RESUMO
BACKGROUND: New precision medicine therapies are urgently required for glioblastoma (GBM). However, to date, efforts to subtype patients based on molecular profiles have failed to direct treatment strategies. We hypothesised that interrogation of the GBM tumour microenvironment (TME) and identification of novel TME-specific subtypes could inform new precision immunotherapy treatment strategies. MATERIALS AND METHODS: A refined and validated microenvironment cell population (MCP) counter method was applied to >800 GBM patient tumours (GBM-MCP-counter). Specifically, partition around medoids (PAM) clustering of GBM-MCP-counter scores in the GLIOTRAIN discovery cohort identified three novel patient clusters, uniquely characterised by TME composition, functional orientation markers and immune checkpoint proteins. Validation was carried out in three independent GBM-RNA-seq datasets. Neoantigen, mutational and gene ontology analysis identified mutations and uniquely altered pathways across subtypes. The longitudinal Glioma Longitudinal AnalySiS (GLASS) cohort and three immunotherapy clinical trial cohorts [treatment with neoadjuvant/adjuvant anti-programmed cell death protein 1 (PD-1) or PSVRIPO] were further interrogated to assess subtype alterations between primary and recurrent tumours and to assess the utility of TME classifiers as immunotherapy biomarkers. RESULTS: TMEHigh tumours (30%) displayed elevated lymphocyte, myeloid cell immune checkpoint, programmed cell death protein 1 (PD-1) and cytotoxic T-lymphocyte-associated protein 4 transcripts. TMEHigh/mesenchymal+ patients featured tertiary lymphoid structures. TMEMed (46%) tumours were enriched for endothelial cell gene expression profiles and displayed heterogeneous immune populations. TMELow (24%) tumours were manifest as an 'immune-desert' group. TME subtype transitions upon recurrence were identified in the longitudinal GLASS cohort. Assessment of GBM immunotherapy trial datasets revealed that TMEHigh patients receiving neoadjuvant anti-PD-1 had significantly increased overall survival (P = 0.04). Moreover, TMEHigh patients treated with adjuvant anti-PD-1 or oncolytic virus (PVSRIPO) showed a trend towards improved survival. CONCLUSIONS: We have established a novel TME-based classification system for application in intracranial malignancies. TME subtypes represent canonical 'termini a quo' (starting points) to support an improved precision immunotherapy treatment approach.
Assuntos
Neoplasias Encefálicas , Glioblastoma , Humanos , Glioblastoma/tratamento farmacológico , Microambiente Tumoral , Recidiva Local de Neoplasia , Imunoterapia/métodos , Neoplasias Encefálicas/tratamento farmacológicoRESUMO
BACKGROUND: The need to unfold the underlying mechanisms of lung cancer aggressiveness, the deadliest cancer in the world, is of prime importance. Because Fas-associated death domain protein (FADD) is the key adaptor molecule transmitting the apoptotic signal delivered by death receptors, we studied the presence and correlation of intra- and extracellular FADD protein with development and aggressiveness of non-small cell lung cancer (NSCLC). METHODS: Fifty NSCLC patients were enrolled in this prospective study. Intracellular FADD was detected in patients' tissue by immunohistochemistry. Tumours and distant non-tumoural lung biopsies were cultured through trans-well membrane in order to analyse extracellular FADD. Correlation between different clinical/histological parameters with level/localisation of FADD protein has been investigated. RESULTS: Fas-associated death domain protein could be specifically downregulated in tumoural cells and FADD loss correlated with the presence of extracellular FADD. Indeed, human NSCLC released FADD protein, and tumoural samples released significantly more FADD than non-tumoural (NT) tissue (P=0.000003). The release of FADD by both tumoural and NT tissue increased significantly with the cancer stage, and was correlated with both early and late steps of the metastasis process. CONCLUSION: The release of FADD by human NSCLC could be a new marker of poor prognosis as it correlates positively with both tumour progression and aggressiveness.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Proteína de Domínio de Morte Associada a Fas/metabolismo , Neoplasias Pulmonares/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/análise , Carcinoma Pulmonar de Células não Pequenas/patologia , Progressão da Doença , Espaço Extracelular/metabolismo , Feminino , Humanos , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , Metástase Neoplásica , Prognóstico , Estudos ProspectivosRESUMO
