RESUMEN
Continuous evolution of Omicron has led to a rapid and simultaneous emergence of numerous variants that display growth advantages over BA. 5. Despite their divergent evolutionary courses, mutations on their receptor-binding domain (RBD) converge on several hotspots. The driving force and destination of such convergent evolution and its impact on humoral immunity remain unclear. Here, we demonstrate that these convergent mutations can cause striking evasion of neutralizing antibody (NAb) drugs and convalescent plasma, including those from BA.5 breakthrough infection, while maintaining sufficient ACE2 binding capability. BQ.1.1.10, BA.4.6.3, XBB, and CH. 1.1 are the most antibody-evasive strain tested, even exceeding SARS-CoV-1 level. To delineate the origin of the convergent evolution, we determined the escape mutation profiles and neutralization activity of monoclonal antibodies (mAbs) isolated from BA.2 and BA.5 breakthrough-infection convalescents. Importantly, due to humoral immune imprinting, BA.2 and especially BA.5 breakthrough infection caused significant reductions in the epitope diversity of NAbs and increased proportion of non-neutralizing mAbs, which in turn concentrated humoral immune pressure and promoted convergent evolution. Moreover, we showed that the convergent RBD mutations could be accurately inferred by integrated deep mutational scanning (DMS) profiles, and the evolution trends of BA.2.75/BA.5 subvariants could be well-simulated through constructed convergent pseudovirus mutants. Together, our results suggest current herd immunity and BA.5 vaccine boosters may not provide good protection against infection. Broad-spectrum SARS-CoV-2 vaccines and NAb drugs development should be highly prioritized, and the constructed mutants could help to examine their effectiveness in advance.
RESUMEN
Multiple BA.4 and BA.5 subvariants with R346 mutations on the spike glycoprotein have been identified in various countries, such as BA.4.6/BF.7 harboring R346T, BA.4.7 harboring R346S, and BA.5.9 harboring R346I. These subvariants, especially BA.4.6, exhibit substantial growth advantages compared to BA.4/BA.5. In this study, we showed that BA.4.6, BA.4.7, and BA.5.9 displayed higher humoral immunity evasion capability than BA.4/BA.5, causing 1.5 to 1.9-fold decrease in NT50 of the plasma from BA.1 and BA.2 breakthrough-infection convalescents compared to BA.4/BA.5. Importantly, plasma from BA.5 breakthrough-infection convalescents also exhibits significant neutralization activity decrease against BA.4.6, BA.4.7, and BA.5.9 than BA.4/BA.5, showing on average 2.4 to 2.6-fold decrease in NT50. For neutralizing antibody drugs, Bebtelovimab remains potent, while Evusheld is completely escaped by these subvariants. Together, our results rationalize the prevailing advantages of the R346 mutated BA.4/BA.5 subvariants and urge the close monitoring of these mutants, which could lead to the next wave of the pandemic.
RESUMEN
SARS-CoV-2 Omicron sublineages have escaped most RBD-targeting therapeutic neutralizing antibodies (NAbs), which proves the previous NAb drug screening strategies deficient against the fast-evolving SARS-CoV-2. Better broad NAb drug candidate selection methods are needed. Here, we describe a rational approach for identifying RBD-targeting broad SARS-CoV-2 NAb cocktails. Based on high-throughput epitope determination, we propose that broad NAb drugs should target non-immunodominant RBD epitopes to avoid herd immunity-directed escape mutations. Also, their interacting antigen residues should focus on sarbecovirus conserved sites and associate with critical viral functions, making the antibody-escaping mutations less likely to appear. Following the criteria, a featured non-competing antibody cocktail, SA55+SA58, is identified from a large collection of broad sarbecovirus NAbs isolated from SARS convalescents. SA55+SA58 potently neutralizes ACE2-utilizing sarbecoviruses, including circulating Omicron variants, and could serve as broad SARS-CoV-2 prophylactics to offer long-term protection. Our screening strategy can also be applied to identify broad-spectrum NAb drugs against other fast-evolving viruses, such as influenza viruses.
RESUMEN
Recently emerged SARS-CoV-2 Omicron subvariant, BA.2.75, displayed a local growth advantage over BA.2.38, BA.2.76 and BA.5 in India. The underlying mechanism of BA.2.75s enhanced infectivity, especially compared to BA.5, remains unclear. Here, we show that BA.2.75 exhibits substantially higher ACE2-binding affinity than BA.5. Also, BA.2.75 spike shows decreased thermostability and increased "up" RBD conformation in acidic conditions, suggesting enhanced low-pH-endosomal cell-entry pathway utilization. BA.2.75 is less humoral immune evasive than BA.4/BA.5 in BA.1/BA.2 breakthrough-infection convalescents; however, BA.2.75 shows heavier neutralization evasion in Delta breakthrough-infection convalescents. Importantly, plasma from BA.5 breakthrough infection exhibit significantly weaker neutralization against BA.2.75 than BA.5, mainly due to BA.2.75s distinct RBD and NTD-targeting antibody escaping pattern from BA.4/BA.5. Additionally, Evusheld and Bebtelovimab remain effective against BA.2.75, and Sotrovimab recovered RBD-binding affinity. Together, our results suggest BA.2.75 may prevail after the global BA.4/BA.5 wave, and its increased receptor-binding capability could allow further incorporation of immune-evasive mutations.
