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1.
Int J Mol Sci ; 22(13)2021 Jun 22.
Artigo em Inglês | MEDLINE | ID: mdl-34206399

RESUMO

Key features of chronic lymphocytic leukemia (CLL) are defects in the immune system and the ability of leukemic cells to evade immune defenses and induce immunosuppression, resulting in increased susceptibility to infections and disease progression. Several immune effectors are impaired in CLL, including T and natural killer (NK) cells. The role of T cells in defense against CLL and in CLL progression and immunotherapy has been extensively studied. Less is known about the role of NK cells in this leukemia, and data on NK cell alterations in CLL are contrasting. Besides studies showing that NK cells have intrinsic defects in CLL, there is a large body of evidence indicating that NK cell dysfunctions in CLL mainly depend on the escape mechanisms employed by leukemic cells. In keeping, it has been shown that NK cell functions, including antibody-dependent cellular cytotoxicity (ADCC), can be retained and/or restored after adequate stimulation. Therefore, due to their preserved ADCC function and the reversibility of CLL-related dysfunctions, NK cells are an attractive source for novel immunotherapeutic strategies in this disease, including chimeric antigen receptor (CAR) therapy. Recently, satisfying clinical responses have been obtained in CLL patients using cord blood-derived CAR-NK cells, opening new possibilities for further exploring NK cells in the immunotherapy of CLL. However, notwithstanding the promising results of this clinical trial, more evidence is needed to fully understand whether and in which CLL cases NK cell-based immunotherapy may represent a valid, alternative/additional therapeutic option for this leukemia. In this review, we provide an overview of the current knowledge about phenotypic and functional alterations of NK cells in CLL and the mechanisms by which CLL cells circumvent NK cell-mediated immunosurveillance. Additionally, we discuss the potential relevance of using NK cells in CLL immunotherapy.


Assuntos
Suscetibilidade a Doenças , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Leucemia Linfocítica Crônica de Células B/etiologia , Leucemia Linfocítica Crônica de Células B/metabolismo , Biomarcadores , Comunicação Celular , Gerenciamento Clínico , Humanos , Sistema Imunitário/imunologia , Sistema Imunitário/metabolismo , Imunoterapia/efeitos adversos , Imunoterapia/métodos , Leucemia Linfocítica Crônica de Células B/diagnóstico , Leucemia Linfocítica Crônica de Células B/terapia , Ligantes , Ligação Proteica , Receptores de Células Matadoras Naturais/genética , Receptores de Células Matadoras Naturais/metabolismo , Resultado do Tratamento , Evasão Tumoral/genética , Evasão Tumoral/imunologia
2.
Int J Mol Sci ; 22(13)2021 Jun 29.
Artigo em Inglês | MEDLINE | ID: mdl-34209845

RESUMO

The vulnerability of humankind to SARS-CoV-2 in the absence of a pre-existing immunity, the unpredictability of the infection outcome, and the high transmissibility, broad tissue tropism, and ability to exploit and subvert the immune response pose a major challenge and are likely perpetuating the COVID-19 pandemic. Nevertheless, this peculiar infectious scenario provides researchers with a unique opportunity for studying, with the latest immunological techniques and understandings, the immune response in SARS-CoV-2 naïve versus recovered subjects as well as in SARS-CoV-2 vaccinees. Interestingly, the current understanding of COVID-19 indicates that the combined action of innate immune cells, cytokines, and chemokines fine-tunes the outcome of SARS-CoV-2 infection and the related immunopathogenesis. Indeed, the emerging picture clearly shows that the excessive inflammatory response against this virus is among the main causes of disease severity in COVID-19 patients. In this review, the innate immune response to SARS-CoV-2 infection is described not only in light of its capacity to influence the adaptive immune response towards a protective phenotype but also with the intent to point out the multiple strategies exploited by SARS-CoV-2 to antagonize host antiviral response and, finally, to outline inborn errors predisposing individuals to COVID-19 disease severity.


Assuntos
COVID-19/patologia , Imunidade Inata , COVID-19/imunologia , COVID-19/virologia , Quimiocinas/metabolismo , Citocinas/metabolismo , Interações Hospedeiro-Patógeno , Humanos , Células Matadoras Naturais/citologia , Células Matadoras Naturais/metabolismo , Monócitos/citologia , Monócitos/metabolismo , SARS-CoV-2/isolamento & purificação , Índice de Gravidade de Doença
3.
Front Immunol ; 12: 619069, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34108958