Immune checkpoint inhibitors (ICIs) show limited clinical activity in patients with advanced soft-tissue sarcomas (STSs). Retrospective analysis suggests that intratumoral tertiary lymphoid structures (TLSs) are associated with improved outcome in these patients. PEMBROSARC is a multicohort phase 2 study of pembrolizumab combined with low-dose cyclophosphamide in patients with advanced STS (NCT02406781). The primary endpoint was the 6-month non-progression rate (NPR). Secondary endpoints included objective response rate (ORR), progression-free survival (PFS), overall survival (OS) and safety. The 6-month NPR and ORRs for cohorts in this trial enrolling all comers were previously reported; here, we report the results of a cohort enrolling patients selected based on the presence of TLSs (n = 30). The 6-month NPR was 40% (95% confidence interval (CI), 22.7-59.4), so the primary endpoint was met. The ORR was 30% (95% CI, 14.7-49.4). In comparison, the 6-month NPR and ORR were 4.9% (95% CI, 0.6-16.5) and 2.4% (95% CI, 0.1-12.9), respectively, in the all-comer cohorts. The most frequent toxicities were grade 1 or 2 fatigue, nausea, dysthyroidism, diarrhea and anemia. Exploratory analyses revealed that the abundance of intratumoral plasma cells (PCs) was significantly associated with improved outcome. These results suggest that TLS presence in advanced STS is a potential predictive biomarker to improve patients' selection for pembrolizumab treatment.
Assuntos
Sarcoma , Neoplasias de Tecidos Moles , Estruturas Linfoides Terciárias , Anticorpos Monoclonais Humanizados , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Humanos , Estudos Retrospectivos , Sarcoma/tratamento farmacológico , Sarcoma/etiologia , Neoplasias de Tecidos Moles/tratamento farmacológico , Neoplasias de Tecidos Moles/etiologia , Estruturas Linfoides Terciárias/etiologiaRESUMO
A large body of evidence indicates that the immune microenvironment controls tumour development. Primary central nervous system lymphomas (PCNSL) are aggressive tumours growing in the central nervous system (CNS). To evaluate the role and characteristics of this immune-privileged site in anti-tumour defences, we compared the cellular and molecular immune microenvironments of growing murine lymphoma B cells injected into the brain or the spleen. In the brain, immune cells, including dendritic cells and T lymphocytes with a large proportion of CD4(+) forkhead box P3 (FoxP3(+)) regulatory T cells, rapidly infiltrated the tumour microenvironment. These populations also increased in number in the spleen. The T cell cytokine profiles in tumour-bearing mice were similar in the two sites, with predominant T helper type 1 (Th1)/Th17 polarization after polyclonal stimulation, although some interleukin (IL)-4 could also be found. We demonstrated that these T cells have anti-tumour activity in the CNS, although less than in the spleen: nude mice that received lymphoma cells intracerebrally died significantly earlier than immunocompetent animals. These results demonstrate that the brain is able to recruit all the major actors to mount a specific anti-tumour immune response against lymphoma.
Assuntos
Neoplasias Encefálicas/imunologia , Linfoma de Células B/imunologia , Neoplasias Esplênicas/imunologia , Microambiente Tumoral/imunologia , Animais , Células Apresentadoras de Antígenos/patologia , Neoplasias Encefálicas/patologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD4-Positivos/patologia , Linfócitos T CD8-Positivos/patologia , Linhagem Celular Tumoral , Movimento Celular/imunologia , Proliferação de Células , Citocinas/metabolismo , Células Dendríticas/patologia , Feminino , Ativação Linfocitária/imunologia , Linfoma de Células B/patologia , Camundongos , Camundongos Nus , Neoplasias Esplênicas/patologia , Análise de Sobrevida , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Subpopulações de Linfócitos T/patologia , Linfócitos T Reguladores/patologia , Células Th1/imunologia , Células Th1/metabolismo , Células Th17/imunologia , Células Th17/metabolismo , Células Th2/imunologia , Células Th2/metabolismoRESUMO
Using a snake toxin as a proteic antigen (Ag), two murine toxin-specific monoclonal antibodies (mAbs), splenocytes, and two murine Ag-specific T cell hybridomas, we showed that soluble protein A (SpA) from Staphylococcus aureus and protein G from Streptococcus subspecies, two Ig binding proteins (IBPs), not only abolish the capacity of the mAbs to decrease Ag presentation but also increase Ag presentation 20-100-fold. Five lines of evidence suggest that this phenomenon results from binding of an IBP-Ab-Ag complex to B cells possessing IBP receptors. First, we showed that SpA is likely to boost presentation of a free mAb, suggesting that the IBP-boosted presentation of an Ag in an immune complex results from the binding of IBP to the mAb. Second, FACS analyses showed that an Ag-Ab complex is preferentially targeted by SpA to a subpopulation of splenocytes mainly composed of B cells. Third, SpA-dependent boosted presentation of an Ag-Ab complex is further enhanced when splenocytes are enriched in cells containing SpA receptors. Fourth, the boosting effect largely diminishes when splenocytes are depleted of cells containing SpA receptors. Fifth, the boosting effect occurs only when IBP simultaneously contains a Fab and an Fc binding site. Altogether, our data suggest that soluble IBPs can bridge immune complexes to APCs containing IBP receptors, raising the possibility that during an infection process by bacteria secreting these IBPs, Ag-specific T cells may activate IBP receptor-containing B cells by a mechanism of intermolecular help, thus leading to a nonspecific immune response.
