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Muscle fatigue represents the most prevalent symptom of long-term COVID, with elusive pathogenic mechanisms. We performed a longitudinal study to characterize histopathological and transcriptional changes in skeletal muscle in a hamster model of respiratory SARS-CoV-2 infection and compared them with influenza A virus (IAV) and mock infections. Histopathological and bulk RNA sequencing analyses of leg muscles derived from infected animals at days 3, 30, and 60 post-infection showed no direct viral invasion but myofiber atrophy in the SARS-CoV-2 group, which was accompanied by persistent downregulation of the genes related to myofibers, ribosomal proteins, fatty acid ß-oxidation, tricarboxylic acid cycle, and mitochondrial oxidative phosphorylation complexes. While both SARS-CoV-2 and IAV infections induced acute and transient type I and II interferon responses in muscle, only the SARS-CoV-2 infection upregulated TNF-α/NF-κB but not IL-6 signaling in muscle. Treatment of C2C12 myotubes, a skeletal muscle cell line, with combined IFN-γ and TNF-α but not with IFN-γ or TNF-α alone markedly impaired mitochondrial function. We conclude that a respiratory SARS-CoV-2 infection can cause myofiber atrophy and persistent energy metabolism suppression without direct viral invasion. The effects may be induced by the combined systemic interferon and TNF-α responses at the acute phase and may contribute to post-COVID-19 persistent muscle fatigue.
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Type I interferons (IFN-I) are cytokines with potent antiviral and inflammatory capacities. IFN-I signaling drives the expression of hundreds of IFN-I stimulated genes (ISGs), whose aggregate function results in the control of viral infection. A few of these ISGs are tasked with negatively regulating the IFN-I response to prevent overt inflammation. ISG15 is a negative regulator whose absence leads to persistent, low-grade elevation of ISG expression and concurrent, self-resolving mild autoinflammation. The limited breadth and low-grade persistence of ISGs expressed in ISG15 deficiency are sufficient to confer broad-spectrum antiviral resistance. Inspired by ISG15 deficiency, we have identified a nominal collection of 10 ISGs that recapitulate the broad antiviral potential of the IFN-I system. The expression of the 10 ISG collection in an IFN-I non-responsive cell line increased cellular resistance to Zika, Vesicular Stomatitis, Influenza A (IAV), and SARS-CoV-2 viruses. A deliverable prophylactic formulation of this syndicate of 10 ISGs significantly inhibited IAV PR8 replication in vivo in mice and protected hamsters against a lethal SARS-CoV-2 challenge, suggesting its potential as a broad-spectrum antiviral against many current and future emerging viral pathogens. One-Sentence Summary: Human inborn error of immunity-guided discovery and development of a broad-spectrum RNA antiviral therapy.
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We describe a previously-unappreciated role for Bruton's tyrosine kinase (BTK) in fungal immune surveillance against aspergillosis, an unforeseen complication of BTK inhibitors (BTKi) used for treating B-cell lymphoid malignancies. We studied BTK-dependent fungal responses in neutrophils from diverse populations, including healthy donors, BTKi-treated patients, and X-linked agammaglobulinemia patients. Upon fungal exposure, BTK was activated in human neutrophils in a TLR2-, Dectin-1-, and FcγR-dependent manner, triggering the oxidative burst. BTK inhibition selectively impeded neutrophil-mediated damage to Aspergillus hyphae, primary granule release, and the fungus-induced oxidative burst by abrogating NADPH oxidase subunit p40phox and GTPase RAC2 activation. Moreover, neutrophil-specific Btk deletion in mice enhanced aspergillosis susceptibility by impairing neutrophil function, not recruitment or lifespan. Conversely, GM-CSF partially mitigated these deficits by enhancing p47phox activation. Our findings underline the crucial role of BTK signaling in neutrophils for antifungal immunity and provide a rationale for GM-CSF use to offset these deficits in susceptible patients.
