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1.
Sci Rep ; 11(1): 17234, 2021 08 26.
Artigo em Inglês | MEDLINE | ID: mdl-34446788

RESUMO

Over the past two decades, there has been a great interest in the study of HLA-E-restricted αß T cells during bacterial and viral infections, including recently SARS-CoV-2 infection. Phenotyping of these specific HLA-E-restricted T cells requires new tools such as tetramers for rapid cell staining or sorting, as well as for the identification of new peptides capable to bind to the HLA-E pocket. To this aim, we have developed an optimal photosensitive peptide to generate stable HLA-E/pUV complexes allowing high-throughput production of new HLA-E/peptide complexes by peptide exchange. We characterized the UV exchange by ELISA and improved the peptide exchange readout using size exclusion chromatography. This novel approach for complex quantification is indeed very important to perform tetramerization of MHC/peptide complexes with the high quality required for detection of specific T cells. Our approach allows the rapid screening of peptides capable of binding to the non-classical human HLA-E allele, paving the way for the development of new therapeutic approaches based on the detection of HLA-E-restricted T cells.


Assuntos
Epitopos de Linfócito T/química , Antígenos de Histocompatibilidade Classe I/química , Complexo Principal de Histocompatibilidade/imunologia , Peptídeos/química , Sequência de Aminoácidos , Epitopos de Linfócito T/imunologia , Ensaios de Triagem em Larga Escala , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Técnicas Imunológicas , Processos Fotoquímicos , Ligação Proteica , Conformação Proteica , Linfócitos T Citotóxicos/imunologia
2.
Front Immunol ; 12: 698193, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34381451

RESUMO

HLA molecules are key restrictive elements to present intracellular antigens at the crossroads of an effective T-cell response against SARS-CoV-2. To determine the impact of the HLA genotype on the severity of SARS-CoV-2 courses, we investigated data from 6,919 infected individuals. HLA-A, -B, and -DRB1 allotypes grouped into HLA supertypes by functional or predicted structural similarities of the peptide-binding grooves did not predict COVID-19 severity. Further, we did not observe a heterozygote advantage or a benefit from HLA diplotypes with more divergent physicochemical peptide-binding properties. Finally, numbers of in silico predicted viral T-cell epitopes did not correlate with the severity of SARS-CoV-2 infections. These findings suggest that the HLA genotype is no major factor determining COVID-19 severity. Moreover, our data suggest that the spike glycoprotein alone may allow for abundant T-cell epitopes to mount robust T-cell responses not limited by the HLA genotype.


Assuntos
Infecções por Coronavirus/genética , Antígenos de Histocompatibilidade Classe II/imunologia , Antígenos de Histocompatibilidade Classe I/imunologia , Adulto , Simulação por Computador , Estudos Transversais , Epitopos de Linfócito T/genética , Epitopos de Linfócito T/imunologia , Feminino , Genótipo , Antígenos de Histocompatibilidade Classe I/genética , Antígenos de Histocompatibilidade Classe II/genética , Humanos , Masculino , Pessoa de Meia-Idade , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus/imunologia
3.
Nat Commun ; 12(1): 4365, 2021 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-34272369

RESUMO

Activating RAS missense mutations are among the most prevalent genomic alterations observed in human cancers and drive oncogenesis in the three most lethal tumor types. Emerging evidence suggests mutant KRAS (mKRAS) may be targeted immunologically, but mKRAS epitopes remain poorly defined. Here we employ a multi-omics approach to characterize HLA class I-restricted mKRAS epitopes. We provide proteomic evidence of mKRAS epitope processing and presentation by high prevalence HLA class I alleles. Select epitopes are immunogenic enabling mKRAS-specific TCRαß isolation. TCR transfer to primary CD8+ T cells confers cytotoxicity against mKRAS tumor cell lines independent of histologic origin, and the kinetics of lytic activity correlates with mKRAS peptide-HLA class I complex abundance. Adoptive transfer of mKRAS-TCR engineered CD8+ T cells leads to tumor eradication in a xenograft model of metastatic lung cancer. This study validates mKRAS peptides as bona fide epitopes facilitating the development of immune therapies targeting this oncoprotein.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Carcinogênese/imunologia , Epitopos de Linfócito T/imunologia , Neoplasias Pulmonares/imunologia , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteínas Proto-Oncogênicas p21(ras)/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/imunologia , Transferência Adotiva , Alelos , Animais , Carcinogênese/genética , Linhagem Celular Tumoral , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Camundongos , Mutação , Peptídeos/genética , Peptídeos/imunologia , Proteômica , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Ensaios Antitumorais Modelo de Xenoenxerto
4.
Genes (Basel) ; 12(6)2021 06 10.
Artigo em Inglês | MEDLINE | ID: mdl-34200642

