The ongoing emergence of new variants of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) underscores the urgent need for developing antivirals targeting both SARS-CoV-2 variants and related sarbecoviruses. To this end, we designed novel trispecific antibodies, Tri-1 and Tri-2, engineered by fusing the single-chain variable fragments (scFvs) of a potent antibody (PW5-570) to the Fc region of "Knob-into-Hole" bispecific antibodies (bsAbs) composed of two distinct broad antibodies (PW5-5 and PW5-535). Compared with the parental antibodies, Tri-1 and Tri-2 displayed enhanced binding affinities to the receptor-binding domains of SARS-CoV, SARS-CoV-2 wild type, and Omicron XBB.1.16, with each arm contributed to the overall enhancement. Furthermore, pseudovirus neutralization assays revealed that Tri-1 and Tri-2 effectively neutralized all tested SARS-CoV, SARS-CoV-2 variants, and related sarbecoviruses (Pangolin-GD, RaTG13, WIV1, and SHC014), demonstrating potency and breadth superior to any single parental antibody. Consistently, Tri-1 and Tri-2 were found to effectively neutralize authentic SARS-CoV and SARS-CoV-2 variants with IC50 values comparable to or better than those of parental antibodies. Taken together, this study highlights the potential effectiveness of Tri-1 and Tri-2 as novel formats for harnessing cross-neutralizing antibodies in the development of multivalent agents to combat both current and future SARS-like coronaviruses.

The global impact of emerging and re-emerging viral agents during epidemics and pandemics leads to serious health and economic burdens. Among the major emerging or re-emerging viruses include SARS-CoV-2, Ebola virus (EBOV), Monkeypox virus (Mpox), Hepatitis viruses, Zika virus, Avian flu, Influenza virus, Chikungunya virus (CHIKV), Dengue fever virus (DENV), West Nile virus, Rhabdovirus, Sandfly fever virus, Crimean-Congo hemorrhagic fever (CCHF) virus, and Rift Valley fever virus (RVFV). A comprehensive literature search was performed to identify existing studies, clinical trials, and reviews that discuss drug repositioning strategies for the treatment of emerging and re-emerging viral infections using databases, such as PubMed, Scholar Google, Scopus, and Web of Science. By utilizing drug repositioning, pharmaceutical companies can take advantage of a cost-effective, accelerated, and effective strategy, which in turn leads to the discovery of innovative treatment options for patients. In light of antiviral drug resistance and the high costs of developing novel antivirals, drug repositioning holds great promise for more rapid substitution of approved drugs. Main repositioned drugs have included chloroquine, ivermectin, dexamethasone, Baricitinib, tocilizumab, Mab114 (Ebanga™), ZMapp (pharming), Artesunate, imiquimod, saquinavir, capmatinib, naldemedine, Trametinib, statins, celecoxib, naproxen, metformin, ruxolitinib, nitazoxanide, gemcitabine, Dorzolamide, Midodrine, Diltiazem, zinc acetate, suramin, 5-fluorouracil, quinine, minocycline, trifluoperazine, paracetamol, berbamine, Nifedipine, and chlorpromazine. This succinct review will delve into the topic of repositioned drugs that have been utilized to combat emerging and re-emerging viral pathogens.

The continuous evolution of SARS-CoV-2, particularly the emergence of the BA.2.86/JN.1 lineage replacing XBB, necessitates re-evaluation of vaccine compositions1-3. Here, we provide a comprehensive analysis of the humoral immune response to XBB and JN.1 human exposure. We demonstrate the antigenic distinctiveness of XBB and JN.1 lineages in SARS-CoV-2-naive individuals and show that infection with JN.1 elicits superior plasma neutralization against its subvariants. We highlight the strong immune evasion and receptor-binding capability of KP.3, supporting its foreseeable prevalence. Extensive analysis of the B cell receptor repertoire, in which we isolate approximately 2,000 receptor-binding-domain-specific antibodies, with targeting epitopes characterized by deep mutational scanning, underscores the superiority of JN.1-elicited memory B cells4,5. Class 1 IGHV3-53/3-66-derived neutralizing antibodies (NAbs) are important contributors to the wild-type reactivity of NAbs against JN.1. However, KP.2 and KP.3 evade a substantial subset of these antibodies, even those induced by JN.1, supporting a need for booster updates. JN.1-induced Omicron-specific antibodies also demonstrate high potency across Omicron. Escape hotspots for these NAbs have already been mutated, resulting in a higher immune barrier to escape and indicating probable recovery of escaped NAbs. In addition, the prevalence of IGHV3-53/3-66-derived antibodies and their ability to compete with all Omicron-specific NAbs suggests that they have an inhibitory effect on the activation of Omicron-specific naive B cells, potentially explaining the heavy immune imprinting in mRNA-vaccinated individuals6-8. These findings delineate the evolving antibody response to the antigenic shift of Omicron from XBB to JN.1 and highlight the importance of developing the JN.1 lineage, especially KP.2- and KP.3-based vaccine boosters.