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Morbilliviruses are among the most contagious viral pathogens of mammals. Although previous metagenomic surveys have identified morbillivirus sequences in bats, full-length morbilliviruses from bats are limited. Here we characterize the myotis bat morbillivirus (MBaMV) from a bat surveillance programme in Brazil, whose full genome was recently published. We demonstrate that the fusion and receptor binding protein of MBaMV utilize bat CD150 and not human CD150, as an entry receptor in a mammalian cell line. Using reverse genetics, we produced a clone of MBaMV that infected Vero cells expressing bat CD150. Electron microscopy of MBaMV-infected cells revealed budding of pleomorphic virions, a characteristic morbillivirus feature. MBaMV replication reached 103-105 plaque-forming units ml-1 in human epithelial cell lines and was dependent on nectin-4. Infection of human macrophages also occurred, albeit 2-10-fold less efficiently than measles virus. Importantly, MBaMV is restricted by cross-neutralizing human sera elicited by measles, mumps and rubella vaccination and is inhibited by orally bioavailable polymerase inhibitors in vitro. MBaMV-encoded P/V genes did not antagonize human interferon induction. Finally, we show that MBaMV does not cause disease in Jamaican fruit bats. We conclude that, while zoonotic spillover into humans may theoretically be plausible, MBaMV replication would probably be controlled by the human immune system.
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Quirópteros , Morbillivirus , Animales , Chlorocebus aethiops , Humanos , Células Vero , Zoonosis , Morbillivirus/genética , Línea CelularRESUMEN
Systemic candidiasis is a common, high-mortality, nosocomial fungal infection. Unexpectedly, it has emerged as a complication of anti-complement C5-targeted monoclonal antibody treatment, indicating a critical niche for C5 in antifungal immunity. We identified transcription of complement system genes as the top biological pathway induced in candidemic patients and as predictive of candidemia. Mechanistically, C5a-C5aR1 promoted fungal clearance and host survival in a mouse model of systemic candidiasis by stimulating phagocyte effector function and ERK- and AKT-dependent survival in infected tissues. C5ar1 ablation rewired macrophage metabolism downstream of mTOR, promoting their apoptosis and enhancing mortality through kidney injury. Besides hepatocyte-derived C5, local C5 produced intrinsically by phagocytes provided a key substrate for antifungal protection. Lower serum C5a concentrations or a C5 polymorphism that decreases leukocyte C5 expression correlated independently with poor patient outcomes. Thus, local, phagocyte-derived C5 production licenses phagocyte antimicrobial function and confers innate protection during systemic fungal infection.
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Antifúngicos , Candidiasis , Animales , Ratones , Complemento C5/metabolismo , Fagocitos/metabolismoRESUMEN
Rationale: SARS-CoV-2 entry into host cells is facilitated by endogenous and exogenous proteases that proteolytically activate the spike glycoprotein and antiproteases inhibiting this process. Understanding the key actors in viral entry is crucial for advancing knowledge of virus tropism, pathogenesis, and potential therapeutic targets. Objectives: We aimed to investigate the role of naïve serum and alpha-1-antitrypsin (AAT) in inhibiting protease-mediated SARS-CoV-2 entry and explore the implications of AAT deficiency on susceptibility to different SARS-CoV-2 variants. Findings: Our study demonstrates that naïve serum exhibits significant inhibition of SARS-CoV-2 entry, with AAT identified as the major serum protease inhibitor potently restricting entry. Using pseudoparticles, replication-competent pseudoviruses, and authentic SARS-CoV-2, we show that AAT inhibition occurs at low concentrations compared with those in serum and bronchoalveolar tissues, suggesting physiological relevance. Furthermore, sera from subjects with an AAT-deficient genotype show reduced ability to inhibit entry of both Wuhan-Hu-1 (WT) and B.1.617.2 (Delta) but exhibit no difference in inhibiting B.1.1.529 (Omicron) entry. Conclusions: AAT may have a variant-dependent therapeutic potential against SARS-CoV-2. Our findings highlight the importance of further investigating the complex interplay between proteases, antiproteases, and spike glycoprotein activation in SARS-CoV-2 and other respiratory viruses to identify potential therapeutic targets and improve understanding of disease pathogenesis.
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CCR5 is among the most studied chemokine receptors due to its profound significance in human health and disease. The notion that CCR5 is a functionally redundant receptor was challenged through the demonstration of its unique protective role in the context of West Nile virus in both mice and humans. In the nearly two decades since this initial discovery, numerous studies have investigated the role of CCR5 in the context of other medically important neurotropic flaviviruses, most of which appear to support a broad neuroprotective role for this receptor, although how CCR5 exerts its protective effect has been remarkably varied. In this review, we summarize the mechanisms by which CCR5 controls neurotropic flaviviruses, as well as results from human studies evaluating a genetic link to CCR5, and propose unexplored areas of research that are needed to unveil even more exciting roles for this important receptor.