RESUMO

FENDRR (Fetal-lethal non-coding developmental regulatory RNA, LncRNA FOXF1-AS1) is a recently identified tumor suppressor long non-coding (LncRNA) RNA, and its expression has been linked with epigenetic modulation of the target genes involved in tumor immunity. In this study, we aimed to understand the role of FENDRR in predicting immune-responsiveness and the inflammatory tumor environment. Briefly, FENDRR expression and its relationship to immune activation signals were assessed in murine cell lines. Data suggested that tumor cells (e.g., C26 colon, 4T1 breast) that typically upregulate immune activation genes and the MHC class I molecule exhibited high FENDRR expression levels. Conversely, tumor cells with a generalized downregulation of immune-related gene expression (e.g., B16F10 melanoma) demonstrated low to undetectable FENDRR levels. Mechanistically, the modulation of FENDRR expression enhanced the inflammatory and WNT signaling pathways in tumors. Our early data suggest that FENDRR can play an important role in the development of immune-relevant phenotypes in tumors, and thereby improve cancer immunotherapy.


Assuntos
Neoplasias do Colo/genética , Melanoma/genética , RNA Longo não Codificante/genética , Animais , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/imunologia , Linhagem Celular Tumoral , Neoplasias do Colo/imunologia , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Melanoma/imunologia , Camundongos , Camundongos Endogâmicos C57BL , RNA Longo não Codificante/metabolismo , Via de Sinalização Wnt
5.
Nat Genet ; 53(8): 1196-1206, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34253920

RESUMO

To systematically define molecular features in human tumor cells that determine their degree of sensitivity to human allogeneic natural killer (NK) cells, we quantified the NK cell responsiveness of hundreds of molecularly annotated 'DNA-barcoded' solid tumor cell lines in multiplexed format and applied genome-scale CRISPR-based gene-editing screens in several solid tumor cell lines, to functionally interrogate which genes in tumor cells regulate the response to NK cells. In these orthogonal studies, NK cell-sensitive tumor cells tend to exhibit 'mesenchymal-like' transcriptional programs; high transcriptional signature for chromatin remodeling complexes; high levels of B7-H6 (NCR3LG1); and low levels of HLA-E/antigen presentation genes. Importantly, transcriptional signatures of NK cell-sensitive tumor cells correlate with immune checkpoint inhibitor (ICI) resistance in clinical samples. This study provides a comprehensive map of mechanisms regulating tumor cell responses to NK cells, with implications for future biomarker-driven applications of NK cell immunotherapies.


Assuntos
Citotoxicidade Imunológica/genética , Resistencia a Medicamentos Antineoplásicos/genética , Regulação Neoplásica da Expressão Gênica , Inibidores de Checkpoint Imunológico/farmacologia , Células Matadoras Naturais/fisiologia , Células Alógenas/fisiologia , Animais , Antígenos B7/genética , Linhagem Celular Tumoral , Montagem e Desmontagem da Cromatina/fisiologia , Testes Imunológicos de Citotoxicidade/métodos , Citotoxicidade Imunológica/fisiologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Feminino , Genoma Humano , Antígenos de Histocompatibilidade Classe I/genética , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Camundongos Endogâmicos NOD , Ensaios Antitumorais Modelo de Xenoenxerto
6.
Nat Commun ; 12(1): 4355, 2021 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-34272362

RESUMO

Mucosal-associated Invariant T (MAIT) cells are recognized for their antibacterial functions. The protective capacity of MAIT cells has been demonstrated in murine models of local infection, including in the lungs. Here we show that during systemic infection of mice with Francisella tularensis live vaccine strain results in evident MAIT cell expansion in the liver, lungs, kidney and spleen and peripheral blood. The responding MAIT cells manifest a polarised Th1-like MAIT-1 phenotype, including transcription factor and cytokine profile, and confer a critical role in controlling bacterial load. Post resolution of the primary infection, the expanded MAIT cells form stable memory-like MAIT-1 cell populations, suggesting a basis for vaccination. Indeed, a systemic vaccination with synthetic antigen 5-(2-oxopropylideneamino)-6-D-ribitylaminouracil in combination with CpG adjuvant similarly boosts MAIT cells, and results in enhanced protection against both systemic and local infections with different bacteria. Our study highlights the potential utility of targeting MAIT cells to combat a range of bacterial pathogens.


