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1.
Nature ; 577(7791): 543-548, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31915378

RESUMO

Although maternal antibodies protect newborn babies from infection1,2, little is known about how protective antibodies are induced without prior pathogen exposure. Here we show that neonatal mice that lack the capacity to produce IgG are protected from infection with the enteric pathogen enterotoxigenic Escherichia coli by maternal natural IgG antibodies against the maternal microbiota when antibodies are delivered either across the placenta or through breast milk. By challenging pups that were fostered by either maternal antibody-sufficient or antibody-deficient dams, we found that IgG derived from breast milk was crucial for protection against mucosal disease induced by enterotoxigenic E. coli. IgG also provides protection against systemic infection by E. coli. Pups used the neonatal Fc receptor to transfer IgG from milk into serum. The maternal commensal microbiota can induce antibodies that recognize antigens expressed by enterotoxigenic E. coli and other Enterobacteriaceae species. Induction of maternal antibodies against a commensal Pantoea species confers protection against enterotoxigenic E. coli in pups. This role of the microbiota in eliciting protective antibodies to a specific neonatal pathogen represents an important host defence mechanism against infection in neonates.


Assuntos
Anticorpos/imunologia , Escherichia coli Enterotoxigênica/imunologia , Infecções por Escherichia coli/imunologia , Infecções por Escherichia coli/prevenção & controle , Imunidade Materno-Adquirida/imunologia , Recém-Nascido/imunologia , Microbiota/imunologia , Leite Humano/imunologia , Animais , Anticorpos/sangue , Anticorpos/metabolismo , Aleitamento Materno , Reações Cruzadas/imunologia , Infecções por Escherichia coli/microbiologia , Feminino , Humanos , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Imunoglobulina G/metabolismo , Masculino , Camundongos , Mães , Pantoea/imunologia , Receptores Fc/imunologia , Receptores Fc/metabolismo , Simbiose/imunologia
2.
J Virol ; 94(5)2020 02 14.
Artigo em Inglês | MEDLINE | ID: mdl-31826992

RESUMO

Antibody-dependent enhancement (ADE) of viral entry has been a major concern for epidemiology, vaccine development, and antibody-based drug therapy. However, the molecular mechanism behind ADE is still elusive. Coronavirus spike protein mediates viral entry into cells by first binding to a receptor on the host cell surface and then fusing viral and host membranes. In this study, we investigated how a neutralizing monoclonal antibody (MAb), which targets the receptor-binding domain (RBD) of Middle East respiratory syndrome (MERS) coronavirus spike, mediates viral entry using pseudovirus entry and biochemical assays. Our results showed that MAb binds to the virus surface spike, allowing it to undergo conformational changes and become prone to proteolytic activation. Meanwhile, MAb binds to cell surface IgG Fc receptor, guiding viral entry through canonical viral-receptor-dependent pathways. Our data suggest that the antibody/Fc-receptor complex functionally mimics viral receptor in mediating viral entry. Moreover, we characterized MAb dosages in viral-receptor-dependent, Fc-receptor-dependent, and both-receptors-dependent viral entry pathways, delineating guidelines on MAb usages in treating viral infections. Our study reveals a novel molecular mechanism for antibody-enhanced viral entry and can guide future vaccination and antiviral strategies.IMPORTANCE Antibody-dependent enhancement (ADE) of viral entry has been observed for many viruses. It was shown that antibodies target one serotype of viruses but only subneutralize another, leading to ADE of the latter viruses. Here we identify a novel mechanism for ADE: a neutralizing antibody binds to the surface spike protein of coronaviruses like a viral receptor, triggers a conformational change of the spike, and mediates viral entry into IgG Fc receptor-expressing cells through canonical viral-receptor-dependent pathways. We further evaluated how antibody dosages impacted viral entry into cells expressing viral receptor, Fc receptor, or both receptors. This study reveals complex roles of antibodies in viral entry and can guide future vaccine design and antibody-based drug therapy.


Assuntos
Anticorpos Antivirais/imunologia , Anticorpos Facilitadores , Coronavírus da Síndrome Respiratória do Oriente Médio/imunologia , Coronavírus da Síndrome Respiratória do Oriente Médio/fisiologia , Glicoproteína da Espícula de Coronavírus/imunologia , Glicoproteína da Espícula de Coronavírus/metabolismo , Internalização do Vírus , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/metabolismo , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/metabolismo , Anticorpos Antivirais/metabolismo , Linhagem Celular , Dipeptidil Peptidase 4/metabolismo , Humanos , Fragmentos Fab das Imunoglobulinas/imunologia , Fragmentos Fab das Imunoglobulinas/metabolismo , Coronavírus da Síndrome Respiratória do Oriente Médio/patogenicidade , Peptídeo Hidrolases/metabolismo , Pró-Proteína Convertases/antagonistas & inibidores , Pró-Proteína Convertases/metabolismo , Conformação Proteica , Domínios Proteicos , Multimerização Proteica , Receptores Fc/metabolismo , Receptores de IgG/imunologia , Receptores de IgG/metabolismo , Receptores Virais/metabolismo , Glicoproteína da Espícula de Coronavírus/química , Tripsina/metabolismo
3.
PLoS One ; 14(12): e0226245, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31887144