The continual emergence of highly pathogenic novel coronaviruses and their variants has underscored the importance of neutralizing monoclonal antibodies (mAbs) as a pivotal therapeutic approach. In the present study, we report the specific neutralizing antibodies 13H7 and 9G11, which target the N-terminal domain (NTD) and receptor-binding domain (RBD) of the SARS-CoV-2, respectively. The comparative analysis observed that 13H7 not only neutralizes early variants of concern (VOCs) but also exhibits neutralizing activity against the Omicron sublineage, including BA.4, BA.5, BQ.1, and BQ.1.1. 9G11, as an RBD antibody, also demonstrated remarkable neutralizing efficacy. A cocktail antibody combining 13H7 and 9G11 with the previously reported 3E2 broaden the neutralization spectrum against new variants of the SARS-CoV-2. We elucidated the cryo-EM structure of the complex, clarifying the mechanism of action of the cocktail antibody combination. Additionally, we also emphasized the molecular mechanism between 13H7 and SARS-CoV-2 NTD, revealing the impact of Y144 and H146 residue deletions and mutations on the neutralizing efficacy of 13H7. Taken together, our findings not only offer novel insights into the combination therapy of NTD and RBD neutralizing mAbs but also lay a theoretical foundation for the development of vaccines targeting NTD antibodies, thus providing valuable understanding of alleviating the emergence of SARS-CoV-2 variants.

The recent emergence of a SARS-CoV-2 saltation variant, BA.2.87.1, which features 65 spike mutations relative to BA.2, has attracted worldwide attention. In this study, we elucidate the antigenic characteristics and immune evasion capability of BA.2.87.1. Our findings reveal that BA.2.87.1 is more susceptible to XBB-induced humoral immunity compared to JN.1. Notably, BA.2.87.1 lacks critical escaping mutations in the receptor binding domain (RBD) thus allowing various classes of neutralizing antibodies (NAbs) that were escaped by XBB or BA.2.86 subvariants to neutralize BA.2.87.1, although the deletions in the N-terminal domain (NTD), specifically 15-23del and 136-146del, compensate for the resistance to humoral immunity. Interestingly, several neutralizing antibody drugs have been found to restore their efficacy against BA.2.87.1, including SA58, REGN-10933 and COV2-2196. Hence, our results suggest that BA.2.87.1 may not become widespread until it acquires multiple RBD mutations to achieve sufficient immune evasion comparable to that of JN.1.

Neutralizing antibodies correlate with protection against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Recent studies, however, show that binding antibody titers, in the absence of robust neutralizing activity, also correlate with protection against disease progression. Non-neutralizing antibodies cannot directly protect against infection but may recruit effector cells and thus contribute to the clearance of infected cells. Additionally, they often bind conserved epitopes across multiple variants. Here, we characterize 42 human monoclonal antibodies (mAbs) from coronavirus disease 2019 (COVID-19)-vaccinated individuals. Most of these antibodies exhibit no neutralizing activity in vitro, but several non-neutralizing antibodies provide protection against lethal challenge with SARS-CoV-2 in different animal models. A subset of those mAbs shows a clear dependence on Fc-mediated effector functions. We have determined the structures of three non-neutralizing antibodies, with two targeting the receptor-binding domain and one that binds the subdomain 1 region. Our data confirm the real-world observation in humans that non-neutralizing antibodies to SARS-CoV-2 can be protective.

Increased use of antiviral monoclonal antibodies (mAbs) for treatment and prophylaxis necessitates better understanding of their impact on endogenous immunity to vaccines and viruses. The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic presented an opportunity to study immunity in individuals who received antiviral mAbs and were subsequently immunized with vaccines encoding the mAb-targeted viral spike antigen. Here, we describe the impact of administration of an antibody combination, casirivimab plus imdevimab (CAS+IMD), on immune responses to subsequent SARS-CoV-2 vaccination in humans, nonhuman primates, and mice. The presence of CAS+IMD at the time of vaccination led to a specific diminishment of vaccine-elicited pseudovirus neutralizing antibody titers without overall dampening of spike protein-directed immune responses, including antibody, B cell, and T cell responses. The impact on pseudovirus neutralizing titers extended to other therapeutic anti-spike protein antibodies when used as either monotherapy or combination therapy. The specific reduction in pseudovirus neutralizing titers was the result of epitope masking, a phenomenon where specific epitopes are bound by high-affinity antibodies and blocked from B cell recognition. Encouragingly, this reduction in pseudovirus neutralizing titers was reversible with additional booster vaccination. Moreover, by assessing the antiviral immune response in SARS-CoV-2-infected individuals treated therapeutically with CAS+IMD, we demonstrated alteration of antiviral humoral immunity in those who had received mAb therapy, but only in those individuals who had yet to start mounting their natural immune response at the time of mAb treatment. Together, these data demonstrate that antiviral mAbs can alter endogenous humoral immunity during vaccination or infection.

