Vaccination with SARS-CoV-2 variants of concern protects mice from challenge with wild-type virus

Vaccines against Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) have been highly efficient in protecting against Coronavirus Disease 2019 (COVID-19). However, the emergence of viral variants that are more transmissible and, in some cases, escape from neutralizing antibody responses has raised concerns. Here, we evaluated recombinant protein spike antigens derived from wild-type SARS-CoV-2 and from variants B.1.1.7, B.1.351, and P.1 for their immunogenicity and protective effect in vivo against challenge with wild-type SARS-CoV-2 in the mouse model. All proteins induced high neutralizing antibodies against the respective viruses but also induced high cross-neutralizing antibody responses. The decline in neutralizing titers between variants was moderate, with B.1.1.7-vaccinated animals having a maximum fold reduction of 4.8 against B.1.351 virus. P.1 induced the most cross-reactive antibody responses but was also the least immunogenic in terms of homologous neutralization titers. However, all antigens protected from challenge with wild-type SARS-CoV-2 in a mouse model.

This study explores the immune response induced by wild type and variant SARS-CoV-2 spike proteins, and the protection that these immune responses provide against challenge with wild type virus in the mouse model.     

新型冠状病毒及泛β冠状病毒广谱中和单抗的研究进展

新型冠状病毒的全球大流行,给人类的生命健康和社会秩序带来了巨大的危害。疫苗、小分子药物及各类抗体药物的研发在遏制新型冠状病毒感染传播、降低重症率和死亡风险上发挥了积极的作用。然而,由于新冠病毒庞大的感染基数及自身易突变的特征,当前已经演化出多种能逃逸疫苗及中和抗体的变异株,显著削弱了抗体的保护效果。研发新型冠状病毒广谱甚至泛β冠状病毒广谱的中和抗体对于未来新冠变异株及其他高致病性β冠状病毒的防治具有重要意义。本文从新型冠状病毒中和抗体的筛选制备策略、作用机制、中和效果及广谱性等方面进行了系统综述,并对当前面临的挑战和未来的发展方向进行了讨论和展望,以期为后续相关研究提供参考。     

Nasal delivery of broadly neutralizing antibodies protects mice from lethal challenge with SARS-CoV-2 delta and omicron variants

Multiple new variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have constantly emerged, as the delta and omicron variants, which have developed resistance to currently gained neutralizing antibodies. This highlights a critical need to discover new therapeutic agents to overcome the variants mutations. Despite the availability of vaccines against coronavirus disease 2019 (COVID-19), the use of broadly neutralizing antibodies has been considered as an alternative way for the prevention or treatment of SARS-CoV-2 variants infection. Here, we show that the nasal delivery of two previously characterized broadly neutralizing antibodies (F61 and H121) protected K18-hACE2 mice against lethal challenge with SARS-CoV-2 variants. The broadly protective efficacy of the F61 or F61/F121 cocktail antibodies was evaluated by lethal challenge with the wild strain (WIV04) and multiple variants, including beta (B.1.351), delta (B.1.617.2), and omicron (B.1.1.529) at 200 or 1000 TCID50, and the minimum antibody administration doses (5-1.25 mg/kg body weight) were also evaluated with delta and omicron challenge. Fully prophylactic protections were found in all challenged groups with both F61 and F61/H121 combination at the administration dose of 20 mg/kg body weight, and corresponding mice lung viral RNA showed negative, with almost all alveolar septa and cavities remaining normal. Furthermore, low-dose antibody treatment induced significant prophylactic protection against lethal challenge with delta and omicron variants, whereas the F61/H121 combination showed excellent results against omicron infection. Our findings indicated the potential use of broadly neutralizing monoclonal antibodies as prophylactic and therapeutic agent for protection of current emerged SARS-CoV-2 variants infection.     