RESUMO

Natural killer (NK) cells are innate effector lymphocytes with strong antitumor effects against hematologic malignancies such as chronic lymphocytic leukemia (CLL). However, NK cells fail to control CLL progression on the long term. For effective lysis of their targets, NK cells use a specific cell-cell interface, known as the immunological synapse (IS), whose assembly and effector function critically rely on dynamic cytoskeletal changes in NK cells. Here we explored the role of CLL cell actin cytoskeleton during NK cell attack. We found that CLL cells can undergo fast actin cytoskeleton remodeling which is characterized by a NK cell contact-induced accumulation of actin filaments at the IS. Such polarization of the actin cytoskeleton was strongly associated with resistance against NK cell-mediated cytotoxicity and reduced amounts of the cell-death inducing molecule granzyme B in target CLL cells. Selective pharmacological targeting of the key actin regulator Cdc42 abrogated the capacity of CLL cells to reorganize their actin cytoskeleton during NK cell attack, increased levels of transferred granzyme B and restored CLL cell susceptibility to NK cell cytotoxicity. This resistance mechanism was confirmed in primary CLL cells from patients. In addition, pharmacological inhibition of actin dynamics in combination with blocking antibodies increased conjugation frequency and improved CLL cell elimination by NK cells. Together our results highlight the critical role of CLL cell actin cytoskeleton in driving resistance against NK cell cytotoxicity and provide new potential therapeutic point of intervention to target CLL immune escape.


Assuntos
Citoesqueleto de Actina/metabolismo , Citotoxicidade Imunológica , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Leucemia Linfocítica Crônica de Células B/imunologia , Leucemia Linfocítica Crônica de Células B/metabolismo , Proteína cdc42 de Ligação ao GTP/antagonistas & inibidores , Citoesqueleto de Actina/efeitos dos fármacos , Biomarcadores , Linhagem Celular Tumoral , Citotoxicidade Imunológica/efeitos dos fármacos , Imunofluorescência , Antígenos HLA-G/imunologia , Humanos , Sinapses Imunológicas/imunologia , Sinapses Imunológicas/metabolismo , Imunofenotipagem , Células Matadoras Naturais/efeitos dos fármacos , Leucemia Linfocítica Crônica de Células B/tratamento farmacológico , Leucemia Linfocítica Crônica de Células B/patologia , Microambiente Tumoral/genética , Microambiente Tumoral/imunologia
4.
Nat Commun ; 12(1): 3501, 2021 06 09.
Artigo em Inglês | MEDLINE | ID: mdl-34108465

RESUMO

The characteristics of COVID-19 patients with persistent SARS-CoV-2 infection are not yet well described. Here, we compare the clinical and molecular features of patients with long duration of viral shedding (LDs) with those from patients with short duration patients (SDs), and healthy donors (HDs). We find that several cytokines and chemokines, such as interleukin (IL)-2, tumor necrosis factor (TNF) and lymphotoxin α (LT-α) are present at lower levels in LDs than SDs. Single-cell RNA sequencing shows that natural killer (NK) cells and CD14+ monocytes are reduced, while regulatory T cells are increased in LDs; moreover, T and NK cells in LDs are less activated than in SDs. Importantly, most cells in LDs show reduced expression of ribosomal protein (RP) genes and related pathways, with this inversed correlation between RP levels and infection duration further validated in 103 independent patients. Our results thus indicate that immunosuppression and low RP expression may be related to the persistence of the viral infection in COVID-19 patients.


Assuntos
COVID-19/imunologia , SARS-CoV-2/patogenicidade , Linfócitos B/metabolismo , Linfócitos B/patologia , COVID-19/virologia , Citocinas/sangue , Perfilação da Expressão Gênica , Humanos , Células Matadoras Naturais/metabolismo , Células Matadoras Naturais/patologia , Leucócitos Mononucleares/patologia , Ativação Linfocitária/genética , Subpopulações de Linfócitos/metabolismo , Subpopulações de Linfócitos/patologia , Proteínas Ribossômicas/genética , SARS-CoV-2/isolamento & purificação , Transdução de Sinais/genética , Linfócitos T/metabolismo , Linfócitos T/patologia , Eliminação de Partículas Virais
5.
Nat Commun ; 12(1): 3655, 2021 06 16.
Artigo em Inglês | MEDLINE | ID: mdl-34135341

RESUMO

RNA in extracellular vesicles (EVs) are uptaken by cells, where they regulate fundamental cellular functions. EV-derived mRNA in recipient cells can be translated. However, it is still elusive whether "naked nonvesicular extracellular mRNA" (nex-mRNA) that are not packed in EVs can be uptaken by cells and, if so, whether they have any functions in recipient cells. Here, we show the entrance of nex-mRNA in the nucleus, where they exert a translation-independent function. Human nex-interleukin-1ß (IL1ß)-mRNA outside cells proved to be captured by RNA-binding zinc finger CCCH domain containing protein 12D (ZC3H12D)-expressing human natural killer (NK) cells. ZC3H12D recruited to the cell membrane binds to the 3'-untranslated region of nex-IL1ß-mRNA and transports it to the nucleus. The nex-IL1ß-mRNA in the NK cell nucleus upregulates antiapoptotic gene expression, migration activity, and interferon-γ production, leading to the killing of cancer cells and antimetastasis in mice. These results implicate the diverse actions of mRNA.


