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1.
Nat Commun ; 11(1): 5332, 2020 10 21.
Artigo em Inglês | MEDLINE | ID: mdl-33087697

RESUMO

Cytotoxic T lymphocyte (CTL)-based cancer immunotherapies have shown great promise for inducing clinical regressions by targeting tumor-associated antigens (TAA). To expand the TAA landscape of pancreatic ductal adenocarcinoma (PDAC), we performed tandem mass spectrometry analysis of HLA class I-bound peptides from 35 PDAC patient tumors. This identified a shared HLA-A*0101 restricted peptide derived from co-transcriptional activator Vestigial-like 1 (VGLL1) as a putative TAA demonstrating overexpression in multiple tumor types and low or absent expression in essential normal tissues. Here we show that VGLL1-specific CTLs expanded from the blood of a PDAC patient could recognize and kill in an antigen-specific manner a majority of HLA-A*0101 allogeneic tumor cell lines derived not only from PDAC, but also bladder, ovarian, gastric, lung, and basal-like breast cancers. Gene expression profiling reveals VGLL1 as a member of a unique group of cancer-placenta antigens (CPA) that may constitute immunotherapeutic targets for patients with multiple cancer types.


Assuntos
Antígenos de Neoplasias/imunologia , Neoplasias da Mama/imunologia , Proteínas de Ligação a DNA/imunologia , Neoplasias Pancreáticas/imunologia , Fatores de Transcrição/imunologia , Antígenos de Neoplasias/genética , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/imunologia , Neoplasias da Mama/genética , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/imunologia , Carcinoma Ductal Pancreático/terapia , Linhagem Celular Tumoral , Citotoxicidade Imunológica , Proteínas de Ligação a DNA/genética , Feminino , Perfilação da Expressão Gênica , Antígeno HLA-A1/imunologia , Humanos , Imunoterapia Adotiva , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/terapia , Placenta/imunologia , Gravidez , Prognóstico , Linfócitos T Citotóxicos/imunologia , Fatores de Transcrição/genética
2.
Proc Natl Acad Sci U S A ; 117(38): 23695-23706, 2020 09 22.
Artigo em Inglês | MEDLINE | ID: mdl-32907941

RESUMO

Long noncoding RNAs (lncRNAs) involved in the regulation of antiviral innate immune responses need to be further identified. By functionally screening the lncRNAs in macrophages, here we identified lncRNA Malat1, abundant in the nucleus but significantly down-regulated after viral infection, as a negative regulator of antiviral type I IFN (IFN-I) production. Malat1 directly bound to the transactive response DNA-binding protein (TDP43) in the nucleus and prevented activation of TDP43 by blocking the activated caspase-3-mediated TDP43 cleavage to TDP35. The cleaved TDP35 increased the nuclear IRF3 protein level by binding and degrading Rbck1 pre-mRNA to prevent IRF3 proteasomal degradation upon viral infection, thus selectively promoting antiviral IFN-I production. Deficiency of Malat1 enhanced antiviral innate responses in vivo, accompanying the increased IFN-I production and reduced viral burden. Importantly, the reduced MALAT1, augmented IRF3, and increased IFNA mRNA were found in peripheral blood mononuclear cells (PBMCs) from systemic lupus erythematosus (SLE) patients. Therefore, the down-regulation of MALAT1 in virus-infected cells or in human cells from autoimmune diseases will increase host resistance against viral infection or lead to autoinflammatory interferonopathies via the increased type I IFN production. Our results demonstrate that the nuclear Malat1 suppresses antiviral innate responses by targeting TDP43 activation via RNA-RBP interactive network, adding insight to the molecular regulation of innate responses and autoimmune pathogenesis.


Assuntos
Proteínas de Ligação a DNA , Imunidade Inata/imunologia , Fator Regulador 3 de Interferon , RNA Longo não Codificante , Adolescente , Adulto , Animais , Antivirais/imunologia , Antivirais/metabolismo , Células Cultivadas , Proteínas de Ligação a DNA/imunologia , Proteínas de Ligação a DNA/metabolismo , Feminino , Humanos , Fator Regulador 3 de Interferon/imunologia , Fator Regulador 3 de Interferon/metabolismo , Interferon Tipo I/imunologia , Interferon Tipo I/metabolismo , Leucócitos Mononucleares/imunologia , Lúpus Eritematoso Sistêmico/imunologia , Macrófagos Peritoneais/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , RNA Longo não Codificante/imunologia , RNA Longo não Codificante/metabolismo , Viroses/imunologia , Adulto Jovem
3.
Nat Commun ; 11(1): 4225, 2020 08 24.
Artigo em Inglês | MEDLINE | ID: mdl-32839463