Assuntos
Apresentação de Antígeno/imunologia , Complexo Antígeno-Anticorpo/imunologia , Linfócitos B/imunologia , Proteínas de Bactérias/imunologia , Linfocinas/imunologia , Proteínas Secretadas pela Próstata , Animais , Anticorpos Monoclonais/imunologia , Sítios de Ligação , Citometria de Fluxo , Hibridomas/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Ligação Proteica , Receptores de Superfície Celular/imunologia , Baço/imunologia , Proteína Estafilocócica A/imunologia , Linfócitos T/imunologia , Fosfolipases Tipo C/imunologiaRESUMO
BACKGROUND: Deletion of the complement factor H related 1 (CFHR1) gene is a consequence of non-allelic homologous recombination and has been reported to be more frequent in atypical haemolytic uraemic syndrome (aHUS) patients than in the normal population. Therefore, it is considered a susceptibility factor for the disease. aHUS is associated with hereditary or acquired abnormalities that lead to uncontrolled alternative pathway complement activation. We tested the CFHR1 deletion for association with aHUS in a population of French aHUS cases and controls. Furthermore, we examined the effect of the deletion in the context of known aHUS risk factors. METHODS AND RESULTS: 177 aHUS patients and 70 healthy donors were studied. The number of CFHR1 alleles was quantified by multiplex ligation dependant probe amplification (MLPA). The frequency of the deleted allele was significantly higher in aHUS patients than in controls (22.7% vs 8.2%, p<0.001). The highest frequency was in the subgroup of patients exhibiting anti-factor H (FH) autoantibodies (92.9%, p<0.0001 vs controls) and in the group of patients exhibiting a factor I (CFI) gene mutation (31.8%, p<0.001 vs controls). The CFHR1 deletion was not significantly more frequent in the cohort of aHUS patients when patients with anti-FH IgG or CFI mutation were excluded. CONCLUSIONS: The high frequency of CFHR1 deletion in aHUS patients is restricted to the subgroups of patients presenting with anti-FH autoantibodies or, to a lesser degree, CFI mutation. These results suggest that the CFHR1 deletion plays a secondary role in susceptibility to aHUS.
