Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 8 de 8
Filtrar
1.
J Infect Dis ; 220(10): 1558-1567, 2019 10 08.
Artículo en Inglés | MEDLINE | ID: mdl-30911758

RESUMEN

BACKGROUND: Infection control measures have played a major role in limiting human/camel-to-human transmission of Middle East respiratory syndrome coronavirus (MERS-CoV); however, development of effective and safe human or camel vaccines is warranted. METHODS: We extended and optimized our previous recombinant adenovirus 5 (rAd5)-based vaccine platform characterized by in vivo amplified and CD40-mediated specific responses to generate MERS-CoV S1 subunit-based vaccine. We generated rAd5 constructs expressing CD40-targeted S1 fusion protein (rAd5-S1/F/CD40L), untargeted S1 (rAd5-S1), and Green Fluorescent Protein (rAd5-GFP), and evaluated their efficacy and safety in human dipeptidyl peptidase 4 transgenic (hDPP4 Tg+) mice. RESULTS: Immunization of hDPP4 Tg+ mice with a single dose of rAd5-S1/F/CD40L elicited as robust and significant specific immunoglobulin G and neutralizing antibodies as those induced with 2 doses of rAd5-S1. After MERS-CoV challenge, both vaccines conferred complete protection against morbidity and mortality, as evidenced by significantly undetectable/reduced pulmonary viral loads compared to the control group. However, rAd5-S1- but not rAd5-S1/F/CD40L-immunized mice exhibited marked pulmonary perivascular hemorrhage post-MERS-CoV challenge despite the observed protection. CONCLUSIONS: Incorporation of CD40L into rAd5-based MERS-CoV S1 vaccine targeting molecule and molecular adjuvants not only enhances immunogenicity and efficacy but also prevents inadvertent pulmonary pathology after viral challenge, thereby offering a promising strategy to enhance safety and potency of vaccines.


Asunto(s)
Ligando de CD40/farmacología , Infecciones por Coronavirus/prevención & control , Coronavirus del Síndrome Respiratorio de Oriente Medio/inmunología , Glicoproteína de la Espiga del Coronavirus/inmunología , Vacunas Virales/inmunología , Adenovirus Humanos/genética , Adyuvantes Inmunológicos/genética , Adyuvantes Inmunológicos/farmacología , Animales , Anticuerpos Neutralizantes/sangre , Anticuerpos Antivirales/sangre , Ligando de CD40/genética , Infecciones por Coronavirus/inmunología , Dipeptidil Peptidasa 4/genética , Dipeptidil Peptidasa 4/metabolismo , Portadores de Fármacos , Vectores Genéticos , Inmunoglobulina G/sangre , Pulmón/virología , Ratones , Ratones Transgénicos , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/inmunología , Proteínas Recombinantes de Fusión/farmacología , Glicoproteína de la Espiga del Coronavirus/genética , Análisis de Supervivencia , Vacunas de Subunidad/administración & dosificación , Vacunas de Subunidad/genética , Vacunas de Subunidad/inmunología , Vacunas Sintéticas/administración & dosificación , Vacunas Sintéticas/genética , Vacunas Sintéticas/inmunología , Carga Viral , Vacunas Virales/administración & dosificación , Vacunas Virales/genética
2.
J Virol ; 90(1): 57-67, 2016 01 01.
Artículo en Inglés | MEDLINE | ID: mdl-26446606