Mutational escape prevention by combination of four neutralizing antibodies that target RBD conserved regions and stem helix

New variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) appear rapidly every few months. They have showed powerful adaptive ability to circumvent the immune system. To further understand SARS-CoV-2's adaptability so as to seek for strategies to mitigate the emergence of new variants, herein we investigated the viral adaptation in the presence of broadly neutralizing antibodies and their combinations. First, we selected four broadly neutralizing antibodies, including pan-sarbecovirus and pan-betacoronavirus neutralizing antibodies that recognize distinct conserved regions on receptor-binding domain (RBD) or conserved stem-helix region on S2 subunit. Through binding competition analysis, we demonstrated that they were capable of simultaneously binding. Thereafter, a replication-competent vesicular stomatitis virus pseudotyped with SARS-CoV-2 spike protein was employed to study the viral adaptation. Twenty consecutive passages of the virus under the selective pressure of individual antibodies or their combinations were performed. It was found that it was not hard for the virus to adapt to broadly neutralizing antibodies, even for pan-sarbecovirus and pan-betacoronavirus antibodies. The virus was more and more difficult to escape the combinations of two/three/four antibodies. In addition, mutations in the viral population revealed by high-throughput sequencing showed that under the selective pressure of three/four combinational antibodies, viral mutations were not prone to present in the highly conserved region across betacoronaviruses (stem-helix region), while this was not true under the selective pressure of single/two antibodies. Importantly, combining neutralizing antibodies targeting RBD conserved regions and stem helix synergistically prevented the emergence of escape mutations. These studies will guide future vaccine and therapeutic development efforts and provide a rationale for the design of RBD-stem helix tandem vaccine, which may help to impede the generation of novel variants.     

Control of SARS-CoV-2 infection after Spike DNA or Spike DNA+Protein co-immunization in rhesus macaques

The speed of development, versatility and efficacy of mRNA-based vaccines have been amply demonstrated in the case of SARS-CoV-2. DNA vaccines represent an important alternative since they induce both humoral and cellular immune responses in animal models and in human trials. We tested the immunogenicity and protective efficacy of DNA-based vaccine regimens expressing different prefusion-stabilized Wuhan-Hu-1 SARS-CoV-2 Spike antigens upon intramuscular injection followed by electroporation in rhesus macaques. Different Spike DNA vaccine regimens induced antibodies that potently neutralized SARS-CoV-2 in vitro and elicited robust T cell responses. The antibodies recognized and potently neutralized a panel of different Spike variants including Alpha, Delta, Epsilon, Eta and A.23.1, but to a lesser extent Beta and Gamma. The DNA-only vaccine regimens were compared to a regimen that included co-immunization of Spike DNA and protein in the same anatomical site, the latter of which showed significant higher antibody responses. All vaccine regimens led to control of SARS-CoV-2 intranasal/intratracheal challenge and absence of virus dissemination to the lower respiratory tract. Vaccine-induced binding and neutralizing antibody titers and antibody-dependent cellular phagocytosis inversely correlated with transient virus levels in the nasal mucosa. Importantly, the Spike DNA+Protein co-immunization regimen induced the highest binding and neutralizing antibodies and showed the strongest control against SARS-CoV-2 challenge in rhesus macaques.     

新型冠状病毒抗体和小分子抑制剂的研究现状

世界卫生组织已宣布新型冠状病毒感染（coronavirus disease 2019,COVID-19）的爆发为全球大流行。中和抗体和小分子抑制剂在预防及治疗COVID-19中发挥重要作用。尽管已开发出了多种中和抗体以及疫苗,但是随着病原体严重急性呼吸综合征冠状病毒2(severe acute respiratory syndrome coronavirus 2,SARS-CoV-2)的不断变异,现有的抗体及疫苗面临巨大的挑战。小分子抑制剂主要通过干扰病毒与宿主的结合以及病毒自身的复制达到消灭病毒以及抑制病毒感染的作用,并且对SARS-CoV-2突变株具有广谱抑制作用,是当前研究的热点。近年来国内外学者对SARS-CoV-2的小分子抑制剂做了大量的研究工作,本文根据中和抗体识别的抗原表位以及小分子抑制剂的作用机制分别对用于预防及治疗COVID-19的中和抗体和小分子抑制剂进行综述,讨论其研究现状,并展望小分子抑制剂的应用前景,以期为该领域的进一步研究提供参考。     