RESUMO

Gallbladder cancer (GBC) is an aggressive gastrointestinal malignancy with no approved targeted therapy. Here, we analyze exomes (n = 160), transcriptomes (n = 115), and low pass whole genomes (n = 146) from 167 gallbladder cancers (GBCs) from patients in Korea, India and Chile. In addition, we also sequence samples from 39 GBC high-risk patients and detect evidence of early cancer-related genomic lesions. Among the several significantly mutated genes not previously linked to GBC are ETS domain genes ELF3 and EHF, CTNNB1, APC, NSD1, KAT8, STK11 and NFE2L2. A majority of ELF3 alterations are frame-shift mutations that result in several cancer-specific neoantigens that activate T-cells indicating that they are cancer vaccine candidates. In addition, we identify recurrent alterations in KEAP1/NFE2L2 and WNT pathway in GBC. Taken together, these define multiple targetable therapeutic interventions opportunities for GBC treatment and management.


Assuntos
Proteínas de Ligação a DNA/genética , Mutação da Fase de Leitura , Neoplasias da Vesícula Biliar/genética , Predisposição Genética para Doença/genética , Proteínas Proto-Oncogênicas c-ets/genética , Fatores de Transcrição/genética , Vacinas Anticâncer/genética , Vacinas Anticâncer/imunologia , Chile , Proteínas de Ligação a DNA/imunologia , Proteínas de Ligação a DNA/metabolismo , Neoplasias da Vesícula Biliar/diagnóstico , Neoplasias da Vesícula Biliar/metabolismo , Perfilação da Expressão Gênica/métodos , Regulação Neoplásica da Expressão Gênica , Genômica/métodos , Humanos , Índia , Proteína 1 Associada a ECH Semelhante a Kelch/genética , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Proteínas Proto-Oncogênicas c-ets/imunologia , Proteínas Proto-Oncogênicas c-ets/metabolismo , República da Coreia , Fatores de Transcrição/imunologia , Fatores de Transcrição/metabolismo , Via de Sinalização Wnt/genética , beta Catenina/genética , beta Catenina/metabolismo
4.
BMC Med Genet ; 21(1): 141, 2020 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-32611331

RESUMO

BACKGROUND: Lynch syndrome (LS), which is known as a hereditary cancer syndrome, is distinguished by microsatellite instability, represented by the altered number of repetitive sequences in the coding and/or non-coding region. Immunohistochemical staining (IHC) of DNA mismatch repair (MMR) proteins (e.g., MLH1, MSH2, MSH6, and PMS2) has been recognized as an useful technique for screening of LS. Previous study has shown that the assessment of IHC, however, requires specific caution due to variable staining patterns even without germline mutations in MMR genes. CASE PRESENTATION: A 48-year-old man, who had been treated for anaplastic astrocytoma, was referred to our department for the precise examination of progressing anemia. Whole-body examination revealed two advanced carcinomas in descending colon and stomach. A hypo-vascular mass lesion was detected in liver as well. Pathological diagnosis (on surgical specimens) was poorly differentiated adenocarcinoma in descending colon, moderately differentiated tubular adenocarcinoma in stomach, and liver metastasis, which is possibly from colon. It was suspected that this case would be Turcot's syndrome-type-1 due to its specific family history having two cases of colon cancer within the second relatives. Pathogenic frameshift mutations in codon 618 of MLH1 gene was identified. Immunohistochemical analyses (IHC) demonstrated complete loss of MLH1 immuno-expression as well as of PMS2 except for those in brain tumor. Although frameshift mutation was not found in MSH6 gene, histological expression of MSH6 was patchy in primary colon carcinoma and was completely lost in the metastatic site in liver. MSH6 expression in gastric carcinoma, a coincidental cancer in this case, was intact. An abnormal (C)8 region was identified by the cloned PCR of colon and liver tumors but not from gastric cancer. Frameshift mutation in a (C)8 tract in exon 5 of the MSH6 gene was also detected in liver metastasis. CONCLUSION: This case supports a plausible mechanism, proposed by a previous literature, for the reduced expression of MSH6 in a somatic mutation manner, which might preferentially happen in colon cancer rather than in stomach carcinoma in MLH1/PMS2-deficient type of Turcot's syndrome type 1.