RESUMEN
SARS-CoV-2 Omicron sublineages BA.2.12.1, BA.4 and BA.5 exhibit higher transmissibility over BA.21. The new variants receptor binding and immune evasion capability require immediate investigation. Here, coupled with Spike structural comparisons, we show that BA.2.12.1 and BA.4/BA.5 exhibit comparable ACE2-binding affinities to BA.2. Importantly, BA.2.12.1 and BA.4/BA.5 display stronger neutralization evasion than BA.2 against the plasma from 3-dose vaccination and, most strikingly, from post-vaccination BA.1 infections. To delineate the underlying antibody evasion mechanism, we determined the escaping mutation profiles2, epitope distribution3 and Omicron neutralization efficacy of 1640 RBD-directed neutralizing antibodies (NAbs), including 614 isolated from BA.1 convalescents. Interestingly, post-vaccination BA.1 infection mainly recalls wildtype-induced humoral memory. The resulting elicited antibodies could neutralize both wildtype and BA.1 and are enriched on non-ACE2-competing epitopes. However, most of these cross-reactive NAbs are heavily escaped by L452Q, L452R and F486V. BA.1 infection can also induce new clones of BA.1-specific antibodies that potently neutralize BA.1; nevertheless, these NAbs are largely escaped by BA.2/BA.4/BA.5 due to D405N and F486V, and react weakly to pre-Omicron variants, exhibiting poor neutralization breadths. As for therapeutic NAbs, Bebtelovimab4 and Cilgavimab5 can effectively neutralize BA.2.12.1 and BA.4/BA.5, while the S371F, D405N and R408S mutations would undermine most broad sarbecovirus NAbs. Together, our results indicate that Omicron may evolve mutations to evade the humoral immunity elicited by BA.1 infection, suggesting that BA.1-derived vaccine boosters may not achieve broad-spectrum protection against new Omicron variants.
RESUMEN
Omicron sub-lineage BA.2 has rapidly surged globally, accounting for over 60% of recent SARS-CoV-2 infections. Newly acquired RBD mutations and high transmission advantage over BA.1 urge the investigation of BA.2s immune evasion capability. Here, we show that BA.2 causes strong neutralization resistance, comparable to BA.1, in vaccinated individuals plasma. However, BA.2 displays more severe antibody evasion in BA.1 convalescents, and most prominently, in vaccinated SARS convalescents plasma, suggesting a substantial antigenicity difference between BA.2 and BA.1. To specify, we determined the escaping mutation profiles1,2 of 714 SARS-CoV-2 RBD neutralizing antibodies, including 241 broad sarbecovirus neutralizing antibodies isolated from SARS convalescents, and measured their neutralization efficacy against BA.1, BA.1.1, BA.2. Importantly, BA.2 specifically induces large-scale escape of BA.1/BA.1.1-effective broad sarbecovirus neutralizing antibodies via novel mutations T376A, D405N, and R408S. These sites were highly conserved across sarbecoviruses, suggesting that Omicron BA.2 arose from immune pressure selection instead of zoonotic spillover. Moreover, BA.2 reduces the efficacy of S309 (Sotrovimab)3,4 and broad sarbecovirus neutralizing antibodies targeting the similar epitope region, including BD55-5840. Structural comparisons of BD55-5840 in complexes with BA.1 and BA.2 spike suggest that BA.2 could hinder antibody binding through S371F-induced N343-glycan displacement. Intriguingly, the absence of G446S mutation in BA.2 enabled a proportion of 440-449 linear epitope targeting antibodies to retain neutralizing efficacy, including COV2-2130 (Cilgavimab)5. Together, we showed that BA.2 exhibits distinct antigenicity compared to BA.1 and provided a comprehensive profile of SARS-CoV-2 antibody escaping mutations. Our study offers critical insights into the humoral immune evading mechanism of current and future variants.