RESUMO

Antibody therapies for Alzheimer's Disease (AD) hold promise but have been limited by the inability of these proteins to migrate efficiently across the blood brain barrier (BBB). Central nervous system (CNS) gene transfer by vectors like adeno-associated virus (AAV) overcome this barrier by allowing the bodies' own cells to produce the therapeutic protein, but previous studies using this method to target amyloid-ß have shown success only with truncated single chain antibodies (Abs) lacking an Fc domain. The Fc region mediates effector function and enhances antigen clearance from the brain by neonatal Fc receptor (FcRn)-mediated reverse transcytosis and is therefore desirable to include for such treatments. Here, we show that single chain Abs fused to an Fc domain retaining FcRn binding, but lacking Fc gamma receptor (FcγR) binding, termed a silent scFv-IgG, can be expressed and released into the CNS following gene transfer with AAV. While expression of canonical IgG in the brain led to signs of neurotoxicity, this modified Ab was efficiently secreted from neuronal cells and retained target specificity. Steady state levels in the brain exceeded peak levels obtained by intravenous injection of IgG. AAV-mediated expression of this scFv-IgG reduced cortical and hippocampal plaque load in a transgenic mouse model of progressive ß-amyloid plaque accumulation. These findings suggest that CNS gene delivery of a silent anti-Aß scFv-IgG was well-tolerated, durably expressed and functional in a relevant disease model, demonstrating the potential of this modality for the treatment of Alzheimer's disease.


Assuntos
Doença de Alzheimer/terapia , Sistema Nervoso Central/metabolismo , Vetores Genéticos/administração & dosagem , Fragmentos Fc das Imunoglobulinas/genética , Anticorpos de Cadeia Única/genética , Doença de Alzheimer/genética , Animais , Barreira Hematoencefálica , Linhagem Celular , Dependovirus/genética , Modelos Animais de Doenças , Progressão da Doença , Terapia Genética , Antígenos de Histocompatibilidade Classe I/metabolismo , Humanos , Fragmentos Fc das Imunoglobulinas/química , Fragmentos Fc das Imunoglobulinas/metabolismo , Camundongos , Camundongos Transgênicos , Domínios Proteicos , Receptores Fc/metabolismo , Receptores de IgG/metabolismo , Anticorpos de Cadeia Única/química , Anticorpos de Cadeia Única/metabolismo
4.
Pharm Res ; 36(11): 157, 2019 Sep 06.
Artigo em Inglês | MEDLINE | ID: mdl-31493066

RESUMO

PURPOSE: Although pharmacokinetic (PK) interaction effects of methotrexate (MTX) on adalimumab have been found, the mechanism of these effects is still unclear. In this work, effects of MTX on the concentration of neonatal Fc receptor (FcRn) and the role of FcRn in the interaction between MTX and adalimumab were investigated. METHODS: The experiment was performed in rats whose FcRn had normal physiological function and also in rats whose FcRn was blocked with FcRn antibody. Rats were randomly assigned to receive placebo or 0.2 mg/kg MTX orally every week while taking one abdominal subcutaneous injection of 0.5 mg/kg adalimumab. The FcRn concentration in tissues and the PK parameters of adalimumab were compared between MTX-treated and placebo groups. RESULTS: In rats with normally functioning FcRn, the concentrations of FcRn were significantly increased in the liver (F=105.5, p=0.000) and kidney (F=996.312, p=0.000) after treatment with MTX, and the clearance (CL/F) of adalimumab was decreased accordingly (F=4.423, p=0.048). However, in rats injected with FcRn antibody, the concentrations of FcRn in MTX-treated rats were close to that of the placebo rats in the tissues of the liver (F=1.279, p=0.268) and kidney (F=0.661, p=0.424). The CL/F of adalimumab in rats was also not affected by MTX (F=0.002, p=0.961). CONCLUSIONS: FcRn may play a vital role in the interaction between adalimumab and MTX.


Assuntos
Adalimumab/farmacocinética , Antígenos de Histocompatibilidade Classe I/metabolismo , Metotrexato/metabolismo , Receptores Fc/metabolismo , Adalimumab/administração & dosagem , Animais , Feminino , Humanos , Injeções Subcutâneas , Rim/metabolismo , Fígado/metabolismo , Masculino , Metotrexato/administração & dosagem , Metotrexato/farmacologia , Ratos Sprague-Dawley
5.
Immunity ; 51(4): 750-765.e10, 2019 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-31492649

RESUMO

Immunity that controls parasitemia and inflammation during Plasmodium falciparum (Pf) malaria can be acquired with repeated infections. A limited understanding of this complex immune response impedes the development of vaccines and adjunctive therapies. We conducted a prospective systems biology study of children who differed in their ability to control parasitemia and fever following Pf infection. By integrating whole-blood transcriptomics, flow-cytometric analysis, and plasma cytokine and antibody profiles, we demonstrate that a pre-infection signature of B cell enrichment, upregulation of T helper type 1 (Th1) and Th2 cell-associated pathways, including interferon responses, and p53 activation associated with control of malarial fever and coordinated with Pf-specific immunoglobulin G (IgG) and Fc receptor activation to control parasitemia. Our hypothesis-generating approach identified host molecules that may contribute to differential clinical outcomes during Pf infection. As a proof of concept, we have shown that enhanced p53 expression in monocytes attenuated Plasmodium-induced inflammation and predicted protection from fever.