Assuntos
Neoplasias Encefálicas/genética , Neoplasias do Colo/genética , Neoplasias Colorretais/genética , Proteínas de Ligação a DNA/imunologia , Predisposição Genética para Doença , Neoplasias Hepáticas/secundário , Mutação/genética , Síndromes Neoplásicas Hereditárias/genética , Adulto , Sequência de Bases , Reparo de Erro de Pareamento de DNA/genética , Feminino , Humanos , Masculino , Repetições de Microssatélites/genética , Pessoa de Meia-Idade , Linhagem
5.
Dermatol Online J ; 26(3)2020 Mar 20.
Artigo em Inglês | MEDLINE | ID: mdl-32609448

RESUMO

Dermatomyositis is a clinically heterogenous inflammatory myopathy with unique cutaneous features. Myositis-specific antibodies can aid in diagnosis and anticipation of patient prognosis. Herein, we report a 22-year-old man who presented with multifocal erythematous plaques with violaceous papules on his bilateral elbows, neck, and face. He was diagnosed with biopsy-proven dermatomyositis and determined to be seropositive for nuclear matrix protein 2 antibody (NXP-2). He was treated with systemic corticosteroids, then intravenous methylprednisolone and azathioprine, and ultimately achieved greatest treatment response with intravenous immune globulin therapy.


Assuntos
Corticosteroides/uso terapêutico , Autoanticorpos/sangue , Proteínas de Ligação a DNA/imunologia , Dermatomiosite/imunologia , Dermatomiosite/terapia , Imunoglobulinas Intravenosas/uso terapêutico , Fatores de Transcrição/imunologia , Algoritmos , Biópsia , Dermatomiosite/diagnóstico , Dermatomiosite/patologia , Diagnóstico Diferencial , Humanos , Masculino , Adulto Jovem
6.
Nat Immunol ; 21(8): 950-961, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32572241

RESUMO

A contribution of epigenetic modifications to B cell tolerance has been proposed but not directly tested. Here we report that deficiency of ten-eleven translocation (Tet) DNA demethylase family members Tet2 and Tet3 in B cells led to hyperactivation of B and T cells, autoantibody production and lupus-like disease in mice. Mechanistically, in the absence of Tet2 and Tet3, downregulation of CD86, which normally occurs following chronic exposure of self-reactive B cells to self-antigen, did not take place. The importance of dysregulated CD86 expression in Tet2- and Tet3-deficient B cells was further demonstrated by the restriction, albeit not complete, on aberrant T and B cell activation following anti-CD86 blockade. Tet2- and Tet3-deficient B cells had decreased accumulation of histone deacetylase 1 (HDAC1) and HDAC2 at the Cd86 locus. Thus, our findings suggest that Tet2- and Tet3-mediated chromatin modification participates in repression of CD86 on chronically stimulated self-reactive B cells, which contributes, at least in part, to preventing autoimmunity.


Assuntos
Autoimunidade/imunologia , Linfócitos B/imunologia , Antígeno B7-2/imunologia , Proteínas de Ligação a DNA/imunologia , Dioxigenases/imunologia , Proteínas Proto-Oncogênicas/imunologia , Animais , Doenças Autoimunes/imunologia , Epigênese Genética/imunologia , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos
7.
PLoS Pathog ; 16(5): e1008618, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32453758

RESUMO

The genomic instability associated with adult T cell leukemia/lymphoma (ATL) is causally linked to Tax, the HTLV-1 viral oncoprotein, but the underlying mechanism is not fully understood. We have previously shown that Tax hijacks and aberrantly activates ring finger protein 8 (RNF8) - a lysine 63 (K63)-specific ubiquitin E3 ligase critical for DNA double-strand break (DSB) repair signaling - to assemble K63-linked polyubiquitin chains (K63-pUbs) in the cytosol. Tax and the cytosolic K63-pUbs, in turn, initiate additional recruitment of linear ubiquitin assembly complex (LUBAC) to produce hybrid K63-M1 pUbs, which trigger a kinase cascade that leads to canonical IKK:NF-κB activation. Here we demonstrate that HTLV-1-infected cells are impaired in DNA damage response (DDR). This impairment correlates with the induction of microscopically visible nuclear speckles by Tax known as the Tax-speckle structures (TSS), which act as pseudo DNA damage signaling scaffolds that sequester DDR factors such as BRCA1, DNA-PK, and MDC1. We show that TSS co-localize with Tax, RNF8 and K63-pUbs, and their formation depends on RNF8. Tax mutants defective or attenuated in inducing K63-pUb assembly are deficient or tempered in TSS induction and DDR impairment. Finally, our results indicate that loss of RNF8 expression reduces HTLV-1 viral gene expression and frequently occurs in ATL cells. Thus, during HTLV-1 infection, Tax activates RNF8 to assemble nuclear K63-pUbs that sequester DDR factors in Tax speckles, disrupting DDR signaling and DSB repair. Down-regulation of RNF8 expression is positively selected during infection and progression to disease, and further exacerbates the genomic instability of ATL.