Assuntos
Proteínas Inativadoras do Complemento C3b/genética , Deleção de Genes , Síndrome Hemolítico-Urêmica/genética , Adulto , Autoanticorpos , Distribuição de Qui-Quadrado , Criança , Estudos de Coortes , Fator H do Complemento/imunologia , Dosagem de Genes , Frequência do Gene , Predisposição Genética para Doença , Humanos , Mutação , Técnicas de Amplificação de Ácido Nucleico/métodosRESUMO
PURPOSE: There are some lines of evidence suggesting a potential role of immunotherapy for treating patients with osteosarcomas. PATIENTS AND METHODS: This was an open-label, multicentre, phase 2 study of pembrolizumab in combination with metronomic cyclophosphamide in patients with advanced osteosarcomas. All patients received 50 mg b.i.d. of cyclophosphamide one week on and one week off and 200 mg of intravenous pembrolizumab (every 3 weeks). There was a dual primary end-point, encompassing both the non-progression and objective responses at 6 months per Response Evaluation Criteria in Solid Tumours (RECIST), version 1.1. An objective response rate of 20% and/or a 6-month non-progression rate of 60% were determined as reasonable objectives for treatment with meaningful effect. Correlative studies of immune biomarkers were planned from the patients' tumour samples. RESULTS: Between October 13 2015 and July 3 2017, 17 patients were included. Fifty were assessable for the efficacy end-point. Four patients experienced tumour shrinkage, resulting in a partial response (PR) in one patient (6.7%). The 6-month non-progression rate was 13.3% (95% confidence interval [CI]: 1.7-40.5). The most frequent adverse events were grade I or II nausea, anaemia, anorexia and fatigue. programmed death-ligand 1 (PD-L1) expression rate was low, observed in only 2 cases of 14 with available tumour material. The only patient who experienced PR had a PD-L1-negative tumour. CONCLUSION: Programmed cell death 1 (PD-1) inhibition has limited activity in osteosarcomas. Further studies investigating PD-1 inhibitor in combination with agents modulating the microenvironment are warranted. TRIAL REGISTRATION: This study is registered with ClinicalTrials.gov, number NCT02406781.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Osteossarcoma/tratamento farmacológico , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Microambiente Tumoral/efeitos dos fármacos , Administração Metronômica , Adulto , Idoso , Idoso de 80 Anos ou mais , Anticorpos Monoclonais Humanizados/administração & dosagem , Anticorpos Monoclonais Humanizados/efeitos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Ciclofosfamida/administração & dosagem , Ciclofosfamida/efeitos adversos , Esquema de Medicação , Feminino , Humanos , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Náusea/induzido quimicamente , Osteossarcoma/metabolismo , Osteossarcoma/patologia , Receptor de Morte Celular Programada 1/metabolismo , Critérios de Avaliação de Resposta em Tumores Sólidos , Adulto JovemRESUMO
Tumor necrosis factor receptor (TNFR) associated factor 4 (TRAF4) was initially identified as a gene amplified and overexpressed in breast carcinomas. Our aim was to evaluate whether TRAF4 protein overexpression exists in other cancer types. Immunohistochemistry analysis of tumor samples from 623 patients with 20 different tumor types showed that TRAF4 was overexpressed in 268 tumors (43%), including 82 of 137 lung adenocarcinomas (60%). Interestingly, 32 primary tumors and their matching metastases exhibited mostly similar TRAF4 expression pattern. TRAF4 protein overexpression was limited to cancer cells and the subcellular localization was consistently cytoplasmic in a large majority of cases. To investigate changes in TRAF4 gene copy number, 125 cases from six different types of carcinomas were also analysed by fluorescence in situ hybridization. Out of the 28 cases (22%) showing an increased TRAF4 gene copy number, 23 (82%) were overexpressing the protein. Thus, TRAF4 gene amplification is one of the mechanisms responsible for TRAF4 protein overexpression in human cancers. Considering that TRAF4 is located at 17q11.2 in a region of amplification devoid of known oncogenes and is commonly overexpressed in cancer, our data support an oncogenic role for TRAF4.
Assuntos
Neoplasias/genética , Fator 4 Associado a Receptor de TNF/genética , Humanos , Imuno-Histoquímica , Hibridização in Situ Fluorescente , Neoplasias/classificação , Fator 4 Associado a Receptor de TNF/metabolismoRESUMO
Matrix metalloproteinases (MMPs) are extracellular enzymes. Some of them are known to be involved in tumor development and/or progression. Several cellular functions have been proposed for MMPs during malignant processes. Notably, they may be involved in tissue-remodeling processes through their ability to digest matrix components or to participate in tumor neoangiogenesis and, subsequently, in cancer cell proliferation. One of these MMPs, stromelysin-3 (ST3/MMP11), although devoid of enzymatic activity against the matrix components, is associated with human tumor progression and poor patient clinical outcome. Using several in vivo experimental models, it has been demonstrated that ST3 expression by the fibroblastic cells surrounding malignant epithelial cells promotes tumorigenesis in a paracrine manner. The present study was devoted to the identification of the cellular function underlying this ST3-induced tumor promotion using a syngeneic tumorigenesis model in mice. Our results show that ST3 exhibits a new and unexpected role for a MMP, because ST3-increased tumorigenesis does not result from increased neoangiogenesis or cancer cell proliferation but from decreased cancer cell death through apoptosis and necrosis. Thus, during malignancy, the cellular function of ST3 is to favor cancer cell survival in the stromal environment.