Assuntos
Núcleo Celular/metabolismo , Espaço Extracelular/metabolismo , RNA Mensageiro/metabolismo , Regiões 3' não Traduzidas , Animais , Antineoplásicos/farmacologia , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Meios de Cultivo Condicionados/metabolismo , Endorribonucleases/metabolismo , Humanos , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Células Matadoras Naturais/metabolismo , Camundongos , Ligação Proteica , RNA Mensageiro/genética , RNA Mensageiro/farmacologia , Proteínas de Ligação a RNA/metabolismo
6.
Int J Mol Sci ; 22(11)2021 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-34072732

RESUMO

CAR-T (chimeric antigen receptor T) cells have emerged as a milestone in the treatment of patients with refractory B-cell neoplasms. However, despite having unprecedented efficacy against hematological malignancies, the treatment is far from flawless. Its greatest drawbacks arise from a challenging and expensive production process, strict patient eligibility criteria and serious toxicity profile. One possible solution, supported by robust research, is the replacement of T lymphocytes with NK cells for CAR expression. NK cells seem to be an attractive vehicle for CAR expression as they can be derived from multiple sources and safely infused regardless of donor-patient matching, which greatly reduces the cost of the treatment. CAR-NK cells are known to be effective against hematological malignancies, and a growing number of preclinical findings indicate that they have activity against non-hematological neoplasms. Here, we present a thorough overview of the current state of knowledge regarding the use of CAR-NK cells in treating various solid tumors.


Assuntos
Imunoterapia Adotiva , Células Matadoras Naturais/imunologia , Neoplasias/terapia , Receptores de Antígenos de Linfócitos T/imunologia , Animais , Antígenos de Neoplasias/imunologia , Técnicas de Cultura de Células , Ensaios Clínicos como Assunto , Terapia Combinada/métodos , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Engenharia Genética , Humanos , Células Matadoras Naturais/metabolismo , Neoplasias/diagnóstico , Neoplasias/etiologia , Neoplasias/metabolismo , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos Quiméricos/genética , Receptores de Antígenos Quiméricos/imunologia , Resultado do Tratamento
7.
Nat Commun ; 12(1): 3908, 2021 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-34162850

RESUMO

Though AsCas12a fills a crucial gap in the current genome editing toolbox, it exhibits relatively poor editing efficiency, restricting its overall utility. Here we isolate an engineered variant, "AsCas12a Ultra", that increased editing efficiency to nearly 100% at all sites examined in HSPCs, iPSCs, T cells, and NK cells. We show that AsCas12a Ultra maintains high on-target specificity thereby mitigating the risk for off-target editing and making it ideal for complex therapeutic genome editing applications. We achieved simultaneous targeting of three clinically relevant genes in T cells at >90% efficiency and demonstrated transgene knock-in efficiencies of up to 60%. We demonstrate site-specific knock-in of a CAR in NK cells, which afforded enhanced anti-tumor NK cell recognition, potentially enabling the next generation of allogeneic cell-based therapies in oncology. AsCas12a Ultra is an advanced CRISPR nuclease with significant advantages in basic research and in the production of gene edited cell medicines.


Assuntos
Acidaminococcus/enzimologia , Proteínas de Bactérias/metabolismo , Proteínas Associadas a CRISPR/metabolismo , Sistemas CRISPR-Cas , Endonucleases/metabolismo , Edição de Genes/métodos , Acidaminococcus/genética , Proteínas de Bactérias/genética , Proteínas Associadas a CRISPR/genética , Células Cultivadas , Endonucleases/genética , Células HEK293 , Células-Tronco Hematopoéticas/metabolismo , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Células Jurkat , Células Matadoras Naturais/metabolismo , Reprodutibilidade dos Testes , Linfócitos T/metabolismo
8.
FEBS Lett ; 595(13): 1797-1805, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33982279

RESUMO

Fatty acid-binding protein (FABP) 5 is highly expressed in various types of tumors and is strongly correlated with tumor growth, development, and metastasis. However, it is unclear how the expression of FABP5 in the host affects tumor progression. In this study, using a lung tumor metastasis model in mice, we found that FABP5-deficient mice were more susceptible to tumor metastasis, which is accompanied by infiltration of a lower frequency of activated natural killer (NK) cells in the lung. Additionally, FABP5 deficiency leads to impaired maturation of NK cells in the lungs, but not in the bone marrow and spleen. Taken together, our results provide the first evidence that FABP5 in the host regulates lung tumor metastasis through controlling NK cell maturation.


