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1.
APMIS ; 127(10): 671-680, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31344276

RESUMO

Regardless of the communal impact of Shiga toxins, till today neither a specific treatment nor licensed vaccine is available. Lactococcus lactis (L. lactis), generally regarded as safe organism, is well known to provide a valuable approach regarding the oral delivery of vaccines. This study was undertaken to evaluate the protective efficacy of Stx2a1 expressed in nisin-inducible L. lactis, against Shiga toxins (Stx1, Stx2) in mouse model. Oral immunization of BALB/c mice with LL-Stx2a1 elicited significant serum antibody titer with elevated fecal and serum IgA, along with minimized intestinal and kidney damage resulting in survival of immunized animals at 84% and 100% when challenged with 10 × LD50 of Escherichia coli O157 and Shigella dysenteriae toxins, respectively. HeLa cells incubated with immune sera and toxin mixture revealed high neutralizing capacity with 90% cell survivability against both the toxins. Mice immunized passively with both toxins and antibody mixture survived the observation period of 15 days, and the controls administered with sham sera and toxins were succumbed to death within 3 days. Our results revealed protective efficacy and toxin neutralization ability of LL-Stx2a1, proposing it as an oral vaccine candidate against Shiga toxicity mediated by E. coli O157 and S. dysenteriae.


Assuntos
Vacinas Bacterianas/administração & dosagem , Vacinas Bacterianas/imunologia , Escherichia coli O157/imunologia , Envenenamento/prevenção & controle , Toxina Shiga/imunologia , Toxina Shiga/toxicidade , Shigella dysenteriae/imunologia , Administração Oral , Animais , Anticorpos Antibacterianos/administração & dosagem , Anticorpos Antibacterianos/sangue , Anticorpos Neutralizantes/administração & dosagem , Anticorpos Neutralizantes/sangue , Antitoxinas/administração & dosagem , Antitoxinas/sangue , Vacinas Bacterianas/genética , Sobrevivência Celular/efeitos dos fármacos , Modelos Animais de Doenças , Portadores de Fármacos/administração & dosagem , Escherichia coli O157/genética , Vetores Genéticos/administração & dosagem , Células HeLa , Humanos , Lactococcus lactis/genética , Camundongos , Camundongos Endogâmicos BALB C , Toxina Shiga/genética , Shigella dysenteriae/genética , Análise de Sobrevida , Resultado do Tratamento , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/genética , Vacinas Sintéticas/imunologia
2.
Nat Commun ; 10(1): 2699, 2019 06 20.
Artigo em Inglês | MEDLINE | ID: mdl-31221976

RESUMO

Human cytomegalovirus (CMV) causes a wide array of disease to diverse populations of immune-compromised individuals. Thus, a more comprehensive understanding of how CMV enters numerous host cell types is necessary to further delineate the complex nature of CMV pathogenesis and to develop targeted therapeutics. To that end, we establish a vaccination strategy utilizing membrane vesicles derived from epithelial cells to generate a library of monoclonal antibodies (mAbs) targeting cell surface proteins in their native conformation. A high-throughput inhibition assay is employed to screen these antibodies for their ability to limit infection, and mAbs targeting CD46 are identified. In addition, a significant reduction of viral proliferation in CD46-KO epithelial cells confirms a role for CD46 function in viral dissemination. Further, we demonstrate a CD46-dependent entry pathway of virus infection in trophoblasts, but not in fibroblasts, highlighting the complexity of CMV entry and identifying CD46 as an entry factor in congenital infection.


Assuntos
Infecções por Citomegalovirus/imunologia , Citomegalovirus/imunologia , Interações Hospedeiro-Patógeno/imunologia , Proteína Cofatora de Membrana/imunologia , Internalização do Vírus , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/administração & dosagem , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/administração & dosagem , Anticorpos Antivirais/imunologia , Linhagem Celular , Infecções por Citomegalovirus/prevenção & controle , Infecções por Citomegalovirus/virologia , Células Epiteliais/imunologia , Células Epiteliais/virologia , Fibroblastos/imunologia , Fibroblastos/virologia , Técnicas de Inativação de Genes , Humanos , Proteína Cofatora de Membrana/genética , RNA Interferente Pequeno/metabolismo , Trofoblastos/imunologia , Trofoblastos/virologia , Vacinas Virais/administração & dosagem , Vacinas Virais/imunologia
3.
Nat Microbiol ; 4(7): 1231-1241, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-30936489

RESUMO

Rift Valley fever virus (RVFV) is a mosquito-borne pathogen that causes substantial morbidity and mortality in livestock and humans. To date, there are no licensed human vaccines or therapeutics available. Here, we report the isolation of monoclonal antibodies from a convalescent patient, targeting the RVFV envelope proteins Gn and Gc. The Gn-specific monoclonal antibodies exhibited much higher neutralizing activities in vitro and protection efficacies in mice against RVFV infection, compared to the Gc-specific monoclonal antibodies. The Gn monoclonal antibodies were found to interfere with soluble Gn binding to cells and prevent infection by blocking the attachment of virions to host cells. Structural analysis of Gn complexed with four Gn-specific monoclonal antibodies resulted in the definition of three antigenic patches (A, B and C) on Gn domain I. Both patches A and B are major neutralizing epitopes. Our results highlight the potential of antibody-based therapeutics and provide a structure-based rationale for designing vaccines against RVFV.


Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Febre do Vale de Rift/prevenção & controle , Vírus da Febre do Vale do Rift/imunologia , Animais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Neutralizantes/administração & dosagem , Anticorpos Antivirais/administração & dosagem , Cercopithecus aethiops , Cristalografia por Raios X , Epitopos , Feminino , Humanos , Camundongos Endogâmicos BALB C , Testes de Neutralização , Domínios Proteicos , Febre do Vale de Rift/imunologia , Células Vero , Proteínas do Envelope Viral/química , Proteínas do Envelope Viral/imunologia , Proteínas do Envelope Viral/metabolismo , Ligação Viral
4.
Nat Microbiol ; 4(5): 766-773, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30833732

RESUMO

The human microbiota provides tonic signals that calibrate the host immune response1,2, but their identity is unknown. Bacterial peptidoglycan (PGN) subunits are likely candidates since they are well-known immunity-enhancing adjuvants, released by most bacteria during growth, and have been found in the blood of healthy people3-7. We developed a monoclonal antibody (mAb), 2E7, that targets muramyl-L-alanyl-D-isoglutamine (MDP), a conserved and minimal immunostimulatory structure of PGN. Using 2E7-based assays, we detected PGN ubiquitously in human blood at a broad range of concentrations that is relatively stable in each individual. We also detected PGN in the serum of several warm-blooded animals. However, PGN is barely detectable in the serum of germ-free mice, indicating that its origin is the host microbiota. Neutralization of circulating PGN via intraperitoneal administration of 2E7 suppressed the development of autoimmune arthritis and experimental autoimmune encephalomyelitis in mice. Arthritic NOD2-/- mice lacking the MDP sensor did not respond to 2E7, indicating that 2E7 dampens inflammation by blocking nucleotide-binding oligomerization domain-containing protein 2 (NOD2)-mediated pathways. We propose that circulating PGN acts as a natural immune potentiator that tunes the host immune response; altering its level is a promising therapeutic strategy for immune-mediated diseases.


Assuntos
Anticorpos Neutralizantes/administração & dosagem , Anticorpos Neutralizantes/imunologia , Artrite/tratamento farmacológico , Autoimunidade/efeitos dos fármacos , Bactérias/imunologia , Encefalomielite/tratamento farmacológico , Microbiota , Peptidoglicano/imunologia , Animais , Artrite/genética , Artrite/imunologia , Encefalomielite/genética , Encefalomielite/imunologia , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos DBA , Proteína Adaptadora de Sinalização NOD2/genética , Proteína Adaptadora de Sinalização NOD2/imunologia , Peptidoglicano/sangue
5.
Emerg Microbes Infect ; 8(1): 155-168, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30866770

RESUMO

Influenza viruses of the H4 subtype are widespread in wild birds, circulate in domestic poultry, readily infect mammals, and tolerate the insertion of a polybasic cleavage site. In addition, serological evidence suggests that humans working with poultry are exposed to these viruses. While H4 viruses are not of immediate pandemic concern, there is a lack of knowledge regarding their antigenicity. In order to study viruses of the H4 subtype, we generated and characterized a panel of antibodies that bind a wide variety of H4 hemagglutinins from avian and swine isolates of both the Eurasian and North American lineage. We further characterized these antibodies using novel recombinant H4N6 viruses that were found to be lethal in DBA/2J mice. Non-neutralizing antibodies, which had activity in an antibody dependent cell-mediated cytotoxicity reporter assay in vitro, protected mice against challenge in vivo, highlighting the importance of effector functions. Our data suggest a high degree of antigenic conservation of the H4 hemagglutinin.


Assuntos
Anticorpos Neutralizantes/administração & dosagem , Aves/virologia , Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Vírus da Influenza A/imunologia , Infecções por Orthomyxoviridae/prevenção & controle , Suínos/virologia , Animais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/administração & dosagem , Anticorpos Antivirais/imunologia , Aves/imunologia , Reações Cruzadas , Modelos Animais de Doenças , Feminino , Vírus da Influenza A/genética , Vírus da Influenza A/isolamento & purificação , Vírus da Influenza A/patogenicidade , Camundongos , Camundongos Endogâmicos DBA , Infecções por Orthomyxoviridae/imunologia , Suínos/imunologia
6.
Nat Immunol ; 20(3): 362-372, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30742080

RESUMO

The present vaccine against influenza virus has the inevitable risk of antigenic discordance between the vaccine and the circulating strains, which diminishes vaccine efficacy. This necessitates new approaches that provide broader protection against influenza. Here we designed a vaccine using the hypervariable receptor-binding domain (RBD) of viral hemagglutinin displayed on a nanoparticle (np) able to elicit antibody responses that neutralize H1N1 influenza viruses spanning over 90 years. Co-display of RBDs from multiple strains across time, so that the adjacent RBDs are heterotypic, provides an avidity advantage to cross-reactive B cells. Immunization with the mosaic RBD-np elicited broader antibody responses than those induced by an admixture of nanoparticles encompassing the same set of RBDs as separate homotypic arrays. Furthermore, we identified a broadly neutralizing monoclonal antibody in a mouse immunized with mosaic RBD-np. The mosaic antigen array signifies a unique approach that subverts monotypic immunodominance and allows otherwise subdominant cross-reactive B cell responses to emerge.


Assuntos
Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Vírus da Influenza A Subtipo H1N1/imunologia , Vacinas contra Influenza/imunologia , Influenza Humana/imunologia , Nanopartículas/química , Infecções por Orthomyxoviridae/imunologia , Animais , Anticorpos Neutralizantes/administração & dosagem , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Linfócitos B/efeitos dos fármacos , Linfócitos B/imunologia , Linfócitos B/virologia , Reações Cruzadas/efeitos dos fármacos , Reações Cruzadas/imunologia , Feminino , Glicoproteínas de Hemaglutininação de Vírus da Influenza/química , Humanos , Imunização , Vírus da Influenza A Subtipo H1N1/metabolismo , Vírus da Influenza A Subtipo H1N1/fisiologia , Vacinas contra Influenza/administração & dosagem , Vacinas contra Influenza/química , Influenza Humana/prevenção & controle , Influenza Humana/virologia , Camundongos Endogâmicos BALB C , Infecções por Orthomyxoviridae/prevenção & controle , Infecções por Orthomyxoviridae/virologia
7.
EBioMedicine ; 40: 574-582, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30638863

RESUMO

BACKGROUND: VIS410, a broadly neutralizing monoclonal antibody that binds the hemagglutinin stem of influenza A viruses, was safe and efficacious in a human H1N1 virus challenge study. This study evaluated the safety and tolerability of VIS410 in non-hospitalized adult patients with uncomplicated influenza A. METHODS: Patients 18 to 65 years of age with symptom onset within 72 h were randomized 1:1:1 to receive a single intravenous infusion of VIS410 4000 mg, 2000 mg, or placebo. Neuraminidase inhibitor therapy was prohibited. Treatment-emergent adverse events (TEAEs) were evaluated up to 100 days post-infusion. Influenza symptoms were assessed daily for 10 days using the FLU-PRO tool. Nasopharyngeal virus shedding was assessed by quantitative reverse-transcription PCR (qRT-PCR) and viral culture through Day 7. FINDINGS: Of the 150 patients randomized, 148 received study drug, and 138 were confirmed influenza A positive. Median age was 42 years; median time from symptom onset to treatment was 42 h; 93% had influenza A subtype H3N2. SAFETY: TEAEs, most commonly diarrhea of mild severity, were dose-related, occurring in 55%, 35%, and 24% of the 4000 mg, 2000 mg, and placebo patients, respectively. Two serious adverse events occurred, both in placebo patients. SYMPTOM ANALYSES: Baseline FLU-PRO symptom scores were balanced among groups. Mean scores were lower by Days 3 and 4 in the pooled VIS410 treatment group versus placebo (p < 0.023), with a tendency toward faster resolution by Kaplan-Meier analysis. VIROLOGY ANALYSES: VIS410 was associated with reduced median nasopharyngeal viral load TCID50 AUCDay7 (days × log10 TCID50/mL) (3.66 pooled VIS410 vs 4.78 placebo, p = 0.08) and in the subset of patients with baseline hemagglutination inhibition (HAI) titer ≤40 (overall, 74% of patients) was significantly reduced vs placebo (4.218 pooled VIS410 vs 6.152 placebo, p = 0.009). Kaplan-Meier estimated time to resolution of viral shedding was reduced (1.9 vs 3.6 days, p = 0.03) in VIS410 treated patients. There was a trend toward greater proportion of culture-negative patients by Day 3 (66.7% vs 51.1%, p = 0.11); when this analysis was limited to the subset of patients with positive baseline cultures, this difference became more pronounced (63.2% vs 42.5%, p = 0.053). No differences were observed in nasopharyngeal influenza qRT-PCR profiles, which represent both live and neutralized virus. INTERPRETATION: VIS410 was safe and well tolerated in adults with uncomplicated influenza A, with favorable effects on symptom resolution and virus replication. TRIAL REGISTRATION: Clinical Trials: NCT02989194. FUNDING: This project was funded in part with Federal funds from the Department of Health and Human Services; Office of the Assistant Secretary for Preparedness and Response; Biomedical Advanced Research and Development Authority (BARDA), under Contract No. HHSO100201500018C.


Assuntos
Anticorpos Monoclonais/uso terapêutico , Anticorpos Neutralizantes/uso terapêutico , Antivirais/uso terapêutico , Vírus da Influenza A , Influenza Humana/tratamento farmacológico , Adulto , Anticorpos Monoclonais/administração & dosagem , Anticorpos Neutralizantes/administração & dosagem , Anticorpos Antivirais/administração & dosagem , Anticorpos Antivirais/uso terapêutico , Antivirais/administração & dosagem , Feminino , Humanos , Imunoterapia , Vírus da Influenza A/classificação , Vírus da Influenza A/genética , Vírus da Influenza A/imunologia , Influenza Humana/diagnóstico , Influenza Humana/imunologia , Influenza Humana/virologia , Masculino , Avaliação de Sintomas , Resultado do Tratamento , Carga Viral , Eliminação de Partículas Virais/efeitos dos fármacos
8.
Cell Host Microbe ; 25(1): 49-58.e5, 2019 01 09.
Artigo em Inglês | MEDLINE | ID: mdl-30629918

RESUMO

Recent and ongoing outbreaks of Ebola virus disease (EVD) underscore the unpredictable nature of ebolavirus reemergence and the urgent need for antiviral treatments. Unfortunately, available experimental vaccines and immunotherapeutics are specific for a single member of the Ebolavirus genus, Ebola virus (EBOV), and ineffective against other ebolaviruses associated with EVD, including Sudan virus (SUDV) and Bundibugyo virus (BDBV). Here we show that MBP134AF, a pan-ebolavirus therapeutic comprising two broadly neutralizing human antibodies (bNAbs), affords unprecedented effectiveness and potency as a therapeutic countermeasure to antigenically diverse ebolaviruses. MBP134AF could fully protect ferrets against lethal EBOV, SUDV, and BDBV infection, and a single 25-mg/kg dose was sufficient to protect NHPs against all three viruses. The development of MBP134AF provides a successful model for the rapid discovery and translational advancement of immunotherapeutics targeting emerging infectious diseases.


Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/uso terapêutico , Anticorpos Antivirais/imunologia , Anticorpos Antivirais/uso terapêutico , Ebolavirus/patogenicidade , Furões/virologia , Doença pelo Vírus Ebola/imunologia , Doença pelo Vírus Ebola/prevenção & controle , Bem-Estar do Animal , Animais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/isolamento & purificação , Anticorpos Neutralizantes/administração & dosagem , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/isolamento & purificação , Anticorpos Neutralizantes/uso terapêutico , Anticorpos Antivirais/administração & dosagem , Linhagem Celular , Cercopithecus aethiops , Modelos Animais de Doenças , Feminino , Filoviridae/imunologia , Infecções por Filoviridae/imunologia , Infecções por Filoviridae/prevenção & controle , Infecções por Filoviridae/virologia , Glicoproteínas/imunologia , Cobaias , Células HEK293 , Doença pelo Vírus Ebola/virologia , Humanos , Células Matadoras Naturais , Macaca , Macaca fascicularis , Masculino , Primatas , Análise de Sobrevida , Resultado do Tratamento , Proteínas Virais/imunologia
9.
Cell Host Microbe ; 25(1): 39-48.e5, 2019 01 09.
Artigo em Inglês | MEDLINE | ID: mdl-30629917

RESUMO

Passive administration of monoclonal antibodies (mAbs) is a promising therapeutic approach for Ebola virus disease (EVD). However, all mAbs and mAb cocktails that have entered clinical development are specific for a single member of the Ebolavirus genus, Ebola virus (EBOV), and ineffective against outbreak-causing Bundibugyo virus (BDBV) and Sudan virus (SUDV). Here, we advance MBP134, a cocktail of two broadly neutralizing human mAbs, ADI-15878 from an EVD survivor and ADI-23774 from the same survivor but specificity-matured for SUDV GP binding affinity, as a candidate pan-ebolavirus therapeutic. MBP134 potently neutralized all ebolaviruses and demonstrated greater protective efficacy than ADI-15878 alone in EBOV-challenged guinea pigs. A second-generation cocktail, MBP134AF, engineered to effectively harness natural killer (NK) cells afforded additional improvement relative to its precursor in protective efficacy against EBOV and SUDV in guinea pigs. MBP134AF is an optimized mAb cocktail suitable for evaluation as a pan-ebolavirus therapeutic in nonhuman primates.


Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Antivirais/imunologia , Ebolavirus/imunologia , Doença pelo Vírus Ebola/imunologia , Doença pelo Vírus Ebola/prevenção & controle , Bem-Estar do Animal , Animais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/isolamento & purificação , Anticorpos Monoclonais/uso terapêutico , Anticorpos Neutralizantes/administração & dosagem , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/isolamento & purificação , Anticorpos Neutralizantes/uso terapêutico , Anticorpos Antivirais/administração & dosagem , Anticorpos Antivirais/uso terapêutico , Antivirais , Modelos Animais de Doenças , Ebolavirus/patogenicidade , Epitopos/imunologia , Feminino , Filoviridae/imunologia , Cobaias , Doença pelo Vírus Ebola/virologia , Humanos , Imunoterapia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Proteínas Recombinantes/imunologia , Resultado do Tratamento
10.
Drugs ; 79(1): 99-103, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30623346

RESUMO

Emapalumab-Izsg (hereafter referred to as emapalumab) [Gamifant®] is a monoclonal antibody directed against interferon gamma that is available as an intravenous infusion. Emapalumab is being developed by Novimmune and Swedish Orphan Biovitrum for the treatment of haemophagocytic lymphohistiocytosis (HLH). In November 2018, emapalumab received its first global approval in the USA, for the treatment of paediatric (newborn and older) and adult patients with primary HLH, who have refractory, recurrent or progressive disease or intolerance to conventional HLH therapy. Emapalumab is under regulatory review in the EU for the treatment of primary HLH. This article summarizes the milestones in the development of emapalumab leading to this first global approval for HLH in the USA.


Assuntos
Anticorpos Monoclonais , Anticorpos Neutralizantes , Interferon gama , Linfo-Histiocitose Hemofagocítica/tratamento farmacológico , Linfo-Histiocitose Hemofagocítica/imunologia , Adolescente , Adulto , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/efeitos adversos , Anticorpos Neutralizantes/administração & dosagem , Anticorpos Neutralizantes/efeitos adversos , Anticorpos Neutralizantes/imunologia , Criança , Pré-Escolar , Aprovação de Drogas , Feminino , Humanos , Lactente , Recém-Nascido , Infusões Intravenosas , Interferon gama/antagonistas & inibidores , Interferon gama/imunologia , Masculino , Pessoa de Meia-Idade , Resultado do Tratamento
11.
Viruses ; 10(9)2018 08 23.
Artigo em Inglês | MEDLINE | ID: mdl-30142928

RESUMO

Infection by the Middle East respiratory syndrome coronavirus (MERS-CoV) causes respiratory illness and has a high mortality rate (~35%). The requirement for the virus to be manipulated in a biosafety level three (BSL-3) facility has impeded development of urgently-needed antiviral agents. Here, we established anovel mouse model by inserting human dipeptidyl peptidase 4 (hDPP4) into the Rosa26 locus using CRISPR/Cas9, resulting in global expression of the transgene in a genetically stable mouse line. The mice were highly susceptible to infection by MERS-CoV clinical strain hCoV-EMC, which induced severe diffuse pulmonary disease in the animals, and could also be infected by an optimized pseudotyped MERS-CoV. Administration of the neutralizing monoclonal antibodies, H111-1 and m336, as well as a fusion inhibitor peptide, HR2P-M2, protected mice from challenge with authentic and pseudotyped MERS-CoV. These results confirmed that the hDPP4-knockin mouse is a novel model for studies of MERS-CoV pathogenesis and anti-MERS-CoV antiviral agents in BSL-3 and BSL-2facilities, respectively.


Assuntos
Infecções por Coronavirus/imunologia , Dipeptidil Peptidase 4/metabolismo , Modelos Animais de Doenças , Suscetibilidade a Doenças , Técnicas de Introdução de Genes , Coronavírus da Síndrome Respiratória do Oriente Médio/crescimento & desenvolvimento , Coronavírus da Síndrome Respiratória do Oriente Médio/imunologia , Animais , Anticorpos Neutralizantes/administração & dosagem , Anticorpos Antivirais/administração & dosagem , Proteínas de Bactérias/metabolismo , Proteína 9 Associada à CRISPR , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Infecções por Coronavirus/prevenção & controle , Infecções por Coronavirus/virologia , Dipeptidil Peptidase 4/genética , Endonucleases/metabolismo , Humanos , Camundongos , Recombinação Genética
12.
Malar J ; 17(1): 304, 2018 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-30126436

RESUMO

BACKGROUND: Plasmodium enolase is a target for the growth neutralizing antibodies. Interestingly, the three invasive stages i.e. sporozoites, merozoites, and ookinetes express this protein on their cell surface. Polyclonal anti-Plasmodium falciparum enolase (Pfeno) antibodies disrupt traversal of ookinete through mosquito mid-gut wall as well as have inhibitory effect on parasite growth at erythrocytic stage. In a recent study, it was observed that immunization with a unique epitope of parasite enolase (EWGWS) could confer partial protection against mouse malaria. Further validation is needed for the protective potential of this unique epitope in otherwise highly conserved enolase. METHODS: In order to investigate the efficacy of growth inhibitory potential of the epitope of P falciparum enolase, a monoclonal antibody specific to EWGWS is generated. In vitro parasite growth inhibition assays and passive immunization of Plasmodium yoelii (or Plasmodium berghei) infected mice were used to assess the parasite growth neutralizing activity of the antibody. RESULTS: Screening a panel of monoclonal antibodies raised against recombinant Pfeno that were specific to EWGWS resulted in isolation of H12E1. This antibody recognized only EWGWS epitope containing enolases. H12E1 strongly inhibited parasite growth in culture. This inhibition was strain transcending. Passive infusion of this antibody in P. yoelii or P. berghei infected mice showed significant reduction in parasitemia as compared to controls (p < 0.001). Surface Plasmon Resonance measurements indicated high affinity binding of H12E1 to P. falciparum enolase (KD ~ 7.6 × 10-9M). CONCLUSIONS: A monoclonal antibody directed against EWGWS epitope of Pfeno was shown to inhibit the growth of blood stage malarial parasites. This inhibition was species/strain transcending and is likely to arise due to blockade of enolase on the surface of merozoites, functionally implicating Pfeno in invasion related events. Presence of enolase on the cell surface of merozoites and ookinetes could potentially result in inhibition of host cell invasions at erythrocytic and transmission stages in the parasite life cycle. It is suggested that antibodies against EWGWS epitope have the potential to confer dual stage, species and strain transcending protection against malaria.


Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Antiprotozoários/imunologia , Malária/prevenção & controle , Fosfopiruvato Hidratase/imunologia , Plasmodium falciparum/enzimologia , Plasmodium falciparum/imunologia , Animais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Neutralizantes/administração & dosagem , Anticorpos Antiprotozoários/administração & dosagem , Modelos Animais de Doenças , Imunização Passiva , Malária/imunologia , Masculino , Camundongos , Plasmodium berghei/imunologia , Plasmodium yoelii/imunologia
13.
J Exp Clin Cancer Res ; 37(1): 200, 2018 Aug 22.
Artigo em Inglês | MEDLINE | ID: mdl-30134951

RESUMO

BACKGROUND: The function of preadipocytes in the progression of early stage breast cancer has not been fully elucidated at the molecular level. To delineate the role of preadipocytes in breast cancer progression, we investigated the cross-talk between human breast ductal carcinoma in situ (DCIS) cells and preadipocytes with both an in vitro culture and xenograft tumor model. METHODS: GFP or RFP was transduced into human DCIS cell line MCF10DCIS.com cells or preadipocytes using lentivirus. Cell sorter was used to separate pure, viable populations of GFP- or RFP-transduced cells. Cell viability and proliferation was assessed by crystal violet assays and cell migration and invasion capability was assayed by the transwell strategy. Gene and protein levels were measured by western blot, RT-PCR and immunostaining. Adipokines and cytokines were quantified using ELISA. Human tumor xenografts in a nude mice model were used. Ultrasound imaging of tumors was performed to evaluate the therapeutic potential of a IL-6 neutralizing antibody. RESULTS: In the co-culture system with the MCF10DCIS.com and preadipocytes, MCF10DCIS.com proliferation, migration and invasion were enhanced by preadipocytes. Preadipocytes exhibited in an increased IL-6 secretion and cancer-associated fibroblast markers expression, FSP1 and α-SMC in co-culture with MCF10DCIS.com or in MCF10DCIS.com conditioned media, whereas the adipocyte differentiation capacity was suppressed by co-culture with MCF10DCIS.com. A neutralizing antibody of IL-6 or IL-6R suppressed the promotion of MCF10DCIS.com proliferation and migration by co-culture with preadipocytes. In the xenograft tumor model, the tumor growth of MCF10DCIS.com was enhanced by the co-injection of preadipocytes, and the administration of IL-6 neutralizing antibodies resulted in potent effects on tumor inhibition. CONCLUSIONS: Our findings suggest that IL-6-mediated cross-talk between preadipocytes and breast DCIS cells can promote the progression of early stage breast cancer. Therefore, blocking IL-6 signaling might be a potential therapeutic strategy for breast DCIS characterized by pathological IL-6 overproduction.


Assuntos
Anticorpos Neutralizantes/administração & dosagem , Neoplasias da Mama/genética , Carcinoma Intraductal não Infiltrante/genética , Interleucina-6/genética , Adipócitos/metabolismo , Adipócitos/patologia , Animais , Anticorpos Neutralizantes/imunologia , Neoplasias da Mama/imunologia , Neoplasias da Mama/patologia , Carcinoma Intraductal não Infiltrante/imunologia , Carcinoma Intraductal não Infiltrante/patologia , Diferenciação Celular/genética , Movimento Celular/genética , Técnicas de Cocultura , Feminino , Fibroblastos/metabolismo , Fibroblastos/patologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/imunologia , Humanos , Interleucina-6/antagonistas & inibidores , Interleucina-6/imunologia , Camundongos , Receptores de Interleucina-6/antagonistas & inibidores , Receptores de Interleucina-6/imunologia , Transdução de Sinais/genética , Ensaios Antitumorais Modelo de Xenoenxerto
14.
Cell Host Microbe ; 23(5): 618-627.e6, 2018 05 09.
Artigo em Inglês | MEDLINE | ID: mdl-29746833

RESUMO

Following the recent emergence of Zika virus (ZIKV), many murine and human neutralizing anti-ZIKV antibodies have been reported. Given the risk of virus escape mutants, engineering antibodies that target mutationally constrained epitopes with therapeutically relevant potencies can be valuable for combating future outbreaks. Here, we applied computational methods to engineer an antibody, ZAb_FLEP, that targets a highly networked and therefore mutationally constrained surface formed by the envelope protein dimer. ZAb_FLEP neutralized a breadth of ZIKV strains and protected mice in distinct in vivo models, including resolving vertical transmission and fetal mortality in infected pregnant mice. Serial passaging of ZIKV in the presence of ZAb_FLEP failed to generate viral escape mutants, suggesting that its epitope is indeed mutationally constrained. A single-particle cryo-EM reconstruction of the Fab-ZIKV complex validated the structural model and revealed insights into ZAb_FLEP's neutralization mechanism. ZAb_FLEP has potential as a therapeutic in future outbreaks.


Assuntos
Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/química , Anticorpos Antivirais/imunologia , Epitopos/imunologia , Engenharia de Proteínas , Infecção por Zika virus/imunologia , Zika virus/genética , Zika virus/imunologia , Animais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/uso terapêutico , Anticorpos Neutralizantes/administração & dosagem , Anticorpos Neutralizantes/uso terapêutico , Anticorpos Antivirais/administração & dosagem , Anticorpos Antivirais/uso terapêutico , Vírus da Dengue/imunologia , Modelos Animais de Doenças , Epitopos/química , Epitopos/genética , Feminino , Masculino , Camundongos , Modelos Moleculares , Testes de Neutralização/métodos , Gravidez , Estrutura Quaternária de Proteína , Resultado do Tratamento , Proteínas do Envelope Viral/química , Proteínas do Envelope Viral/genética , Proteínas do Envelope Viral/imunologia , Viremia/tratamento farmacológico , Infecção por Zika virus/tratamento farmacológico , Infecção por Zika virus/virologia
15.
Biochem Biophys Res Commun ; 501(1): 145-151, 2018 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-29705699

RESUMO

Perivascular adipose tissue (PAT) is associated with vascular homeostasis; however, its causal effect on atherosclerosis currently remains undefined. Here, we investigated the effect of experimental PAT transplantation on atherosclerosis. The thoracic periaortic adipose tissue (tPAT) was dissected from 16-week-old wild-type mice and transplanted over the infrarenal aorta of 20-week-old apoE deficient (apoE-/-) mice fed high-cholesterol diet for 3 months. Oil-red O staining after 4 weeks showed a significant 20% decrease in the atherosclerotic lesion of suprarenal aorta compared with that of sham control mice, while that of infrarenal aorta showed no difference between the two groups. TGF-ß1 mRNA expression was significantly higher in grafted tPAT than donor tPAT, accompanied by a significant increase in serum TGF-ß1 concentration, which was inversely correlated with the suprarenal lesion area (r = -0.63, P = 0.012). Treatment with neutralizing TGF-ß antibody abrogated the anti-atherogenic effect of tPAT transplantation. Immunofluorescent analysis of grafted tPAT showed that TGF-ß-positive cells were co-localized with Mac-2-positive cells and this number was significantly increased compared with donor tPAT. There was also marked increase in mRNA expression of alternatively activated macrophages-related genes. Furthermore, the percentage of eosinophils in stromal vascular fraction of donor tPAT was much higher than that in epididymal white adipose tissue, concomitant with the significantly higher protein level of IL-4. IL-4 mRNA expression levels in grafted tPAT were increased in a time-dependent manner after tPAT transplantation. Our findings show that tPAT transplantation inhibits atherosclerosis development by exerting TGF-ß1-mediated anti-inflammatory response, which may involve alternatively activated macrophages.


Assuntos
Tecido Adiposo/transplante , Aterosclerose/prevenção & controle , Fator de Crescimento Transformador beta1/metabolismo , Tecido Adiposo/metabolismo , Tecido Adiposo/patologia , Animais , Anticorpos Neutralizantes/administração & dosagem , Aorta Torácica/metabolismo , Aorta Torácica/patologia , Apolipoproteínas E/deficiência , Apolipoproteínas E/genética , Aterosclerose/metabolismo , Aterosclerose/patologia , Eosinófilos/patologia , Inflamação/metabolismo , Inflamação/patologia , Inflamação/prevenção & controle , Interleucina-4/metabolismo , Ativação de Macrófagos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout para ApoE , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fator de Crescimento Transformador beta1/antagonistas & inibidores , Fator de Crescimento Transformador beta1/genética
16.
Viruses ; 10(4)2018 03 24.
Artigo em Inglês | MEDLINE | ID: mdl-29587363

RESUMO

Western equine encephalitis virus (WEEV) causes symptoms in humans ranging from mild febrile illness to life-threatening encephalitis, and no human medical countermeasures are licensed. A previous study demonstrated that immune serum from vaccinated mice protected against lethal WEEV infection, suggesting the utility of antibodies for pre- and post-exposure treatment. Here, three neutralizing and one binding human-like monoclonal antibodies were evaluated against WEEV aerosol challenge. Dose-dependent protection was observed with two antibodies administered individually, ToR69-3A2 and ToR68-2C3. In vitro neutralization was not a critical factor for protection in this murine model, as ToR69-3A2 is a strong neutralizing antibody, and ToR68-2C3 is a non-neutralizing antibody. This result highlights the importance of both neutralizing and non-neutralizing antibodies in the protection of mice from WEEV lethality.


Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Vírus da Encefalite Equina do Oeste/imunologia , Encefalomielite Equina/imunologia , Encefalomielite Equina/prevenção & controle , Aerossóis , Animais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Neutralizantes/administração & dosagem , Anticorpos Antivirais/administração & dosagem , Modelos Animais de Doenças , Encefalomielite Equina/mortalidade , Encefalomielite Equina/virologia , Imunização , Camundongos , Morbidade , Mortalidade
17.
Consult Pharm ; 33(2): 89-97, 2018 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-29409575

RESUMO

OBJECTIVE: To review the pharmacology, pharmacokinetics, pharmacodynamics, clinical efficacy, tolerability, dosing, and administration of bezlotoxumab (BEZ), as well as its place in the prevention of Clostridium difficile infection (CDI) recurrence. DATA SOURCES: A search of PubMed and Google Scholar using the terms "bezlotoxumab," "CDB1," "MDX-1388," and "MK-6072" was performed. The manufacturer's website was also reviewed to further identify relevant information. STUDY SELCTION: All English-language articles from 2006 to May 2017 appearing in these searches were reviewed for relevance to this paper. In addition, their bibliographies were reviewed to identify any articles not identified in the searches. DATA SYNTHESIS: BEZ is a human monoclonal antibody that binds to Clostridium difficile toxin B. It is approved by the Food and Drug Administration to reduce CDI recurrence in adult patients who are receiving antibiotic therapy for CDI and are at high risk for CDI recurrence. It is given as a single dose of 10 mg/kg via an intravenous infusion. It is eliminated by catabolism. Phase III clinical trials demonstrated that BEZ was associated with significantly lower rates of CDI recurrence, compared with placebo. The most common adverse events reported during clinical trials were diarrhea and nausea. There is a warning regarding the use of BEZ in patients with a history of congestive heart failure. The most common adverse reactions associated with BEZ are nausea, pyrexia, and headache. CONCLUSION: BEZ has been proven safe and effective in preventing CDI recurrence. Given its high cost, it should be reserved for patients at high risk for CDI recurrence.


Assuntos
Anticorpos Monoclonais/administração & dosagem , Anticorpos Neutralizantes/administração & dosagem , Infecções por Clostridium/prevenção & controle , Clostridium difficile/efeitos dos fármacos , Animais , Anticorpos Monoclonais/efeitos adversos , Anticorpos Monoclonais/farmacologia , Anticorpos Neutralizantes/efeitos adversos , Anticorpos Neutralizantes/farmacologia , Proteínas de Bactérias/imunologia , Toxinas Bacterianas/imunologia , Humanos , Infusões Intravenosas , Prevenção Secundária/métodos
18.
Brain Behav Immun ; 67: 24-35, 2018 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-28780000

RESUMO

Perinatal hypoxic-ischemic reperfusion (I/R)-related brain injury is a leading cause of neurologic morbidity and life-long disability in children. Infants exposed to I/R brain injury develop long-term cognitive and behavioral deficits, placing a large burden on parents and society. Therapeutic strategies are currently not available for infants with I/R brain damage, except for hypothermia, which can only be used in full term infants with hypoxic-ischemic encephalopathy (HIE). Moreover, hypothermia is only partially protective. Pro-inflammatory cytokines are key contributors to the pathogenesis of perinatal I/R brain injury. Interleukin-1ß (IL-1ß) is a critical pro-inflammatory cytokine, which has been shown to predict the severity of HIE in infants. We have previously shown that systemic infusions of mouse anti-ovine IL-1ß monoclonal antibody (mAb) into fetal sheep resulted in anti-IL-1ß mAb penetration into brain, reduced I/R-related increases in IL-1ß expression and blood-brain barrier (BBB) dysfunction in fetal brain. The purpose of the current study was to examine the effects of systemic infusions of anti-IL-1ß mAb on short-term I/R-related parenchymal brain injury in the fetus by examining: 1) histopathological changes, 2) apoptosis and caspase-3 activity, 3) neuronal degeneration 4) reactive gliosis and 5) myelin basic protein (MBP) immunohistochemical staining. The study groups included non-ischemic controls, placebo-treated ischemic, and anti-IL-1ß mAb treated ischemic fetal sheep at 127days of gestation. The systemic intravenous infusions of anti-IL-1ß mAb were administered at fifteen minutes and four hours after in utero brain ischemia. The duration of each infusion was two hours. Parenchymal brain injury was evaluated by determining pathological injury scores, ApopTag® positive cells/mm2, caspase-3 activity, Fluoro-Jade B positive cells/mm2, glial fibrillary acidic protein (GFAP) and MBP staining in the brains of fetal sheep 24h after 30min of ischemia. Treatment with anti-IL-1ß mAb reduced (P<0.05) the global pathological injury scores, number of apoptotic positive cells/mm2, and caspase-3 activity after ischemia in fetal sheep. The regional pathological scores and Fluoro-Jade B positive cells/mm2 did not differ between the placebo- and anti-IL-1ß mAb treated ischemic fetal sheep. The percent of the cortical area stained for GFAP was lower (P<0.05) in the placebo ischemic treated than in the non-ischemic group, but did not differ between the placebo- and anti-IL-1ß mAb treated ischemic groups. MBP immunohistochemical expression did not differ among the groups. In conclusion, infusions of anti-IL-1ß mAb attenuate short-term I/R-related histopathological tissue injury, apoptosis, and reduce I/R-related increases in caspase-3 activity in ovine fetal brain. Therefore, systemic infusions of anti-IL-1ß mAb attenuate short-term I/R-related parenchymal brain injury in the fetus.