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Flavivirus , Virus del Nilo Occidental , Humanos , Animales , Ratones , Virus del Nilo Occidental/genética , Receptores de Quimiocina , Receptores CCR5/genéticaRESUMEN
SARS-CoV-2, the etiological agent of COVID-19, is characterized by a delay in type I interferon (IFN-I)-mediated antiviral defenses alongside robust cytokine production. Here, we investigate the underlying molecular basis for this imbalance and implicate virus-mediated activation of NF-κB in the absence of other canonical IFN-I-related transcription factors. Epigenetic and single-cell transcriptomic analyses show a selective NF-κB signature that was most prominent in infected cells. Disruption of NF-κB signaling through the silencing of the NF-κB transcription factor p65 or p50 resulted in loss of virus replication that was rescued upon reconstitution. These findings could be further corroborated with the use of NF-κB inhibitors, which reduced SARS-CoV-2 replication in vitro. These data suggest that the robust cytokine production in response to SARS-CoV-2, despite a diminished IFN-I response, is the product of a dependency on NF-κB for viral replication. IMPORTANCE The COVID-19 pandemic has caused significant mortality and morbidity around the world. Although effective vaccines have been developed, large parts of the world remain unvaccinated while new SARS-CoV-2 variants keep emerging. Furthermore, despite extensive efforts and large-scale drug screenings, no fully effective antiviral treatment options have been discovered yet. Therefore, it is of the utmost importance to gain a better understanding of essential factors driving SARS-CoV-2 replication to be able to develop novel approaches to target SARS-CoV-2 biology.
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COVID-19/metabolismo , Citocinas/metabolismo , Interferón Tipo I/metabolismo , SARS-CoV-2 , Factor de Transcripción ReIA/metabolismo , Transcriptoma , Replicación Viral , Células A549 , Animales , COVID-19/virología , Chlorocebus aethiops , Epigenómica , Regulación de la Expresión Génica , Células HEK293 , Células HeLa , Interacciones Microbiota-Huesped , Humanos , Transducción de Señal , Análisis de la Célula Individual , Factor de Transcripción ReIA/antagonistas & inhibidores , Factor de Transcripción ReIA/genética , Factores de Transcripción/metabolismo , Células VeroRESUMEN
Puel and Casanova and Kisand et al. challenge our conclusions that interferonopathy and not IL-17/IL-22 autoantibodies promote candidiasis in autoimmune polyendocrinopathycandidiasisectodermal dystrophy. We acknowledge that conclusive evidence for causation is difficult to obtain in complex human diseases. However, our studies clearly document interferonopathy driving mucosal candidiasis with intact IL-17/IL-22 responses in Aire-deficient mice, with strong corroborative evidence in patients.
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Inmunidad Mucosa , Micosis , Humanos , Membrana Mucosa , Animales , RatonesRESUMEN
Heart injury has been reported in up to 20% of COVID-19 patients, yet the cause of myocardial histopathology remains unknown. Here, using an established in vivo hamster model, we demonstrate that SARS-CoV-2 can be detected in cardiomyocytes of infected animals. Furthermore, we found damaged cardiomyocytes in hamsters and COVID-19 autopsy samples. To explore the mechanism, we show that both human pluripotent stem cell-derived cardiomyocytes (hPSC-derived CMs) and adult cardiomyocytes (CMs) can be productively infected by SARS-CoV-2, leading to secretion of the monocyte chemoattractant cytokine CCL2 and subsequent monocyte recruitment. Increased CCL2 expression and monocyte infiltration was also observed in the hearts of infected hamsters. Although infected CMs suffer damage, we find that the presence of macrophages significantly reduces SARS-CoV-2-infected CMs. Overall, our study provides direct evidence that SARS-CoV-2 infects CMs in vivo and suggests a mechanism of immune cell infiltration and histopathology in heart tissues of COVID-19 patients.