Assuntos
Proteínas de Ligação a DNA/imunologia , Regulação para Baixo/imunologia , Instabilidade Genômica/imunologia , Infecções por HTLV-I/imunologia , Vírus Linfotrópico T Tipo 1 Humano/imunologia , Leucemia-Linfoma de Células T do Adulto/imunologia , Proteínas de Neoplasias/imunologia , Ubiquitina-Proteína Ligases/imunologia , Quebras de DNA de Cadeia Dupla , Reparo do DNA/genética , Reparo do DNA/imunologia , Proteínas de Ligação a DNA/genética , Produtos do Gene tax/genética , Produtos do Gene tax/imunologia , Infecções por HTLV-I/genética , Infecções por HTLV-I/patologia , Células HeLa , Vírus Linfotrópico T Tipo 1 Humano/genética , Humanos , Leucemia-Linfoma de Células T do Adulto/genética , Leucemia-Linfoma de Células T do Adulto/patologia , Proteínas de Neoplasias/genética , Ubiquitina-Proteína Ligases/genética
8.
PLoS Pathog ; 16(5): e1008244, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32365082

RESUMO

Viral escape from CD8+ cytotoxic T lymphocyte responses correlates with disease progression and represents a significant challenge for vaccination. Here, we demonstrate that CD8+ T cell recognition of the naturally occurring MHC-I-restricted LCMV-associated immune escape variant Y4F is restored following vaccination with a proline-altered peptide ligand (APL). The APL increases MHC/peptide (pMHC) complex stability, rigidifies the peptide and facilitates T cell receptor (TCR) recognition through reduced entropy costs. Structural analyses of pMHC complexes before and after TCR binding, combined with biophysical analyses, revealed that although the TCR binds similarly to all complexes, the p3P modification alters the conformations of a very limited amount of specific MHC and peptide residues, facilitating efficient TCR recognition. This approach can be easily introduced in peptides restricted to other MHC alleles, and can be combined with currently available and future vaccination protocols in order to prevent viral immune escape.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Vírus da Coriomeningite Linfocítica/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Animais , Antivirais/metabolismo , Linfócitos T CD8-Positivos/fisiologia , Proteínas de Ligação a DNA/imunologia , Epitopos/imunologia , Epitopos de Linfócito T/imunologia , Genes RAG-1/imunologia , Ligantes , Ativação Linfocitária/imunologia , Vírus da Coriomeningite Linfocítica/genética , Vírus da Coriomeningite Linfocítica/patogenicidade , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Peptídeos/metabolismo , Prolina/metabolismo , Ligação Proteica , Linfócitos T Citotóxicos/imunologia , Vacinação/métodos
9.
Dermatol Online J ; 26(2)2020 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-32239891

RESUMO

Dermatomyositis is an auto-immune inflammatory myopathy that primarily affects the skin and muscle and can be triggered by exposure to various environmental factors. We present a patient with active syphilis infection who developed dermatomyositis and discuss the significance of anti-NXP2 autoantibody positivity.


Assuntos
Adenosina Trifosfatases/imunologia , Doenças Autoimunes/etiologia , Proteínas de Ligação a DNA/imunologia , Dermatomiosite/etiologia , Pele/patologia , Sífilis/complicações , Adulto , Autoanticorpos/sangue , Doenças Autoimunes/patologia , Dermatomiosite/imunologia , Dermatomiosite/patologia , Humanos , Masculino
10.
Sci Rep ; 10(1): 3709, 2020 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-32111888

RESUMO

Mycobacterium tuberculosis infection causes high rates of morbidity and mortality. Host-directed therapy may enhance the immune response, reduce tissue damage and shorten treatment duration. The inflammasome is integral to innate immune responses but over-activation has been described in tuberculosis (TB) pathology and TB-immune reconstitution syndrome. Here we explore how clinical isolates differentially activate the inflammasome and how inflammasome inhibition can lead to enhanced bacterial clearance. Wild-type, Nlrp3-/-/Aim2-/-, Casp1/11-/- and Asc-/- murine bone-marrow derived macrophages (BMDMs) were infected with laboratory strain M. tuberculosis H37Rv or clinical isolates from various lineages. Inflammasome activation and bacterial numbers were measured, and pharmacological inhibition of NLRP3 was achieved using MCC950. Clinical isolates of M. tuberculosis differed in their ability to activate inflammasomes. Beijing isolates had contrasting effects on IL-1ß and caspase-1 activation, but all clinical isolates induced lower IL-1ß release than H37Rv. Our studies suggest the involvement of NLRP3, AIM2 and an additional unknown sensor in IL-1ß maturation. Pharmacological blockade of NLRP3 with MCC950 reduced bacterial survival, and combined treatment with the antimycobacterial drug rifampicin enhanced the effect. Modulating the inflammasome is an attractive adjunct to current anti-mycobacterial therapy that warrants further investigation.


Assuntos
Inflamassomos/imunologia , Macrófagos/imunologia , Mycobacterium tuberculosis/crescimento & desenvolvimento , Tuberculose/imunologia , Animais , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/imunologia , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Humanos , Inflamassomos/efeitos dos fármacos , Inflamassomos/genética , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Macrófagos/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/imunologia , Sulfonas/farmacologia , Tuberculose/genética , Tuberculose/microbiologia
11.
Sci Adv ; 6(10): eaax6346, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32181339

RESUMO

Placental malaria (PM) is associated with severe inflammation leading to abortion, preterm delivery, and intrauterine growth restriction. Innate immunity responses play critical roles, but the mechanisms underlying placental immunopathology are still unclear. Here, we investigated the role of inflammasome activation in PM by scrutinizing human placenta samples from an endemic area and ablating inflammasome components in a PM mouse model. The reduction in birth weight in babies from infected mothers is paralleled by increased placental expression of AIM2 and NLRP3 inflammasomes. Using genetic dissection, we reveal that inflammasome activation pathways are involved in the production and detrimental action of interleukin-1ß (IL-1ß) in the infected placenta. The IL-1R pharmacological antagonist Anakinra improved pregnancy outcomes by restoring fetal growth and reducing resorption in an experimental model. These findings unveil that IL-1ß-mediated signaling is a determinant of PM pathogenesis, suggesting that IL-1R antagonists can improve clinical outcomes of malaria infection in pregnancy.


Assuntos
Inflamassomos/efeitos dos fármacos , Interleucina-1beta/imunologia , Malária Falciparum/imunologia , Malária/imunologia , Plasmodium falciparum/patogenicidade , Complicações Parasitárias na Gravidez/imunologia , Transdução de Sinais/efeitos dos fármacos , Animais , Caspase 1/genética , Caspase 1/imunologia , Linhagem Celular , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/imunologia , Feminino , Regulação da Expressão Gênica , Humanos , Imunidade Inata , Fatores Imunológicos/farmacologia , Inflamassomos/genética , Inflamassomos/imunologia , Interferon gama/genética , Interferon gama/imunologia , Proteína Antagonista do Receptor de Interleucina 1/farmacologia , Interleucina-1beta/antagonistas & inibidores , Interleucina-1beta/genética , Malária/tratamento farmacológico , Malária/genética , Malária/parasitologia , Malária Falciparum/genética , Malária Falciparum/parasitologia , Malária Falciparum/patologia , Camundongos , Camundongos Knockout , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/imunologia , Plasmodium berghei/imunologia , Plasmodium berghei/patogenicidade , Plasmodium falciparum/imunologia , Gravidez , Complicações Parasitárias na Gravidez/genética , Complicações Parasitárias na Gravidez/parasitologia , Complicações Parasitárias na Gravidez/prevenção & controle , Receptores de Interleucina-1/genética , Receptores de Interleucina-1/imunologia , Transdução de Sinais/imunologia , Células THP-1 , Trofoblastos/efeitos dos fármacos , Trofoblastos/imunologia , Trofoblastos/parasitologia , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologia
12.
PLoS One ; 15(3): e0229672, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32214362

RESUMO

More than 170 types of human papilloma viruses (HPV) exist with many causing proliferative diseases linked to malignancy in indications such as cervical cancer and head and neck squamous cell carcinoma. Characterization of antibody levels toward HPV serology is challenging due to complex biology of oncoproteins, pre-existing titers to multiple HPV types, cross-reactivity, and low affinity, polyclonal responses. Using multiplex technology from MSD, we have developed an assay that simultaneously characterizes antibodies against E6 and E7 oncoproteins of HPV16 and 18, the primary drivers of HPV-associated oncogenesis. We fusion tagged our E6 and E7 proteins with MBP via two-step purification, spot-printed an optimized concentration of protein into wells of MSD 96-well plates, and assayed various cynomolgus monkey, human and HPV+ cervical cancer patient serum to validate the assay. The dynamic range of the assay covered 4-orders of magnitude and antibodies were detected in serum at a dilution up to 100,000-fold. The assay was very precise (n = 5 assay runs) with median CV of human serum samples ~ 5.3% and inter-run variability of 11.4%. The multiplex serology method has strong cross-reactivity between E6 oncoproteins from human serum samples as HPV18 E6 antigens neutralized 5 of 6 serum samples as strongly as HPV16 E6. Moderate concordance (Spearman's Rank = 0.775) was found between antibody responses against HPV16 E7 in the multiplex assay compared to standard ELISA serology methods. These results demonstrate the development of a high-throughput, multi-plex assay that requires lower sample quantity input with greater dynamic range to detect type-specific anti-HPV concentrations to E6 and E7 oncoproteins of HPV16 and 18.


Assuntos
Anticorpos Antivirais/sangue , Papillomavirus Humano 16/imunologia , Papillomavirus Humano 18/imunologia , Imunoensaio/métodos , Imunoglobulina G/sangue , Animais , Especificidade de Anticorpos , Reações Cruzadas , Proteínas de Ligação a DNA/imunologia , Técnicas Eletroquímicas , Ensaio de Imunoadsorção Enzimática , Feminino , Ensaios de Triagem em Larga Escala/métodos , Ensaios de Triagem em Larga Escala/estatística & dados numéricos , Humanos , Imunoensaio/estatística & dados numéricos , Limite de Detecção , Medições Luminescentes/métodos , Medições Luminescentes/estatística & dados numéricos , Macaca fascicularis , Proteínas Oncogênicas Virais/imunologia , Proteínas E7 de Papillomavirus/imunologia , Proteínas Repressoras/imunologia , Neoplasias do Colo do Útero/imunologia , Neoplasias do Colo do Útero/virologia
13.
Cell Prolif ; 53(4): e12791, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32162380

RESUMO

Hepatocellular carcinoma (HCC) is a primary liver malignancy with a high global prevalence and a dismal prognosis. Studies are urgently needed to examine the molecular pathogenesis and biological characteristics of HCC. Chromatin remodelling, an integral component of the DNA damage response, protects against DNA damage-induced genome instability and tumorigenesis by triggering the signalling events that activate the interconnected DNA repair pathways. The SWI/SNF complexes are one of the most extensively investigated adenosine triphosphate-dependent chromatin remodelling complexes, and mutations in genes encoding SWI/SNF subunits are frequently observed in various human cancers, including HCC. The mutated SWI/SNF complex subunits exert dual functions by accelerating or inhibiting HCC initiation and progression. Furthermore, the abnormal SWI/SNF complexes influence the transcription of interferon-stimulated genes, as well as the differentiation, activation and recruitment of several immune cell types. In addition, they exhibit synergistic effects with immune checkpoint inhibitors in the treatment of diverse tumour types. Therefore, understanding the mutations and deficiencies of the SMI/SNF complexes, together with the associated functional mechanisms, may provide a novel strategy to treat HCC through targeting the related genes or modulating the tumour microenvironment.


Assuntos
Carcinoma Hepatocelular/genética , Neoplasias Hepáticas/genética , Mutação , Proteínas Nucleares/genética , Fatores de Transcrição/genética , Animais , Carcinogênese/genética , Carcinogênese/imunologia , Carcinoma Hepatocelular/imunologia , Carcinoma Hepatocelular/terapia , Montagem e Desmontagem da Cromatina , Dano ao DNA , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/imunologia , Regulação Neoplásica da Expressão Gênica , Instabilidade Genômica , Humanos , Imunidade , Imunoterapia , Neoplasias Hepáticas/imunologia , Neoplasias Hepáticas/terapia , Proteínas Nucleares/imunologia , Fatores de Transcrição/imunologia
14.
Proc Natl Acad Sci U S A ; 117(9): 4653-4663, 2020 03 03.
Artigo em Inglês | MEDLINE | ID: mdl-32060122

RESUMO

The LIN28:pre-let-7:TUTase ternary complex regulates pluripotency and oncogenesis by controlling processing of the let-7 family of microRNAs. The complex oligouridylates the 3' ends of pre-let-7 molecules, leading to their degradation via the DIS3L2 exonuclease. Previous studies suggest that components of this complex are potential therapeutic targets in malignancies that aberrantly express LIN28. In this study we developed a functional epitope selection approach to identify nanobody inhibitors of the LIN28:pre-let-7:TUT4 complex. We demonstrate that one of the identified nanobodies, Nb-S2A4, targets the 106-residue LIN28:let-7 interaction (LLI) fragment of TUT4. Nb-S2A4 can effectively inhibit oligouridylation and monouridylation of pre-let-7g in vitro. Expressing Nb-S2A4 allows maturation of the let-7 species in cells expressing LIN28, highlighting the therapeutic potential of targeting the LLI fragment.


Assuntos
Proteínas de Ligação a DNA/imunologia , MicroRNAs/metabolismo , Processamento de Terminações 3' de RNA , Anticorpos de Domínio Único/imunologia , Animais , Sítios de Ligação , Proteínas de Ligação a DNA/química , Células HEK293 , Células HeLa , Humanos , Camundongos , MicroRNAs/genética , Ligação Proteica , Estabilidade de RNA , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Células Sf9 , Spodoptera
15.
Gene ; 739: 144496, 2020 May 20.
Artigo em Inglês | MEDLINE | ID: mdl-32088242

RESUMO

Early larval developmental stages of fish are highly susceptible to opportunistic pathogens until the complete maturation of the lymphoid organs. Knowledge of the expression pattern of important markers of adaptive immune system during the ontogenetic development is essential before vaccinating the fish. In the present study, Pterophyllum scalare (angelfish) was taken to explore the relative expression profile of developmental markers of adaptive immunity, recombination activating gene-2 (RAG-2) and immunoglobulin M (IgM). The fishes were bred and early developmental stages (0-45 days post-hatched) were used to assess the expression profile. The genes, RAG-2 and IgM were cloned and sequenced with the base pair lengths of 1958 bp and 225 bp respectively. The mRNA expression of RAG-2 appeared at insignificant level at the first day of hatching, but the expression was significantly increased from 24 dph (days post-hatching) onwards and reached its peak at 27 dph. The results proved that the maturation of lymphoid organs was completed at 27 dph as the respective protein is involved in the V(D)J recombination, important for the maturation of lymphoid organs. A similar trend was also observed in the mRNA transcript levels of IgM gene and a significantly high expression was detected from 27 dph onwards. The present study suggested that the suitable time for vaccination in P. scalare could be taken at 27 dph, as the maturation and development of lymphoid organs is completed thus helps in stimulating the adaptive response of immunity against any pathogen.


Assuntos
Imunidade Adaptativa/genética , Ciclídeos/genética , Proteínas de Ligação a DNA/imunologia , Vacinação/veterinária , Animais , Ciclídeos/imunologia , Clonagem Molecular , Proteínas de Ligação a DNA/genética , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Imunoglobulina M/genética , Imunoglobulina M/imunologia , Rim/imunologia , Tecido Linfoide/imunologia , Baço/imunologia
16.
Blood ; 135(9): 610-619, 2020 02 27.
Artigo em Inglês | MEDLINE | ID: mdl-31942628

RESUMO

Traditionally, primary immune deficiencies have been defined based on increased susceptibility to recurrent and/or severe infections. However, immune dysregulation, manifesting with autoimmunity or hyperinflammatory disease, has emerged as a common feature. This is especially true in patients affected by combined immune deficiency (CID), a group of disorders caused by genetic defects that impair, but do not completely abolish, T-cell function. Hypomorphic mutations in the recombination activating genes RAG1 and RAG2 represent the prototype of the broad spectrum of clinical and immunological phenotypes associated with CID. The study of patients with RAG deficiency and with other forms of CID has revealed distinct abnormalities in central and peripheral T- and B-cell tolerance as the key mechanisms involved in immune dysregulation. Understanding the pathophysiology of autoimmunity and hyperinflammation in these disorders may also permit more targeted therapeutic interventions.