Modeling coronavirus spike protein dynamics: implications for immunogenicity and immune escape

The ongoing COVID-19 pandemic is a global public health emergency requiring urgent development of efficacious vaccines. While concentrated research efforts have focused primarily on antibody-based vaccines that neutralize SARS-CoV-2, and several first-generation vaccines have either been approved or received emergency use authorization, it is forecasted that COVID-19 will become an endemic disease requiring updated second-generation vaccines. The SARS-CoV-2 surface spike (S) glycoprotein represents a prime target for vaccine development because antibodies that block viral attachment and entry, i.e., neutralizing antibodies, bind almost exclusively to the receptor-binding domain. Here, we develop computational models for a large subset of S proteins associated with SARS-CoV-2, implemented through coarse-grained elastic network models and normal mode analysis. We then analyze local protein domain dynamics of the S protein systems and their thermal stability to characterize structural and dynamical variability among them. These results are compared against existing experimental data and used to elucidate the impact and mechanisms of SARS-CoV-2 S protein mutations and their associated antibody binding behavior. We construct a SARS-CoV-2 antigenic map and offer predictions about the neutralization capabilities of antibody and S mutant combinations based on protein dynamic signatures. We then compare SARS-CoV-2 S protein dynamics to SARS-CoV and MERS-CoV S proteins to investigate differing antibody binding and cellular fusion mechanisms that may explain the high transmissibility of SARS-CoV-2. The outbreaks associated with SARS-CoV, MERS-CoV, and SARS-CoV-2 over the last two decades suggest that the threat presented by coronaviruses is ever-changing and long term. Our results provide insights into the dynamics-driven mechanisms of immunogenicity associated with coronavirus S proteins and present a new, to our knowledge, approach to characterize and screen potential mutant candidates for immunogen design, as well as to characterize emerging natural variants that may escape vaccine-induced antibody responses.     

De novo design and Rosetta-based assessment of high-affinity antibody variable regions (Fv) against the SARS-CoV-2 spike receptor binding domain (RBD)

The continued emergence of new SARS-CoV-2 variants has accentuated the growing need for fast and reliable methods for the design of potentially neutralizing antibodies (Abs) to counter immune evasion by the virus. Here, we report on the de novo computational design of high-affinity Ab variable regions (Fv) through the recombination of VDJ genes targeting the most solvent-exposed hACE2-binding residues of the SARS-CoV-2 spike receptor binding domain (RBD) protein using the software tool OptMAVEn-2.0. Subsequently, we carried out computational affinity maturation of the designed variable regions through amino acid substitutions for improved binding with the target epitope. Immunogenicity of designs was restricted by preferring designs that match sequences from a 9-mer library of “human Abs” based on a human string content score. We generated 106 different antibody designs and reported in detail on the top five that trade-off the greatest computational binding affinity for the RBD with human string content scores. We further describe computational evaluation of the top five designs produced by OptMAVEn-2.0 using a Rosetta-based approach. We used Rosetta SnugDock for local docking of the designs to evaluate their potential to bind the spike RBD and performed “forward folding” with DeepAb to assess their potential to fold into the designed structures. Ultimately, our results identified one designed Ab variable region, P1.D1, as a particularly promising candidate for experimental testing. This effort puts forth a computational workflow for the de novo design and evaluation of Abs that can quickly be adapted to target spike epitopes of emerging SARS-CoV-2 variants or other antigenic targets.     

