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1.
Front Immunol ; 12: 614436, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33790892

RESUMEN

The novel coronavirus named severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused a global pandemic of the coronavirus disease 2019 (COVID-19), which elicits a wide variety of symptoms, ranging from mild to severe, with the potential to lead to death. Although used as the standard method to screen patients for SARS-CoV-2 infection, real-time PCR has challenges in dealing with asymptomatic patients and those with an undetectable viral load. Serological tests are therefore considered potent diagnostic tools to complement real-time PCR-based diagnosis and are used for surveillance of seroprevalence in populations. However, the dynamics of the antibody response against SARS-CoV-2 currently remain to be investigated. Here, through analysis of plasma samples from 84 patients with COVID-19, we observed that the response of virus-specific antibodies against three important antigens, RBD, N and S, dynamically changed over time and reached a peak 5-8 weeks after the onset of symptoms. The antibody responses were irrespective of sex. Severe cases were found to have higher levels of antibody response, larger numbers of inflammatory cells and C-reactive protein levels. Within the mild/moderate cases, pairwise comparison indicated moderate association between anti-RBD vs. anti-N, anti-RBD vs. anti-S1S2, and anti-N vs. anti-S1S2. Furthermore, the majority of cases could achieve IgM and IgG seroconversion at 2 weeks since the disease onset. Analysis of neutralizing antibodies indicated that these responses were able to last for more than 112 days but decline significantly after the peak. In summary, our findings demonstrate the longitudinally dynamic changes in antibody responses against SARS-CoV-2, which can contribute to the knowledge of humoral immune response after SARS-CoV-2 infection and are informative for future development of vaccine and antibody-based therapies.


Asunto(s)
Anticuerpos Neutralizantes/sangre , Anticuerpos Antivirales/sangre , /inmunología , Glicoproteína de la Espiga del Coronavirus/inmunología , Adulto , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Beijing , China , Femenino , Humanos , Inmunoglobulina G/sangre , Inmunoglobulina M/sangre , Masculino , Persona de Mediana Edad , Fosfoproteínas/inmunología , Dominios Proteicos/inmunología , Seroconversión , Índice de Severidad de la Enfermedad , Centros de Atención Terciaria
2.
Science ; 372(6537)2021 04 02.
Artículo en Inglés | MEDLINE | ID: mdl-33795432

RESUMEN

Multivalent display of receptor-engaging antibodies or ligands can enhance their activity. Instead of achieving multivalency by attachment to preexisting scaffolds, here we unite form and function by the computational design of nanocages in which one structural component is an antibody or Fc-ligand fusion and the second is a designed antibody-binding homo-oligomer that drives nanocage assembly. Structures of eight nanocages determined by electron microscopy spanning dihedral, tetrahedral, octahedral, and icosahedral architectures with 2, 6, 12, and 30 antibodies per nanocage, respectively, closely match the corresponding computational models. Antibody nanocages targeting cell surface receptors enhance signaling compared with free antibodies or Fc-fusions in death receptor 5 (DR5)-mediated apoptosis, angiopoietin-1 receptor (Tie2)-mediated angiogenesis, CD40 activation, and T cell proliferation. Nanocage assembly also increases severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pseudovirus neutralization by α-SARS-CoV-2 monoclonal antibodies and Fc-angiotensin-converting enzyme 2 (ACE2) fusion proteins.


Asunto(s)
Anticuerpos/química , Anticuerpos/inmunología , Nanoestructuras , Ingeniería de Proteínas , Transducción de Señal , Angiopoyetinas/química , Angiopoyetinas/inmunología , Angiopoyetinas/metabolismo , Anticuerpos Monoclonales/química , Anticuerpos Monoclonales/inmunología , Anticuerpos Neutralizantes/química , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/química , Anticuerpos Antivirales/inmunología , Linfocitos B/inmunología , Antígenos CD40/química , Antígenos CD40/inmunología , Antígenos CD40/metabolismo , Línea Celular Tumoral , Proliferación Celular , Simulación por Computador , Genes Sintéticos , Humanos , Fragmentos Fc de Inmunoglobulinas/química , Activación de Linfocitos , Modelos Moleculares , Unión Proteica , Receptor TIE-2/metabolismo , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/inmunología , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Linfocitos T/inmunología , Linfocitos T/fisiología
3.
Cell Rep ; 35(1): 108950, 2021 04 06.
Artículo en Inglés | MEDLINE | ID: mdl-33794145

