoalveolar tissues, suggesting that AAT effects are physiologically relevant. Moreover, AAT deficiency affects up to 20% of the population and its symptomatic manifestations coincides with many risk factors associated with severe COVID-19 disease. In addition to the effects that AAT may have on viral entry itself, we argue that the anti-inflammatory and coagulation regulatory activity of AAT have implications for coronavirus disease 2019 (COVID-19) pathogenicity, SARS-CoV-2 tissue restriction, convalescent plasma therapies, and even potentially AAT therapy.
ABSTRACT
SARS-CoV-2 has infected millions of people and is on a trajectory to kill more than one million globally. Virus entry depends on the receptor-binding domain (RBD) of the spike protein. Although previous studies demonstrated anti-spike and -RBD antibodies as essential for protection and convalescent plasma as a promising therapeutic option, little is known about the immunoglobulin (Ig) isotypes capable of blocking virus entry. Here, we studied spike- and RBD-specific Ig isotypes in plasma/sera from two acutely infected and 29 convalescent individuals. Spike- and RBD-specific IgM, IgG1, and IgA1 antibodies were produced by all or nearly all subjects at varying levels and detected at 7-8 days post-disease onset. IgG2, IgG3, IgG4, and IgA2 were also present but at much lower levels. All samples also displayed neutralizing activity. IgM, IgG, and IgA were capable of mediating neutralization, but neutralization titers correlated better with binding levels of IgM and IgA1 than IgG.
ABSTRACT
The global COVID-19 pandemic has mobilized efforts to develop vaccines and antibody-based therapeutics, including convalescent plasma therapy, that inhibit viral entry by inducing or transferring neutralizing antibodies (nAbs) against the SARS-CoV-2 spike glycoprotein (CoV2-S). However, rigorous efficacy testing requires extensive screening with live virus under onerous BSL3 conditions which limits high throughput screening of patient and vaccine sera. Myriad BSL-2 compatible surrogate virus neutralization assays (VNAs) have been developed to overcome this barrier. Yet, there is marked variability between VNAs and how their results are presented, making inter-group comparisons difficult. To address these limitations, we developed a standardized VNA using VSVAG-based CoV-2-S pseudotyped particles (CoV2pp) that can be robustly produced at scale and generate accurate neutralizing titers within 18 hours post-infection. Our standardized CoV2pp VNA showed a strong positive correlation with CoV2-S ELISA and live virus neutralizations in confirmed convalescent patient sera. Three independent groups subsequently validated our standardized CoV2pp VNA (n>120). Our data show that absolute (abs) IC50, IC80, and IC90 values can be legitimately compared across diverse cohorts, highlight the substantial but consistent variability in neutralization potency across these cohorts, and support the use of absIC80 as a more meaningful metric for assessing the neutralization potency of vaccine or convalescent sera. Lastly, we used our CoV2pp in a screen to identify ultra-permissive 293T clones that stably express ACE2 or ACE2+TMPRSS2. When used in combination with our CoV2pp, we can now produce CoV2pp sufficient for 150,000 standardized VNA/week. ImportanceVaccines and antibody-based therapeutics like convalescent plasma therapy are premised upon inducing or transferring neutralizing antibodies that inhibit SARS-CoV-2 entry into cells. Virus neutralization assays (VNAs) for measuring neutralizing antibody titers (NATs) is an essential part of determining vaccine or therapeutic efficacy. However, such efficacy testing is limited by the inherent dangers of working with the live virus, which requires specialized high-level biocontainment facilities. We therefore developed a standardized replication-defective pseudotyped particle system that mimics entry of live SARS-CoV-2. This tool allows for the safe and efficient measurement of NATs, determination of other forms of entry inhibition, and thorough investigation of virus entry mechanisms. Four independent labs across the globe validated our standardized VNA using diverse cohorts. We argue that a standardized and scalable assay is necessary for meaningful comparisons of the myriad of vaccines and antibody-based therapeutics becoming available. Our data provide generalizable metrics for assessing their efficacy.