Assuntos
Proteínas de Ligação a Ácido Graxo/genética , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/secundário , Linfócitos do Interstício Tumoral/metabolismo , Melanoma/genética , Proteínas de Neoplasias/genética , Animais , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Células Matadoras Naturais/metabolismo , Metabolismo dos Lipídeos , Pulmão/imunologia , Pulmão/patologia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/imunologia , Melanoma/imunologia , Melanoma/patologia , Camundongos , Transplante de Neoplasias , Fatores de Transcrição/genética
9.
Cancer Sci ; 112(7): 2664-2678, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33934451

RESUMO

Immunotherapy targeting the PD-L1/PD-1 pathway is a novel type of clinical cancer treatment, but only small subsets of patients can benefit from it because of multiple factors. PD-L1/PD-1 expression is a biomarker for predicting the efficacy of anti-PD-L1/PD-1 therapy, which highlights the importance of understanding the regulatory mechanisms of PD-L1 expression in cancer cells. Casp8 is an apical caspase protease involved in mediating cell apoptosis, but it also has multiple nonapoptotic functions. Casp8 mutations are associated with increased risks of cancer, and low expression of Casp8 is closely connected with poor prognosis in patients with cancer. In addition, mutations of Casp8 in lymphocytes also lead to human immunodeficiency, thereby causing dysfunction of the innate immune system, but the roles of Casp8 in antitumor immunity remain unclear. Here, we found that knocking down Casp8 in mouse melanoma cells promoted tumor progression in an immune system-dependent manner. Mechanistically, Casp8 induced PD-L1 degradation by upregulating TNFAIP3 (A20) expression, a ubiquitin-editing enzyme that results in PD-L1 ubiquitination. In addition, compared with Casp8fl/fl mice, mice with conditional deletion of Casp8 in natural killer (NK) cells (Ncr1iCre/+ Casp8fl/fl mice) showed a decreased frequency of IFN-γ+ and CD107a+ NK cells but an increased frequency of PD-1+ and CTLA-4+ NK cells. Melanoma cells with Casp8 knocked down exhibited sensitivity to anti-PD-1 or anti-CTLA-4 antibody treatments, particularly in Ncr1iCre/+Casp8fl/fl mice. Together, the results indicate that Casp8 induces PD-L1 degradation by upregulating A20 expression and that decreased Casp8 expression is a potential biomarker for predicting the sensitivity to anti-PD-L1/PD-1 immunotherapy.


Assuntos
Antígeno B7-H1/metabolismo , Caspase 8/fisiologia , Imunoterapia Adotiva/métodos , Melanoma/terapia , Proteína 3 Induzida por Fator de Necrose Tumoral alfa/metabolismo , Animais , Antígeno B7-H1/genética , Antígeno CTLA-4/metabolismo , Caspase 8/genética , Linhagem Celular Tumoral , Progressão da Doença , Regulação para Baixo , Proteínas Ativadoras de GTPase/metabolismo , Interferon gama/metabolismo , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Glicoproteínas de Membrana Associadas ao Lisossomo/metabolismo , Melanoma/imunologia , Melanoma/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Nus , NF-kappa B/metabolismo , Ubiquitinação , Regulação para Cima
10.
Brain Behav Immun ; 95: 477-488, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33989745

RESUMO

Physical and social environments influence immune homeostasis within adipose tissue, yet the mechanisms remain poorly defined. We report that an enriched environment (EE) housing modulates the immune cell population in white adipose tissue of mice including an increase in the abundance of natural killer (NK) cells. EE upregulates the expression of IL-15 and its receptor IL-15Rα specifically within mature adipocytes. Mechanistically, we show that hypothalamic brain-derived neurotrophic factor (BDNF) upregulates IL-15 production in adipocytes via sympathetic ß-adrenergic signaling. Overexpressing BDNF mediated by recombinant adeno-associated virus (rAAV) vector in the hypothalamus expands adipose NK cells. Conversely, inhibition of hypothalamic BDNF signaling via gene transfer of a dominant negative TrkB receptor suppresses adipose NK cells. In white adipose tissue, overexpression of IL-15 using an adipocyte-specific rAAV vector stimulates adipose NK cells and inhibits the progression of subcutaneous melanoma, whereas local IL-15 knockdown blocks the EE effect. These results suggest that bio-behavioral factors regulate adipose NK cells via a hypothalamic BDNF-sympathoneural-adipocyte IL-15 axis. Targeting this pathway may have therapeutic significance for cancer.