RESUMEN

Antibodies with heavy chains that derive from the VH1-2 gene constitute some of the most potent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-neutralizing antibodies yet identified. To provide insight into whether these genetic similarities inform common modes of recognition, we determine the structures of the SARS-CoV-2 spike in complex with three VH1-2-derived antibodies: 2-15, 2-43, and H4. All three use VH1-2-encoded motifs to recognize the receptor-binding domain (RBD), with heavy-chain N53I-enhancing binding and light-chain tyrosines recognizing F486RBD. Despite these similarities, class members bind both RBD-up and -down conformations of the spike, with a subset of antibodies using elongated CDRH3s to recognize glycan N343 on a neighboring RBD-a quaternary interaction accommodated by an increase in RBD separation of up to 12 Å. The VH1-2 antibody class, thus, uses modular recognition encoded by modular genetic elements to effect potent neutralization, with the VH-gene component specifying recognition of RBD and the CDRH3 component specifying quaternary interactions.


Asunto(s)
Anticuerpos Neutralizantes , Anticuerpos Antivirales , Región Variable de Inmunoglobulina , /inmunología , Anticuerpos Neutralizantes/genética , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/genética , Anticuerpos Antivirales/inmunología , /inmunología , Células HEK293 , Humanos , Región Variable de Inmunoglobulina/genética , Región Variable de Inmunoglobulina/inmunología
4.
Int J Mol Sci ; 22(5)2021 Mar 08.
Artículo en Inglés | MEDLINE | ID: mdl-33800363

RESUMEN

Quantitative and robust serology assays are critical measurements underpinning global COVID-19 response to diagnostic, surveillance, and vaccine development. Here, we report a proof-of-concept approach for the development of quantitative, multiplexed flow cytometry-based serological and neutralization assays. The serology assays test the IgG and IgM against both the full-length spike antigens and the receptor binding domain (RBD) of the spike antigen. Benchmarking against an RBD-specific SARS-CoV IgG reference standard, the anti-SARS-CoV-2 RBD antibody titer was quantified in the range of 37.6 µg/mL to 31.0 ng/mL. The quantitative assays are highly specific with no correlative cross-reactivity with the spike proteins of MERS, SARS1, OC43 and HKU1 viruses. We further demonstrated good correlation between anti-RBD antibody titers and neutralizing antibody titers. The suite of serology and neutralization assays help to improve measurement confidence and are complementary and foundational for clinical and epidemiologic studies.


Asunto(s)
/métodos , /sangre , Pruebas de Neutralización/métodos , Pruebas de Neutralización/normas , /inmunología , Anticuerpos Neutralizantes/sangre , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/sangre , Anticuerpos Antivirales/inmunología , Reacciones Cruzadas , Citometría de Flujo/métodos , Fluorescencia , Humanos , Inmunoglobulina G/sangre , Inmunoglobulina G/inmunología , Inmunoglobulina M/sangre , Inmunoglobulina M/inmunología , Microesferas , Receptores Virales/química , Receptores Virales/inmunología , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/inmunología
5.
Viruses ; 13(3)2021 03 09.
Artículo en Inglés | MEDLINE | ID: mdl-33803400

RESUMEN

SARS-CoV-2 (Severe Acute Respiratory Syndrome-Coronavirus 2) has accumulated multiple mutations during its global circulation. Recently, three SARS-CoV-2 lineages, B.1.1.7 (501Y.V1), B.1.351 (501Y.V2) and B.1.1.28.1 (P.1), have emerged in the United Kingdom, South Africa and Brazil, respectively. Here, we have presented global viewpoint on implications of emerging SARS-CoV-2 variants based on structural-function impact of crucial mutations occurring in its spike (S), ORF8 and nucleocapsid (N) proteins. While the N501Y mutation was observed in all three lineages, the 501Y.V1 and P.1 accumulated a different set of mutations in the S protein. The missense mutational effects were predicted through a COVID-19 dedicated resource followed by atomistic molecular dynamics simulations. Current findings indicate that some mutations in the S protein might lead to higher affinity with host receptors and resistance against antibodies, but not all are due to different antibody binding (epitope) regions. Mutations may, however, result in diagnostic tests failures and possible interference with binding of newly identified anti-viral candidates against SARS-CoV-2, likely necessitating roll out of recurring "flu-like shots" annually for tackling COVID-19. The functional relevance of these mutations has been described in terms of modulation of host tropism, antibody resistance, diagnostic sensitivity and therapeutic candidates. Besides global economic losses, post-vaccine reinfections with emerging variants can have significant clinical, therapeutic and public health impacts.