Assuntos
Linfócitos B/imunologia , Proteínas Sanguíneas/metabolismo , Inflamação/metabolismo , Malária Falciparum/metabolismo , Plasmodium falciparum/fisiologia , Células Th1/imunologia , Células Th2/imunologia , Proteína Supressora de Tumor p53/metabolismo , Adolescente , Adulto , Animais , Anticorpos Antiprotozoários/metabolismo , Criança , Pré-Escolar , Resistência à Doença , Feminino , Perfilação da Expressão Gênica , Humanos , Lactente , Interferons/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Estudos Prospectivos , Receptores Fc/metabolismo , Transdução de Sinais , Proteína Supressora de Tumor p53/genética , Adulto Jovem
6.
J Biol Chem ; 294(38): 13995-14008, 2019 09 20.
Artigo em Inglês | MEDLINE | ID: mdl-31362986

RESUMO

Human immunoglobulin A (IgA) is the most prevalent antibody class at mucosal sites with an important role in mucosal defense. Little is known about the impact of N-glycan modifications of IgA1 and IgA2 on binding to the Fcα receptor (FcαRI), which is also heavily glycosylated at its extracellular domain. Here, we transiently expressed human epidermal growth factor receptor 2 (HER2)-binding monomeric IgA1, IgA2m(1), and IgA2m(2) variants in Nicotiana benthamiana ΔXT/FT plants lacking the enzymes responsible for generating nonhuman N-glycan structures. By coinfiltrating IgA with the respective glycan-modifying enzymes, we generated IgA carrying distinct homogenous N-glycans. We demonstrate that distinctly different N-glycan profiles did not influence antigen binding or the overall structure and integrity of the IgA antibodies but did affect their thermal stability. Using size-exclusion chromatography, differential scanning and isothermal titration calorimetry, surface plasmon resonance spectroscopy, and molecular modeling, we probed distinct IgA1 and IgA2 glycoforms for binding to four different FcαRI glycoforms and investigated the thermodynamics and kinetics of complex formation. Our results suggest that different N-glycans on the receptor significantly contribute to binding affinities for its cognate ligand. We also noted that full-length IgA and FcαRI form a mixture of 1:1 and 1:2 complexes tending toward a 1:1 stoichiometry due to different IgA tailpiece conformations that make it less likely that both binding sites are simultaneously occupied. In conclusion, N-glycans of human IgA do not affect its structure and integrity but its thermal stability, and FcαRI N-glycans significantly modulate binding affinity to IgA.


Assuntos
Imunoglobulina A/metabolismo , Polissacarídeos/química , Receptores Fc/metabolismo , Sítios de Ligação , Glicosilação , Células HEK293 , Humanos , Imunoglobulina A/química , Cinética , Simulação de Dinâmica Molecular , Estabilidade Proteica , Estrutura Quaternária de Proteína , Receptores Fc/química , Receptores Fc/genética , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Termodinâmica , Tabaco/metabolismo
7.
PLoS One ; 14(8): e0220867, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31393930

RESUMO

Phagocytosis is a receptor-mediated process critical to innate immune clearance of pathogens. It proceeds in a regulated sequence of stages: (a) migration of phagocytes towards pathogens, (b) recognition of PAMPs and binding through PRRs, (c) engulfment and internalisation into phagosomes, (d) phagosome maturation, and (e) killing of pathogen or host cells. However, little is known about the role that individual receptors play in these discrete stages in the recognition of fungal cells. In a previous study, we found that dectin-2 deficiency impacted some but not all stages of macrophage-mediated phagocytosis of Candida glabrata. Because the C-type lectin receptor dectin-2 critically requires coupling to the FcRγ chain for signalling, we hypothesised that this coupling may be important for regulating phagocytosis of fungal cargo. We therefore examined how deficiency in FcRγ itself or two receptors to which it couples (dectin-2 and mincle) impacts phagocytosis of six fungal organisms representing three different fungal taxa. Our data show that deficiency in these proteins impairs murine bone marrow-derived macrophage migration, engulfment, and phagosome maturation, but not macrophage survival. Therefore, FcRγ engagement with selective C-type lectin receptors (CLRs) critically affects the spatio-temporal dynamics of fungal phagocytosis.


Assuntos
Fungos/imunologia , Fagocitose , Receptores de Reconhecimento de Padrão/imunologia , Animais , Candida/imunologia , Movimento Celular , Lectinas Tipo C/imunologia , Lectinas Tipo C/metabolismo , Macrófagos/citologia , Malassezia/imunologia , Proteínas de Membrana/imunologia , Proteínas de Membrana/metabolismo , Camundongos , Mucor/imunologia , Ligação Proteica , Receptores Fc/imunologia , Receptores Fc/metabolismo , Receptores de Reconhecimento de Padrão/metabolismo , Saccharomyces/imunologia
8.
Nat Commun ; 10(1): 3020, 2019 07 09.
Artigo em Inglês | MEDLINE | ID: mdl-31289263

RESUMO

Human cytomegalovirus (HCMV) can persistently infect humans, but how HCMV avoids humoral immunity is not clear. The neonatal Fc receptor (FcRn) controls IgG transport from the mother to the fetus and prolongs IgG half-life. Here we show that US11 inhibits the assembly of FcRn with ß2m and retains FcRn in the endoplasmic reticulum (ER), consequently blocking FcRn trafficking to the endosome. Furthermore, US11 recruits the ubiquitin enzymes Derlin-1, TMEM129 and UbE2J2 to engage FcRn, consequently initiating the dislocation of FcRn from the ER to the cytosol and facilitating its degradation. Importantly, US11 inhibits IgG-FcRn binding, resulting in a reduction of IgG transcytosis across intestinal or placental epithelial cells and IgG degradation in endothelial cells. Hence, these results identify the mechanism by which HCMV infection exploits an ER-associated degradation pathway through US11 to disable FcRn functions. These results have implications for vaccine development and immune surveillance.