Assuntos
Citocinas/metabolismo , Francisella tularensis/imunologia , Imunidade Inata , Células T Invariantes Associadas à Mucosa/imunologia , Adjuvantes Imunológicos , Animais , Antígenos de Histocompatibilidade Classe I/genética , Antígenos de Histocompatibilidade Classe I/imunologia , Fígado/imunologia , Pulmão/imunologia , Camundongos , Camundongos Knockout , Antígenos de Histocompatibilidade Menor/genética , Antígenos de Histocompatibilidade Menor/imunologia , Células T Invariantes Associadas à Mucosa/metabolismo , Fenótipo , RNA-Seq , Ribitol/análogos & derivados , Ribitol/imunologia , Análise de Célula Única , Baço/imunologia , Células Th1/imunologia , Células Th1/metabolismo , Transcriptoma/genética , Uracila/análogos & derivados , Uracila/imunologia , Vacinas Atenuadas/imunologia
7.
Nat Commun ; 12(1): 4372, 2021 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-34272370

RESUMO

Intrarenal B cells in human renal allografts indicate transplant recipients with a poor prognosis, but how these cells contribute to rejection is unclear. Here we show using single-cell RNA sequencing that intrarenal class-switched B cells have an innate cell transcriptional state resembling mouse peritoneal B1 or B-innate (Bin) cells. Antibodies generated by Bin cells do not bind donor-specific antigens nor are they enriched for reactivity to ubiquitously expressed self-antigens. Rather, Bin cells frequently express antibodies reactive with either renal-specific or inflammation-associated antigens. Furthermore, local antigens can drive Bin cell proliferation and differentiation into plasma cells expressing self-reactive antibodies. These data show a mechanism of human inflammation in which a breach in organ-restricted tolerance by infiltrating innate-like B cells drives local tissue destruction.


Assuntos
Aloenxertos/imunologia , Linfócitos B/metabolismo , Rejeição de Enxerto/imunologia , Inflamação/metabolismo , Transplante de Rim/efeitos adversos , Animais , Autoanticorpos/imunologia , Linfócitos B/imunologia , Linfócitos B/patologia , Regulação da Expressão Gênica/genética , Regulação da Expressão Gênica/imunologia , Ontologia Genética , Antígenos de Histocompatibilidade Classe I/imunologia , Antígenos de Histocompatibilidade Classe II/imunologia , Humanos , Imunoglobulina G/imunologia , Rim/imunologia , Rim/metabolismo , Camundongos , Tonsila Palatina/imunologia , Tonsila Palatina/metabolismo , RNA-Seq , Análise de Célula Única , Transplante Homólogo
8.
Int J Mol Sci ; 22(14)2021 Jul 06.
Artigo em Inglês | MEDLINE | ID: mdl-34298868

RESUMO

Cancer eradication and clinical outcome of immunotherapy depend on tumor cell immunogenicity, including HLA class I (HLA-I) and PD-L1 expression on malignant cells, and on the characteristics of the tumor microenvironment, such as tumor immune infiltration and stromal reaction. Loss of tumor HLA-I is a common mechanism of immune escape from cytotoxic T lymphocytes and is linked to cancer progression and resistance to immunotherapy with the inhibitors of PD-L1/PD-1 signaling. Here we observed that HLA-I loss in bladder tumors is associated with T cell exclusion and tumor encapsulation with stromal elements rich in FAP-positive cells. In addition, PD-L1 upregulation in HLA-I negative tumors demonstrated a correlation with high tumor grade and worse overall- and cancer-specific survival of the patients. These changes define common immuno-morphological signatures compatible with cancer immune escape and acquired resistance to therapeutic interventions across different types of malignancy. They also may contribute to the search of new targets for cancer treatment, such as FAP-expressing cancer-associated fibroblasts, in refractory bladder tumors.