Asunto(s)
/virología , /genética , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , /terapia , /inmunología , Humanos , Simulación de Dinámica Molecular , Mutación , Salud Pública , /inmunología , Glicoproteína de la Espiga del Coronavirus/genética , Glicoproteína de la Espiga del Coronavirus/inmunología
6.
Viruses ; 13(3)2021 03 20.
Artículo en Inglés | MEDLINE | ID: mdl-33804667

RESUMEN

While SARS-CoV-2 specific neutralizing antibodies have been developed for therapeutic purposes, the specific viral triggers that drive the generation of SARS-CoV-2 specific IgG and IgM antibodies remain only partially characterized. Moreover, it is unknown whether endogenously derived antibodies drive viral clearance that might result in mitigation of clinical severity during natural infection. We developed a series of non-linear mathematical models to investigate whether SARS-CoV-2 viral and antibody kinetics are coupled or governed by separate processes. Patients with severe disease had a higher production rate of IgG but not IgM antibodies. Maximal levels of both isotypes were governed by their production rate rather than different saturation levels between people. Our results suggest that an exponential surge in IgG levels occurs approximately 5-10 days after symptom onset with no requirement for continual antigenic stimulation. SARS-CoV-2 specific IgG antibodies appear to have limited to no effect on viral dynamics but may enhance viral clearance late during primary infection resulting from the binding effect of antibody to virus, rather than neutralization. In conclusion, SARS-CoV-2 specific IgG antibodies may play only a limited role in clearing infection from the nasal passages despite providing long-term immunity against infection following vaccination or prior infection.


Asunto(s)
Anticuerpos Antivirales/inmunología , Inmunoglobulina G/inmunología , Nariz/virología , /fisiología , Anticuerpos Neutralizantes/inmunología , Humanos , Inmunoglobulina M/inmunología , Estudios Longitudinales , Nariz/inmunología , /inmunología , Esparcimiento de Virus
7.
Viruses ; 13(3)2021 03 23.
Artículo en Inglés | MEDLINE | ID: mdl-33806922

RESUMEN

Since the coronavirus disease (COVID-19) pandemic was first identified in early 2020, rare cases of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection in pet cats have been reported worldwide. Some reports of cats with SARS-CoV-2 showed self-limiting respiratory or gastrointestinal disease after suspected human-to-feline transmission via close contact with humans with SARS-CoV-2. In the present study, we investigated a cat with SARS-CoV-2 that was presented to a private animal clinic in Northern Italy in May 2020 in a weak clinical condition due to an underlying intestinal B-cell lymphoma. The cat developed signs of respiratory tract disease, including a sneeze, a cough and ocular discharge, three days after an oropharyngeal swab tested positive for SARS-CoV-2 viral RNA using two real-time reverse transcriptase polymerase chain reaction (RT-qPCR) assays for the envelope (E) and RNA-dependent RNA polymerase (RdRp) gene. Thus, SARS-CoV-2 viral RNA was detectable prior to the onset of clinical signs. Five and six months after positive molecular results, the serological testing substantiated the presence of a SARS-CoV-2 infection in the cat with the detection of anti-SARS-CoV-2 receptor binding domain (RBD) immunoglobulin (IgG) antibodies and neutralizing activity in a surrogate virus neutralization assay (sVNT). To the best of our knowledge, this extends the known duration of seropositivity of SARS-CoV-2 in a cat. Our study provides further evidence that cats are susceptible to SARS-CoV-2 under natural conditions and strengthens the assumption that comorbidities may play a role in the development of clinical disease.