Assuntos
Adipócitos , Fator Neurotrófico Derivado do Encéfalo , Interleucina-15 , Células Matadoras Naturais , Adipócitos/metabolismo , Tecido Adiposo/metabolismo , Animais , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Hipotálamo/metabolismo , Interleucina-15/metabolismo , Células Matadoras Naturais/metabolismo , Camundongos , Camundongos Endogâmicos C57BL
11.
Front Immunol ; 12: 609762, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33968014

RESUMO

Cancer is one of the leading causes of death and a major public health problem all over the world. Immunotherapy is becoming a revolutionary clinical management for various cancer types. Restoration of aberrant immune surveillance on cancers has achieved markable progress in the past years by either in vivo or ex vivo engineering of the immune cells. Here, we summarized the central roles of immune cells in tumor progression and regression, and the existing and emerging strategies for different immune cell-based immunotherapies. In addition, the current challenges and the potential solutions in translating the immunotherapies into the clinic are also discussed.


Assuntos
Reprogramação Celular/imunologia , Imunomodulação , Neoplasias/imunologia , Neoplasias/terapia , Animais , Reprogramação Celular/genética , Engenharia Genética , Humanos , Imunoterapia/efeitos adversos , Imunoterapia/métodos , Imunoterapia Adotiva/efeitos adversos , Imunoterapia Adotiva/métodos , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/metabolismo , Neoplasias/diagnóstico , Neoplasias/metabolismo , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/imunologia , Receptores de Antígenos Quiméricos/genética , Receptores de Antígenos Quiméricos/imunologia , Linfócitos T/imunologia , Linfócitos T/metabolismo , Microambiente Tumoral/genética , Microambiente Tumoral/imunologia , Macrófagos Associados a Tumor/imunologia , Macrófagos Associados a Tumor/metabolismo
12.
Biomed Res Int ; 2021: 6673427, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33997038

RESUMO

Results: KIR2DL1 and ILT-2 expression on idNK cells was higher in healthy women than in RPL patients. Sildenafil enhanced NKG2A expression in RPL patients. VEGF concentration was higher in fertile woman idNK cell cultures. idNK cells were more sensitive for necrosis in RPL than in fertile women. SC did not influence VEGF production or idNK cell apoptosis. Conclusions: A combination of hypoxia, IL-15, and AZA promotes the conversion of pbNK into idNK cells CD56+CD16--expressing KIR receptors and produces VEGF. Alterations in KIR2DL1 and ILT-2 expression as well as impaired VEGF production were associated with RPL. SC affects NKG2A expression on RPL idNK cells. SC had no effect on VEGF release or idNK cell apoptosis.


Assuntos
Aborto Habitual , Antígenos CD/análise , Células Matadoras Naturais , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/análise , Receptores KIR2DL1/análise , Fator A de Crescimento do Endotélio Vascular/análise , Aborto Habitual/sangue , Aborto Habitual/metabolismo , Adulto , Antígenos CD/metabolismo , Apoptose , Células Cultivadas , Feminino , Humanos , Células Matadoras Naturais/citologia , Células Matadoras Naturais/metabolismo , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/metabolismo , Receptores KIR2DL1/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo
13.
J Med Chem ; 64(11): 7371-7389, 2021 06 10.
Artigo em Inglês | MEDLINE | ID: mdl-34029463

RESUMO

The previous virtual screening of ten million compounds yielded two novel nonlipopeptide-like chemotypes as TLR2 agonists. Herein, we present the chemical optimization of our initial hit, 1-phenyl-3-(thiophen-2-yl)urea, which resulted in the identification of SMU-C80 (EC50 = 31.02 ± 1.01 nM) as a TLR2-specific agonist with a 370-fold improvement in bioactivity. Mechanistic studies revealed that SMU-C80, through TLR1/2, recruits the adaptor protein MyD88 and triggers the NF-κB pathway to release cytokines such as TNF-α and IL-1ß from human, but not murine, cells. To the best of our knowledge, it is the first species-specific TLR1/2 agonist reported until now. Moreover, SMU-C80 increased the percentage of T, B, and NK cells ex vivo and activated the immune cells, which suppressed cancer cell growth in vitro. In summary, we obtained a highly efficient and specific human TLR1/2 agonist that acts through the MyD88 and NF-κB pathway, facilitating cytokine release and the simultaneous activation of immune cells that in turn affects the apoptosis of cancer cells.


