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1.
Antiviral Res ; 196: 105197, 2021 12.
Artigo em Inglês | MEDLINE | ID: mdl-34774603

RESUMO

SARS-CoV-2 enters host cells after binding through its spike glycoprotein to the angiotensin-converting enzyme 2 (ACE2) receptor. Soluble ACE2 ectodomains bind and neutralize the virus, yet their short in vivo half-live limits their therapeutic use. This limitation can be overcome by fusing the fragment crystallizable (Fc) part of human immunoglobulin G (IgG) to the ACE2 ectodomain, but this bears the risk of Fc-receptor activation and antibody-dependent cellular cytotoxicity. Here, we describe optimized ACE2-IgG4-Fc fusion constructs that avoid Fc-receptor activation, preserve the desired ACE2 enzymatic activity and show promising pharmaceutical properties. The engineered ACE2-IgG4-Fc fusion proteins neutralize the original SARS-CoV, pandemic SARS-CoV-2 as well as the rapidly spreading SARS-CoV-2 alpha, beta and delta variants of concern. Importantly, these variants of concern are inhibited at picomolar concentrations proving that ACE2-IgG4 maintains - in contrast to therapeutic antibodies - its full antiviral potential. Thus, ACE2-IgG4-Fc fusion proteins are promising candidate anti-antivirals to combat the current and future pandemics.


Assuntos
Enzima de Conversão de Angiotensina 2 , Antivirais/síntese química , Tratamento Farmacológico da COVID-19 , Imunoglobulina G , Internalização do Vírus/efeitos dos fármacos , Enzima de Conversão de Angiotensina 2/química , Enzima de Conversão de Angiotensina 2/uso terapêutico , Antivirais/uso terapêutico , Humanos , Ligação Proteica , SARS-CoV-2/efeitos dos fármacos
2.
Cancer Cell ; 31(6): 771-789.e6, 2017 06 12.
Artigo em Inglês | MEDLINE | ID: mdl-28609656

RESUMO

Intrahepatic cholangiocarcinoma (ICC) is a highly malignant, heterogeneous cancer with poor treatment options. We found that mitochondrial dysfunction and oxidative stress trigger a niche favoring cholangiocellular overgrowth and tumorigenesis. Liver damage, reactive oxygen species (ROS) and paracrine tumor necrosis factor (Tnf) from Kupffer cells caused JNK-mediated cholangiocellular proliferation and oncogenic transformation. Anti-oxidant treatment, Kupffer cell depletion, Tnfr1 deletion, or JNK inhibition reduced cholangiocellular pre-neoplastic lesions. Liver-specific JNK1/2 deletion led to tumor reduction and enhanced survival in Akt/Notch- or p53/Kras-induced ICC models. In human ICC, high Tnf expression near ICC lesions, cholangiocellular JNK-phosphorylation, and ROS accumulation in surrounding hepatocytes are present. Thus, Kupffer cell-derived Tnf favors cholangiocellular proliferation/differentiation and carcinogenesis. Targeting the ROS/Tnf/JNK axis may provide opportunities for ICC therapy.


Assuntos
Neoplasias dos Ductos Biliares/metabolismo , Colangiocarcinoma/metabolismo , Células de Kupffer/metabolismo , Sistema de Sinalização das MAP Quinases , Fator de Necrose Tumoral alfa/metabolismo , Animais , Neoplasias dos Ductos Biliares/patologia , Hidroxianisol Butilado/uso terapêutico , Carcinogênese , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Transformação Celular Neoplásica/efeitos dos fármacos , Colangiocarcinoma/patologia , Humanos , Células de Kupffer/efeitos dos fármacos , Fígado/efeitos dos fármacos , Fígado/patologia , Camundongos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Mitocôndrias/fisiologia , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Receptores do Fator de Necrose Tumoral/genética , Receptores do Fator de Necrose Tumoral/metabolismo , Transdução de Sinais , Microambiente Tumoral
3.
Hepatology ; 61(1): 238-48, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-25131778