Asunto(s)
Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Enfermedades de los Gatos/inmunología , Linfoma de Células B/veterinaria , Animales , Formación de Anticuerpos , /virología , Enfermedades de los Gatos/virología , Gatos , Inmunoglobulina G/inmunología , Italia , Linfoma de Células B/inmunología , Linfoma de Células B/virología , Glicoproteína de la Espiga del Coronavirus
8.
Front Immunol ; 12: 653189, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33828563

RESUMEN

After the pandemic of COVID-19, neutralizing antibodies (NAbs) against SARS-CoV-2 have been developed for the prophylactic and therapeutic purposes. However, few methodologies are described in detail on how to rapidly and efficiently generate effective NAbs to SARS-CoV-2. Here, we integrated and optimized a strategically screening method for NAbs, which has enabled us to obtain SARS-CoV-2 receptor-binding domain (RBD) specific NAbs within 6 days, followed by additional 9 days for antibody production and function analysis. Using this method, we obtained 198 specific Abs against SARS-CoV-2 RBD from the blood samples of COVID-19 convalescent patients, and 96 of them showed neutralizing activity. At least 20% of these NAbs exhibited advanced neutralizing potency and high affinity, with the top two NAbs showing half-maximal inhibitory concentration (IC50) to block authentic SARS-CoV-2 at 9.88 and 11.13 ng/ml, respectively. Altogether, our study provides an effective methodology with high applicable value for discovering potential preventative and therapeutic NAbs for the emerging infectious diseases.


Asunto(s)
Anticuerpos Neutralizantes , Anticuerpos Antivirales , Anticuerpos Neutralizantes/sangre , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/sangre , Anticuerpos Antivirales/inmunología , /inmunología , Humanos , /metabolismo
9.
PLoS Pathog ; 17(4): e1009453, 2021 04.
Artículo en Inglés | MEDLINE | ID: mdl-33831132

RESUMEN

There is intense interest in antibody immunity to coronaviruses. However, it is unknown if coronaviruses evolve to escape such immunity, and if so, how rapidly. Here we address this question by characterizing the historical evolution of human coronavirus 229E. We identify human sera from the 1980s and 1990s that have neutralizing titers against contemporaneous 229E that are comparable to the anti-SARS-CoV-2 titers induced by SARS-CoV-2 infection or vaccination. We test these sera against 229E strains isolated after sera collection, and find that neutralizing titers are lower against these "future" viruses. In some cases, sera that neutralize contemporaneous 229E viral strains with titers >1:100 do not detectably neutralize strains isolated 8-17 years later. The decreased neutralization of "future" viruses is due to antigenic evolution of the viral spike, especially in the receptor-binding domain. If these results extrapolate to other coronaviruses, then it may be advisable to periodically update SARS-CoV-2 vaccines.


Asunto(s)
Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Coronavirus Humano 229E/genética , Coronavirus Humano 229E/inmunología , Evasión Inmune , Humanos , /inmunología
10.
Viruses ; 13(4)2021 03 26.
Artículo en Inglés | MEDLINE | ID: mdl-33810465

RESUMEN

Monoclonal antibodies represent an important avenue for COVID-19 therapy and are routinely used for rapid and accessible diagnosis of SARS-CoV-2 infection. The recent emergence of SARS-CoV-2 genetic variants emphasized the need to enlarge the repertoire of antibodies that target diverse epitopes, the combination of which may improve immune-diagnostics, augment the efficiency of the immunotherapy and prevent selection of escape-mutants. Antigen-specific controlled immunization of experimental animals may elicit antibody repertoires that significantly differ from those generated in the context of the immune response mounted in the course of disease. Accordingly, rabbits were immunized by several recombinant antigens representing distinct domains of the viral spike protein and monoclonal antibodies were isolated from single cells obtained by cell sorting. Characterization of a panel of successfully isolated anti-receptor binding domain (RBD) and anti-N-terminal domain (NTD) antibodies demonstrated that they exhibit high specificity and affinity profiles. Anti-RBD antibodies revealing significant neutralizing potency against SARS-CoV-2 in vitro were found to target at least three distinct epitopes. Epitope mapping established that two of these antibodies recognized a novel epitope located on the surface of the RBD. We suggest that the antibodies isolated in this study are useful for designing SARS-CoV-2 diagnosis and therapy approaches.


Asunto(s)
Anticuerpos Antivirales/inmunología , /inmunología , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/inmunología , Animales , Anticuerpos Neutralizantes/inmunología , Mapeo Epitopo , Epítopos/química , Epítopos/genética , Epítopos/inmunología , Femenino , Humanos , Pruebas de Neutralización , Conejos , /genética , Glicoproteína de la Espiga del Coronavirus/genética
11.
MAbs ; 13(1): 1893426, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33666135

RESUMEN

Numerous neutralizing antibodies that target SARS-CoV-2 have been reported, and most directly block binding of the viral Spike receptor-binding domain (RBD) to angiotensin-converting enzyme II (ACE2). Here, we deliberately exploit non-neutralizing RBD antibodies, showing they can dramatically assist in neutralization when linked to neutralizing binders. We identified antigen-binding fragments (Fabs) by phage display that bind RBD, but do not block ACE2 or neutralize virus as IgGs. When these non-neutralizing Fabs were assembled into bispecific VH/Fab IgGs with a neutralizing VH domain, we observed a ~ 25-fold potency improvement in neutralizing SARS-CoV-2 compared to the mono-specific bi-valent VH-Fc alone or the cocktail of the VH-Fc and IgG. This effect was epitope-dependent, reflecting the unique geometry of the bispecific antibody toward Spike. Our results show that a bispecific antibody that combines both neutralizing and non-neutralizing epitopes on Spike-RBD is a promising and rapid engineering strategy to improve the potency of SARS-CoV-2 antibodies.


Asunto(s)
Anticuerpos Biespecíficos/inmunología , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Epítopos/inmunología , Fragmentos Fab de Inmunoglobulinas/inmunología , Glicoproteína de la Espiga del Coronavirus/inmunología , Anticuerpos Biespecíficos/genética , Anticuerpos Biespecíficos/uso terapéutico , Anticuerpos Neutralizantes/genética , Anticuerpos Neutralizantes/uso terapéutico , Anticuerpos Antivirales/genética , Anticuerpos Antivirales/uso terapéutico , /genética , Epítopos/genética , Células HEK293 , Humanos , Fragmentos Fab de Inmunoglobulinas/genética , Fragmentos Fab de Inmunoglobulinas/uso terapéutico , Glicoproteína de la Espiga del Coronavirus/genética
12.
Nat Commun ; 12(1): 1346, 2021 03 01.
Artículo en Inglés | MEDLINE | ID: mdl-33649323

RESUMEN

SARS-CoV-2 is the underlying cause for the COVID-19 pandemic. Like most enveloped RNA viruses, SARS-CoV-2 uses a homotrimeric surface antigen to gain entry into host cells. Here we describe S-Trimer, a native-like trimeric subunit vaccine candidate for COVID-19 based on Trimer-Tag technology. Immunization of S-Trimer with either AS03 (oil-in-water emulsion) or CpG 1018 (TLR9 agonist) plus alum adjuvants induced high-level of neutralizing antibodies and Th1-biased cellular immune responses in animal models. Moreover, rhesus macaques immunized with adjuvanted S-Trimer were protected from SARS-CoV-2 challenge compared to vehicle controls, based on clinical observations and reduction of viral loads in lungs. Trimer-Tag may be an important platform technology for scalable production and rapid development of safe and effective subunit vaccines against current and future emerging RNA viruses.


Asunto(s)
/uso terapéutico , /prevención & control , /patogenicidad , Animales , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Western Blotting , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , Inmunidad Celular/fisiología , Inmunización Pasiva , Inmunohistoquímica , Macaca mulatta , Ratones , Ratones Endogámicos BALB C , Microscopía Electrónica , /inmunología
13.
J Exp Med ; 218(5)2021 05 03.
Artículo en Inglés | MEDLINE | ID: mdl-33656515

RESUMEN

The neutralizing antibody response to SARS-CoV-2 is dominated by antibodies deriving from germlines IGHV3-53/IGHV3-66, which are also associated with self-reacting antibodies. Could vaccines avoid the expansion of this immunodominant response, decrease the risk of autoimmunity, and still protect against emerging SARS-CoV-2 variants?