Assuntos
Antígenos de Histocompatibilidade Classe I/imunologia , Linfócitos do Interstício Tumoral/imunologia , Linfócitos T Citotóxicos/imunologia , Evasão Tumoral/imunologia , Neoplasias da Bexiga Urinária/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígeno B7-H1/imunologia , Feminino , Humanos , Imunoterapia/métodos , Masculino , Pessoa de Meia-Idade , Fenótipo , Microambiente Tumoral/imunologia , Adulto Jovem
9.
Viruses ; 13(7)2021 06 27.
Artigo em Inglês | MEDLINE | ID: mdl-34199077

RESUMO

Many viruses, especially RNA viruses, utilize programmed ribosomal frameshifting and/or stop codon readthrough in their expression, and in the decoding of a few a UGA is dynamically redefined to specify selenocysteine. This recoding can effectively increase viral coding capacity and generate a set ratio of products with the same N-terminal domain(s) but different C-terminal domains. Recoding can also be regulatory or generate a product with the non-universal 21st directly encoded amino acid. Selection for translation speed in the expression of many viruses at the expense of fidelity creates host immune defensive opportunities. In contrast to host opportunism, certain viruses, including some persistent viruses, utilize recoding or adventitious frameshifting as part of their strategy to evade an immune response or specific drugs. Several instances of recoding in small intensively studied viruses escaped detection for many years and their identification resolved dilemmas. The fundamental importance of ribosome ratcheting is consistent with the initial strong view of invariant triplet decoding which however did not foresee the possibility of transitory anticodon:codon dissociation. Deep level dynamics and structural understanding of recoding is underway, and a high level structure relevant to the frameshifting required for expression of the SARS CoV-2 genome has just been determined.


Assuntos
Vírus de DNA/genética , Vírus de DNA/imunologia , Antígenos de Histocompatibilidade Classe I/imunologia , Evasão da Resposta Imune , Vírus de RNA/genética , Antivirais/farmacologia , Códon de Terminação , Vírus de DNA/efeitos dos fármacos , Mudança da Fase de Leitura do Gene Ribossômico , Antígenos de Histocompatibilidade Classe I/genética , Conformação de Ácido Nucleico , Peptídeos/imunologia , Biossíntese de Proteínas , Vírus de RNA/efeitos dos fármacos , Vírus de RNA/imunologia
10.
Int J Mol Sci ; 22(13)2021 Jun 23.
Artigo em Inglês | MEDLINE | ID: mdl-34201655

RESUMO

It is now well accepted that the immune system can control cancer growth. However, tumors escape immune-mediated control through multiple mechanisms and the downregulation or loss of major histocompatibility class (MHC)-I molecules is a common immune escape mechanism in many cancers. MHC-I molecules present antigenic peptides to cytotoxic T cells, and MHC-I loss can render tumor cells invisible to the immune system. In this review, we examine the dysregulation of MHC-I expression in cancer, explore the nature of MHC-I-bound antigenic peptides recognized by immune cells, and discuss therapeutic strategies that can be used to overcome MHC-I deficiency in solid tumors, with a focus on the role of natural killer (NK) cells and CD4 T cells.


Assuntos
Antígenos de Neoplasias/imunologia , Antígenos de Histocompatibilidade Classe I/genética , Antígenos de Histocompatibilidade Classe I/metabolismo , Neoplasias/terapia , Evasão Tumoral/fisiologia , Animais , Linfócitos T CD4-Positivos/imunologia , Regulação para Baixo , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Imunoterapia/métodos , Células Matadoras Naturais/imunologia , Mutação , Neoplasias/imunologia
11.
J Immunol ; 207(4): 1211-1221, 2021 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-34312257

RESUMO

Long half-life of therapeutic Abs and Fc fusion proteins is crucial to their efficacy and is, in part, regulated by their interaction with neonatal Fc receptor (FcRn). However, the current methods (e.g., surface plasmon resonance and biolayer interferometry) for measurement of interaction between IgG and FcRn (IgG/FcRn) require either FcRn or IgG to be immobilized on the surface, which is known to introduce experimental artifacts and have led to conflicting data. To study IgG/FcRn interactions in solution, without a need for surface immobilization, we developed a novel (to our knowledge), solution-based homogeneous binding immunoassay based on NanoBiT luminescent protein complementation technology. We optimized the assay (NanoBiT FcRn assay) for human FcRn, mouse FcRn, rat FcRn, and cynomolgus FcRn and used them to determine the binding affinities of a panel of eight Abs. Assays could successfully capture the modulation in IgG/FcRn binding based on changes in Fc fragment of the Abs. We also looked at the individual contribution of Fc and F(ab)2 on the IgG/FcRn interaction and found that Fc is the main driver for the interaction at pH 6. Our work highlights the importance of using orthogonal methods to validate affinity data generated using biosensor platforms. Moreover, the simple add-and-read format of the NanoBiT FcRn assay is amenable for high-throughput screening during early Ab discovery phase.