The COVID-19 epidemic has become a major international health emergency. Millions of people have died as a result of this phenomenon since it began. Has there been any successful pharmacological treatment for COVID-19 since the initial report on the virus? How many searches are undertaken to address the impact of the infection? What is the number of drugs that have undergone investigation? What are the mechanisms of action and adverse effects associated with the investigated pharmaceuticals used to treat COVID-19? Has the Food and Drug Administration (FDA) approved any medication to treat COVID-19? To date, our understanding is based on a restricted corpus of published investigations into the treatment of COVID-19. It is important to note that no single study comprehensively encompasses all pharmacological interventions for COVID-19. This paper provides an introductory summary of a bibliometric analysis conducted on the data about COVID-19, sourced explicitly from two platforms, namely PubMed and ScienceDirect. The analysis encompasses the period spanning from 2019 to 2022. Furthermore, this study examines the published literature about the pharmacological interventions for the novel coronavirus disease 2019 (COVID-19), explicitly focusing on the safety and effectiveness of different medications such as Remdesivir (marketed as Veklury®), Lopinavir/Ritonavir (commercially known as Kaletra® or Aluvia®), Ribavirin, Favipiravir (marketed as Avigan®), Ivermectin, Casirivimab and Imdevimab (branded as Ronapreve®), Sotrovimab (marketed as Xevudy®), Anakinra, Molnupiravir, Nirmatrelvir/Ritonavir (marketed as Paxlovid®), and Galidesivir. Findings indicate that while Remdesivir and Nirmatrelvir/Ritonavir show significant efficacy in reducing hospitalization and severe outcomes, drugs like Lopinavir/Ritonavir and Ivermectin have inconsistent results. Our insights suggest a multifaceted approach incorporating these therapies can significantly improve patient outcomes. Repurposing drugs has been critical in rapidly responding to COVID-19, allowing existing medications to be used in new ways to combat the virus. Combination therapies and further research are essential to optimize treatment strategies.

Protein aggregation plays key roles in age-related degenerative diseases, but how different proteins coalesce to form inclusions that vary in composition, morphology, molecular dynamics and confer physiological consequences is poorly understood. Here we employ a general reporter based on mutant Hsp104 to identify proteins forming aggregates in human cells under common proteotoxic stress. We identify over 300 proteins that form different inclusions containing subsets of aggregating proteins. In particular, TDP43, implicated in Amyotrophic Lateral Sclerosis (ALS), partitions dynamically between two distinct types of aggregates: stress granule and a previously unknown non-dynamic (solid-like) inclusion at the ER exit sites (ERES). TDP43-ERES co-aggregation is induced by diverse proteotoxic stresses and observed in the motor neurons of ALS patients. Such aggregation causes retention of secretory cargos at ERES and therefore delays ER-to-Golgi transport, providing a link between TDP43 aggregation and compromised cellular function in ALS patients.

Pregnant women with COVID-19 are at elevated risk for severe outcomes, but clinical data on management of these patients are limited. Monoclonal antibodies, such as casirivimab plus imdevimab (CAS+IMD), have proven effective in treating non-pregnant adults with COVID-19, prompting further evaluation in pregnant women.

A phase 3 portion of an adaptive, multicentre, randomised, double-blind, placebo-controlled trial evaluated the safety, clinical outcomes, pharmacokinetics and immunogenicity of CAS+IMD (1200 mg or 2400 mg) in the treatment of pregnant outpatients with COVID-19 (NCT04425629). Participants were enrolled between December 2020 and November 2021, prior to the emergence of Omicron-lineage variants against which CAS+IMD is not active. Safety was evaluated in randomised participants who received study drug (n=80); clinical outcomes were evaluated in all randomised participants (n=82). Only two pregnant participants received placebo, limiting conclusions regarding treatment effect. Infants born to pregnant participants were followed for developmental outcomes ≤1 year of age.