Assuntos
Desenho de Fármacos , Tioureia/análogos & derivados , Receptor 1 Toll-Like/agonistas , Receptor 2 Toll-Like/agonistas , Animais , Apoptose/efeitos dos fármacos , Citocinas/metabolismo , Humanos , Imunoterapia , Células Matadoras Naturais/citologia , Células Matadoras Naturais/efeitos dos fármacos , Células Matadoras Naturais/metabolismo , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Fator 88 de Diferenciação Mieloide/metabolismo , NF-kappa B/metabolismo , Neoplasias/terapia , Transdução de Sinais/efeitos dos fármacos , Relação Estrutura-Atividade , Linfócitos T/citologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/metabolismo , Tioureia/metabolismo , Tioureia/uso terapêutico , Receptor 1 Toll-Like/metabolismo , Receptor 2 Toll-Like/metabolismo
14.
Front Immunol ; 12: 532484, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33897679

RESUMO

Human cytomegalovirus (HCMV) infection often leads to systemic disease in immunodeficient patients and congenitally infected children. Despite its clinical significance, the exact mechanisms contributing to HCMV pathogenesis and clinical outcomes have yet to be determined. One of such mechanisms involves HCMV-mediated NK cell immune response, which favors viral immune evasion by hindering NK cell-mediated cytolysis. This process appears to be dependent on the extent of HCMV genetic variation as high levels of variability in viral genes involved in immune escape have an impact on viral pathogenesis. However, the link between viral genome variations and their functional effects has so far remained elusive. Thus, here we sought to determine whether inter-host genetic variability of HCMV influences its ability to modulate NK cell responses to infection. For this purpose, five HCMV clinical isolates from a previously characterized cohort of pediatric patients with confirmed HCMV congenital infection were evaluated by next-generation sequencing (NGS) for genetic polymorphisms, phylogenetic relationships, and multiple-strain infection. We report variable levels of genetic characteristics among the selected clinical strains, with moderate variations in genome regions associated with modulation of NK cell functions. Remarkably, we show that different HCMV clinical strains differentially modulate the expression of several ligands for the NK cell-activating receptors NKG2D, DNAM-1/CD226, and NKp30. Specifically, the DNAM-1/CD226 ligand PVR/CD155 appears to be predominantly upregulated by fast-replicating ("aggressive") HCMV isolates. On the other hand, the NGK2D ligands ULBP2/5/6 are downregulated regardless of the strain used, while other NK cell ligands (i.e., MICA, MICB, ULBP3, Nectin-2/CD112, and B7-H6) are not significantly modulated. Furthermore, we show that IFN-γ; production by NK cells co-cultured with HCMV-infected fibroblasts is directly proportional to the aggressiveness of the HCMV clinical isolates employed. Interestingly, loss of NK cell-modulating genes directed against NK cell ligands appears to be a common feature among the "aggressive" HCMV strains, which also share several gene variants across their genomes. Overall, even though further studies based on a higher number of patients would offer a more definitive scenario, our findings provide novel mechanistic insights into the impact of HCMV genetic variability on NK cell-mediated immune responses.


Assuntos
Infecções por Citomegalovirus/imunologia , Citomegalovirus/imunologia , Peptídeos e Proteínas de Sinalização Intercelular/imunologia , Interferon gama/imunologia , Células Matadoras Naturais/imunologia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/imunologia , Células Cultivadas , Citomegalovirus/genética , Citomegalovirus/fisiologia , Infecções por Citomegalovirus/metabolismo , Infecções por Citomegalovirus/virologia , Citotoxicidade Imunológica/genética , Citotoxicidade Imunológica/imunologia , Proteínas Ligadas por GPI/genética , Proteínas Ligadas por GPI/imunologia , Proteínas Ligadas por GPI/metabolismo , Expressão Gênica , Variação Genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Evasão da Resposta Imune/genética , Evasão da Resposta Imune/imunologia , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Interferon gama/metabolismo , Células Matadoras Naturais/metabolismo , Células Matadoras Naturais/virologia , Ligantes , Masculino , Subfamília K de Receptores Semelhantes a Lectina de Células NK/genética , Subfamília K de Receptores Semelhantes a Lectina de Células NK/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa
15.
Front Immunol ; 12: 610789, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33815365

RESUMO

Natural killer (NK) cells are lymphocytes primarily involved in innate immunity and possess important functional properties in anti-viral and anti-tumor responses; thus, these cells have broad potential for clinical utilization. NK cells originate from hematopoietic stem cells (HSCs) through the following two independent and continuous processes: early commitment from HSCs to IL-15-responsive NK cell progenitors (NKPs) and subsequent differentiation into mature NK cells in response to IL-15. IL-15 is the most important cytokine for NK cell development, is produced by both hematopoietic and nonhematopoietic cells, and functions through a distinct delivery process termed transpresentation. Upon being transpresented to NK cells, IL-15 contributes to NK cell development via the activation of several downstream signaling pathways, including the Ras-MEK-MAPK, JAK-STAT5, and PI3K-ATK-mTOR pathways. Nonetheless, the exact role of IL-15 in NK cell development has not been discussed in a consecutive and comprehensive manner. Here, we review current knowledge about the indispensable role of IL-15 in NK cell development and address which cells produce IL-15 to support NK cell development and when IL-15 exerts its function during multiple developmental stages. Specifically, we highlight how IL-15 supports NK cell development by elucidating the distinct transpresentation of IL-15 to NK cells and revealing the downstream target of IL-15 signaling during NK cell development.