Assuntos
Anticorpos Monoclonais/imunologia , Antígenos de Histocompatibilidade Classe I/imunologia , Imunoensaio/métodos , Medições Luminescentes/métodos , Receptores Fc/imunologia , Sequência de Aminoácidos , Animais , Técnicas Biossensoriais/métodos , Humanos , Fragmentos Fc das Imunoglobulinas/imunologia , Testes Imunológicos/métodos , Camundongos , Ligação Proteica/imunologia , Ratos
12.
Int J Mol Sci ; 22(13)2021 Jun 23.
Artigo em Inglês | MEDLINE | ID: mdl-34201864

RESUMO

Fetal and neonatal alloimmune thrombocytopenia (FNAIT) is a disease that causes thrombocytopenia and a risk of bleeding in the (unborn) child that result from maternal alloantibodies directed against fetal, paternally inherited, human platelet antigens (HPA). It is hypothesized that these alloantibodies can also bind to the placenta, causing placental damage. This study aims to explore signs of antibody-mediated placental damage in FNAIT. We performed a retrospective study that included pregnant women, their newborns, and placentas. It comprised 23 FNAIT cases, of which nine were newly diagnosed (14 samples) and 14 were antenatally treated with intravenous immune globulins (IVIg) (21 samples), and 20 controls, of which 10 had anti-HLA-class I antibodies. Clinical information was collected from medical records. Placental samples were stained for complement activation markers (C1q, C4d, SC5b-9, and mannose-binding lectin) using immunohistochemistry. Histopathology was examined according to the Amsterdam criteria. A higher degree of C4d deposition was present in the newly diagnosed FNAIT cases (10/14 samples), as compared to the IVIg-treated FNAIT cases (2/21 samples, p = 0.002) and anti-HLA-negative controls (3/20 samples, p = 0.006). A histopathological examination showed delayed maturation in four (44%) placentas in the newly diagnosed FNAIT cases, five (36%) in the IVIg-treated FNAIT cases, and one in the controls (NS). C4d deposition at the syncytiotrophoblast was present in combination with low-grade villitis of unknown etiology in three newly diagnosed FNAIT cases that were born SGA. We conclude that a higher degree of classical pathway-induced complement activation is present in placentas from pregnancies with untreated FNAIT. This may affect placental function and fetal growth.


Assuntos
Ativação do Complemento/imunologia , Feto/patologia , Antígenos de Histocompatibilidade Classe I/imunologia , Imunoglobulinas Intravenosas/imunologia , Placenta/patologia , Trombocitopenia Neonatal Aloimune/patologia , Adulto , Anticorpos/imunologia , Estudos de Casos e Controles , Feminino , Feto/imunologia , Humanos , Recém-Nascido , Masculino , Placenta/imunologia , Gravidez , Estudos Retrospectivos , Trombocitopenia Neonatal Aloimune/imunologia
13.
Int J Mol Sci ; 22(13)2021 Jun 22.
Artigo em Inglês | MEDLINE | ID: mdl-34206395

RESUMO

The innate immune system's natural killer (NK) cells exert their cytolytic function against a variety of pathological challenges, including tumors and virally infected cells. Their activation depends on net signaling mediated via inhibitory and activating receptors that interact with specific ligands displayed on the surfaces of target cells. The CD94/NKG2C heterodimer is one of the NK activating receptors and performs its function by interacting with the trimeric ligand comprised of the HLA-E/ß2m/nonameric peptide complex. Here, simulations of the all-atom multi-microsecond molecular dynamics in five immune complexes provide atomistic insights into the receptor-ligand molecular recognition, as well as the molecular events that facilitate the NK cell activation. We identify NKG2C, the HLA-Eα2 domain, and the nonameric peptide as the key elements involved in the molecular machinery of signal transduction via an intertwined hydrogen bond network. Overall, the study addresses the complex intricacies that are necessary to understand the mechanisms of the innate immune system.