RESUMEN
The SARS-CoV-2 B.1.1.529 variant (Omicron) contains 15 mutations on the receptor-binding domain (RBD). How Omicron would evade RBD neutralizing antibodies (NAbs) requires immediate investigation. Here, we used high-throughput yeast display screening1,2 to determine the RBD escaping mutation profiles for 247 human anti-RBD NAbs and showed that the NAbs could be unsupervised clustered into six epitope groups (A-F), which is highly concordant with knowledge-based structural classifications3-5. Strikingly, various single mutations of Omicron could impair NAbs of different epitope groups. Specifically, NAbs in Group A-D, whose epitope overlap with ACE2-binding motif, are largely escaped by K417N, G446S, E484A, and Q493R. Group E (S309 site)6 and F (CR3022 site)7 NAbs, which often exhibit broad sarbecovirus neutralizing activity, are less affected by Omicron, but still, a subset of NAbs are escaped by G339D, N440K, and S371L. Furthermore, Omicron pseudovirus neutralization showed that single mutation tolerating NAbs could also be escaped due to multiple synergetic mutations on their epitopes. In total, over 85% of the tested NAbs are escaped by Omicron. Regarding NAb drugs, the neutralization potency of LY-CoV016/LY-CoV555, REGN10933/REGN10987, AZD1061/AZD8895, and BRII-196 were greatly reduced by Omicron, while VIR-7831 and DXP-604 still function at reduced efficacy. Together, data suggest Omicron would cause significant humoral immune evasion, while NAbs targeting the sarbecovirus conserved region remain most effective. Our results offer instructions for developing NAb drugs and vaccines against Omicron and future variants.
RESUMEN
Emergence of variants of concern (VOC) with altered antigenic structures and waning humoral immunity to SARS-CoV-2 are harbingers of a long pandemic. Administration of a third dose of an inactivated virus vaccine can boost the immune response. Here, we have dissected the immunogenic profiles of antibodies from 3-dose vaccinees, 2-dose vaccinees and convalescents. Better neutralization breadth to VOCs, expeditious recall and long-lasting humoral response bolster 3-dose vaccinees in warding off COVID-19. Analysis of 171 complex structures of SARS-CoV-2 neutralizing antibodies identified structure-activity correlates, revealing ultrapotent, VOCs-resistant and broad-spectrum antigenic patches. Construction of immunogenic and mutational heat maps revealed a direct relationship between "hot" immunogenic sites and areas with high mutation frequencies. Ongoing antibody somatic mutation, memory B cell clonal turnover and antibody composition changes in B cell repertoire driven by prolonged and repeated antigen stimulation confer development of monoclonal antibodies with enhanced neutralizing potency and breadth. Our findings rationalize the use of 3-dose immunization regimens for inactivated vaccines. One sentence summaryA third booster dose of inactivated vaccine produces a highly sifted humoral immune response via a sustained evolution of antibodies capable of effectively neutralizing SARS-CoV-2 variants of concern.
RESUMEN
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its emerging variants of concern (VOC), such as Delta (B.1.617.2) and Omicron (B.1.1.529), has continued to drive the worldwide pandemic. Therefore, there is a high demand for vaccines with enhanced efficacy, high thermostability, superior design flexibility, and fast manufacturing speed. Here, we report a circular RNA (circRNA) vaccine that encodes the trimeric RBD of SARS-CoV-2 Spike protein. Without the need of nucleotide modification, 5-capping or 3-polyadenylation, circRNA could be rapidly produced via in vitro transcription and is highly thermostable whether stored in naked or lipid-nanoparticle (LNP)-encapsulated format. LNP-encapsulated circRNARBD elicited potent neutralizing antibodies and T cell responses, providing robust protection against Beta (B.1.351) and native viruses in mice and rhesus macaques, respectively. Notably, circRNA vaccine enabled higher and more durable antigen production than 1m{Psi}-modified mRNA vaccine, eliciting a higher proportion of neutralizing antibodies and stronger Th1-biased immune responses. Importantly, we found that circRNARBD-Omicron vaccine induced effective neutralizing antibodies against only Omicron but not Delta variant. By contrast, circRNARBD-Delta could elicit high level of neutralizing antibodies against both Delta and Omicron. Following two doses of either native- or Delta-specific vaccination, circRNARBD-Delta, but not Omicron or Beta vaccines, could effectively boost the neutralizing antibodies against both Delta and Omicron variants. These results suggest that circRNARBD-Delta is a favorable choice for vaccination to provide a broad-spectrum protection against the current variants of concern of SARS-CoV-2.
RESUMEN
Understanding the mechanism of neutralizing antibodies (NAbs) against SARS-CoV-2 is critical for effective vaccines and therapeutics development. We recently reported an exceptionally potent NAb, BD-368-2, and revealed the existence of VH3-53/VH3-66 convergent NAbs in COVID-19. Here we report the 3.5-[A] cryo-EM structure of BD-368-2s Fabs in complex with a mutation-induced prefusion-state-stabilized spike trimer. Unlike VH3-53/VH3-66 NAbs, BD-368-2 fully blocks ACE2 binding by occupying all three receptor-binding domains (RBDs) simultaneously, regardless of their "up" and "down" positions. BD-368-2 also triggers fusogenic-like structural rearrangements of the spike trimer, which could impede viral entry. Moreover, BD-368-2 completely avoids the common epitope of VH3-53/VH3-66 NAbs, evidenced by multiple crystal structures of their Fabs in tripartite complexes with RBD, suggesting a new way of pairing potent NAbs to prevent neutralization escape. Together, these results rationalize a unique epitope that leads to exceptional neutralization potency, and provide guidance for NAb therapeutics and vaccine designs against SARS-CoV-2.