Assuntos
Apoptose/fisiologia , Neoplasias do Colo/enzimologia , Metaloendopeptidases/deficiência , Animais , Divisão Celular/fisiologia , Neoplasias do Colo/irrigação sanguínea , Neoplasias do Colo/patologia , Endogamia , Macrófagos/imunologia , Macrófagos/patologia , Metaloproteinase 11 da Matriz , Metaloendopeptidases/genética , Camundongos , Camundongos Endogâmicos BALB C , Neovascularização Patológica/enzimologia , Neutrófilos/imunologia , Neutrófilos/patologiaRESUMO
Interleukin (IL) 17 is a proinflammatory cytokine secreted mainly by activated human memory CD4 T cells that induces IL-6, IL-8, and nitric oxide. Because IL-6 and IL-8 have been implicated in the pathogenesis of cervical cancer, we investigated the action of IL-17 on human cervical tumor cell lines in vitro and in vivo. We showed that in vitro, IL-17 increases IL-6 and IL-8 secretion by cervical carcinoma cell lines at both protein and mRNA levels. No direct effect of IL-17 on in vitro proliferation of cervical tumor cell lines could be demonstrated. However, two cervical cell lines transfected with a cDNA encoding IL-17 exhibited a significant increase in tumor size as compared to the parent tumor when transplanted in nude mice. This enhanced tumor growth elicited by IL-17 was associated with increased expression of IL-6 and macrophage recruitment at the tumor site. A potential role of IL-17 in modulation of the human cervical tumor phenotype was also supported by its expression on the cervical tumor in patients with CD4 infiltration. IL-17 therefore behaves like a T-cell-specific cytokine with paradoxical tumor-promoting activity. This may partially explain previous reports concerning the deleterious effect of CD4 T cells in cancer.
Assuntos
Carcinógenos/toxicidade , Interleucina-17/toxicidade , Linfócitos T/metabolismo , Neoplasias do Colo do Útero/patologia , Animais , Linfócitos T CD4-Positivos/patologia , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Células HeLa/efeitos dos fármacos , Humanos , Interleucina-17/genética , Interleucina-6/biossíntese , Interleucina-6/genética , Interleucina-6/metabolismo , Interleucina-8/biossíntese , Interleucina-8/genética , Interleucina-8/metabolismo , Masculino , Melanoma/patologia , Camundongos , Camundongos Nus , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Transplante de Neoplasias , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/toxicidade , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transfecção , Células Tumorais Cultivadas/efeitos dos fármacos , Células Tumorais Cultivadas/metabolismo , Células Tumorais Cultivadas/transplante , Neoplasias do Colo do Útero/metabolismoRESUMO
The binding of multivalent antigen-antibody complexes to receptors for the Fc portion of IgG (FcgammaR) induces the clustering of the FcgammaR and triggers cell activation leading to defence reactions against pathogens. The Fc portion of IgG consists of two identical polypeptide chains which are related to each other by a 2-fold axis and are folded in two structural domains, the C(H)2 domain, near the flexible hinge region of the IgG molecule, and the C(H)3 domain. We studied the interaction in solution between the Fc fragment of mouse IgG2b and the extracellular region of mouse FcgammaRII. We find that one Fc molecule binds one FcgammaRII molecule only. Using NMR spectroscopy, we show that FcgammaRII binds to a negatively charged area of the C(H)2 domain, corresponding to the lower hinge region, and that the binding of FcgammaRII onto one of the two symmetrically related sites on the Fc induces a conformational change in the other site. We therefore propose a model that explains why IgG molecules are unable to trigger FcgammaR-mediated cellular responses spontaneously in the absence of crosslinking by multivalent antigens.
Assuntos
Receptores de IgG/química , Receptores de IgG/metabolismo , Animais , Espectroscopia de Ressonância Magnética , Camundongos , Modelos Moleculares , Ligação Proteica , Conformação Proteica , Dobramento de Proteína , Receptores de IgG/classificação , Eletricidade Estática , Relação Estrutura-AtividadeRESUMO
CD34(+)-derived dendritic cells (DCs) can be infected by the T cell-tropic HIVLAI strain, but are poorly permissive for efficient virus production. However, HIVLAI-infected DCs are able to transmit a vigorous cytopathic infection to activated CD4(+) T cells. We show that DCs differentiated from CD34(+) cells can be efficiently transduced by a retroviral vector carrying the IFN-beta coding sequence. This results in resistance to infection by HIV as shown by a threefold reduction in the HIV DNA copy number per cell, and by inhibition of HIV transmission from DCs to CD4(+) T cells. Moreover, constitutive IFN-beta production by DCs increases the synthesis of IL-12 and IFN-gamma Th1-type cytokines and of the beta-chemokines MIP-1alpha, MIP-1beta, and RANTES. This indicates that IFN-beta transduction of DCs blocks HIV infection and viral transmission to CD4(+) T cells, and could favor cellular immune responses in HIV-infected patients.