In pregnant participants, CAS+IMD was well tolerated, with no grade ≥2 hypersensitivity or infusion-related reactions reported. There were no participant deaths, and only one COVID-19-related medically attended visit. Although two pregnancies (3%) reported issues in the fetus/neonate, they were confounded by maternal history or considered to be due to an alternate aetiology. No adverse developmental outcomes in infants ≤1 year of age were considered related to in utero exposure to the study drug. CAS+IMD 1200 mg and 2400 mg rapidly and similarly reduced viral loads, with a dose-proportional increase in concentrations of CAS+IMD in serum. Pharmacokinetics were consistent with that reported in the general population. Immunogenicity incidence was low.

CAS+IMD treatment of pregnant outpatients with COVID-19 showed similar safety, clinical outcomes and pharmacokinetic profiles to that observed in non-pregnant adults. There was no evidence of an impact on developmental outcomes in infants ≤1 year of age.

NCT04425629.

Recombinant SARS-CoV-2 S protein expression was examined in Vero cells by imaging using the human monoclonal antibody panel (PD4, PD5, sc23, and sc29). The PD4 and sc29 antibodies recognised conformational specific epitopes in the S2 protein subunit at the Endoplasmic reticulum and Golgi complex. While PD5 and sc23 detected conformationally specific epitopes in the S1 protein subunit at the Golgi complex, only PD5 recognised the receptor binding domain (RBD). A comparison of the staining patterns of PD5 with non-conformationally specific antibodies that recognises the S1 subunit and RBD suggested the PD5 recognised a conformational structure within the S1 protein subunit. Our data suggests the antibody binding epitopes recognised by the human monoclonal antibodies formed at different locations in the secretory pathway during S protein transport, but a conformational change in the S1 protein subunit at the Golgi complex formed antibody binding epitopes that are recognised by virus neutralising antibodies.

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) belongs to the genus Beta coronavirus and the family of Coronaviridae. It is a positive-sense, non-segmented single-strand RNA virus. Four common types of human coronaviruses circulate globally, particularly in the fall and winter seasons. They are responsible for 10%-30% of all mild upper respiratory tract infections in adults. These are 229E, NL63 of the Alfacoronaviridae family, OC43, and HKU1 of the Betacoronaviridae family. However, there are three highly pathogenic human coronaviruses: SARS-CoV-2, Middle East respiratory syndrome coronavirus, and the latest pandemic caused by the SARS-CoV-2 infection. All viruses, including SARS-CoV-2, have the inherent tendency to evolve. SARS-CoV-2 is still evolving in humans. Additionally, due to the development of herd immunity, prior infection, use of medication, vaccination, and antibodies, the viruses are facing immune pressure. During the replication process and due to immune pressure, the virus may undergo mutations. Several SARS-CoV-2 variants, including the variants of concern (VOCs), such as B.1.1.7 (Alpha), B.1.351 (Beta), B.1.617/B.1.617.2 (Delta), P.1 (Gamma), and B.1.1.529 (Omicron) have been reported from various parts of the world. These VOCs contain several important mutations; some of them are on the spike proteins. These mutations may lead to enhanced infectivity, transmissibility, and decreased neutralization efficacy by monoclonal antibodies, convalescent sera, or vaccines. Mutations may also lead to a failure of detection by molecular diagnostic tests, leading to a delayed diagnosis, increased community spread, and delayed treatment. We searched PubMed, EMBASE, Covariant, the Stanford variant Database, and the CINAHL from December 2019 to February 2023 using the following search terms: VOC, SARS-CoV-2, Omicron, mutations in SARS-CoV-2, etc. This review discusses the various mutations and their impact on infectivity, transmissibility, and neutralization efficacy.

Effective treatment and immunoprophylaxis of viral respiratory infections with neutralizing monoclonal antibodies (mAbs) require maintaining inhibitory concentrations of mAbs at the airway surface. While engineered mAbs with increased affinity to the neonatal Fc receptor (FcRn) are increasingly employed, little is known how increased affinity of Fc to FcRn influences basal-to-apical transepithelial transport (transcytosis) of mAbs across the airway epithelium. To investigate this, we utilized a model of well-differentiated human airway epithelium (WD-HAE) that exhibited robust FcRn expression, and measured the transepithelial transport of a mAb against SARS-CoV-2 Spike protein (CR3022) with either wildtype IgG1-Fc or Fc modified with YTE or LS mutations known to increase affinity for FcRn. Despite the marked differences in the affinity of these CR3022 variants for FcRn, we did not find substantial differences in basal-to-apical transport reflective of systemic dosing, or apical-to-basal transport reflective of inhaled dosing, compared to the transport of wildtype IgG1-Fc. These results suggest increasing FcRn affinity may only have limited influence over transcytosis rates of systemically dosed mAbs across the human airway epithelium over short time scales. Over longer time scales, the elevated circulating levels of mAbs with greater FcRn affinity, due to more effective FcRn-mediated recycling, may better resupply mAb into the respiratory tract, leading to more effective extended immunoprophylaxis.