RESUMEN

UNLABELLED: Characterized animal models are needed for studying the pathogenesis of and evaluating medical countermeasures for persisting Middle East respiratory syndrome-coronavirus (MERS-CoV) infections. Here, we further characterized a lethal transgenic mouse model of MERS-CoV infection and disease that globally expresses human CD26 (hCD26)/DPP4. The 50% infectious dose (ID50) and lethal dose (LD50) of virus were estimated to be <1 and 10 TCID50 of MERS-CoV, respectively. Neutralizing antibody developed in the surviving mice from the ID50/LD50 determinations, and all were fully immune to challenge with 100 LD50 of MERS-CoV. The tissue distribution and histopathology in mice challenged with a potential working dose of 10 LD50 of MERS-CoV were subsequently evaluated. In contrast to the overwhelming infection seen in the mice challenged with 10(5) LD50 of MERS-CoV, we were able to recover infectious virus from these mice only infrequently, although quantitative reverse transcription-PCR (qRT-PCR) tests indicated early and persistent lung infection and delayed occurrence of brain infection. Persistent inflammatory infiltrates were seen in the lungs and brain stems at day 2 and day 6 after infection, respectively. While focal infiltrates were also noted in the liver, definite pathology was not seen in other tissues. Finally, using a receptor binding domain protein vaccine and a MERS-CoV fusion inhibitor, we demonstrated the value of this model for evaluating vaccines and antivirals against MERS. As outcomes of MERS-CoV infection in patients differ greatly, ranging from asymptomatic to overwhelming disease and death, having available both an infection model and a lethal model makes this transgenic mouse model relevant for advancing MERS research. IMPORTANCE: Fully characterized animal models are essential for studying pathogenesis and for preclinical screening of vaccines and drugs against MERS-CoV infection and disease. When given a high dose of MERS-CoV, our transgenic mice expressing hCD26/DPP4 viral receptor uniformly succumbed to death within 6 days, making it difficult to evaluate host responses to infection and disease. We further characterized this model by determining both the ID50 and the LD50 of MERS-CoV in order to establish both an infection model and a lethal model for MERS and followed this by investigating the antibody responses and immunity of the mice that survived MERS-CoV infection. Using the estimated LD50 and ID50 data, we dissected the kinetics of viral tissue distribution and pathology in mice challenged with 10 LD50 of virus and utilized the model for preclinical evaluation of a vaccine and drug for treatment of MERS-CoV infection. This further-characterized transgenic mouse model will be useful for advancing MERS research.


Asunto(s)
Infecciones por Coronavirus/patología , Infecciones por Coronavirus/virología , Modelos Animales de Enfermedad , Coronavirus del Síndrome Respiratorio de Oriente Medio/crecimiento & desarrollo , Estructuras Animales/patología , Estructuras Animales/virología , Animales , Anticuerpos Neutralizantes/sangre , Anticuerpos Antivirales/sangre , Antivirales/administración & dosificación , Encéfalo/patología , Encéfalo/virología , Infecciones por Coronavirus/tratamiento farmacológico , Infecciones por Coronavirus/prevención & control , Evaluación Preclínica de Medicamentos/métodos , Histocitoquímica , Humanos , Dosificación Letal Mediana , Hígado/patología , Hígado/virología , Pulmón/patología , Pulmón/virología , Ratones Transgénicos , Coronavirus del Síndrome Respiratorio de Oriente Medio/inmunología , Análisis de Supervivencia , Resultado del Tratamiento , Vacunas Virales/administración & dosificación , Vacunas Virales/inmunología
3.
J Virol ; 89(7): 3659-70, 2015 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-25589660