Assuntos
Linfócitos T CD4-Positivos/virologia , Células Dendríticas/virologia , Técnicas de Transferência de Genes , Infecções por HIV/transmissão , HIV/patogenicidade , Interferon beta/genética , Antígenos CD34/metabolismo , Células Cultivadas , Quimiocina CCL3 , Quimiocina CCL4 , Quimiocina CCL5/biossíntese , Efeito Citopatogênico Viral , Células Dendríticas/metabolismo , Ensaio de Imunoadsorção Enzimática , Feminino , Sangue Fetal/citologia , Humanos , Interferon beta/metabolismo , Interferon gama/biossíntese , Interleucina-2/biossíntese , Proteínas Inflamatórias de Macrófagos/biossíntese , Reação em Cadeia da PolimeraseRESUMO
The class IIa of low-affinity receptors for the Fc region of IgG, Fc gamma RIIa, are expressed on immune cells. The cross-linking of Fc gamma RIIa by complexed IgG triggers activation of protein tyrosine kinase and internalization of immune complexes. In this report, we demonstrate the expression of Fc gamma RIIa by a human melanoma cell line (VIO) derived from a metastasis of a patient with regressive melanoma. The analysis of Fc gamma RIIa functions was performed in VIO cells and Fc gamma RlIa- or Fc gamma RIlb-transfected human melanoma cells (A375). The Fc gamma RIIa cross-linking induced protein tyrosine phosphorylation, including Fc gamma RIIa phosphorylation, and led to its internalization in a clathrin-independent way in human melanoma cells. Moreover, we showed that a part of internalized Fc gamma RIIa migrates in late endosomes, lysosomes and class II-containing compartments. These results suggest that melanoma cells can express functional Fc gamma RII, which might play a role in tumor-host relationships.
Assuntos
Melanoma/imunologia , Melanoma/secundário , Receptores de IgG/metabolismo , Western Blotting , Reagentes de Ligações Cruzadas/metabolismo , Imunofluorescência , Humanos , Fosforilação , Testes de Precipitina , Proteínas Tirosina Quinases/metabolismo , Receptores de IgG/genética , Células Tumorais CultivadasRESUMO
CD16 (Fc gamma R type III), a low affinity IgG Fc receptor, is found in two forms, a transmembrane Fc gamma RIIIa expressed by NK cells and monocytes and a phosphatidylinositol-linked Fc gamma RIIIb present on neutrophils. Exposure of neutrophils to inflammatory signals induces a rapid loss of CD16 expression and release of a soluble form of CD16 (sCD16). Soluble CD16 circulates in plasma, levels being reduced in sera from patients with multiple myeloma. In the present manuscript the authors summarize work that aimed to better understand: (i) the role of proteinases in sCD16 production and CD16 membrane shedding; and (ii) the regulation of sCD16 levels in multiple myeloma patients and the possible biological consequences of its decrease in this disease. Soluble CD16 was purified from human serum. Its N-terminal sequencing demonstrated that it originates from neutrophil CD16 and its C-terminal sequencing showed that the cleavage site was between Val 196 and Ser 197, close to the membrane anchor. Analysis of the effect of protease inhibitors revealed that the cleavage leading to sCD16 production by PMA-activated neutrophils was metalloproteinase-dependent. In addition, membrane and sCD16 were sensitive to serine proteinases released by azurophil granules or added under purified form. The reduction of sCD16 levels that occurs in patients with multiple myeloma was associated with a slight decrease in circulating neutrophils, but not with a significant defect in sCD16 production by neutrophils, as detected in vitro. Moreover, addition of a recombinant sCD16 to plasmocytoma lines did not significantly modify their proliferation and Ig secretion.
Assuntos
Receptores de IgG/biossíntese , Receptores de IgG/metabolismo , Sequência de Aminoácidos , Humanos , Dados de Sequência Molecular , Receptores de IgG/fisiologia , SolubilidadeRESUMO
The local immune reactions may influence the clinical outcome of human tumors. In carcinoma of the cervix, high gene expression of IL6 with tumor invasiveness whereas lack of gene expression of IFNbeta is correlated with poor prognosis. In colorectal cancer, lack of expression of IFNbeta is associated with the presence of distant metastasis and poor survival. The production of IL17 and IL18, inducers of IL6 and IFNbeta respectively is regulated in these tumors and may control the levels of the effector cytokines, i.e. IL6 and IFNbeta. The mechanisms by which these cytokines act are linked to the recruitment of effector cells such as macrophages.
Assuntos
Transformação Celular Neoplásica/imunologia , Citocinas/fisiologia , Neoplasias/imunologia , Animais , Transformação Celular Neoplásica/patologia , Humanos , Neoplasias/etiologia , Neoplasias/patologia , Neoplasias Experimentais/etiologia , Neoplasias Experimentais/imunologia , Neoplasias Experimentais/patologiaAssuntos
Antígenos CD/metabolismo , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Linfoma de Células B/tratamento farmacológico , Linfoma Difuso de Grandes Células B/tratamento farmacológico , Receptores de IgG/metabolismo , Idoso , Idoso de 80 Anos ou mais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais Murinos , Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Ciclofosfamida/administração & dosagem , Doxorrubicina/administração & dosagem , Humanos , Prednisona/administração & dosagem , Prognóstico , Rituximab , Taxa de Sobrevida , Vincristina/administração & dosagemRESUMO
The natural history of a tumor includes phases of 'in situ' growth, invasion, extravasation and metastasis. During these phases, tumor cells interact with their microenvironment and are influenced by signals coming from stromal, endothelial, inflammatory and immune cells. Indeed, tumors are often infiltrated by various numbers of lymphocytes, macrophages or mast cells. It is generally believed that the latter produce factors that maintain chronic inflammation and promote tumor growth, whereas lymphocytes may control cancer outcome, as evidenced in mouse models. In this study, we analyze data from large cohorts of human tumors, clearly establishing that infiltration of the primary tumor by memory T cells, particularly of the Th1 and cytotoxic types, is the strongest prognostic factor in terms of freedom from disease and overall survival at all stages of clinical disease. We review data suggesting that tertiary lymphoid structures adjacent to tumors and composed of mature dendritic cells (T and B cells organized as germinal centers) may be the site of an antitumor reaction. We propose an immune scoring based on the type, density and location of lymphocyte infiltrates as a novel prognostic factor for use in addition to tumor node metastasis staging to predict disease-free survival and to aid in decisions regarding adjuvant therapies in early stage human cancers.
Assuntos
Neoplasias Colorretais/diagnóstico , Metástase Linfática/diagnóstico , Estadiamento de Neoplasias , Prognóstico , Animais , Neoplasias da Mama/diagnóstico , Neoplasias Colorretais/fisiopatologia , HumanosRESUMO
Measles virus (MV) nucleoprotein (N) is a cytosolic protein that is released into the extracellular compartment after apoptosis and/or secondary necrosis of MV-infected cells in vitro. Thus, MV-N becomes accessible to inhibitory cell-surface receptors: FcgammaRIIB and an uncharacterized nucleoprotein receptor (NR). MV-N is composed of two domains: NCORE (aa 1-400) and NTAIL (aa 401-525). To assess the contribution of MV-N domains and of these two receptors in suppression of cell proliferation, a human melanoma HT144 cell line expressing (HT144IIB1) or lacking FcgammaRIIB1 was used as a model. Specific and exclusive NCORE-FcgammaRIIB1 and NTAIL-NR interactions were shown. Moreover, NTAIL binding to human NR predominantly led to suppression of cell proliferation by arresting cells in the G0/G1 phases of the cell cycle, rather than to apoptosis. NCORE binding to HT144IIB1 cells primarily triggered caspase-3 activation, in contrast to HT144IIB1/IC- cells lacking the FcgammaRIIB1 intra-cytoplasmic tail, thus demonstrating the specific inhibitory effect of the NCORE-FcgammaRIIB1 interaction. MV-N- and NCORE-mediated apoptosis through FcgammaRIIB1 was inhibited by the pan-caspase inhibitor zVAD-FMK, indicating that apoptosis was dependent on caspase activation. By using NTAIL deletion proteins, it was also shown that the region of NTAIL responsible for binding to human NR and for cell growth arrest maps to one of the three conserved boxes (Box1, aa 401-420) found in N of Morbilliviruses. This work unveils novel mechanisms by which distinct domains of MV-N may display different immunosuppressive activities, thus contributing to our comprehension of the immunosuppressive state associated with MV infection. Finally, MV-N domains may be good tools to target tumour cell proliferation and/or apoptosis.