Structural basis for SARS-CoV-2 neutralizing antibodies with novel binding epitopes

The ongoing Coronavirus Disease 2019 (COVID-19) pandemic caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) threatens global public health and economy unprecedentedly, requiring accelerating development of prophylactic and therapeutic interventions. Molecular understanding of neutralizing antibodies (NAbs) would greatly help advance the development of monoclonal antibody (mAb) therapy, as well as the design of next generation recombinant vaccines. Here, we applied H2L2 transgenic mice encoding the human immunoglobulin variable regions, together with a state-of-the-art antibody discovery platform to immunize and isolate NAbs. From a large panel of isolated antibodies, 25 antibodies showed potent neutralizing activities at sub-nanomolar levels by engaging the spike receptor-binding domain (RBD). Importantly, one human NAb, termed PR1077, from the H2L2 platform and 2 humanized NAb, including PR953 and PR961, were further characterized and subjected for subsequent structural analysis. High-resolution X-ray crystallography structures unveiled novel epitopes on the receptor-binding motif (RBM) for PR1077 and PR953, which directly compete with human angiotensin-converting enzyme 2 (hACE2) for binding, and a novel non-blocking epitope on the neighboring site near RBM for PR961. Moreover, we further tested the antiviral efficiency of PR1077 in the Ad5-hACE2 transduction mouse model of COVID-19. A single injection provided potent protection against SARS-CoV-2 infection in either prophylactic or treatment groups. Taken together, these results shed light on the development of mAb-related therapeutic interventions for COVID-19.

This study characterizes novel neutralizing antibodies against the SARS-CoV-2 spike protein. Co-crystal structures of the spike protein receptor-binding domain and humanised mouse antibodies identify novel epitopes on the spike protein; binding to these epitopes competes with the ACE2 receptor, and one of the antibodies provides protection against SARS-CoV-2 infection in a mouse model of COVID-19.     

Humoral Immunity against SARS-CoV-2 and the Impact on COVID-19 Pathogenesis

It has been more than a year since severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) first emerged. Many studies have provided insights into the various aspects of the immune response in coronavirus disease 2019 (COVID-19). Especially for antibody treatment and vaccine development, humoral immunity to SARS-CoV-2 has been studied extensively, though there is still much that is unknown and controversial. Here, we introduce key discoveries on the humoral immune responses in COVID-19, including the immune dynamics of antibody responses and correlations with disease severity, neutralizing antibodies and their cross-reactivity, how long the antibody and memory B-cell responses last, aberrant autoreactive antibodies generated in COVID-19 patients, and the efficacy of currently available therapeutic antibodies and vaccines against circulating SARS-CoV-2 variants, and highlight gaps in the current knowledge.     

The humoral and cellular immune evasion of SARS-CoV-2 Omicron and sub-lineages

The recently discovered SARS-CoV-2 variant Omicron (B.1.1.529) has rapidly become a global public health issue. The substantial mutations in the spike protein in this new variant have raised concerns about its ability to escape from pre-existing immunity established by natural infection or vaccination. In this review, we give a summary of current knowledge concerning the antibody evasion properties of Omicron and its subvariants (BA.2, BA.2.12.1, BA.4/5, and BA.2.75) from therapeutic monoclonal antibodies and the sera of SARS-CoV-2 vaccine recipients or convalescent patients. We also summarize whether vaccine-induced cellular immunity (memory B cell and T cell response) can recognize Omicron specifically. In brief, the Omicron variants demonstrated remarkable antibody evasion, with even more striking antibody escape seen in the Omicron BA.4 and BA.5 sub-lineages. Luckily, the third booster vaccine dose significantly increased the neutralizing antibodies titers, and the vaccine-induced cellular response remains conserved and provides second-line defense against the Omicron.     

Specificity tuning of antibody fragments to neutralize two human chemokines with a single agent

Chemokines are important mediators of the immune response that are responsible for the trafficking of immune cells between lymphoid organs and migration towards sites of inflammation. Using phage display selection and a functional screening approach, we have isolated a panel of single-chain fragment variable (scFv) capable of neutralizing the activity of the human chemokine CXCL10 (hCXCL10). One of the isolated scFv was weakly cross-reactive against another human chemokine CXCL9, but was unable to block its biological activity. We diversified the complementarity determining region 3 (CDR3) of the light chain variable domain (VL) of this scFv and combined phage display with high throughput antibody array screening to identify variants capable of neutralizing both chemokines. Using this approach it is therefore possible to engineer pan-specific antibodies that could prove very useful to antagonize redundant signaling pathways such as the chemokine signaling network.Key words: cross-reactive antibody, antibody arrays, chemotaxis, multiple targeting, affinity maturation, phage display     