RESUMO

UNLABELLED: Due to its ability to inhibit prometastatic matrix metalloproteinases, tissue inhibitor of metalloproteinases (TIMP)-1 has been thought to suppress tumor metastasis. However, elevated systemic levels of TIMP-1 correlate with poor prognosis in cancer patients, suggesting a metastasis-stimulating role of TIMP-1. In colorectal cancer patients, tumor as well as plasma TIMP-1 levels were correlated with synchronous liver metastasis or distant metastasis-associated disease relapse. In mice, high systemic TIMP-1 levels increased the liver susceptibility towards metastasis by triggering the formation of a premetastatic niche. This promoted hepatic metastasis independent of origin or intrinsic metastatic potential of tumor cells. High systemic TIMP-1 led to increased hepatic SDF-1 levels, which in turn promoted recruitment of neutrophils to the liver. Both inhibition of SDF-1-mediated neutrophil recruitment and systemic depletion of neutrophils reduced TIMP-1-induced increased liver susceptibility towards metastasis. This indicates a crucial functional role of neutrophils in the TIMP-1-induced premetastatic niche. CONCLUSION: Our results identify TIMP-1 as an essential promoter of hepatic premetastatic niche formation.


Assuntos
Carcinoma/secundário , Quimiocina CXCL12/metabolismo , Neoplasias Hepáticas/secundário , Infiltração de Neutrófilos , Receptores CXCR4/metabolismo , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Animais , Carcinoma/sangue , Linhagem Celular Tumoral , Humanos , Fígado/imunologia , Fígado/metabolismo , Neoplasias Hepáticas/sangue , Camundongos , Camundongos Endogâmicos , Células NIH 3T3 , Inibidor Tecidual de Metaloproteinase-1/sangue
4.
Int J Cancer ; 136(10): 2304-15, 2015 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-25354204

RESUMO

The tetraspanin CD63 is implicated in pro-metastatic signaling pathways but, so far, it is unclear, how CD63 levels affect the tumor cell phenotype. Here, we investigated the effect of CD63 modulation in different metastatic tumor cell lines. In vitro, knock down of CD63 induced a more epithelial-like phenotype concomitant with increased E-cadherin expression, downregulation of its repressors Slug and Zeb1, and decreased N-cadherin. In addition, ß-catenin protein was markedly reduced, negatively affecting expression of the target genes MMP-2 and PAI-1. ß-catenin inhibitors mimicked the epithelial phenotype induced by CD63 knock down. Inhibition of ß-catenin upstream regulators PI3K/AKT or GSK3ß could rescue the mesenchymal phenotype underlining the importance of the ß-catenin pathway in CD63-regulated cell plasticity. CD63 knock down-induced phenotypical changes correlated with a decrease of experimental metastasis whereas CD63 overexpression enhanced the tumor cell-intrinsic metastatic potential. Taken together, our data show that CD63 is a crucial player in the regulation of the tumor cell-intrinsic metastatic potential by affecting cell plasticity.


Assuntos
Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Tetraspanina 30/metabolismo , beta Catenina/metabolismo , Linhagem Celular Tumoral , Transição Epitelial-Mesenquimal , Feminino , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Dados de Sequência Molecular , Transdução de Sinais , Tetraspanina 30/genética
5.
Cancer Cell ; 26(4): 549-64, 2014 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-25314080

RESUMO

Hepatocellular carcinoma (HCC), the fastest rising cancer in the United States and increasing in Europe, often occurs with nonalcoholic steatohepatitis (NASH). Mechanisms underlying NASH and NASH-induced HCC are largely unknown. We developed a mouse model recapitulating key features of human metabolic syndrome, NASH, and HCC by long-term feeding of a choline-deficient high-fat diet. This induced activated intrahepatic CD8(+) T cells, NKT cells, and inflammatory cytokines, similar to NASH patients. CD8(+) T cells and NKT cells but not myeloid cells promote NASH and HCC through interactions with hepatocytes. NKT cells primarily cause steatosis via secreted LIGHT, while CD8(+) and NKT cells cooperatively induce liver damage. Hepatocellular LTßR and canonical NF-κB signaling facilitate NASH-to-HCC transition, demonstrating that distinct molecular mechanisms determine NASH and HCC development.


Assuntos
Ativação Metabólica , Linfócitos T CD8-Positivos/imunologia , Fígado Gorduroso/imunologia , Hepatócitos/imunologia , Células Matadoras Naturais/imunologia , Neoplasias Hepáticas/imunologia , Animais , Humanos , Camundongos , Camundongos Endogâmicos C57BL
6.
Cancer Cell ; 22(1): 91-105, 2012 Jul 10.
Artigo em Inglês | MEDLINE | ID: mdl-22789541

RESUMO

Increased expression of the chemokine CCL2 in tumor cells correlates with enhanced metastasis, poor prognosis, and recruitment of CCR2(+)Ly6C(hi) monocytes. However, the mechanisms driving tumor cell extravasation through the endothelium remain elusive. Here, we describe CCL2 upregulation in metastatic UICC stage IV colon carcinomas and demonstrate that tumor cell-derived CCL2 activates the CCR2(+) endothelium to increase vascular permeability in vivo. CCR2 deficiency prevents colon carcinoma extravasation and metastasis. Of note, CCR2 expression on radio-resistant cells or endothelial CCR2 expression restores extravasation and metastasis in Ccr2(-/-) mice. Reduction of CCR2 expression on myeloid cells decreases but does not prevent metastasis. CCL2-induced vascular permeability and metastasis is dependent on JAK2-Stat5 and p38MAPK signaling. Our study identifies potential targets for treating CCL2-dependent metastasis.