Respiratory syncytial virus (RSV) is a leading cause of acute respiratory infection amongst all ages, causing a significant global health burden. Preventative and therapeutic options for RSV infection have long been under development, and recently, several widely-publicised vaccines targeting older adult and maternal populations have become available. Promising monoclonal antibody (mAb) and antiviral (AV) therapies are also progressing in clinical trials, with the prophylactic mAb nirsevimab recently approved for clinical use in infant populations. A systematic review on current progress in this area is lacking. We performed a systematic literature search (PubMed, Embase, Web of Science, ClinicalTrials.gov, EudraCT, ANZCTR-searched Nov 29th, 2023) to identify studies on all RSV-specific mAbs and AV therapies that has undergone human clinical trials since year 2000. Data extraction focused on outcomes related to the therapeutic efficacy and safety of the intervention on trial, and all studies were graded against the OCEBM Levels of Evidence Table. Results from 59 studies were extracted, covering efficacy and safety data on six mAbs (motavizumab, motavizumab-YTE, nirsevimab, ALX-0171, suptavumab, clesrovimab) and 12 AV therapies (ALN-RSV01, RSV604, presatovir, MDT-637, lumicitabine, IFN-α1b, rilematovir, enzaplatovir, AK0529, sisunatovir, PC786, EDP-938). Of the mAbs reviewed, nirsevimab and clesrovimab hold considerable promise. The timeline for RSV-specific AV availability is less advanced, although EDP-938 and AK0529 have reported promising phase 2 efficacy and safety data. Moving forward, passive immunisation and treatment options for RSV infection will play a significant role in reducing the health burden of RSV, complementing recent advancements in vaccine development. TRIAL REGISTRATION: PROSPERO registration: CRD42022376633.

This study aimed to assess the effects of a monoclonal antibody (mAb) combination on symptoms, daily function, and overall health-related quality of life.

We analyzed patient-reported outcomes data from symptomatic outpatients in a phase 1/2/3 trial. Patients with confirmed SARS-CoV-2 infection and ≥ 1 risk factor for severe COVID-19 received mAb treatment (casirivimab plus imdevimab 1200 mg) or placebo. Prespecified exploratory assessments included time to sustained symptoms resolution, usual health, and return to usual activities (assessed daily for 29 days). The trial was conducted from September 2020 to February 2021, prior to widespread COVID-19 vaccination programs and Omicron-lineage variants against which casirivimab + imdevimab is not active.

In this analysis 736 outpatients received mAb and 1341 received placebo. Median time to sustained symptoms resolution was consistently shorter with mAb versus placebo (≥ 2 consecutive days: 14 vs 17 days, [nominal p = 0.0017]; ≥ 3 consecutive days: 17 vs 21 days, [nominal p = 0.0046]). Median time to sustained return to usual health and usual activities were both consistently shorter with mAb versus placebo (≥ 2 consecutive days: 12 vs 15 days [nominal p = 0.0001] and 9 vs 11 days [nominal p = 0.0001], respectively; ≥ 3 consecutive days: 14 vs 18 days [nominal p = 0.0003] and 10 vs 13 days [nominal p = 0.0041], respectively).

mAb treatment against susceptible SARS-CoV-2 strains improved how patients feel and function, as evidenced by shortened time to sustained symptoms resolution and return to usual health and activities. Future studies are warranted to assess the patient experience with next generation mAbs.

GOV: Registration number, NCT04425629; Submission date June 11, 2020.

CD8+ T cell immunity, mediated by human leukocyte antigen (HLA) and T cell receptor (TCR), plays a critical role in conferring immune memory and protection against viral pathogens. The emergence of SARS-CoV-2 variants poses a serious challenge to the efficacy of current vaccines. Whereas numerous SARS-CoV-2 mutations associated with immune escape from CD8+ T cells have been documented, the molecular effects of most mutations on epitope-specific TCR recognition remain largely unexplored. Here, we studied an HLA-A24-restricted NYN epitope (Spike448-456) that elicits broad CD8+ T cell responses in COVID-19 patients characterized by a common TCR repertoire. Four natural mutations, N450K, L452Q, L452R, and Y453F, arose within the NYN epitope and have been transmitted in certain viral lineages. Our findings indicate that these mutations have minimal impact on the epitope's presentation by cell surface HLA, yet they diminish the affinities of their respective peptide-HLA complexes (pHLAs) for NYN peptide-specific TCRs, particularly L452R and Y453F. Furthermore, we determined the crystal structure of HLA-A24 loaded with the Y453F peptide (NYNYLFRLF), and subsequently a ternary structure of the public TCRNYN-I complexed to the original NYN-HLA-A24 (NYNYLYRLF). Our structural analysis unveiled that despite competent presentation by HLA, the mutant Y453F peptide failed to establish a stable TCR-pHLA ternary complex due to reduced peptide: TCR contacts. This study supports the idea that cellular immunity restriction is an important driving force behind viral evolution.