Assuntos
Complexo Antígeno-Anticorpo/química , Antígenos de Histocompatibilidade Classe I/química , Modelos Moleculares , Subfamília C de Receptores Semelhantes a Lectina de Células NK/química , Subfamília D de Receptores Semelhantes a Lectina de Células NK/química , Peptídeos/química , Sequência de Aminoácidos , Complexo Antígeno-Anticorpo/imunologia , Complexo Antígeno-Anticorpo/metabolismo , Sítios de Ligação , Antígenos de Histocompatibilidade Classe I/imunologia , Antígenos de Histocompatibilidade Classe I/metabolismo , Humanos , Ligação de Hidrogênio , Ligantes , Subfamília C de Receptores Semelhantes a Lectina de Células NK/metabolismo , Subfamília D de Receptores Semelhantes a Lectina de Células NK/metabolismo , Peptídeos/metabolismo , Ligação Proteica , Conformação Proteica , Domínios e Motivos de Interação entre Proteínas , Transdução de Sinais , Relação Estrutura-Atividade
14.
Cell Rep ; 35(13): 109305, 2021 06 29.
Artigo em Inglês | MEDLINE | ID: mdl-34166618

RESUMO

The human leukocyte antigen (HLA)-bound viral antigens serve as an immunological signature that can be selectively recognized by T cells. As viruses evolve by acquiring mutations, it is essential to identify a range of presented viral antigens. Using HLA peptidomics, we are able to identify severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-derived peptides presented by highly prevalent HLA class I (HLA-I) molecules by using infected cells as well as overexpression of SARS-CoV-2 genes. We find 26 HLA-I peptides and 36 HLA class II (HLA-II) peptides. Among the identified peptides, some are shared between different cells and some are derived from out-of-frame open reading frames (ORFs). Seven of these peptides were previously shown to be immunogenic, and we identify two additional immunoreactive peptides by using HLA multimer staining. These results may aid the development of the next generation of SARS-CoV-2 vaccines based on presented viral-specific antigens that span several of the viral genes.


Assuntos
Antígenos Virais/imunologia , COVID-19/imunologia , COVID-19/virologia , Peptídeos/imunologia , SARS-CoV-2/imunologia , Apresentação do Antígeno , Antígenos Virais/metabolismo , Vacinas contra COVID-19 , Linhagem Celular , Epitopos de Linfócito T/imunologia , Células HEK293 , Antígenos de Histocompatibilidade Classe I/imunologia , Antígenos de Histocompatibilidade Classe II/imunologia , Humanos , Peptidomiméticos , SARS-CoV-2/genética , Linfócitos T
15.
Cell ; 184(15): 3962-3980.e17, 2021 07 22.
Artigo em Inglês | MEDLINE | ID: mdl-34171305

RESUMO

T cell-mediated immunity plays an important role in controlling SARS-CoV-2 infection, but the repertoire of naturally processed and presented viral epitopes on class I human leukocyte antigen (HLA-I) remains uncharacterized. Here, we report the first HLA-I immunopeptidome of SARS-CoV-2 in two cell lines at different times post infection using mass spectrometry. We found HLA-I peptides derived not only from canonical open reading frames (ORFs) but also from internal out-of-frame ORFs in spike and nucleocapsid not captured by current vaccines. Some peptides from out-of-frame ORFs elicited T cell responses in a humanized mouse model and individuals with COVID-19 that exceeded responses to canonical peptides, including some of the strongest epitopes reported to date. Whole-proteome analysis of infected cells revealed that early expressed viral proteins contribute more to HLA-I presentation and immunogenicity. These biological insights, as well as the discovery of out-of-frame ORF epitopes, will facilitate selection of peptides for immune monitoring and vaccine development.


Assuntos
Epitopos de Linfócito T/imunologia , Antígenos de Histocompatibilidade Classe I/imunologia , Fases de Leitura Aberta/genética , Peptídeos/imunologia , Proteoma/imunologia , SARS-CoV-2/imunologia , Células A549 , Alelos , Sequência de Aminoácidos , Animais , Apresentação do Antígeno/imunologia , COVID-19/imunologia , COVID-19/virologia , Feminino , Células HEK293 , Humanos , Cinética , Masculino , Camundongos , Peptídeos/química , Linfócitos T/imunologia
16.
Nature ; 595(7866): 309-314, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33953401

RESUMO

Epigenetic dysregulation is a defining feature of tumorigenesis that is implicated in immune escape1,2. Here, to identify factors that modulate the immune sensitivity of cancer cells, we performed in vivo CRISPR-Cas9 screens targeting 936 chromatin regulators in mouse tumour models treated with immune checkpoint blockade. We identified the H3K9 methyltransferase SETDB1 and other members of the HUSH and KAP1 complexes as mediators of immune escape3-5. We also found that amplification of SETDB1 (1q21.3) in human tumours is associated with immune exclusion and resistance to immune checkpoint blockade. SETDB1 represses broad domains, primarily within the open genome compartment. These domains are enriched for transposable elements (TEs) and immune clusters associated with segmental duplication events, a central mechanism of genome evolution6. SETDB1 loss derepresses latent TE-derived regulatory elements, immunostimulatory genes, and TE-encoded retroviral antigens in these regions, and triggers TE-specific cytotoxic T cell responses in vivo. Our study establishes SETDB1 as an epigenetic checkpoint that suppresses tumour-intrinsic immunogenicity, and thus represents a candidate target for immunotherapy.