RESUMEN

UNLABELLED: The emergence of Middle East respiratory syndrome-coronavirus (MERS-CoV) in the Middle East since 2012 has caused more than 900 human infections with ∼40% mortality to date. Animal models are needed for studying pathogenesis and for development of preventive and therapeutic agents against MERS-CoV infection. Nonhuman primates (rhesus macaques and marmosets) are expensive models of limited availability. Although a mouse lung infection model has been described using adenovirus vectors expressing human CD26/dipeptidyl peptidase 4 (DPP4), it is believed that a transgenic mouse model is needed for MERS-CoV research. We have developed this transgenic mouse model as indicated in this study. We show that transgenic mice globally expressing hCD26/DPP4 were fully permissive to MERS-CoV infection, resulting in relentless weight loss and death within days postinfection. High infectious virus titers were recovered primarily from the lungs and brains of mice at 2 and 4 days postinfection, respectively, whereas viral RNAs were also detected in the heart, spleen, and intestine, indicating a disseminating viral infection. Infected Tg(+) mice developed a progressive pneumonia, characterized by extensive inflammatory infiltration. In contrast, an inconsistent mild perivascular cuffing was the only pathological change associated with the infected brains. Moreover, infected Tg(+) mice were able to activate genes encoding for many antiviral and inflammatory mediators within the lungs and brains, coinciding with the high levels of viral replication. This new and unique transgenic mouse model will be useful for furthering knowledge of MERS pathogenesis and for the development of vaccine and treatments against MERS-CoV infection. IMPORTANCE: Small and economical animal models are required for the controlled and extensive studies needed for elucidating pathogenesis and development of vaccines and antivirals against MERS. Mice are the most desirable small-animal species for this purpose because of availability and the existence of a thorough knowledge base, particularly of genetics and immunology. The standard small animals, mice, hamsters, and ferrets, all lack the functional MERS-CoV receptor and are not susceptible to infection. So, initial studies were done with nonhuman primates, expensive models of limited availability. A mouse lung infection model was described where a mouse adenovirus was used to transfect lung cells for receptor expression. Nevertheless, all generally agree that a transgenic mouse model expressing the DPP4 receptor is needed for MERS-CoV research. We have developed this transgenic mouse model as indicated in this study. This new and unique transgenic mouse model will be useful for furthering MERS research.


Asunto(s)
Infecciones por Coronavirus/patología , Infecciones por Coronavirus/virología , Modelos Animales de Enfermedad , Coronavirus del Síndrome Respiratorio de Oriente Medio/fisiología , Estructuras Animales/virología , Animales , Dipeptidil Peptidasa 4/biosíntesis , Dipeptidil Peptidasa 4/genética , Expresión Génica , Humanos , Ratones Transgénicos , Factores de Tiempo , Carga Viral
4.
Vaccine ; 38(47): 7533-7541, 2020 11 03.
Artículo en Inglés | MEDLINE | ID: mdl-33039209

RESUMEN

We developed a severe acute respiratory syndrome (SARS) subunit recombinant protein vaccine candidate based on a high-yielding, yeast-engineered, receptor-binding domain (RBD219-N1) of the SARS beta-coronavirus (SARS-CoV) spike (S) protein. When formulated with Alhydrogel®, RBD219-N1 induced high levels of neutralizing antibodies against both pseudotyped virus and a clinical (mouse-adapted) isolate of SARS-CoV. Here, we report that mice immunized with RBD219-N1/Alhydrogel® were fully protected from lethal SARS-CoV challenge (0% mortality), compared to ~30% mortality in mice immunized with the SARS S protein formulated with Alhydrogel®, and 100% mortality in negative controls. An RBD219-N1 formulation with Alhydrogel® was also superior to the S protein, unadjuvanted RBD, and AddaVax (MF59-like adjuvant)-formulated RBD in inducing specific antibodies and preventing cellular infiltrates in the lungs upon SARS-CoV challenge. Specifically, a formulation with a 1:25 ratio of RBD219-N1 to Alhydrogel® provided high neutralizing antibody titers, 100% protection with non-detectable viral loads with minimal or no eosinophilic pulmonary infiltrates. As a result, this vaccine formulation is under consideration for further development against SARS-CoV and potentially other emerging and re-emerging beta-CoVs such as SARS-CoV-2.


Asunto(s)
Betacoronavirus/inmunología , Infecciones por Coronavirus/prevención & control , Pandemias/prevención & control , Neumonía Viral/prevención & control , Glicoproteína de la Espiga del Coronavirus/inmunología , Vacunas de Subunidad/inmunología , Vacunas Sintéticas/inmunología , Vacunas Virales/inmunología , Hidróxido de Aluminio/administración & dosificación , Animales , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , COVID-19 , Vacunas contra la COVID-19 , Infecciones por Coronavirus/inmunología , Femenino , Humanos , Ratones , Ratones Endogámicos BALB C , Dominios Proteicos/inmunología , Proteínas Recombinantes/inmunología , SARS-CoV-2 , Síndrome Respiratorio Agudo Grave/prevención & control , Glicoproteína de la Espiga del Coronavirus/genética , Carga Viral/inmunología
5.
Sci Adv ; 5(4): eaav4580, 2019 04.
Artículo en Inglés | MEDLINE | ID: mdl-30989115