Assuntos
Diferenciação Celular , Interleucina-15/metabolismo , Células Matadoras Naturais/citologia , Células Matadoras Naturais/metabolismo , Fatores de Transcrição/metabolismo , Animais , Biomarcadores , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Regulação da Expressão Gênica , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Humanos , Interleucina-15/genética , Células Matadoras Naturais/imunologia , Linfopoese/genética , Linfopoese/imunologia , Transdução de Sinais , Fatores de Transcrição/genética
16.
Transplant Proc ; 53(5): 1423-1432, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-33888343

RESUMO

INTRODUCTION: The usage of extended-criteria donors (ECD) became a routinely accepted manner in the last decade. ECD is a potential risk factor for antibody-mediated rejection. Analysis of lymphocyte subsets might be a complementary diagnostic toolkit because there is limited knowledge about this term. METHOD: Between May 12, 2016, and September 4, 2019, a total of 130 patients who had undergone kidney transplant were investigated. Patients were divided in ECD and standard criteria donor (SCD) groups. Blood samples were collected before the operation, then in the first week and after 30, 60, 180, and 365 days. Besides routine laboratory tests, multicolor flow cytometry was performed for lymphocyte subsets. RESULTS: ECD grafts were transplanted to older recipients. The number of CD4+ cells increased in the SCDs from the first week to until the end of first month, and then decreased. The number of CD4+ cells decreased from the beginning of the study until the end of first year to 66% of its original value in ECDs. At the first month, the number of CD19+ cells was higher in SCD compared with ECD cases; the number then decreased in both groups. T-regulatory cells had a drop at the first week that lasted until the first month. A bigger increase in SCD and a moderate increase in ECD group were then observed. The kinetics of CD19+ and CD19+ naive cells are similar in the ECD and SCD groups. In the SCD group, cell count decreased in both CD19+ (13%) and CD19+ naive (12%) between third and sixth month. The count of CD19+ cells decreased by 9%, but the count of CD19+ naive cells increased by 11% between the sixth month and first year. DISCUSSION: The prolonged postoperative uremic state caused by the poorer initial function, together with an aging immune system, explains the weaker immune response in ECD patients, which may be the cause of the decreased number of memory and regulatory T cells. Older patients with an ECD graft need a tailored, personalized, and less aggressive immunosuppressive treatment.


Assuntos
Subpopulações de Linfócitos T/metabolismo , Adulto , Idoso , Antígenos CD19/metabolismo , Linfócitos B/citologia , Linfócitos B/metabolismo , Feminino , Humanos , Transplante de Rim , Células Matadoras Naturais/citologia , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Cinética , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Subpopulações de Linfócitos T/citologia , Subpopulações de Linfócitos T/imunologia , Fatores de Tempo , Doadores de Tecidos , Transplantados
17.
Front Immunol ; 12: 608625, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33790890

RESUMO

B-cell receptors, multiple receptor tyrosine kinases, and downstream effectors are constitutively active in chronic lymphocytic leukemia (CLL) B cells. Activation of these pathways results in resistance to apoptosis and enhanced survival of the leukemic cells. Idelalisib is a highly selective inhibitor of the PI3K p110∂ isoform and is approved for the treatment of CLL in patients with relapsed/refractory disease or in those harboring 17p deletions or tp53 mutations. Despite the initial excitement centered around high response rates in clinical trials of idelalisib, its therapeutic success has been hindered by the incidence of severe opportunistic infections. To examine the potential contribution of idelalisib to the increased risk of infection, we investigated the effects of idelalisib on the immune cell compartments of healthy donors (HDs) and CLL patients. PI3K∂ blockade by idelalisib reduced the expression levels of inhibitory checkpoint molecules in T cells isolated from both HDs and CLL patients. In addition, the presence of idelalisib in cultures significantly decreased T-cell-mediated cytotoxicity and granzyme B secretion, as well as cytokine secretion levels in both cohorts. Furthermore, idelalisib reduced the proliferation and cytotoxicity of HD NK cells. Collectively, our data demonstrate that both human T and NK cells are highly sensitive to PI3K∂ inhibition. Idelalisib interfered with the functions of T and NK cell cells from both HDs and CLL patients. Therefore, idelalisib might contribute to an increased risk of infections regardless of the underlying B-cell malignancy.