Assuntos
Inativação Gênica , Histona-Lisina N-Metiltransferase/metabolismo , Neoplasias/genética , Neoplasias/imunologia , Animais , Antígenos Virais/imunologia , Sistemas CRISPR-Cas/genética , Cromatina/genética , Cromatina/metabolismo , Elementos de DNA Transponíveis/genética , Modelos Animais de Doenças , Feminino , Antígenos de Histocompatibilidade Classe I/genética , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Camundongos , Neoplasias/tratamento farmacológico , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Linfócitos T Citotóxicos/citologia , Linfócitos T Citotóxicos/imunologia
17.
Nat Commun ; 12(1): 2593, 2021 05 10.
Artigo em Inglês | MEDLINE | ID: mdl-33972535

RESUMO

The COVID-19 pandemic caused by SARS-CoV-2 is a continuous challenge worldwide, and there is an urgent need to map the landscape of immunogenic and immunodominant epitopes recognized by CD8+ T cells. Here, we analyze samples from 31 patients with COVID-19 for CD8+ T cell recognition of 500 peptide-HLA class I complexes, restricted by 10 common HLA alleles. We identify 18 CD8+ T cell recognized SARS-CoV-2 epitopes, including an epitope with immunodominant features derived from ORF1ab and restricted by HLA-A*01:01. In-depth characterization of SARS-CoV-2-specific CD8+ T cell responses of patients with acute critical and severe disease reveals high expression of NKG2A, lack of cytokine production and a gene expression profile inhibiting T cell re-activation and migration while sustaining survival. SARS-CoV-2-specific CD8+ T cell responses are detectable up to 5 months after recovery from critical and severe disease, and these responses convert from dysfunctional effector to functional memory CD8+ T cells during convalescence.


Assuntos
Linfócitos T CD8-Positivos/imunologia , COVID-19/imunologia , Epitopos Imunodominantes/imunologia , SARS-CoV-2/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Alelos , Linfócitos T CD8-Positivos/patologia , COVID-19/patologia , Epitopos de Linfócito T/imunologia , Feminino , Antígenos de Histocompatibilidade Classe I/genética , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Epitopos Imunodominantes/química , Memória Imunológica , Ativação Linfocitária , Masculino , Pessoa de Meia-Idade , Poliproteínas/imunologia , Proteínas Virais/imunologia
18.
Nat Commun ; 12(1): 2637, 2021 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-33976146

RESUMO

Checkpoint inhibitors and T-cell therapies have highlighted the critical role of T cells in anti-cancer immunity. However, limitations associated with these treatments drive the need for alternative approaches. Here, we engineer red blood cells into artificial antigen-presenting cells (aAPCs) presenting a peptide bound to the major histocompatibility complex I, the costimulatory ligand 4-1BBL, and interleukin (IL)-12. This leads to robust, antigen-specific T-cell expansion, memory formation, additional immune activation, tumor control, and antigen spreading in tumor models in vivo. The presence of 4-1BBL and IL-12 induces minimal toxicities due to restriction to the vasculature and spleen. The allogeneic aAPC, RTX-321, comprised of human leukocyte antigen-A*02:01 presenting the human papilloma virus (HPV) peptide HPV16 E711-19, 4-1BBL, and IL-12 on the surface, activates HPV-specific T cells and promotes effector function in vitro. Thus, RTX-321 is a potential 'off-the-shelf' in vivo cellular immunotherapy for treating HPV + cancers, including cervical and head/neck cancers.


Assuntos
Células Apresentadoras de Antígenos/transplante , Engenharia Celular/métodos , Eritrócitos/imunologia , Imunoterapia Adotiva/métodos , Neoplasias/terapia , Ligante 4-1BB/genética , Ligante 4-1BB/imunologia , Ligante 4-1BB/metabolismo , Animais , Células Apresentadoras de Antígenos/imunologia , Células Apresentadoras de Antígenos/metabolismo , Linhagem Celular Tumoral , Técnicas de Cocultura , Modelos Animais de Doenças , Eritrócitos/metabolismo , Feminino , Antígeno HLA-A2/genética , Antígeno HLA-A2/imunologia , Antígeno HLA-A2/metabolismo , Antígenos de Histocompatibilidade Classe I/genética , Antígenos de Histocompatibilidade Classe I/imunologia , Antígenos de Histocompatibilidade Classe I/metabolismo , Humanos , Interleucina-12/genética , Interleucina-12/imunologia , Interleucina-12/metabolismo , Ativação Linfocitária , Neoplasias/imunologia , Proteínas E7 de Papillomavirus/genética , Proteínas E7 de Papillomavirus/imunologia , Proteínas E7 de Papillomavirus/metabolismo , Cultura Primária de Células , Linfócitos T/imunologia , Linfócitos T/transplante , Transplante Homólogo/métodos
19.
Proc Natl Acad Sci U S A ; 118(23)2021 06 08.
Artigo em Inglês | MEDLINE | ID: mdl-34021074

RESUMO

COVID-19, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has become a global pandemic and has claimed over 2 million lives worldwide. Although the genetic sequences of SARS-CoV and SARS-CoV-2 have high homology, the clinical and pathological characteristics of COVID-19 differ significantly from those of SARS. How and whether SARS-CoV-2 evades (cellular) immune surveillance requires further elucidation. In this study, we show that SARS-CoV-2 infection leads to major histocompability complex class Ι (MHC-Ι) down-regulation both in vitro and in vivo. The viral protein encoded by open reading frame 8 (ORF8) of SARS-CoV-2, which shares the least homology with SARS-CoV among all viral proteins, directly interacts with MHC-Ι molecules and mediates their down-regulation. In ORF8-expressing cells, MHC-Ι molecules are selectively targeted for lysosomal degradation via autophagy. Thus, SARS-CoV-2-infected cells are much less sensitive to lysis by cytotoxic T lymphocytes. Because ORF8 protein impairs the antigen presentation system, inhibition of ORF8 could be a strategy to improve immune surveillance.


Assuntos
Apresentação do Antígeno , COVID-19/imunologia , Regulação para Baixo/imunologia , Antígenos de Histocompatibilidade Classe I/imunologia , Evasão da Resposta Imune , SARS-CoV-2/imunologia , Proteínas Virais/imunologia , Animais , Autofagia/genética , Autofagia/imunologia , COVID-19/genética , Chlorocebus aethiops , Células HEK293 , Antígenos de Histocompatibilidade Classe I/genética , Humanos , Lisossomos/genética , Lisossomos/imunologia , Lisossomos/virologia , Camundongos , Camundongos Transgênicos , SARS-CoV-2/genética , Células Vero , Proteínas Virais/genética
20.
Cancer Metastasis Rev ; 40(2): 501-517, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33860434

RESUMO

In recent years, immunotherapy has proven to be an effective treatment against cancer. Cytotoxic T lymphocytes perform an important role in this anti-tumor immune response, recognizing cancer cells as foreign, through the presentation of tumor antigens by MHC class I molecules. However, tumors and metastases develop escape mechanisms for evading this immunosurveillance and may lose the expression of these polymorphic molecules to become invisible to cytotoxic T lymphocytes. In other situations, they may maintain MHC class I expression and promote immunosuppression of cytotoxic T lymphocytes. Therefore, the analysis of the expression of MHC class I molecules in tumors and metastases is important to elucidate these escape mechanisms. Moreover, it is necessary to determine the molecular mechanisms involved in these alterations to reverse them and recover the expression of MHC class I molecules on tumor cells. This review discusses the role and regulation of MHC class I expression in tumor progression. We focus on altered MHC class I phenotypes present in tumors and metastases, as well as the molecular mechanisms responsible for MHC-I alterations, emphasizing the mechanisms of recovery of the MHC class I molecules expression on cancer cells. The individualized study of the HLA class I phenotype of the tumor and the metastases of each patient will allow choosing the most appropriate immunotherapy treatment based on a personalized medicine.


Assuntos
Antígenos de Histocompatibilidade Classe I/imunologia , Imunoterapia/métodos , Neoplasias/imunologia , Neoplasias/terapia , Animais , Antígenos de Histocompatibilidade Classe I/biossíntese , Humanos , Metástase Neoplásica , Neoplasias/patologia , Fenótipo , Evasão Tumoral
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