RESUMEN

Continuously emerging highly pathogenic human coronaviruses (HCoVs) remain a major threat to human health, as illustrated in past SARS-CoV and MERS-CoV outbreaks. The development of a drug with broad-spectrum HCoV inhibitory activity would address this urgent unmet medical need. Although previous studies have suggested that the HR1 of HCoV spike (S) protein is an important target site for inhibition against specific HCoVs, whether this conserved region could serve as a target for the development of broad-spectrum pan-CoV inhibitor remains controversial. Here, we found that peptide OC43-HR2P, derived from the HR2 domain of HCoV-OC43, exhibited broad fusion inhibitory activity against multiple HCoVs. EK1, the optimized form of OC43-HR2P, showed substantially improved pan-CoV fusion inhibitory activity and pharmaceutical properties. Crystal structures indicated that EK1 can form a stable six-helix bundle structure with both short α-HCoV and long ß-HCoV HR1s, further supporting the role of HR1 region as a viable pan-CoV target site.


Asunto(s)
Infecciones por Coronavirus/virología , Coronavirus/efectos de los fármacos , Coronavirus/metabolismo , Dominios y Motivos de Interacción de Proteínas/efectos de los fármacos , Glicoproteína de la Espiga del Coronavirus/antagonistas & inhibidores , Glicoproteína de la Espiga del Coronavirus/química , Inhibidores de Proteínas Virales de Fusión/química , Inhibidores de Proteínas Virales de Fusión/farmacología , Secuencia de Aminoácidos , Animales , Línea Celular , Coronavirus/clasificación , Coronavirus/genética , Infecciones por Coronavirus/tratamiento farmacológico , Modelos Animales de Enfermedad , Humanos , Fusión de Membrana/efectos de los fármacos , Ratones , Modelos Moleculares , Péptidos/química , Péptidos/farmacología , Filogenia , Conformación Proteica , Dominios Proteicos , Solubilidad
6.
Sci Rep ; 6: 31629, 2016 08 19.
Artículo en Inglés | MEDLINE | ID: mdl-27538452

RESUMEN

Middle East Respiratory Syndrome coronavirus (MERS-CoV) has repeatedly caused outbreaks in the Arabian Peninsula. To date, no approved medical countermeasures (MCM) are available to combat MERS-CoV infections. Several neutralizing human monoclonal antibodies (mAbs), including m336, a germline-like human mAb, have been chosen as promising MCM for MERS-CoV. However, their clinical development has been hindered by the lack of a robust animal model that recapitulate the morbidity and mortality of human infections. We assessed the prophylactic and therapeutic efficacy of m336 by using well-characterized transgenic mice shown to be highly sensitive to MERS-CoV infection and disease. We found that mice treated with m336 prior to or post lethal MERS-CoV challenging were fully protected, compared to control mice which sufferered from profound weight loss and uniform death within days after infection. Taken together, these results support further development of m336 and other human monoclonal antibodies as potential therapeutics for MERS-CoV infection.


Asunto(s)
Anticuerpos Monoclonales/farmacología , Anticuerpos Neutralizantes/farmacología , Anticuerpos Antivirales/farmacología , Infecciones por Coronavirus/tratamiento farmacológico , Inmunización Pasiva , Coronavirus del Síndrome Respiratorio de Oriente Medio/inmunología , Animales , Anticuerpos Monoclonales/inmunología , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Infecciones por Coronavirus/inmunología , Humanos , Ratones , Ratones Transgénicos
7.
Hum Vaccin Immunother ; 12(9): 2351-6, 2016 09.
Artículo en Inglés | MEDLINE | ID: mdl-27269431