The emergence of SARS-CoV-2 variants raises concerns about the efficacy of existing COVID-19 vaccines and therapeutics. Previously, we identified a conserved cryptic class 5 epitope of SARS-CoV-2 receptor binding domain (RBD) by two cross-neutralizing antibodies 7D6 and 6D6. Intriguingly, this site remains resistant to substantial mutations occurred in ever-changing SARS-CoV-2 subvariants. As compared to class 3 antibody S309, 6D6 maintains broad and consistent neutralizing activities against SARS-CoV-2 variants. Furthermore, 6D6 effectively protected hamster from the virulent Beta strain. Sequence alignment of approximately 6 million documented SARS-CoV-2 isolates revealed that 6D6 epitope maintains an exceptionally high conservation rate (99.92%). Structural analysis demonstrated that all 33 mutations accumulated in XBB.1.5 since the original strain do not perturb the binding 6D6 to RBD, in line with the sequence analysis throughout the antigenicity evolution of SARS-CoV-2. These findings suggest the potential of this epitope serving as a critical determinant for vaccines and therapeutic design.

H1N1 influenza virus is a significant global public health concern. Monoclonal antibodies (mAbs) targeting specific viral proteins such as hemagglutinin (HA) have become an important therapeutic strategy, offering highly specific targeting to block viral transmission and infection. This study focused on the development of mAbs targeting HA of the A/Victoria/2570/2019 (H1N1pdm09, VIC-19) strain by utilizing hybridoma technology to produce two mAbs with high binding capacity. Notably, mAb 2B2 has demonstrated a strong affinity for HA proteins in recent H1N1 influenza vaccine strains. In vitro assessments showed that both mAbs exhibited broad-spectrum hemagglutination inhibition and potent neutralizing effects against various vaccine strains of H1N1pdm09 viruses. 2B2 was also effective in animal models, offering both preventive and therapeutic protection against infections caused by recent H1N1 strains, highlighting its potential for clinical application. By individually co-cultivating each of the aforementioned mAbs with the virus in chicken embryos, four amino acid substitution sites in HA (H138Q, G140R, A141E/V, and D187E) were identified in escape mutants, three in the antigenic site Ca2, and one in Sb. The identification of such mutations is pivotal, as it compels further investigation into how these alterations could undermine the binding efficacy and neutralization capacity of antibodies, thereby impacting the design and optimization of mAb therapies and influenza vaccines. This research highlights the necessity for continuous exploration into the dynamic interaction between viral evolution and antibody response, which is vital for the formulation of robust therapeutic and preventive strategies against influenza.

The emergence of the Middle East Respiratory Syndrome Coronavirus (MERS-CoV) has posed a significant global health concern due to its severe respiratory illness and high fatality rate. Currently, despite the potential for resurgence, there are no specific treatments for MERS-CoV, and only supportive care is available. Our study aimed to address this therapeutic gap by developing a potent neutralizing bispecific antibody (bsAb) against MERS-CoV. Initially, we isolated four human monoclonal antibodies (mAbs) that specifically target the MERS-CoV receptor-binding domain (RBD) using phage display technology and an established human antibody library. Among these four selected mAbs, our intensive in vitro functional analyses showed that the MERS-CoV RBD-specific mAb K111.3 exhibited the most potent neutralizing activity against MERS-CoV pseudoviral infection and the molecular interaction between MERS-CoV RBD and human dipeptidyl peptidase 4. Consequently, we engineered a novel bsAb, K207.C, by utilizing K111.3 as the IgG base and fusing it with the single-chain variable fragment of its non-competing pair, K111.1. This engineered bsAb showed significantly enhanced neutralization potential against MERS-CoV compared to its parental mAb. These findings suggest that K207.C may serve as a potential candidate for effective MERS-CoV neutralization, further highlighting the promise of the bsAb dual-targeting approach in MERS-CoV neutralization.