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COVID-19/patología , Quimiocina CCL2/metabolismo , Lesiones Cardíacas/virología , Monocitos/inmunología , Miocitos Cardíacos/metabolismo , Animales , Comunicación Celular/fisiología , Línea Celular , Chlorocebus aethiops , Cricetinae , Modelos Animales de Enfermedad , Humanos , Macrófagos/inmunología , Masculino , Miocitos Cardíacos/virología , Células Madre Pluripotentes/citología , Células VeroRESUMEN
Antibody-dependent enhancement (ADE) is a phenomenon that occurs when cross-reactive antibodies generated from a previous flaviviral infection increase the pathogenesis of a related virus. Zika virus (ZIKV) is the most recent flavivirus introduced to the Western Hemisphere and has become a significant public health threat due to the unanticipated impact on the developing fetus. West Nile virus (WNV) is the primary flavivirus that circulates in North America, and we and others have shown that antibodies against WNV are cross-reactive to ZIKV. Thus, there is concern that WNV immunity could increase the risk of severe ZIKV infection, particularly during pregnancy. In this study, we examined the extent to which WNV antibodies could impact ZIKV pathogenesis in a murine pregnancy model. To test this, we passively transferred WNV antibodies into pregnant Stat2-/- mice on E6.5 prior to infection with ZIKV. Evaluation of pregnant dams showed weight loss following ZIKV infection; however, no differences in maternal weights or viral loads in the maternal brain, spleen, or spinal cord were observed in the presence of WNV antibodies. Resorption rates, and other fetal parameters, including fetal and placental size, were similarly unaffected. Further, the presence of WNV antibodies did not significantly alter the viral load or the inflammatory response in the placenta or the fetus in response to ZIKV. Our data suggest that pre-existing WNV immunity may not significantly impact the pathogenesis of ZIKV infection during pregnancy. Our findings are promising for the safety of implementing WNV vaccines in the continental US.
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Reacciones Cruzadas/inmunología , Virus del Nilo Occidental/inmunología , Infección por el Virus Zika/inmunología , Infección por el Virus Zika/prevención & control , Virus Zika/inmunología , Animales , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Acrecentamiento Dependiente de Anticuerpo/inmunología , Femenino , Masculino , Ratones , Ratones Noqueados , Pruebas de Neutralización , Embarazo , Factor de Transcripción STAT2/genética , Carga Viral , Fiebre del Nilo Occidental/inmunologíaRESUMEN
Recent clinical data have suggested a correlation between coronavirus disease 2019 (COVID-19) and diabetes. Here, we describe the detection of SARS-CoV-2 viral antigen in pancreatic beta cells in autopsy samples from individuals with COVID-19. Single-cell RNA sequencing and immunostaining from ex vivo infections confirmed that multiple types of pancreatic islet cells were susceptible to SARS-CoV-2, eliciting a cellular stress response and the induction of chemokines. Upon SARS-CoV-2 infection, beta cells showed a lower expression of insulin and a higher expression of alpha and acinar cell markers, including glucagon and trypsin1, respectively, suggesting cellular transdifferentiation. Trajectory analysis indicated that SARS-CoV-2 induced eIF2-pathway-mediated beta cell transdifferentiation, a phenotype that could be reversed with trans-integrated stress response inhibitor (trans-ISRIB). Altogether, this study demonstrates an example of SARS-CoV-2 infection causing cell fate change, which provides further insight into the pathomechanisms of COVID-19.
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COVID-19/virología , Transdiferenciación Celular , Células Secretoras de Insulina/virología , SARS-CoV-2/patogenicidad , Acetamidas/farmacología , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Animales , COVID-19/mortalidad , Transdiferenciación Celular/efectos de los fármacos , Chlorocebus aethiops , Ciclohexilaminas/farmacología , Citocinas/metabolismo , Factor 2 Eucariótico de Iniciación/metabolismo , Femenino , Glucagón , Interacciones Huésped-Patógeno , Humanos , Insulina/metabolismo , Células Secretoras de Insulina/efectos de los fármacos , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/patología , Masculino , Persona de Mediana Edad , Fenotipo , Transducción de Señal , Técnicas de Cultivo de Tejidos , Tripsina/metabolismo , Células Vero , Adulto JovenRESUMEN
[Figure: see text].