Assuntos
Infecções por Citomegalovirus/imunologia , Citomegalovirus/imunologia , Antígenos de Histocompatibilidade Classe I/metabolismo , Evasão da Resposta Imune , Imunidade Humoral , Proteínas de Ligação a RNA/metabolismo , Receptores Fc/metabolismo , Proteínas Virais/metabolismo , Linhagem Celular , Citomegalovirus/patogenicidade , Infecções por Citomegalovirus/virologia , Degradação Associada com o Retículo Endoplasmático/imunologia , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Imunoglobulina G/imunologia , Imunoglobulina G/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Mutagênese Sítio-Dirigida , RNA Interferente Pequeno/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/imunologia , Receptores Fc/imunologia , Enzimas de Conjugação de Ubiquitina/genética , Enzimas de Conjugação de Ubiquitina/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , Proteínas Virais/genética , Proteínas Virais/imunologia
9.
J Chromatogr A ; 1603: 15-22, 2019 Oct 11.
Artigo em Inglês | MEDLINE | ID: mdl-31213362

RESUMO

Affinity chromatography technologies play an important role in the purification of antibodies. To prepare affinity materials, prior isolation and purification of affinity ligands are required before coupling onto solid supports, which is quite expensive and laborious in large-scale applications. In this study, a one-step approach which circumvents the ligand purification procedures was developed to fabricate affinity gel for purifying immunoglobulin G (IgG). A self-labeling tag, haloalkane dehalogenase, was fused to the C-terminal of an anti-Fc variable domain of the heavy chain of the heavy-chain antibody (AFV) which was isolated in previous work. The AFV binds to various sources of IgG and is highly thermal stable. The fusion protein, namely HAFV, was expressed in Escherichia coli as a soluble protein. The binding affinity of HAFV to the Fc region of IgG was characterized and compared with the untagged anti-Fc nanobody. Next, the HAFV was immobilized directly from the crude cell lysate of isopropylthio-ß-D-galactoside (IPTG) induced E. coli. The effects of NaCl concentrations and pH on the capacity of the HAFV resin were investigated. In addition, the one-step coupled HAFV resin was compared with the AFV resin and commercial resins (Protein A and Protein G) by evaluating the static capacity and stability. Though the Protein A (8.34 ±â€¯0.37 mg/ml) and Protein G (9.19 ±â€¯0.28 mg/ml) showed higher static capacity, the static capacity of HAFV resin (8.21 ±â€¯0.30 mg/ml) was better than that of the untagged AFV gel (6.48 ±â€¯0.56 mg/ml). The recovery results calculated for the reusability and stability show that there is no significant difference between the results obtained for the HAFV gel with those of the untagged AFV gel and commercial Protein A and G. After stored at 37 ℃ for 7 days and recycled 10 times, the static capacity of HAFV gel remains above 78%. Our strategy is site-specific, cost-effective, reproducible, and has the potential to dramatically cut down the costs of affinity materials for IgG purification.


Assuntos
Cromatografia de Afinidade/métodos , Escherichia coli/metabolismo , Imunoglobulina G/isolamento & purificação , Proteínas Recombinantes/metabolismo , Anticorpos de Domínio Único/metabolismo , Imunoglobulina G/imunologia , Imunoglobulina G/metabolismo , Ligantes , Ligação Proteica , Receptores Fc/química , Receptores Fc/metabolismo , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/isolamento & purificação , Anticorpos de Domínio Único/imunologia , Anticorpos de Domínio Único/isolamento & purificação , Proteína Estafilocócica A/química , Proteína Estafilocócica A/metabolismo
10.
Cell ; 178(1): 202-215.e14, 2019 06 27.
Artigo em Inglês | MEDLINE | ID: mdl-31204102

RESUMO

Despite the worldwide success of vaccination, newborns remain vulnerable to infections. While neonatal vaccination has been hampered by maternal antibody-mediated dampening of immune responses, enhanced regulatory and tolerogenic mechanisms, and immune system immaturity, maternal pre-natal immunization aims to boost neonatal immunity via antibody transfer to the fetus. However, emerging data suggest that antibodies are not transferred equally across the placenta. To understand this, we used systems serology to define Fc features associated with antibody transfer. The Fc-profile of neonatal and maternal antibodies differed, skewed toward natural killer (NK) cell-activating antibodies. This selective transfer was linked to digalactosylated Fc-glycans that selectively bind FcRn and FCGR3A, resulting in transfer of antibodies able to efficiently leverage innate immune cells present at birth. Given emerging data that vaccination may direct antibody glycosylation, our study provides insights for the development of next-generation maternal vaccines designed to elicit antibodies that will most effectively aid neonates.


Assuntos
Antígenos de Histocompatibilidade Classe I/imunologia , Antígenos de Histocompatibilidade Classe I/metabolismo , Imunoglobulina G/metabolismo , Placenta/metabolismo , Polissacarídeos/metabolismo , Receptores Fc/imunologia , Receptores Fc/metabolismo , Adolescente , Adulto , Bélgica , Degranulação Celular , Estudos de Coortes , Feminino , Glicosilação , Humanos , Recém-Nascido , Células Matadoras Naturais/imunologia , Ativação Linfocitária/imunologia , Masculino , Gravidez , Receptores de IgG/metabolismo , Células THP-1 , Estados Unidos , Vacinação , Adulto Jovem
11.
Cell ; 177(6): 1553-1565.e16, 2019 05 30.
Artigo em Inglês | MEDLINE | ID: mdl-31104841

RESUMO

Enterovirus B (EV-B), a major proportion of the genus Enterovirus in the family Picornaviridae, is the causative agent of severe human infectious diseases. Although cellular receptors for coxsackievirus B in EV-B have been identified, receptors mediating virus entry, especially the uncoating process of echovirus and other EV-B remain obscure. Here, we found that human neonatal Fc receptor (FcRn) is the uncoating receptor for major EV-B. FcRn binds to the virus particles in the "canyon" through its FCGRT subunit. By obtaining multiple cryo-electron microscopy structures at different stages of virus entry at atomic or near-atomic resolution, we deciphered the underlying mechanisms of enterovirus attachment and uncoating. These structures revealed that different from the attachment receptor CD55, binding of FcRn to the virions induces efficient release of "pocket factor" under acidic conditions and initiates the conformational changes in viral particle, providing a structural basis for understanding the mechanisms of enterovirus entry.


Assuntos
Enterovirus Humano B/metabolismo , Antígenos de Histocompatibilidade Classe I/metabolismo , Antígenos de Histocompatibilidade Classe I/ultraestrutura , Receptores Fc/metabolismo , Receptores Fc/ultraestrutura , Capsídeo/metabolismo , Microscopia Crioeletrônica , Enterovirus , Enterovirus Humano B/patogenicidade , Infecções por Enterovirus/metabolismo , Antígenos de Histocompatibilidade Classe I/fisiologia , Humanos , Modelos Moleculares , Filogenia , Receptores Fc/fisiologia , Vírion , Internalização do Vírus
12.
AAPS J ; 21(4): 62, 2019 05 06.
Artigo em Inglês | MEDLINE | ID: mdl-31062128

RESUMO

Fusing the human immunoglobulin G1 (IgG1) constant region (Fc-domain) to therapeutic proteins or peptides increases their circulating plasma half-life via neonatal Fc receptor (FcRn) binding and recycling. However, Fc-mediated interactions with other molecules including complement C1q and Fc gamma receptors (FcγRs) can have immunological consequences and the potential to modulate the immunogenicity of Fc-fusion therapeutics. In a comparative study, we carried out a comprehensive assessment of Fc-mediated interactions for five FDA-approved Fc-fusion therapeutics. C1q binding and complement activation were measured by ELISA, while FcγR binding and signaling were evaluated using BW5147:FcγR-ζ reporter cell lines. We demonstrate that FIX-Fc and FVIII-Fc bound C1q as well as activating and inhibitory FcγRs (I, IIA, IIB, IIIA). These coagulation factor Fc-fusions also signaled via FcγRIIIA, and to a lesser extent via FcγRI and FcγRIIB. TNFR-Fc and CTLA4-Fc bound FcγRI, while TNFR-Fc also bound FcγRIIIA, but these interactions did not result in FcγR signaling. Our comprehensive assessment demonstrates that (i) different Fc-fusion drugs have distinct C1q/FcγR binding and signaling properties, (ii) FcγR binding does not predict signaling, and (iii) the fusion partner (effector molecule) can influence Fc-mediated interactions.


Assuntos
Anticorpos Monoclonais/imunologia , Complemento C1q/imunologia , Antígenos de Histocompatibilidade Classe I/imunologia , Fragmentos Fc das Imunoglobulinas/imunologia , Receptores Fc/imunologia , Receptores de IgG/metabolismo , Animais , Anticorpos Monoclonais/metabolismo , Linhagem Celular Tumoral , Complemento C1q/metabolismo , Hipersensibilidade a Drogas/imunologia , Hipersensibilidade a Drogas/metabolismo , Antígenos de Histocompatibilidade Classe I/metabolismo , Humanos , Fragmentos Fc das Imunoglobulinas/metabolismo , Camundongos , Ligação Proteica , Receptores Fc/metabolismo , Transdução de Sinais
13.
Vet Immunol Immunopathol ; 211: 75-84, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31084898

RESUMO

The primary function of the mammary gland is to produce milk to feed the suckling young. In ruminants, ingestion of maternal antibodies in mammary secretions facilitates the transfer of passive immunity from mother to young, providing antibody-mediated immunity to protect the neonate against disease while their own immune system develops. Antibodies in mammary secretions also play a role in protecting the gland itself against infection. Here we provide a brief history of studies on immunoglobulins in ruminant mammary secretions and review recent findings describing the mechanisms by which antibody-producing plasmablasts are recruited to the gland and immunoglobulins are transported into ruminant mammary secretions. An improved understanding of the complex interaction of factors which regulate immunoglobulin production and transfer to the ruminant mammary gland may provide opportunities to enhance antibody concentrations in mammary secretions during normal lactation and in response to immunisation. Strategies aimed at increasing antibody concentrations in ruminant mammary secretions have the potential to improve the ability of animals to resist mammary infections, enhance the transfer of passive immunity from mother to young and increase the feasibility of harvesting antibodies from the mammary secretions for use in commercial therapeutic applications for humans and domesticated animals.