Asunto(s)
Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Células Germinativas/inmunología , Interacciones Huésped-Patógeno/inmunología , /inmunología , Antígenos Virales/inmunología , /virología , Interacciones Huésped-Patógeno/genética , Humanos
14.
Cell Host Microbe ; 29(3): 322-324, 2021 03 10.
Artículo en Inglés | MEDLINE | ID: mdl-33705702

RESUMEN

In the latest issues of Cell Host & Microbe and Cell, three articles describe new mutations in the SARS-CoV-2 Spike receptor binding domain that escape neutralizing responses. These highlight the importance of surveillance of SARS-CoV-2 evolution to anticipate and manage new variants that could impact reinfection, vaccine efficacy, and immunotherapies.


Asunto(s)
Anticuerpos Neutralizantes/inmunología , /inmunología , Anticuerpos Antivirales/inmunología , Sitios de Unión , Humanos , Evasión Inmune , Mutación , Unión Proteica , Dominios Proteicos , Receptores Virales , Glicoproteína de la Espiga del Coronavirus/genética , Glicoproteína de la Espiga del Coronavirus/inmunología
15.
Cell Host Microbe ; 29(3): 327-333, 2021 03 10.
Artículo en Inglés | MEDLINE | ID: mdl-33705704

RESUMEN

Most viral vaccines are based on inducing neutralizing antibodies (NAbs) against the virus envelope or spike glycoproteins. Many viral surface proteins exist as trimers that transition from a pre-fusion state when key NAb epitopes are exposed to a post-fusion form in which the potential for virus-cell fusion no longer exists. For optimal vaccine performance, these viral proteins are often engineered to enhance stability and presentation of these NAb epitopes. The method involves the structure-guided introduction of proline residues at key positions that maintain the trimer in the pre-fusion configuration. We review how this technique emerged during HIV-1 Env vaccine development and its subsequent wider application to other viral vaccines including SARS-CoV-2.


Asunto(s)
Prolina/química , Prolina/inmunología , Vacunas Virales/química , Vacunas Virales/inmunología , Anticuerpos Neutralizantes/inmunología , /genética , Humanos , Modelos Moleculares , Prolina/genética , Ingeniería de Proteínas , Vacunas Virales/genética
16.
PLoS One ; 16(3): e0248348, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33690649

RESUMEN

Pseudoviruses are useful surrogates for highly pathogenic viruses because of their safety, genetic stability, and scalability for screening assays. Many different pseudovirus platforms exist, each with different advantages and limitations. Here we report our efforts to optimize and characterize an HIV-based lentiviral pseudovirus assay for screening neutralizing antibodies for SARS-CoV-2 using a stable 293T cell line expressing human angiotensin converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2). We assessed different target cells, established conditions that generate readouts over at least a two-log range, and confirmed consistent neutralization titers over a range of pseudovirus input. Using reference sera and plasma panels, we evaluated assay precision and showed that our neutralization titers correlate well with results reported in other assays. Overall, our lentiviral assay is relatively simple, scalable, and suitable for a variety of SARS-CoV-2 entry and neutralization screening assays.


Asunto(s)
/metabolismo , Lentivirus/metabolismo , Pruebas de Neutralización/métodos , /genética , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Evaluación Preclínica de Medicamentos/métodos , Células HEK293 , Humanos , /patogenicidad , Glicoproteína de la Espiga del Coronavirus/genética
18.
Cell Rep ; 34(13): 108915, 2021 03 30.
Artículo en Inglés | MEDLINE | ID: mdl-33761319

RESUMEN

To fully decipher the immunogenicity of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Spike protein, it is essential to assess which part is highly immunogenic in a systematic way. We generate a linear epitope landscape of the Spike protein by analyzing the serum immunoglobulin G (IgG) response of 1,051 coronavirus disease 2019 (COVID-19) patients with a peptide microarray. We reveal two regions rich in linear epitopes, i.e., C-terminal domain (CTD) and a region close to the S2' cleavage site and fusion peptide. Unexpectedly, we find that the receptor binding domain (RBD) lacks linear epitope. We reveal that the number of responsive peptides is highly variable among patients and correlates with disease severity. Some peptides are moderately associated with severity and clinical outcome. By immunizing mice, we obtain linear-epitope-specific antibodies; however, no significant neutralizing activity against the authentic virus is observed for these antibodies. This landscape will facilitate our understanding of SARS-CoV-2-specific humoral responses and might be useful for vaccine refinement.