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COVID-19/inmunología , Inmunidad Celular/inmunología , Mediadores de Inflamación/inmunología , Macrófagos/inmunología , Miocitos Cardíacos/inmunología , Adulto , Anciano , Anciano de 80 o más Años , COVID-19/patología , Técnicas de Cocultivo/métodos , Femenino , Humanos , Mediadores de Inflamación/metabolismo , Macrófagos/patología , Masculino , Persona de Mediana Edad , Miocardio/inmunología , Miocardio/patología , Miocitos Cardíacos/patología , Células Madre Pluripotentes/inmunología , Análisis de Secuencia de ARN/métodosRESUMEN
The global coronavirus disease 2019 (COVID-19) pandemic has mobilized efforts to develop vaccines and antibody-based therapeutics, including convalescent-phase plasma therapy, that inhibit viral entry by inducing or transferring neutralizing antibodies (nAbs) against the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike glycoprotein (CoV2-S). However, rigorous efficacy testing requires extensive screening with live virus under onerous biosafety level 3 (BSL3) conditions, which limits high-throughput screening of patient and vaccine sera. Myriad BSL2-compatible surrogate virus neutralization assays (VNAs) have been developed to overcome this barrier. Yet, there is marked variability between VNAs and how their results are presented, making intergroup comparisons difficult. To address these limitations, we developed a standardized VNA using CoV2-S pseudotyped particles (CoV2pp) based on vesicular stomatitis virus bearing the Renilla luciferase gene in place of its G glycoprotein (VSVΔG); this assay can be robustly produced at scale and generate accurate neutralizing titers within 18 h postinfection. Our standardized CoV2pp VNA showed a strong positive correlation with CoV2-S enzyme-linked immunosorbent assay (ELISA) results and live-virus neutralizations in confirmed convalescent-patient sera. Three independent groups subsequently validated our standardized CoV2pp VNA (n > 120). Our data (i) show that absolute 50% inhibitory concentration (absIC50), absIC80, and absIC90 values can be legitimately compared across diverse cohorts, (ii) highlight the substantial but consistent variability in neutralization potency across these cohorts, and (iii) support the use of the absIC80 as a more meaningful metric for assessing the neutralization potency of a vaccine or convalescent-phase sera. Lastly, we used our CoV2pp in a screen to identify ultrapermissive 293T clones that stably express ACE2 or ACE2 plus TMPRSS2. When these are used in combination with our CoV2pp, we can produce CoV2pp sufficient for 150,000 standardized VNAs/week.IMPORTANCE Vaccines and antibody-based therapeutics like convalescent-phase plasma therapy are premised upon inducing or transferring neutralizing antibodies that inhibit SARS-CoV-2 entry into cells. Virus neutralization assays (VNAs) for measuring neutralizing antibody titers (NATs) are an essential part of determining vaccine or therapeutic efficacy. However, such efficacy testing is limited by the inherent dangers of working with the live virus, which requires specialized high-level biocontainment facilities. We therefore developed a standardized replication-defective pseudotyped particle system that mimics the entry of live SARS-CoV-2. This tool allows for the safe and efficient measurement of NATs, determination of other forms of entry inhibition, and thorough investigation of virus entry mechanisms. Four independent labs across the globe validated our standardized VNA using diverse cohorts. We argue that a standardized and scalable assay is necessary for meaningful comparisons of the myriad of vaccines and antibody-based therapeutics becoming available. Our data provide generalizable metrics for assessing their efficacy.
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COVID-19/diagnóstico , COVID-19/inmunología , SARS-CoV-2/inmunología , SARS-CoV-2/patogenicidad , Anticuerpos Neutralizantes/inmunología , Anticuerpos Neutralizantes/metabolismo , Anticuerpos Antivirales/metabolismo , Ensayo de Inmunoadsorción Enzimática , Humanos , Pruebas de NeutralizaciónRESUMEN
The emergence and spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has resulted in significant global morbidity, mortality, and societal disruption. A better understanding of virus-host interactions may potentiate therapeutic insights toward limiting this infection. Here we investigated the dynamics of the systemic response to SARS-CoV-2 in hamsters by histological analysis and transcriptional profiling. Infection resulted in consistently high levels of virus in the upper and lower respiratory tracts and sporadic occurrence in other distal tissues. A longitudinal cohort revealed a wave of inflammation, including a type I interferon (IFN-I) response, that was evident in all tissues regardless of viral presence but was insufficient to prevent disease progression. Bolstering the antiviral response with intranasal administration of recombinant IFN-I reduced viral disease, prevented transmission, and lowered inflammation in vivo. This study defines the systemic host response to SARS-CoV-2 infection and supports use of intranasal IFN-I as an effective means of early treatment.