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) persistence in COVID-19 patients could play a key role in the emergence of variants of concern. The rapid intra-host evolution of SARS-CoV-2 may result in an increased transmissibility, immune and therapeutic escape which could be a direct consequence of COVID-19 epidemic currents. In this context, a longitudinal retrospective study on eight consecutive COVID-19 patients with persistent SARS-CoV-2 infection, from January 2022 to March 2023, was conducted. To characterize the intra- and inter-host viral evolution, whole genome sequencing and phylogenetic analysis were performed on nasopharyngeal samples collected at different time points. Phylogenetic reconstruction revealed an accelerated SARS-CoV-2 intra-host evolution and emergence of antigenically divergent variants. The Bayesian inference and principal coordinate analysis analysis showed a host-based genomic structuring among antigenically divergent variants, that might reflect the positive effect of containment practices, within the critical hospital area. All longitudinal antigenically divergent isolates shared a wide range of amino acidic (aa) changes, particularly in the Spike (S) glycoprotein, that increased viral transmissibility (K417N, S477N, N501Y and Q498R), enhanced infectivity (R346T, S373P, R408S, T478K, Q498R, Y505H, D614G, H655Y, N679K and P681H), caused host immune escape (S371L, S375F, T376A, K417N, and K444T/R) and displayed partial or complete resistance to treatments (G339D, R346K/T, S371F/L, S375F, T376A, D405N, N440K, G446S, N460K, E484A, F486V, Q493R, G496S and Q498R). These results suggest that multiple novel variants which emerge in the patient during persistent infection, might spread to another individual and continue to evolve. A pro-active genomic surveillance of persistent SARS-CoV-2 infected patients is recommended to identify genetically divergent lineages before their diffusion.

High-risk patients, often immunocompromised and not responding to vaccine, continue to experience severe coronavirus disease 2019 (COVID-19) and death. Monoclonal antibodies (mAbs) were shown to be effective to prevent severe COVID-19 for these patients. Nevertheless, concerns about the emergence of resistance mutations were raised.

We conducted a multicentric prospective cohort study, including 264 patients with mild to moderate COVID-19 at high risk for progression to severe COVID-19 and treated early with casirivimab/imdevimab, sotrovimab, or tixagevimab/cilgavimab. We sequenced the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genome during follow-up and searched for emerging spike mutations.

Immunocompromised patients have a 6-fold increased risk of developing mutations, which are associated with a prolonged duration of viral clearance but no clinical worsening. Emerging P337S/R/L/H, E340D/K/A/Q/V/G, and K356T/R substitutions in patients treated with sotrovimab are associated with higher viral RNA loads for up to 14 days post-treatment initiation. Tixagevimab/cilgavimab is associated with a 5-fold increased risk of developing mutations. R346K/I/T/S and K444R/N/M substitutions associated with tixagevimab/cilgavimab have been identified in multiple SARS-CoV-2 lineages, including BQ.1 and XBB.

The probability of emerging mutations arising in response to mAbs is significant, emphasizing the crucial need to investigate these mutations thoroughly and assess their impact on patients and the evolutionary trajectory of SARS-CoV-2.

Over four years have passed since the beginning of the COVID-19 pandemic. The scientific response has been rapid and effective, with many therapeutic monoclonal antibodies and small molecules developed for clinical use. However, given the ability for viruses to become resistant to antivirals, it is perhaps no surprise that the field has identified resistance to nearly all of these compounds. Here, we provide a comprehensive review of the resistance profile for each of these therapeutics. We hope that this resource provides an atlas for mutations to be aware of for each agent, particularly as a springboard for considerations for the next generation of antivirals. Finally, we discuss the outlook and thoughts for moving forward in how we continue to manage this, and the next, pandemic.

Antiviral therapies are integral in the fight against SARS-CoV-2 (i.e. severe acute respiratory syndrome coronavirus 2), the causative agent of COVID-19. Antiviral therapeutics can be divided into categories based on how they combat the virus, including viral entry into the host cell, viral replication, protein trafficking, post-translational processing, and immune response regulation. Drugs that target how the virus enters the cell include: Evusheld, REGEN-COV, bamlanivimab and etesevimab, bebtelovimab, sotrovimab, Arbidol, nitazoxanide, and chloroquine. Drugs that prevent the virus from replicating include: Paxlovid, remdesivir, molnupiravir, favipiravir, ribavirin, and Kaletra. Drugs that interfere with protein trafficking and post-translational processing include nitazoxanide and ivermectin. Lastly, drugs that target immune response regulation include interferons and the use of anti-inflammatory drugs such as dexamethasone. Antiviral therapies offer an alternative solution for those unable or unwilling to be vaccinated and are a vital weapon in the battle against the global pandemic. Learning more about these therapies helps raise awareness in the general population about the options available to them with respect to aiding in the reduction of the severity of COVID-19 infection. In this 'A Guide To' article, we provide an in-depth insight into the development of antiviral therapeutics against SARS-CoV-2 and their ability to help fight COVID-19.

Various therapeutic agents are being developed for the treatment of coronavirus disease 2019 (COVID-19). Therefore, it is crucial to accumulate information regarding the features of drug-resistant viruses to these antiviral drugs.