Assuntos
Imunoglobulinas/metabolismo , Glândulas Mamárias Animais/imunologia , Ruminantes/imunologia , Animais , Transporte Biológico Ativo , Bovinos , Feminino , Imunoglobulina E/metabolismo , Glândulas Mamárias Animais/metabolismo , Leite/imunologia , Receptores Fc/metabolismo , Ruminantes/metabolismo , Ovinos
14.
Am J Health Syst Pharm ; 76(11): 789-794, 2019 May 17.
Artigo em Inglês | MEDLINE | ID: mdl-30951590

RESUMO

PURPOSE: The pharmacology, pharmacokinetics, efficacy, safety, dosing and administration, and place in therapy of fostamatinib, a novel spleen tyrosine kinase inhibitor for the treatment of adult immune thrombocytopenia that has had an insufficient response to a previous treatment are summarized. SUMMARY: Fostamatinib is an oral inhibitor of spleen tyrosine kinase that is expressed on hematopoietic cells and plays a key role in the accelerated destruction of platelets through Fc-receptor activation. Fostamatinib is indicated for the treatment of adults with immune thrombocytopenia that has had an insufficient response to a previous treatment. In 2 parallel, identically designed, placebo-controlled Phase III trials, patients with persistent and chronic immune thrombocytopenia treated with fostamatinib demonstrated clinically meaningful responses in platelet counts with lower rates of moderate and severe bleeding-related adverse events. Overall, fostamatinib therapy is generally well tolerated; the most common adverse events reported in clinical trials were diarrhea, nausea, hypertension, liver function test elevations, and infection. Being primarily metabolized through the CYP3A4 pathway, fostamatinib is subject to drug-drug interactions and concomitant administration with strong CYP3A4 inhibitors or inducers can affect fostamatinib exposure. CONCLUSION: Fostamatinib is a first-in-class spleen tyrosine kinase inhibitor approved for the treatment of adults with immune thrombocytopenia that has had an insufficient response to a previous treatment.


Assuntos
Plaquetas/efeitos dos fármacos , Oxazinas/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Púrpura Trombocitopênica Idiopática/tratamento farmacológico , Piridinas/farmacologia , Quinase Syk/antagonistas & inibidores , Administração Oral , Plaquetas/imunologia , Ensaios Clínicos Fase III como Assunto , Relação Dose-Resposta a Droga , Humanos , Oxazinas/uso terapêutico , Contagem de Plaquetas , Inibidores de Proteínas Quinases/uso terapêutico , Púrpura Trombocitopênica Idiopática/imunologia , Piridinas/uso terapêutico , Receptores Fc/imunologia , Receptores Fc/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/imunologia , Quinase Syk/metabolismo , Resultado do Tratamento
15.
MAbs ; 11(5): 848-860, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-30964743

RESUMO

The maintenance of the homeostasis of immunoglobulin G (IgG) represents a fundamental aspect of humoral immunity that has direct relevance to the successful delivery of antibody-based therapeutics. The ubiquitously expressed neonatal Fc receptor (FcRn) salvages IgG from cellular degradation following pinocytic uptake into cells, conferring prolonged in vivo persistence on IgG. However, the cellular sites of FcRn function are poorly defined. Pinocytic uptake is a prerequisite for FcRn-mediated IgG salvage, prompting us to investigate the consequences of IgG uptake and catabolism by macrophages, which represent both abundant and highly pinocytic cells in the body. Site-specific deletion of FcRn to generate mice harboring FcRn-deficient macrophages results in IgG hypercatabolism and ~threefold reductions in serum IgG levels, whereas these effects were not observed in mice that lack functional FcRn in B cells and dendritic cells. Consistent with the degradative activity of FcRn-deficient macrophages, depletion of these cells in FcRn-deficient mice leads to increased persistence and serum levels of IgG. These studies demonstrate a pivotal role for FcRn-mediated salvage in compensating for the high pinocytic and degradative activities of macrophages to maintain IgG homeostasis.


Assuntos
Antígenos de Histocompatibilidade Classe I/metabolismo , Imunoglobulina G/sangue , Macrófagos/imunologia , Receptores Fc/metabolismo , Animais , Linfócitos B , Linhagem Celular , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Células Endoteliais , Antígenos de Histocompatibilidade Classe I/genética , Homeostase/imunologia , Humanos , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pinocitose/imunologia , Receptores Fc/genética
16.
mBio ; 10(2)2019 04 02.
Artigo em Inglês | MEDLINE | ID: mdl-30940710