RESUMEN

To determine if a hypersensitive-type lung pathology might occur when mice were given an inactivated MERS-CoV vaccine and challenged with infectious virus as was seen with SARS-CoV vaccines, we prepared and vaccinated mice with an inactivated MERS-CoV vaccine. Neutralizing antibody was induced by vaccine with and without adjuvant and lung virus was reduced in vaccinated mice after challenge. Lung mononuclear infiltrates occurred in all groups after virus challenge but with increased infiltrates that contained eosinophils and increases in the eosinophil promoting IL-5 and IL-13 cytokines only in the vaccine groups. Inactivated MERS-CoV vaccine appears to carry a hypersensitive-type lung pathology risk from MERS-CoV infection that is similar to that found with inactivated SARS-CoV vaccines from SARS-CoV infection.


Asunto(s)
Infecciones por Coronavirus/prevención & control , Pulmón/patología , Coronavirus del Síndrome Respiratorio de Oriente Medio/inmunología , Vacunas Virales/inmunología , Animales , Anticuerpos Neutralizantes/sangre , Anticuerpos Antivirales/sangre , Infecciones por Coronavirus/patología , Hipersensibilidad/patología , Ratones Transgénicos , Vacunas de Productos Inactivados/administración & dosificación , Vacunas de Productos Inactivados/efectos adversos , Vacunas de Productos Inactivados/inmunología , Vacunas Virales/administración & dosificación
8.
J Clin Virol ; 45(4): 358-61, 2009 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-19570709

RESUMEN

BACKGROUND: Although recognized as a significant respiratory pathogen, there is a dearth of information on genetic diversity of human respiratory syncytial virus (HRSV) strains among children in India. OBJECTIVES: To study prevalence and genomic diversity of HRSV strains among children (<5 years of age) with acute respiratory tract infection (ARTI) at Dr. B.C. Roy Memorial Hospital for Children, Kolkata, Eastern India. STUDY DESIGN: During September 2005 to August 2008, nasal and/or throat swabs from children with ARTI were screened for presence of HRSVs by RT-PCR of nucleocapsid (N) and glycoprotein (G) genes. Classification of G gene of HRSV strains were achieved with different primer pairs designed for amplification of N'- and/or C'-terminal hypervariable regions (HVR1 and HVR2) of HRSV A and B strains. The HVR1 and HVR2 of G gene of 27 and 22 HRSV B strains were sequenced. RESULTS: One hundred seventy seven of 1720 clinical samples were positive for HRSVs. Group B strains were detected at higher rates (95% against 5%, n=80) than A in 2005-2006, whereas in consecutive years, the rate of detection of group A were higher (94.84% against 5.16%, n=97). The group B strains were genetically related to globally spreading BA genotype, exhibiting conserved nature of stop codon, six nucleotide deletions in HVR1 and formed single phylogenetic clusters for both HVR1 and HVR2. CONCLUSIONS: The detection of significant high rates of group B strains in 2005-2006 followed by increase in prevalence of A strains in subsequent years highlight the dynamic nature of prevalence of HRSV subtypes among children with ARTI in Eastern India.


Asunto(s)
Infecciones por Virus Sincitial Respiratorio/epidemiología , Infecciones por Virus Sincitial Respiratorio/virología , Virus Sincitial Respiratorio Humano/clasificación , Virus Sincitial Respiratorio Humano/genética , Infecciones del Sistema Respiratorio/epidemiología , Infecciones del Sistema Respiratorio/virología , Preescolar , Análisis por Conglomerados , Variación Genética , Genotipo , Hospitales , Humanos , India/epidemiología , Lactante , Recién Nacido , Epidemiología Molecular , Datos de Secuencia Molecular , Filogenia , Prevalencia , ARN Viral/genética , Virus Sincitial Respiratorio Humano/aislamiento & purificación , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Análisis de Secuencia de ADN , Homología de Secuencia , Proteínas Estructurales Virales/genética
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA