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OA15.02 A. Hauser (1); D. Peterhoff (1); T. Hartinger (1); M. Schachtner (1); N. Friedrich (2); A. Trkola (2); C. Moog (3); W. Weissenhorn (4); Q. Sattentau (5); R. Sanders [...]

Neutralizing antibody titers and binding antibody levels are considered correlates of protection against severe SARS-CoV-2 infection. The clinical utility of serology should be reevaluated in light of the emergence of escape variants, as commercial antibody-binding assays have not been adapted to the virus' antigenic evolution. We compared anti-SARS-CoV-2 antibody titers in four quantitative serological tests based on variable ancestral spike antigens (three in-house ELISAs and the prototype VIDAS SARS-CoV-2 IgG QUANT assay) and neutralization assays against the pseudotyped Wuhan, BA.2, BA.4/5, BQ.1.1, and XBB.1.1 viruses in a cohort of 100 patients infected in 2020 or during the Omicron waves. Binding antibody levels correlated well with neutralizing antibody titers for Wuhan, BA.2, and BA.4/5, but the association decreased for BQ.1.1 and XBB.1 (for the VIDAS assay, Spearman's correlation was 0.82 [95% CI 0.74-0.88] and 0.61 [0.46-0.72] for BA.2 and XBB.1, respectively). In 15% of patients with no neutralizing antibodies against XBB.1, the VIDAS assay still yielded binding antibody levels ranging from 74 to 7,652 binding antibody units/mL. Using an adjusted threshold based on receiver operating characteristic (ROC) curve analysis, the specificity of neutralizing antibody detection increased from 0.15 (95% CI 0.02-0.45) and 0.17 (0.04-0.41) to 0.92 (0.64-1.00) and 0.83 (0.59-0.96) against BQ.1.1 and XBB.1, respectively. Serological tests based on receptor-binding domain antigens from the ancestral virus fail to predict neutralizing activity against the latest circulating Omicron variants. Adapting serological tests may improve their clinical utility in immunocompromised patients., Importance: Anti-SARS-CoV-2 serology was developed in 2020 in response to the COVID-19 pandemic to diagnose SARS-CoV-2 infection and monitor an individual's immunity following natural infection or vaccination. Given the relationship between neutralizing antibody titers and protection against severe infection, many studies have evaluated the correlation between serology tests and neutralization assays in the pre-Omicron era. An important potential clinical use of serology, which explores binding antibodies, is estimating an individual's level of protection against new infection, particularly in immunosuppressed individuals and those at risk of severe COVID. However, in the Omicron era, as new viruses evade the immunity induced by previous infections and vaccination, the correlation between binding antibody levels determined by serological assays developed from ancestral antigens and neutralizing antibody titers against new viruses should be re-examined in order to determine whether these assays should be optimized by adapting antigens to the circulating SARS-CoV-2 strains., Competing Interests: The virology laboratory (CHU Grenoble Alpes) received grants from bioMérieux to carry out the study.

HIV-1 budding as well as many other cellular processes require the Endosomal Sorting Complex Required for Transport (ESCRT) machinery. Understanding the architecture of the native ESCRT-III complex at HIV-1 budding sites is limited due to spatial resolution and transient ESCRT-III recruitment. Here, we developed a drug-inducible transient HIV-1 budding inhibitory tool to enhance the ESCRT-III lifetime at budding sites. We generated autocleavable CHMP2A, CHMP3, and CHMP4B fusion proteins with the hepatitis C virus NS3 protease. We characterized the CHMP-NS3 fusion proteins in the absence and presence of protease inhibitor Glecaprevir with regard to expression, stability, localization, and HIV-1 Gag VLP budding. Immunoblotting experiments revealed rapid and stable accumulation of CHMP-NS3 fusion proteins. Notably, upon drug administration, CHMP2A-NS3 and CHMP4B-NS3 fusion proteins substantially decrease VLP release while CHMP3-NS3 exerted no effect but synergized with CHMP2A-NS3. Localization studies demonstrated the relocalization of CHMP-NS3 fusion proteins to the plasma membrane, endosomes, and Gag VLP budding sites. Through the combined use of transmission electron microscopy and video-microscopy, we unveiled drug-dependent accumulation of CHMP2A-NS3 and CHMP4B-NS3, causing a delay in HIV-1 Gag-VLP release. Our findings provide novel insight into the functional consequences of inhibiting ESCRT-III during HIV-1 budding and establish new tools to decipher the role of ESCRT-III at HIV-1 budding sites and other ESCRT-catalyzed cellular processes.

The SARS-CoV-2 pandemic has again shown that structural biology plays an important role in understanding biological mechanisms and exploiting structural data for therapeutic interventions. Notably, previous work on SARS-related glycoproteins has paved the way for the rapid structural determination of the SARS-CoV-2 S glycoprotein, which is the main target for neutralizing antibodies. Therefore, all vaccine approaches aimed to employ S as an immunogen to induce neutralizing antibodies. Like all enveloped virus glycoproteins, SARS-CoV-2 S native prefusion trimers are in a metastable conformation, which primes the glycoprotein for the entry process via membrane fusion. S-mediated entry is associated with major conformational changes in S, which can expose many off-target epitopes that deviate vaccination approaches from the major aim of inducing neutralizing antibodies, which mainly target the native prefusion trimer conformation. Here, we review the viral glycoprotein stabilization methods developed prior to SARS-CoV-2, and applied to SARS-CoV-2 S, in order to stabilize S in the prefusion conformation. The importance of structure-based approaches is highlighted by the benefits of employing stabilized S trimers versus non-stabilized S in vaccines with respect to their protective efficacy.