Assuntos
Neoplasias do Colo/metabolismo , Extravasamento de Materiais Terapêuticos e Diagnósticos , Janus Quinase 2/metabolismo , Receptores CCR2/metabolismo , Fator de Transcrição STAT5/metabolismo , Transdução de Sinais , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Linhagem Celular Tumoral , Neoplasias do Colo/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout
7.
Blood ; 120(7): 1516-27, 2012 Aug 16.
Artigo em Inglês | MEDLINE | ID: mdl-22740442

RESUMO

Blood vessel networks form in a 2-step process of sprouting angiogenesis followed by selective branch regression and stabilization of remaining vessels. Pericytes are known to function in stabilizing blood vessels, but their role in vascular sprouting and selective vessel regression is poorly understood. The endosialin (CD248) receptor is expressed by pericytes associated with newly forming but not stable quiescent vessels. In the present study, we used the Endosialin(-/-) mouse as a means to uncover novel roles for pericytes during the process of vascular network formation. We demonstrate in a postnatal retina model that Endosialin(-/-) mice have normal vascular sprouting but are defective in selective vessel regression, leading to increased vessel density. Examination of the Endosialin(-/-) mouse tumor vasculature revealed an equivalent phenotype, indicating that pericytes perform a hitherto unidentified function to promote vessel destabilization and regression in vivo in both physiologic and pathologic angiogenesis. Mechanistically, Endosialin(-/-) mice have no defect in pericyte recruitment. Rather, endosialin binding to an endothelial associated, but not a pericyte associated, basement membrane component induces endothelial cell apoptosis and detachment. The results of the present study advance our understanding of pericyte biology and pericyte/endothelial cell cooperation during vascular patterning and have implications for the design of both pro- and antiangiogenic therapies.


Assuntos
Vasos Sanguíneos/crescimento & desenvolvimento , Vasos Sanguíneos/patologia , Padronização Corporal , Neovascularização Fisiológica , Pericitos/patologia , Animais , Animais Recém-Nascidos , Antígenos CD/metabolismo , Aorta/crescimento & desenvolvimento , Aorta/patologia , Apoptose , Membrana Basal/metabolismo , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Técnicas In Vitro , Camundongos , Camundongos Endogâmicos C57BL , Proteínas de Neoplasias/deficiência , Proteínas de Neoplasias/metabolismo , Pericitos/metabolismo , Ratos , Retina/metabolismo , Retina/patologia , Vasos Retinianos/crescimento & desenvolvimento , Transdução de Sinais , Fator A de Crescimento do Endotélio Vascular/metabolismo
8.
Mod Pathol ; 21(3): 308-15, 2008 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-18192970

RESUMO

Gliomas are the most frequent primary tumors of the central nervous system in adults. The most prevalent and aggressive subclass of these is glioblastoma multiforme, which is characterized by massive neovascularization. Endosialin (CD248) has generated interest as a target for antiangiogenic therapy following reports that its expression is upregulated on angiogenic endothelial cells. We demonstrate here that endosialin is not expressed in normal human adult brain but is strongly upregulated in the angiogenic vasculature of all high-grade glioma specimens examined. However, by taking advantage of a technique which allows for multiple fluorescent labeling of formalin-fixed paraffin-embedded archival sections, we demonstrate unambiguously that endosialin is not expressed by the glioma endothelial cells but on closely associated perivascular cells. With increasing awareness that targeting pericytes is an attractive adjunct in antiangiogenic therapy, this finding has important implications for understanding the molecular mechanisms regulating angiogenesis in these highly vascularized tumors.