Asunto(s)
/inmunología , Glicoproteína de la Espiga del Coronavirus/inmunología , Adulto , Animales , Anticuerpos Monoclonales/inmunología , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Antígenos Virales/inmunología , /genética , China/epidemiología , Modelos Animales de Enfermedad , Mapeo Epitopo/métodos , Epítopos/inmunología , Femenino , Humanos , Inmunoglobulina G/inmunología , Masculino , Ratones , Ratones Endogámicos BALB C , Glicoproteína de la Espiga del Coronavirus/genética , Glicoproteína de la Espiga del Coronavirus/metabolismo
19.
Diabetes Metab Syndr ; 15(2): 505-508, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33662837

RESUMEN

BACKGROUND AND AIMS: To summarize the available evidence on the use COVID-19 vaccines in patients with diabetes mellitus. METHODS: We performed a thorough literature search with regard to COVID-19 vaccines in patients with type 1 and type 2 diabetes mellitus. RESULTS: The novel coronavirus disease (COVID-19) tends to portend a poor prognosis in patients with diabetes mellitus (DM). Primary prevention remains the mainstay for mitigating the risks associated with COVID-19 in patients with DM. A significant step in primary prevention is timely vaccination. Routine vaccination against pneumococcal pneumonia, influenza, and hepatitis B is recommended in patients with DM with good efficacy and reasonable safety profile. With clinical data supporting a robust neutralizing antibody response in COVID-19 patients with DM, vaccination in individuals with DM is justified. In fact, as the burden of the disease is borne by people with DM, COVID-19 vaccination should be prioritized in individuals with DM. Multiple unresolved issues with regard to preferred vaccine type, vaccine efficacy and durability, frequency of administration, vaccination in children (<18 years) and pregnant/lactating women however remain, and need to be addressed through future research. CONCLUSIONS: Patients with type 1 and type 2 diabetes mellitus are at a high risk of poor prognosis with COVID-19 and vaccination should be prioritized in them. However, many unresolved issues with regard to COVID-19 vaccination need to be addressed through future research.


Asunto(s)
/uso terapéutico , Diabetes Mellitus Tipo 1 , Diabetes Mellitus Tipo 2 , Anticuerpos Neutralizantes/inmunología , Humanos , Factores de Riesgo
20.
Sci Rep ; 11(1): 5558, 2021 03 10.
Artículo en Inglés | MEDLINE | ID: mdl-33692386

RESUMEN

The recent COVID-19 pandemic poses a serious threat to global public health, thus there is an urgent need to define the molecular mechanisms involved in SARS-CoV-2 spike (S) protein-mediated virus entry that is essential for preventing and/or treating this emerging infectious disease. In this study, we examined the blocking activity of human COVID-19 convalescent plasma by cell-cell fusion assays using SARS-CoV-2-S-transfected 293 T as effector cells and ACE2-expressing 293 T as target cells. We demonstrate that the SARS-CoV-2 S protein exhibits a very high capacity for membrane fusion and is efficient in mediating virus fusion and entry into target cells. Importantly, we find that COVID-19 convalescent plasma with high titers of IgG neutralizing antibodies can block cell-cell fusion and virus entry by interfering with the SARS-CoV-2-S/ACE2 or SARS-CoV-S/ACE2 interactions. These findings suggest that COVID-19 convalescent plasma may not only inhibit SARS-CoV-2-S but also cross-neutralize SARS-CoV-S-mediated membrane fusion and virus entry, supporting its potential as a preventive and/or therapeutic agent against SARS-CoV-2 as well as other SARS-CoV infections.


Asunto(s)
/inmunología , Glicoproteína de la Espiga del Coronavirus/inmunología , Adulto , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Fusión Celular/métodos , Femenino , Humanos , Inmunización Pasiva/métodos , Masculino , Fusión de Membrana/efectos de los fármacos , Persona de Mediana Edad , Pandemias/prevención & control , Plasma/química , Receptores Virales/metabolismo , /patogenicidad , Glicoproteína de la Espiga del Coronavirus/metabolismo , Internalización del Virus/efectos de los fármacos
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