Assuntos
Antígenos CD/metabolismo , Vírus do Sarampo/fisiologia , Nucleoproteínas/metabolismo , Receptores de Superfície Celular/metabolismo , Receptores de IgG/metabolismo , Receptores Virais/metabolismo , Proteínas Virais/metabolismo , Replicação Viral , Animais , Apoptose , Ciclo Celular , Linhagem Celular Tumoral , Proliferação de Células , Deleção de Genes , Humanos , Vírus do Sarampo/metabolismo , Proteínas do Nucleocapsídeo , Nucleoproteínas/genética , Estrutura Terciária de Proteína/genética , Proteínas Virais/genéticaRESUMO
Fcgamma receptors mediate antibody-dependent inflammatory responses and cytotoxicity as well as certain autoimmune dysfunctions. Here we report the crystal structure of a human Fc receptor (FcgammaRIIIB) in complex with an Fc fragment of human IgG1 determined from orthorhombic and hexagonal crystal forms at 3.0- and 3.5-A resolution, respectively. The refined structures from the two crystal forms are nearly identical with no significant discrepancies between the coordinates. Regions of the C-terminal domain of FcgammaRIII, including the BC, C'E, FG loops, and the C' beta-strand, bind asymmetrically to the lower hinge region, residues Leu(234)-Pro(238), of both Fc chains creating a 1:1 receptor-ligand stoichiometry. Minor conformational changes are observed in both the receptor and Fc upon complex formation. Hydrophobic residues, hydrogen bonds, and salt bridges are distributed throughout the receptor.Fc interface. Sequence comparisons of the receptor-ligand interface residues suggest a conserved binding mode common to all members of immunoglobulin-like Fc receptors. Structural comparison between FcgammaRIII.Fc and FcepsilonRI.Fc complexes highlights the differences in ligand recognition between the high and low affinity receptors. Although not in direct contact with the receptor, the carbohydrate attached to the conserved glycosylation residue Asn(297) on Fc may stabilize the conformation of the receptor-binding epitope on Fc. An antibody-FcgammaRIII model suggests two possible ligand-induced receptor aggregations.
Assuntos
Antígenos CD/química , Fragmentos Fc das Imunoglobulinas/química , Receptores de IgG/química , Sequência de Aminoácidos , Antígenos CD/imunologia , Sítios de Ligação , Sítios de Ligação de Anticorpos , Membrana Celular/imunologia , Cristalografia por Raios X , Proteínas Ligadas por GPI , Glicosilação , Humanos , Fragmentos Fc das Imunoglobulinas/imunologia , Modelos Moleculares , Dados de Sequência Molecular , Fragmentos de Peptídeos/química , Estrutura Secundária de Proteína , Receptores de IgG/imunologia , Proteínas Recombinantes/química , Alinhamento de Sequência , Homologia de Sequência de AminoácidosRESUMO
We demonstrate that soluble CD16 (sCD16; soluble Fc gamma RIII), a natural ligand of CR3, inhibits the infection of monocytes by primary R5 HIV-1 strain opsonized with serum of seronegative individuals. Inhibition of monocyte infection by sCD16 was similar to that observed with anti-CR3 mAbs, indicating that opsonized HIV may use a CR3-dependent pathway for entry in monocytic cells. Cultured human monocytes express both CR3 (CD11b/CD18) and CCR5 receptors. RANTES, the natural ligand of CCR5, inhibited infection of monocytes with unopsonized HIV particles and partially that of monocytes infected with HIV particles opsonized with complement-derived fragments. Although HIV-infected monocytes from homozygous CCR5 Delta 32/Delta 32 (CCR5(-/-)) individuals produce low levels of p24, cells infected with opsonized particles produced higher levels of p24 than cells infected with unopsonized particles. Our results thus suggest that CR3 may represent an alternative coreceptor to CCR5 of opsonized primary R5 virus entry into monocytes/macrophages. We also observed that the concentration of sCD16 is greatly decreased in sera of HIV-infected patients with low lymphocyte CD4(+) counts. Taken together, our findings suggest that sCD16, present in plasma, may play an important role in controlling HIV-1 spread.