Assuntos
Anticorpos Monoclonais/administração & dosagem , Isquemia Encefálica/imunologia , Encéfalo/imunologia , Interleucina-1beta/imunologia , Animais , Anticorpos Neutralizantes/administração & dosagem , Apoptose , Encéfalo/patologia , Encéfalo/fisiopatologia , Isquemia Encefálica/patologia , Feto/imunologia , Traumatismo por Reperfusão/imunologia , Traumatismo por Reperfusão/patologia , Ovinos
19.
Vaccine ; 36(2): 285-291, 2018 01 04.
Artigo em Inglês | MEDLINE | ID: mdl-29191737

RESUMO

BACKGROUND: Obesity is a risk factor for increased morbidity and mortality associated with many vaccine preventable infectious diseases such as influenza. Moreover, higher volume of passive rabies immunoglobulin (RIG) due to weight based dosing might suppress vaccine-induced immune responses in obese patients. This study aimed to evaluate the effect of obesity on humoral immune responses to combined equine RIG and rabies vaccine treatment among patients with WHO category III exposure to a rabies suspected animal. METHODS: A single centre, prospective, open-labelled study among WHO category III rabies exposed patients was conducted to compare serum rabies virus neutralizing antibody (RVNA) responses measured by rapid fluorescent focus inhibition test between obese (body mass index, BMI > 30 kg/m2) and control (BMI < 25 kg/m2) patients after combined immunization with equine rabies immunoglobulin and purified chick-embryo cell rabies vaccine for post exposure prophylaxis treatment. RESULTS: Post-vaccination geometric mean titer (GMT) of RVNA concentrations between two groups at day 7 were 0.33 (95% CI: 0.23, 0.46) vs 0.39 (95% CI: 0.27, 0.55), 4.61 (95% CI: 3.20, 6.63) vs 3.78 (95% CI: 2.77, 5.16) at day 14, and 7.45 (95% CI: 5.86, 9.49) vs 5.93 (95%CI: 4.46-7.90) at day 28 for obese and control patients, respectively. There was no statistically significant difference of RVNA GMT between two groups. Seroconversion to at least adequate concentration (RVNA titer ≥0.5 IU/mL) rates were 34% at day 7 and 100% at days 14 and 28 in both groups. There were no immediate hypersensitivity reaction and no serious adverse events observed during the study period. CONCLUSIONS: There was no evidence of immunosuppression of antibodies' responses in obese patients. Combined ERIG and rabies virus vaccination for post exposure treatment is safe.


Assuntos
Anticorpos Antivirais/administração & dosagem , Anticorpos Antivirais/sangue , Obesidade/complicações , Profilaxia Pós-Exposição/métodos , Vacinas Antirrábicas/administração & dosagem , Vacinas Antirrábicas/imunologia , Raiva/prevenção & controle , Adulto , Idoso , Animais , Anticorpos Neutralizantes/administração & dosagem , Anticorpos Neutralizantes/sangue , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Testes de Neutralização , Estudos Prospectivos , Adulto Jovem
20.
Vaccine ; 36(1): 155-164, 2018 01 02.
Artigo em Inglês | MEDLINE | ID: mdl-29180028

RESUMO

Clostridium botulinum readily persists in the soil and secretes life-threatening botulinum neurotoxins (BoNTs) that are categorized into serotypes A to H, of which, serotype A (BoNT/A) is the most commonly occurring in nature. An efficacious vaccine with high longevity against BoNT intoxication is urgent. Herein, we developed a dual-route vaccine administered over four consecutive weeks by mucosal and parenteral routes, consisting of the heavy chain (Hc) of BoNT/A targeting dendritic cell peptide (DCpep) expressed by Lactobacillus acidophilus as a secretory immunogenic protein. The administered dual-route vaccine elicited robust and long-lasting memory B cell responses comprising germinal center (GC) B cells and follicular T cells (Tfh) that fully protected mice from lethal oral BoNT/A fatal intoxication. Additionally, passively transferring neutralizing antibodies against BoNT/A into naïve mice induced robust protection against BoNT/A lethal intoxication. Together, a targeted vaccine employing local and systemic administrative routes may represent a novel formulation eliciting protective B cell responses with remarkable longevity against threatening biologic agents such as BoNTs.


Assuntos
Anticorpos Neutralizantes/imunologia , Vacinas Bacterianas/imunologia , Toxinas Botulínicas Tipo A/imunologia , Neurotoxinas/imunologia , Vacinação/métodos , Administração através da Mucosa , Animais , Anticorpos Antibacterianos/imunologia , Anticorpos Neutralizantes/administração & dosagem , Linfócitos B/imunologia , Vacinas Bacterianas/administração & dosagem , Botulismo/prevenção & controle , Clostridium botulinum/imunologia , Células Dendríticas/química , Células Dendríticas/imunologia , Vias de Administração de Medicamentos , Imunização Passiva , Memória Imunológica , Lactobacillus acidophilus/química , Camundongos , Peptídeos/administração & dosagem , Peptídeos/genética , Peptídeos/imunologia , Peptídeos/metabolismo , Sorogrupo
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