Assuntos
Imunodeficiência Combinada Severa/imunologia , Autoimunidade/genética , Autoimunidade/imunologia , Proteínas de Ligação a DNA/deficiência , Proteínas de Ligação a DNA/imunologia , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/imunologia , Humanos , Proteínas Nucleares/deficiência , Proteínas Nucleares/imunologia , Imunodeficiência Combinada Severa/genética
17.
Mol Immunol ; 117: 131-138, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31770676

RESUMO

V(D)J recombination is a process of somatic recombination catalyzed by proteins encoded by RAG1 and RAG2 genes, both restricted to the genome of jawed vertebrates. Their proteins constitute the enzymatic core of V(D)J recombination machinery and are crucial for jawed vertebrate adaptive immunity. Mammals possess great ecological diversity, and their complex evolutionary history associated with radiation to different environments presented many distinct pathogenic challenges from these different habitats. Cetaceans comprise a mammalian order of fully aquatic mammals that have arisen from a complete terrestrial ancestor and, accordingly, was confronted with challenges from changing environmental pathogens while they transitioned from land to sea. In this study we undertook molecular evolutionary analyses of RAG1 and RAG2 genes, exploring the possible role of natural selection acting on these genes focusing on the cetacean lineage. We performed phylogenetic reconstructions on IQ-TREE, together with selection analyses in the codeml program of the PAML package, and in the FITMODEL program for codon evolution and switching on both the RAG1 and RAG2 genes. Our findings demonstrate that RAG1 and RAG2 remained fairly conserved among tetrapods, with purifying selection acting on both genes, with evidence for a few punctuated shifts in nucleotide substitution rates of both genes along tetrapod evolution. We demonstrate differential evolution in the closely linked genes RAG1 and RAG2 specifically in cetaceans.


Assuntos
Evolução Biológica , Cetáceos/genética , Cetáceos/imunologia , Proteínas de Ligação a DNA/genética , Genes RAG-1/genética , Animais , Proteínas de Ligação a DNA/imunologia , Genes RAG-1/imunologia , Filogenia
19.
Arthritis Rheumatol ; 72(1): 89-99, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31342656

RESUMO

OBJECTIVE: Long interspersed nuclear element 1 (LINE-1) encodes 2 proteins, the RNA binding protein p40 and endonuclease and reverse transcriptase (open-reading frame 2p [ORF2p]), which are both required for LINE-1 to retrotranspose. In cells expressing LINE-1, these proteins assemble with LINE-1 RNA and additional RNA binding proteins, some of which are well-known autoantigens in patients with systemic lupus erythematosus (SLE). This study was undertaken to investigate whether SLE patients also produce autoantibodies against LINE-1 p40. METHODS: Highly purified p40 protein was used to quantitate IgG autoantibodies in serum from 172 SLE patients and from disease controls and age-matched healthy subjects by immunoblotting and enzyme-linked immunosorbent assay (ELISA). Preparations of p40 that also contained associated proteins were analyzed by immunoblotting with patient sera. RESULTS: Antibodies reactive with p40 were detected in the majority of patients and many healthy controls. Their levels were higher in patients with SLE, but not those with systemic sclerosis, compared to healthy subjects (P = 0.01). Anti-p40 reactivity was higher in patients during a flare than in patients with disease in remission (P = 0.03); correlated with the SLE Disease Activity Index score (P = 0.0002), type I interferon score (P = 0.006), decrease in complement C3 level (P = 0.0001), the presence of anti-DNA antibodies (P < 0.0001) and anti-C1q antibodies (P = 0.004), and current or past history of nephritis (P = 0.02 and P = 0.003, respectively); and correlated inversely with age (r = -0.49, P < 0.0001). SLE patient sera also reacted with p40-associated proteins. CONCLUSION: Autoantibodies reacting with LINE-1 p40 characterize a population of SLE patients with severe and active disease. These autoantibodies may represent an early immune response against LINE-1 p40 that does not yet by itself imply clinically significant autoimmunity, but may represent an early, and still reversible, step toward SLE pathogenesis.


Assuntos
Autoanticorpos/imunologia , Proteínas de Ligação a DNA/imunologia , Lúpus Eritematoso Sistêmico/imunologia , Fatores Etários , Anticorpos Antinucleares/imunologia , Estudos de Casos e Controles , Complemento C1q/imunologia , Complemento C3/imunologia , Humanos , Elementos Nucleotídeos Longos e Dispersos/imunologia , Nefrite Lúpica/imunologia , Escleroderma Sistêmico/imunologia , Índice de Gravidade de Doença
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