Humoral immune response to circulating SARS-CoV-2 variants elicited by inactivated and RBD-subunit vaccines

SARS-CoV-2 variants could induce immune escape by mutations on the receptor-binding domain (RBD) and N-terminal domain (NTD). Here we report the humoral immune response to circulating SARS-CoV-2 variants, such as 501Y.V2 (B.1.351), of the plasma and neutralizing antibodies (NAbs) elicited by CoronaVac (inactivated vaccine), ZF2001 (RBD-subunit vaccine) and natural infection. Among 86 potent NAbs identified by high-throughput single-cell VDJ sequencing of peripheral blood mononuclear cells from vaccinees and convalescents, near half anti-RBD NAbs showed major neutralization reductions against the K417N/E484K/N501Y mutation combination, with E484K being the dominant cause. VH3-53/VH3-66 recurrent antibodies respond differently to RBD variants, and K417N compromises the majority of neutralizing activity through reduced polar contacts with complementarity determining regions. In contrast, the 242–244 deletion (242–244Δ) would abolish most neutralization activity of anti-NTD NAbs by interrupting the conformation of NTD antigenic supersite, indicating a much less diversity of anti-NTD NAbs than anti-RBD NAbs. Plasma of convalescents and CoronaVac vaccinees displayed comparable neutralization reductions against pseudo- and authentic 501Y.V2 variants, mainly caused by E484K/N501Y and 242–244Δ, with the effects being additive. Importantly, RBD-subunit vaccinees exhibit markedly higher tolerance to 501Y.V2 than convalescents, since the elicited anti-RBD NAbs display a high diversity and are unaffected by NTD mutations. Moreover, an extended gap between the third and second doses of ZF2001 leads to better neutralizing activity and tolerance to 501Y.V2 than the standard three-dose administration. Together, these results suggest that the deployment of RBD-vaccines, through a third-dose boost, may be ideal for combating SARS-CoV-2 variants when necessary, especially for those carrying mutations that disrupt the NTD supersite.Subject terms: Immunology, Cryoelectron microscopy, X-ray crystallography     

A protease-activatable luminescent biosensor and reporter cell line for authentic SARS-CoV-2 infection

Efforts to define serological correlates of protection against COVID-19 have been hampered by the lack of a simple, scalable, standardised assay for SARS-CoV-2 infection and antibody neutralisation. Plaque assays remain the gold standard, but are impractical for high-throughput screening. In this study, we show that expression of viral proteases may be used to quantitate infected cells. Our assays exploit the cleavage of specific oligopeptide linkers, leading to the activation of cell-based optical biosensors. First, we characterise these biosensors using recombinant SARS-CoV-2 proteases. Next, we confirm their ability to detect viral protease expression during replication of authentic virus. Finally, we generate reporter cells stably expressing an optimised luciferase-based biosensor, enabling viral infection to be measured within 24 h in a 96- or 384-well plate format, including variants of concern. We have therefore developed a luminescent SARS-CoV-2 reporter cell line, and demonstrated its utility for the relative quantitation of infectious virus and titration of neutralising antibodies.     

Structural and antigenic variations in the spike protein of emerging SARS-CoV-2 variants

The Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) virus is continuously evolving, and this poses a major threat to antibody therapies and currently authorized Coronavirus Disease 2019 (COVID-19) vaccines. It is therefore of utmost importance to investigate and predict the putative mutations on the spike protein that confer immune evasion. Antibodies are key components of the human immune system’s response to SARS-CoV-2, and the spike protein is a prime target of neutralizing antibodies (nAbs) as it plays critical roles in host cell recognition, fusion, and virus entry. The potency of therapeutic antibodies and vaccines partly depends on how readily the virus can escape neutralization. Recent structural and functional studies have mapped the epitope landscape of nAbs on the spike protein, which illustrates the footprints of several nAbs and the site of escape mutations. In this review, we discuss (1) the emerging SARS-CoV-2 variants; (2) the structural basis for antibody-mediated neutralization of SARS-CoV-2 and nAb classification; and (3) identification of the RBD escape mutations for several antibodies that resist antibody binding and neutralization. These escape maps are a valuable tool to predict SARS-CoV-2 fitness, and in conjunction with the structures of the spike-nAb complex, they can be utilized to facilitate the rational design of escape-resistant antibody therapeutics and vaccines.     

Receptor binding,immune escape,and protein stability direct the natural selection of SARS-CoV-2 variants

Emergence of new severe acute respiratory syndrome coronavirus 2 variants has raised concerns related to the effectiveness of vaccines and antibody therapeutics developed against the unmutated wildtype virus. Here, we examined the effect of the 12 most commonly occurring mutations in the receptor-binding domain of the spike protein on its expression, stability, activity, and antibody escape potential. Stability was measured using thermal denaturation, and the activity and antibody escape potential were measured using isothermal titration calorimetry in terms of binding to the human angiotensin-converting enzyme 2 and to neutralizing human antibody CC12.1, respectively. Our results show that mutants differ in their expression levels. Of the eight best-expressed mutants, two (N501Y and K417T/E484K/N501Y) showed stronger affinity to angiotensin-converting enzyme 2 compared with the wildtype, whereas four (Y453F, S477N, T478I, and S494P) had similar affinity and two (K417N and E484K) had weaker affinity than the wildtype. Compared with the wildtype, four mutants (K417N, Y453F, N501Y, and K417T/E484K/N501Y) had weaker affinity for the CC12.1 antibody, whereas two (S477N and S494P) had similar affinity, and two (T478I and E484K) had stronger affinity than the wildtype. Mutants also differ in their thermal stability, with the two least stable mutants showing reduced expression. Taken together, these results indicate that multiple factors contribute toward the natural selection of variants, and all these factors need to be considered to understand the evolution of the virus. In addition, since not all variants can escape a given neutralizing antibody, antibodies to treat new variants can be chosen based on the specific mutations in that variant.     

DNA vaccination induced protective immunity against SARS CoV-2 infection in hamsterss

The development of efficient vaccines against COVID-19 is an emergent need for global public health. The spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a major target for the COVID-19 vaccine. To quickly respond to the outbreak of the SARS-CoV-2 pandemic, a nucleic acid-based vaccine is a novel option, beyond the traditional inactivated virus vaccine or recombinant protein vaccine. Here, we report a DNA vaccine containing the spike gene for delivery via electroporation. The spike genes of SARS-CoV and SARS-CoV-2 were codon optimized for mammalian cell expression and then cloned into mammalian cell expression vectors, called pSARS-S and pSARS2-S, respectively. Spike protein expression was confirmed by immunoblotting after transient expression in HEK293T cells. After immunization, sera were collected for antigen-specific antibody and neutralizing antibody titer analyses. We found that both pSARS-S and pSARS2-S immunization induced similar levels of antibodies against S2 of SARS-CoV-2. In contrast, only pSARS2-S immunization induced antibodies against the receptor-binding domain of SARS-CoV-2. We further found that pSARS2-S immunization, but not pSARS-S immunization, could induce very high titers of neutralizing antibodies against SARS-CoV-2. We further analyzed SARS-CoV-2 S protein-specific T cell responses and found that the immune responses were biased toward Th1. Importantly, pSARS2-S immunization in hamsters could induce protective immunity against SARS-CoV-2 challenge in vivo. These data suggest that DNA vaccination could be a promising approach for protecting against COVID-19.     