Assuntos
Antígenos CD/biossíntese , Antígenos de Neoplasias/biossíntese , Neoplasias Encefálicas/irrigação sanguínea , Glioblastoma/irrigação sanguínea , Glioma/irrigação sanguínea , Neovascularização Patológica/patologia , Pericitos/metabolismo , Adulto , Idoso , Anticorpos Monoclonais , Antígenos CD/imunologia , Antígenos de Neoplasias/imunologia , Neoplasias Encefálicas/patologia , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Glioblastoma/patologia , Glioma/patologia , Humanos , Imuno-Histoquímica , Pessoa de Meia-Idade , Neovascularização Patológica/metabolismo , Pericitos/imunologia , Pericitos/patologia
9.
J Biol Chem ; 281(45): 34457-64, 2006 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-16966319

RESUMO

Free fatty acids (FFAs) play important physiological roles in many tissues as an energy source and as signaling molecules in various cellular processes. Elevated levels of circulating FFAs are associated with obesity, dyslipidemia, and diabetes. Here we show that GPR84, a previously orphan G protein-coupled receptor, functions as a receptor for medium-chain FFAs with carbon chain lengths of 9-14. Medium-chain FFAs elicit calcium mobilization, inhibit 3',5'-cyclic AMP production, and stimulate [35S]guanosine 5'-O-(3-thiotriphosphate) binding in a GPR84-dependent manner. The activation of GPR84 by medium-chain FFAs couples primarily to a pertussis toxin-sensitive G(i/o) pathway. In addition, we show that GPR84 is selectively expressed in leukocytes and markedly induced in monocytes/macrophages upon activation by lipopolysaccharide. Furthermore, we demonstrate that medium-chain FFAs amplify lipopolysaccharide-stimulated production of the proinflammatory cytokine interleukin-12 p40 through GPR84. Our results indicate a role for GPR84 in directly linking fatty acid metabolism to immunological regulation.


Assuntos
Ácidos Graxos não Esterificados/metabolismo , Receptores de Superfície Celular/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Equorina/metabolismo , Animais , Medula Óssea , Células CHO , Cálcio/metabolismo , Sinalização do Cálcio , Cricetinae , AMP Cíclico/metabolismo , DNA Complementar/genética , DNA Complementar/metabolismo , Citometria de Fluxo , Imunofluorescência , Guanosina 5'-O-(3-Tiotrifosfato)/metabolismo , Humanos , Sistema Imunitário/citologia , Sistema Imunitário/metabolismo , Subunidade p40 da Interleucina-12/metabolismo , Leucócitos/metabolismo , Ligantes , Lipopolissacarídeos/farmacologia , Macrófagos/metabolismo , Camundongos , Monócitos/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de Superfície Celular/genética , Receptores Acoplados a Proteínas G/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa
10.
J Biol Chem ; 281(31): 22021-22028, 2006 Aug 04.
Artigo em Inglês | MEDLINE | ID: mdl-16754668

RESUMO

Local catabolism of the essential amino acid tryptophan is considered an important mechanism in regulating immunological and neurological responses. The kynurenine pathway is the main route for the non-protein metabolism of tryptophan. The intermediates of the kynurenine pathway are present at micromolar concentrations in blood and are regulated by inflammatory stimuli. Here we show that GPR35, a previously orphan G protein-coupled receptor, functions as a receptor for the kynurenine pathway intermediate kynurenic acid. Kynurenic acid elicits calcium mobilization and inositol phosphate production in a GPR35-dependent manner in the presence of G(qi/o) chimeric G proteins. Kynurenic acid stimulates [35S]guanosine 5'-O-(3-thiotriphosphate) binding in GPR35-expressing cells, an effect abolished by pertussis toxin treatment. Kynurenic acid also induces the internalization of GPR35. Expression analysis indicates that GPR35 is predominantly detected in immune cells and the gastrointestinal tract. Furthermore, we show that kynurenic acid inhibits lipopolysaccharide-induced tumor necrosis factor-alpha secretion in peripheral blood mononuclear cells. Our results suggest unexpected signaling functions for kynurenic acid through GPR35 activation.


Assuntos
Ácido Cinurênico/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Animais , Sinalização do Cálcio , Trato Gastrointestinal/citologia , Trato Gastrointestinal/metabolismo , Perfilação da Expressão Gênica , Humanos , Sistema Imunitário/citologia , Sistema Imunitário/metabolismo , Fosfatos de Inositol/biossíntese , Ácido Cinurênico/sangue , Leucócitos Mononucleares , Ligantes , Lipopolissacarídeos/farmacologia , Camundongos , Ratos , Receptores Acoplados a Proteínas G/genética , Triptofano/metabolismo , Fator de Necrose Tumoral alfa/antagonistas & inibidores
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