RESUMO

Zika virus is a mosquito-borne flavivirus which can cause severe disease in humans, including microcephaly and other congenital malformations in newborns and Guillain-Barré syndrome in adults. There are currently no approved prophylactics or therapeutics for Zika virus; the development of a safe and effective vaccine is an urgent priority. Preclinical studies suggest that the envelope glycoprotein can elicit potently neutralizing antibodies. However, such antibodies are implicated in the phenomenon of antibody-dependent enhancement of disease. We have previously shown that monoclonal antibodies targeting the Zika virus nonstructural NS1 protein are protective without inducing antibody-dependent enhancement of disease. Here, we investigated whether the NS1 protein itself is a viable vaccine target. Wild-type mice were vaccinated with an NS1-expressing DNA plasmid followed by two adjuvanted protein boosters, which elicited high antibody titers. Passive transfer of the immune sera was able to significantly protect STAT2 knockout mice against lethal challenge by Zika virus. In addition, long-lasting NS1-specific IgG responses were detected in serum samples from patients in either the acute or the convalescent phase of Zika virus infection. These NS1-specific antibodies were able to functionally engage Fcγ receptors. In contrast, envelope-specific antibodies did not activate Fc-mediated effector functions on infected cells. Our data suggest that the Zika virus NS1 protein, which is expressed on infected cells, is critical for Fc-dependent cell-mediated immunity. The present study demonstrates that the Zika virus NS1 protein is highly immunogenic and can elicit protective antibodies, underscoring its potential for an effective Zika virus vaccine.IMPORTANCE Zika virus is a global public health threat that causes microcephaly and congenital malformations in newborns and Guillain-Barré syndrome in adults. Currently, no vaccines or treatments are available. While antibodies targeting the envelope glycoprotein can neutralize virus, they carry the risk of antibody-dependent enhancement of disease (ADE). In contrast, antibodies generated against the NS1 protein can be protective without eliciting ADE. The present study demonstrates the effectiveness of an NS1-based vaccine in eliciting high titers of protective antibodies against Zika virus disease in a mouse model. Sera generated by this vaccine can elicit Fc-mediated effector functions against Zika virus-infected cells. Lastly, we provide human data suggesting that the antibody response against the Zika virus NS1 protein is long-lasting and functionally active. Overall, our work will inform the development of a safe and effective Zika virus vaccine.


Assuntos
Anticorpos Antivirais/sangue , Proteínas não Estruturais Virais/imunologia , Vacinas Virais/imunologia , Infecção por Zika virus/prevenção & controle , Animais , Linhagem Celular , Modelos Animais de Doenças , Humanos , Imunidade Celular , Esquemas de Imunização , Imunização Passiva , Imunoglobulina G/sangue , Camundongos , Camundongos Knockout , Receptores Fc/metabolismo , Análise de Sobrevida , Vacinas de DNA/administração & dosagem , Vacinas de DNA/imunologia , Vacinas de Subunidades/administração & dosagem , Vacinas de Subunidades/imunologia , Vacinas Virais/administração & dosagem
17.
Placenta ; 78: 36-43, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30955709

RESUMO

INTRODUCTION: Starting from the second trimester of pregnancy, passive immunity is provided to the human fetus by transplacental transfer of maternal IgG. IgG transfer depends on the neonatal Fc receptor, FcRn. While FcRn localization in the placental syncytiotrophoblast (STB) has been demonstrated unequivocally, FcRn expression in placental-fetal endothelial cells (pFECs), which are part of the materno-fetal barrier, is still unclear. Therefore, this study aimed to elucidate the spatio-specific expression pattern of FcRn in placental tissue. METHODS: FcRn expression was investigated by western blotting in term placentas and in isolated human placental arterial and venous endothelial cells (HPAEC, HPVEC) using a validated affinity-purified polyclonal anti-peptide antibody against the cytoplasmic tail of FcRn α-chain. In situ localization of FcRn and IgG was studied by immunofluorescence microscopy on tissue sections of healthy term placentas. RESULTS: FcRn expression was demonstrated in placental vasculature particularly, in HPAEC, and HPVEC. FcRn was localized in cytokeratin 7+ STB and in CD31+ pFECs in terminal as well as stem villi in situ. Additionally, CD68+ placental macrophages exhibited FcRn expression in situ. Endogenous IgG partially co-localized with FcRn in STB, pFECs, and in placental macrophages. DISCUSSION: Placental FcRn expression in endothelial cells and macrophages is analogous to the expression pattern in other organs. FcRn expression in pFECs suggests an involvement of FcRn in IgG transcytosis and/or participation in recycling/salvaging of maternal IgG present in the fetal circulation. FcRn expression in placental macrophages may account for recycling of monomeric IgG and/or processing and presentation of immune complexes.


Assuntos
Células Endoteliais/metabolismo , Endotélio/metabolismo , Feto/metabolismo , Antígenos de Histocompatibilidade Classe I/metabolismo , Sistema Imunitário/metabolismo , Placenta/metabolismo , Receptores Fc/metabolismo , Células Cultivadas , Vilosidades Coriônicas/imunologia , Vilosidades Coriônicas/metabolismo , Células Endoteliais/imunologia , Endotélio/citologia , Endotélio/imunologia , Feminino , Feto/citologia , Células HL-60 , Humanos , Imunoglobulina G/metabolismo , Troca Materno-Fetal , Placenta/citologia , Gravidez , Células Estromais/metabolismo , Trofoblastos/metabolismo
18.
Mol Pharm ; 16(6): 2385-2393, 2019 06 03.
Artigo em Inglês | MEDLINE | ID: mdl-31002261

RESUMO

nab-Paclitaxel ( nab-P), an albumin-bound formulation of paclitaxel, was developed to improve the tolerability and antitumor activity of taxanes. The neonatal Fc receptor (FcRn) is a transport protein that can bind to albumin and regulate the homeostasis of circulating albumin. Therefore, the pharmacokinetics and pharmacodynamics of nab-P may be impacted by FcRn expression. This study aimed to investigate the effects of FcRn on nab-P elimination and distribution to targeted tissues. Wild-type and FcRn-knockout (FcRn-KO) mice were treated with nab-P, mouse-specific nab-P (distribution experiments only), and solvent-based paclitaxel (pac-T). Blood and tissue samples were collected for distribution analyses. Organ, urine, and fecal samples were collected for elimination analyses. The nab-P tissue penetration in the pancreas, fat pad, and kidney of wild-type mice, as reflected by the ratio of tissue/plasma concentration, was significantly higher (ranging from 5 to 80 fold) than that of FcRn-KO mice. In contrast, the tissue penetration of pac-T in these organs of FcRn-KO mice was similar to that of wild-type mice. More importantly, the excretion of nab-P in feces of FcRn-KO mice (45-68%) was significantly higher than that of wild-type mice (26-46%) from 8 to 48 h post treatment. In comparison, the difference of excretion of pac-T in feces between FcRn-KO mice and wild-type mice was smaller than that of nab-P. Furthermore, greater tissue penetration and fecal excretion were observed with nab-P than pac-T in both FcRn-KO and wild-type mice. These findings suggest that FcRn enhances the tissue distribution and penetration of nab-P in the targeted organs, while FcRn prevents excretion of nab-P to feces in the intestinal lumen. The findings support the notion that albumin nanoparticle delivery alters drug distribution and elimination through an FcRn-mediated process to impact drug efficacy and toxicity.


Assuntos
Antígenos de Histocompatibilidade Classe I/metabolismo , Paclitaxel/metabolismo , Receptores Fc/metabolismo , Albuminas/química , Animais , Camundongos , Camundongos Knockout , Nanopartículas/química , Distribuição Tecidual
19.
Int J Mol Sci ; 20(6)2019 Mar 19.
Artigo em Inglês | MEDLINE | ID: mdl-30893823

RESUMO

Monoclonal antibodies (mAbs) are promising therapies to treat airway chronic inflammatory disease (asthma or nasal polyps). To date, no study has specifically assessed, in vitro, the potential function of neonatal Fc receptor (FcRn) in IgG transcytosis through the human nasal airway epithelium. The objective of this study was to report the in vitro expression and function of FcRn in nasal human epithelium. FcRn expression was studied in an air⁻liquid interface (ALI) primary culture model of human nasal epithelial cells (HNEC) from polyps. FcRn expression was characterized by quantitative RT-PCR, western blot, and immunolabeling. The ability of HNECs to support mAb transcytosis via FcRn was assessed by transcytosis assay. This study demonstrates the expression of FcRn mRNA and protein in HNEC. We report a high expression of FcRn in the cytosol of ciliated, mucus, and basal cells by immunohistochemistry with a higher level of FcRn proteins in differentiated HNEC. We also proved in vitro transepithelial delivery of an IgG1 therapeutic mAb with a dose⁻response curve. This is the first time that FcRn expression and mAb transcytosis has been shown in a model of human nasal respiratory epithelium in vitro. This study is a prerequisite for FcRn-dependent nasal administration of mAbs.


Assuntos
Anticorpos Monoclonais/metabolismo , Sistemas de Liberação de Medicamentos , Células Epiteliais/metabolismo , Antígenos de Histocompatibilidade Classe I/metabolismo , Nariz/citologia , Receptores Fc/metabolismo , Transcitose , Diferenciação Celular , Células HEK293 , Humanos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
20.
Immunobiology ; 224(3): 362-370, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30876792

RESUMO

Given the heightened interest in manipulation of co-signaling cascades for cancer immunotherapy, we sought to determine how/whether tumors decorated with therapeutic monoclonal antibodies (mAbs) impact the expression of co-signaling molecules on human NK cells. Stimulation of NK cells with aggregated IgG1 resulted in the upregulation of HAVCR2 - the gene encoding T-cell immunoglobulin and mucin-containing domain (Tim)-3 - known to be involved in the induction of peripheral T cell tolerance. This upregulation of HAVCR2 was recapitulated at the protein level, following NK cell stimulation by either mAb opsonized tumors, recombinant human IgG1 Fc multimer, and/or non-Fc stimuli e.g. IL-12/IL-18. The patterns of Tim-3 expression were temporally distinct from the FcR mediated induction of the co-signaling molecule, 4-1BB (CD137), with Tim-3 increases observed twenty minutes following exposure to Fc multimers and remaining at high levels for at least six hours, while increases in CD137 expression were first observed at the four-hour time point. Importantly, these Tim-3+ NK cells were functionally diverse, as evidenced by the fact that their ability to produce IFN-γ in response to an NK cell responsive tumor was strictly dependent upon the stimuli employed for Tim-3 induction. These data suggest that Tim-3 upregulation is the common end-result of NK cell activation by a variety of unique and overlapping stimuli and is not an independent marker of NK cell exhaustion. Furthermore, our observations potentially explain the diverse functionality attributed to Tim-3+ NK cells and should be considered prior to use of anti-Tim-3 inhibitory mAbs for cancer immunotherapy.


Assuntos
Anticorpos Monoclonais/uso terapêutico , Receptor Celular 2 do Vírus da Hepatite A/metabolismo , Imunoglobulina G/metabolismo , Imunoterapia/métodos , Células Matadoras Naturais/imunologia , Neoplasias/terapia , Células Cultivadas , Receptor Celular 2 do Vírus da Hepatite A/genética , Receptor Celular 2 do Vírus da Hepatite A/imunologia , Humanos , Tolerância Imunológica , Interferon gama/metabolismo , Células K562 , Ativação Linfocitária , Neoplasias/imunologia , Multimerização Proteica , Agregação de Receptores , Receptores Fc/metabolismo , Membro 9 da Superfamília de Receptores de Fatores de Necrose Tumoral/metabolismo , Regulação para Cima
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