Structural and energetic profiling of SARS-CoV-2 receptor binding domain antibody recognition and the impact of circulating variants

The SARS-CoV-2 pandemic highlights the need for a detailed molecular understanding of protective antibody responses. This is underscored by the emergence and spread of SARS-CoV-2 variants, including Alpha (B.1.1.7) and Delta (B.1.617.2), some of which appear to be less effectively targeted by current monoclonal antibodies and vaccines. Here we report a high resolution and comprehensive map of antibody recognition of the SARS-CoV-2 spike receptor binding domain (RBD), which is the target of most neutralizing antibodies, using computational structural analysis. With a dataset of nonredundant experimentally determined antibody-RBD structures, we classified antibodies by RBD residue binding determinants using unsupervised clustering. We also identified the energetic and conservation features of epitope residues and assessed the capacity of viral variant mutations to disrupt antibody recognition, revealing sets of antibodies predicted to effectively target recently described viral variants. This detailed structure-based reference of antibody RBD recognition signatures can inform therapeutic and vaccine design strategies.     

Cross-neutralization of SARS-CoV-2 by HIV-1 specific broadly neutralizing antibodies and polyclonal plasma

Cross-reactive epitopes (CREs) are similar epitopes on viruses that are recognized or neutralized by same antibodies. The S protein of SARS-CoV-2, similar to type I fusion proteins of viruses such as HIV-1 envelope (Env) and influenza hemagglutinin, is heavily glycosylated. Viral Env glycans, though host derived, are distinctly processed and thereby recognized or accommodated during antibody responses. In recent years, highly potent and/or broadly neutralizing human monoclonal antibodies (bnAbs) that are generated in chronic HIV-1 infections have been defined. These bnAbs exhibit atypical features such as extensive somatic hypermutations, long complementary determining region (CDR) lengths, tyrosine sulfation and presence of insertions/deletions, enabling them to effectively neutralize diverse HIV-1 viruses despite extensive variations within the core epitopes they recognize. As some of the HIV-1 bnAbs have evolved to recognize the dense viral glycans and cross-reactive epitopes (CREs), we assessed if these bnAbs cross-react with SARS-CoV-2. Several HIV-1 bnAbs showed cross-reactivity with SARS-CoV-2 while one HIV-1 CD4 binding site bnAb, N6, neutralized SARS-CoV-2. Furthermore, neutralizing plasma antibodies of chronically HIV-1 infected children showed cross neutralizing activity against SARS-CoV-2 pseudoviruses. Collectively, our observations suggest that human monoclonal antibodies tolerating extensive epitope variability can be leveraged to neutralize pathogens with related antigenic profile.     

Hepatitis C virus (HCV)-induced immunoglobulin hypermutation reduces the affinity and neutralizing activities of antibodies against HCV envelope protein

Hepatitis C virus (HCV) often causes persistent infection despite the presence of neutralizing antibodies against the virus in the sera of hepatitis C patients. HCV infects both hepatocytes and B cells through the binding of its envelope glycoprotein E2 to CD81, the putative viral receptor. Previously, we have shown that E2-CD81 interaction induces hypermutation of heavy-chain immunoglobulin (V(H)) in B cells. We hypothesize that if HCV infects antibody-producing B cells, the resultant hypermutation of V(H) may lower the affinity and specificity of the HCV-specific antibodies, enabling HCV to escape from immune surveillance. To test this hypothesis, we infected human hybridoma clones producing either neutralizing or non-neutralizing anti-E2 or anti-E1 antibodies with a lymphotropic HCV (SB strain). All of the hybridoma clones, except for a neutralizing antibody-producing hybridoma, could be infected with HCV and support virus replication for at least 8 weeks after infection. The V(H) sequences in the infected hybridomas had a significantly higher mutation frequency than those in the uninfected hybridomas, with mutations concentrating in complementarity-determining region 3. These mutations lowered the antibody affinity against the targeting protein and also lowered the virus-neutralizing activity of anti-E2 antibodies. Furthermore, antibody-mediated complement-dependent cytotoxicity with the antibodies secreted from the HCV-infected hybridomas was impaired. These results suggest that HCV infection could cause some anti-HCV-antibody-producing hybridoma B cells to make less-protective antibodies.