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COVID-19/metabolismo , COVID-19/virología , Interacciones Huésped-Patógeno , Interferón Tipo I/metabolismo , SARS-CoV-2/fisiología , Animales , Biopsia , COVID-19/genética , COVID-19/inmunología , Cricetinae , Citocinas/genética , Citocinas/metabolismo , Modelos Animales de Enfermedad , Perfilación de la Expresión Génica , Interacciones Huésped-Patógeno/inmunología , Humanos , Inmunidad Innata , Interferón Tipo I/genética , Pulmón/inmunología , Pulmón/metabolismo , Pulmón/patología , Pulmón/virología , Especificidad de Órganos/inmunología , Virulencia , Replicación Viral/inmunologíaRESUMEN
Human monogenic disorders have revealed the critical contribution of type 17 responses in mucosal fungal surveillance. We unexpectedly found that in certain settings, enhanced type 1 immunity rather than defective type 17 responses can promote mucosal fungal infection susceptibility. Notably, in mice and humans with AIRE deficiency, an autoimmune disease characterized by selective susceptibility to mucosal but not systemic fungal infection, mucosal type 17 responses are intact while type 1 responses are exacerbated. These responses promote aberrant interferon-γ (IFN-γ)- and signal transducer and activator of transcription 1 (STAT1)-dependent epithelial barrier defects as well as mucosal fungal infection susceptibility. Concordantly, genetic and pharmacologic inhibition of IFN-γ or Janus kinase (JAK)-STAT signaling ameliorates mucosal fungal disease. Thus, we identify aberrant T cell-dependent, type 1 mucosal inflammation as a critical tissue-specific pathogenic mechanism that promotes mucosal fungal infection susceptibility in mice and humans.
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Candida albicans/inmunología , Candidiasis Mucocutánea Crónica/genética , Candidiasis Mucocutánea Crónica/inmunología , Inmunidad Mucosa/inmunología , Poliendocrinopatías Autoinmunes/genética , Poliendocrinopatías Autoinmunes/inmunología , Adolescente , Adulto , Anciano , Animales , Modelos Animales de Enfermedad , Femenino , Humanos , Inmunidad Mucosa/genética , Vigilancia Inmunológica/genética , Vigilancia Inmunológica/inmunología , Interferón gamma/genética , Interleucinas/genética , Quinasas Janus/genética , Masculino , Ratones , Ratones Endogámicos BALB C , Persona de Mediana Edad , Mucosa Bucal/inmunología , Mucosa Bucal/patología , Receptores de Interleucina-17/genética , Factor de Transcripción STAT1/genética , Linfocitos T/inmunología , Adulto Joven , Interleucina-22RESUMEN
Type I interferon (IFN) signaling in fetal tissues causes developmental abnormalities and fetal demise. Although pathogens that infect fetal tissues can induce birth defects through the local production of type I IFN, it remains unknown why systemic IFN generated during maternal infections only rarely causes fetal developmental defects. Here, we report that activation of the guanine nucleotide-binding protein-coupled estrogen receptor 1 (GPER1) during pregnancy is both necessary and sufficient to suppress IFN signaling and does so disproportionately in reproductive and fetal tissues. Inactivation of GPER1 in mice halted fetal development and promoted fetal demise, but only in the context of maternal inflammation. Thus, GPER1 is a central regulator of IFN signaling during pregnancy that allows dynamic antiviral responses in maternal tissues while also preserving fetal health.