We investigated the emergence of dual-drug resistance in a kidney transplant recipient who received sotrovimab (from day 0) and remdesivir (RDV) (from day 8 to day 17). We sequenced the whole viral genomes from nasopharyngeal swabs taken on day 0 and seven points after starting treatment (on days 12, 19, 23, 37, 43, 48, and 58). The genetic traits of the wild-type (day 0) and descendant viruses (after day 12) were determined by comparing the genomes with those of a Wuhan strain and the day 0 wild-type strain, respectively. Three viral isolates (from samples collected on days 0, 23, and 37) were investigated for their escape ability and growth kinetics in vitro.

The sotrovimab resistant mutation (S:E340K) and the RDV resistant mutation RdRp:V792I (nt: G15814A) emerged within 12 days (day 12) and 11 days (day 19) after the treatment, respectively. The day 23 isolate harboring S:E340K/RdRp:V791I was resistant to both sotrovimab and RDV, showing 364- and 2.73-fold higher resistance respectively, compared with the wild-type. Moreover, compared with the day 23 isolate, the day 37 isolate accumulated multiple additional mutations and had a higher level of resistance to both drugs.

Drug-resistant variants with double mutations (S:E340K/RdRp:V791I) became dominant within 23 days after starting treatment, suggesting that even a combination therapy involving sotrovimab and RDV, dual-drug resistant viruses may emerge rapidly in immunocompromised patients. The dual-resistant variants had lower virus yields than those of the wild-type virus in vitro, suggesting that they paid a fitness cost.

Exposure to severe acute respiratory syndrome-corona virus-2 (SARS-CoV-2) prompts humoral immune responses in the human body. As the auxiliary diagnosis of a current infection, the existence of viral proteins can be checked from specific antibodies (Abs) induced by immunogenic viral proteins. For people with a weakened immune system, Ab treatment can help neutralize viral antigens to resist and treat the disease. On the other hand, highly immunogenic viral proteins can serve as effective markers for detecting prior infections. Additionally, the identification of viral particles or the presence of antibodies may help establish an immune defense against the virus. These immunogenic proteins rather than SARS-CoV-2 can be given to uninfected people as a vaccination to improve their coping ability against COVID-19 through the generation of memory plasma cells. In this work, we review immunogenic and immune-response proteins derived from SARS-CoV-2 with regard to their classification, origin, and diverse applications (e.g., prevention (vaccine development), diagnostic testing, and treatment (via neutralizing Abs)). Finally, advanced immunization strategies against COVID-19 are discussed along with the contemporary circumstances and future challenges.

SARS-CoV-2-neutralizing antibodies (nABs) showed great promise in the early phases of the COVID-19 pandemic. The emergence of resistant strains, however, quickly rendered the majority of clinically approved nABs ineffective. This underscored the imperative to develop nAB cocktails targeting non-overlapping epitopes.

Undertaking a nAB discovery program, we employed a classical workflow, while integrating artificial intelligence (AI)-based prediction to select non-competing nABs very early in the pipeline. We identified and in vivo validated (in female Syrian hamsters) two highly potent nABs.

Despite the promising results, in depth cryo-EM structural analysis demonstrated that the AI-based prediction employed with the intention to ensure non-overlapping epitopes was inaccurate. The two nABs in fact bound to the same receptor-binding epitope in a remarkably similar manner.

Our findings indicate that, even in the Alphafold era, AI-based predictions of paratope-epitope interactions are rough and experimental validation of epitopes remains an essential cornerstone of a successful nAB lead selection.

Full list of funders is provided at the end of the manuscript.

Continual evolution of the severe acute respiratory syndrome coronavirus (SARS-CoV-2) virus has allowed for its gradual evasion of neutralizing antibodies (nAbs) produced in response to natural infection or vaccination. The rapid nature of these changes has incited a need for the development of superior broad nAbs (bnAbs) and/or the rational design of an antibody cocktail that can protect against the mutated virus strain. Here, we report two angiotensin-converting enzyme 2 competing nAbs-8H12 and 3E2-with synergistic neutralization but evaded by some Omicron subvariants. Cryo-electron microscopy reveals the two nAbs synergistic neutralizing virus through a rigorous pairing permitted by rearrangement of the 472-489 loop in the receptor-binding domain to avoid steric clashing. Bispecific antibodies based on these two nAbs tremendously extend the neutralizing breadth and restore neutralization against recent variants including currently dominant XBB.1.5. Together, these findings expand our understanding of the potential strategies for the neutralization of SARS-CoV-2 variants toward the design of broad-acting antibody therapeutics and vaccines.