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Enfermedades Fetales/inmunología , Inflamación/inmunología , Intercambio Materno-Fetal/inmunología , Complicaciones Infecciosas del Embarazo/inmunología , Receptores de Estrógenos/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Animales , Benzodioxoles/farmacología , Sistemas CRISPR-Cas , Femenino , Enfermedades Fetales/virología , Feto/inmunología , Feto/virología , Humanos , Virus de la Influenza A/inmunología , Gripe Humana/inmunología , Interferón Tipo I/inmunología , Ratones , Ratones Endogámicos C57BL , Placenta/inmunología , Placenta/virología , Embarazo , Quinolinas/farmacología , Receptores de Estrógenos/antagonistas & inhibidores , Receptores Acoplados a Proteínas G/antagonistas & inhibidoresRESUMEN
BACKGROUND: A tetravalent live attenuated dengue vaccine, Dengvaxia, sensitised naïve recipients to severe dengue illness upon a subsequent natural dengue infection and is suspected to be due to antibody-dependent enhancement (ADE). ADE has also been implicated in the severe neurological outcomes of Zika virus (ZIKV) infection. It has become evident that cross-reactive antibodies targeting the viral pre-membrane protein and fusion-loop epitope are ADE-competent. A pre-clinical tetravalent dengue sub-unit vaccine candidate, DSV4, eliminates these ADE-competent epitopes. METHODS: We compared protective efficacy and ADE-competence of murine polyclonal antibodies induced by DSV4, Dengvaxia and an 'in house' tetravalent mixture of all four laboratory DENV strains, TV DENV, using established mouse models. FINDINGS: DSV4-induced antibodies, known to be predominantly type-specific, provided significant protection against lethal DENV challenge, but did not promote ADE of either DENV or ZIKV infection in vivo. Antibodies elicited by Dengvaxia and TV DENV, which are predominantly cross-reactive, not only failed to offer protection against lethal DENV challenge, but also promoted ADE of both DENV and ZIKV infection in vivo. INTERPRETATION: Protective efficacy against DENV infection may be linked to the induction of neutralising antibodies which are type-specific rather than cross-reactive. Whole virus-based dengue vaccines may be associated with ADE risk, despite their potent virus-neutralising capacity. Vaccines designed to eliminate ADE-competent epitopes may help eliminate/minimise ADE risk. FUNDING: This study was supported partly by ICGEB, India, the National Biopharma Mission, DBT, Government of India, Sun Pharmaceutical Industries Limited, India, and NIAID, NIH, USA.
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Reacciones Cruzadas/inmunología , Vacunas contra el Dengue/inmunología , Virus del Dengue/inmunología , Dengue/prevención & control , Vacunas Atenuadas/inmunología , Vacunas Sintéticas/inmunología , Infección por el Virus Zika/inmunología , Virus Zika/inmunología , Animales , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales , Dengue/inmunología , Dengue/virología , Vacunas contra el Dengue/genética , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Epítopos/inmunología , Humanos , Inmunización , Inmunogenicidad Vacunal , Ratones , Ratones Noqueados , Vacunas Sintéticas/genética , Proteínas del Envoltorio Viral/inmunología , Carga Viral , Infección por el Virus Zika/virologíaRESUMEN
The global COVID-19 pandemic has mobilized efforts to develop vaccines and antibody-based therapeutics, including convalescent plasma therapy, that inhibit viral entry by inducing or transferring neutralizing antibodies (nAbs) against the SARS-CoV-2 spike glycoprotein (CoV2-S). However, rigorous efficacy testing requires extensive screening with live virus under onerous BSL3 conditions which limits high throughput screening of patient and vaccine sera. Myriad BSL-2 compatible surrogate virus neutralization assays (VNAs) have been developed to overcome this barrier. Yet, there is marked variability between VNAs and how their results are presented, making inter-group comparisons difficult. To address these limitations, we developed a standardized VNA using VSVΔG-based CoV-2-S pseudotyped particles (CoV2pp) that can be robustly produced at scale and generate accurate neutralizing titers within 18 hours post-infection. Our standardized CoV2pp VNA showed a strong positive correlation with CoV2-S ELISA and live virus neutralizations in confirmed convalescent patient sera. Three independent groups subsequently validated our standardized CoV2pp VNA (n>120). Our data show that absolute (abs) IC50, IC80, and IC90 values can be legitimately compared across diverse cohorts, highlight the substantial but consistent variability in neutralization potency across these cohorts, and support the use of absIC80 as a more meaningful metric for assessing the neutralization potency of vaccine or convalescent sera. Lastly, we used our CoV2pp in a screen to identify ultra-permissive 293T clones that stably express ACE2 or ACE2+TMPRSS2. When used in combination with our CoV2pp, we can now produce CoV2pp sufficient for 150,000 standardized VNA/week.