Assuntos
Antineoplásicos/farmacologia , Hospedeiro Imunocomprometido , Células Matadoras Naturais/efeitos dos fármacos , Leucemia Linfocítica Crônica de Células B/imunologia , Inibidores de Fosfoinositídeo-3 Quinase/farmacologia , Purinas/farmacologia , Quinazolinonas/farmacologia , Linfócitos T/efeitos dos fármacos , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais , Aberrações Cromossômicas , Citocinas/metabolismo , Feminino , Humanos , Proteínas de Checkpoint Imunológico/genética , Proteínas de Checkpoint Imunológico/metabolismo , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Leucemia Linfocítica Crônica de Células B/diagnóstico , Leucemia Linfocítica Crônica de Células B/genética , Ativação Linfocitária/efeitos dos fármacos , Ativação Linfocitária/imunologia , Masculino , Pessoa de Meia-Idade , Mutação , Linfócitos T/imunologia , Linfócitos T/metabolismo
18.
J Infect Dis ; 224(2): 229-240, 2021 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-33928374

RESUMO

BACKGROUND: Etiopathogenesis of the clinical variability of the coronavirus disease 2019 (COVID-19) remains mostly unknown. In this study, we investigate the role of killer cell immunoglobulin-like receptor (KIR)/human leukocyte antigen class-I (HLA-I) interactions in the susceptibility and severity of COVID-19. METHODS: We performed KIR and HLA-I genotyping and natural killer cell (NKc) receptors immunophenotyping in 201 symptomatic patients and 210 noninfected controls. RESULTS: The NKcs with a distinctive immunophenotype, suggestive of recent activation (KIR2DS4low CD16low CD226low CD56high TIGIThigh NKG2Ahigh), expanded in patients with severe COVID-19. This was associated with a higher frequency of the functional A-telomeric activating KIR2DS4 in severe versus mild and/or moderate patients and controls (83.7%, 55.7% and 36.2%, P < 7.7 × 10-9). In patients with mild and/or moderate infection, HLA-B*15:01 was associated with higher frequencies of activating B-telomeric KIR3DS1 compared with patients with other HLA-B*15 subtypes and noninfected controls (90.9%, 42.9%, and 47.3%; P < .002; Pc = 0.022). This strongly suggests that HLA-B*15:01 specifically presenting severe acute respiratory syndrome coronavirus 2 peptides could form a neoligand interacting with KIR3DS1. Likewise, a putative neoligand for KIR2DS4 could arise from other HLA-I molecules presenting severe acute respiratory syndrome coronavirus 2 peptides expressed on infected an/or activated lung antigen-presenting cells. CONCLUSIONS: Our results support a crucial role of NKcs in the clinical variability of COVID-19 with specific KIR/ligand interactions associated with disease severity.


Assuntos
COVID-19/genética , Predisposição Genética para Doença/genética , Receptores KIR/genética , Idoso , COVID-19/imunologia , COVID-19/patologia , Estudos Transversais , Feminino , Genótipo , Antígenos HLA/genética , Antígenos HLA/metabolismo , Humanos , Imunofenotipagem , Células Matadoras Naturais/metabolismo , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Receptores KIR/metabolismo , SARS-CoV-2 , Índice de Gravidade de Doença
19.
Nat Immunol ; 22(5): 627-638, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33859404

RESUMO

Cytokine signaling via signal transducer and activator of transcription (STAT) proteins is crucial for optimal antiviral responses of natural killer (NK) cells. However, the pleiotropic effects of both cytokine and STAT signaling preclude the ability to precisely attribute molecular changes to specific cytokine-STAT modules. Here, we employed a multi-omics approach to deconstruct and rebuild the complex interaction of multiple cytokine signaling pathways in NK cells. Proinflammatory cytokines and homeostatic cytokines formed a cooperative axis to commonly regulate global gene expression and to further repress expression induced by type I interferon signaling. These cytokines mediated distinct modes of epigenetic regulation via STAT proteins, and collective signaling best recapitulated global antiviral responses. The most dynamically responsive genes were conserved across humans and mice, which included a cytokine-STAT-induced cross-regulatory program. Thus, an intricate crosstalk exists between cytokine signaling pathways, which governs NK cell responses.


Assuntos
Epigênese Genética/imunologia , Infecções por Herpesviridae/imunologia , Interleucinas/metabolismo , Células Matadoras Naturais/imunologia , Fatores de Transcrição STAT/metabolismo , Animais , Separação Celular , Sequenciamento de Cromatina por Imunoprecipitação , Modelos Animais de Doenças , Feminino , Citometria de Fluxo , Redes Reguladoras de Genes/imunologia , Infecções por Herpesviridae/sangue , Infecções por Herpesviridae/virologia , Humanos , Imunidade Inata/genética , Células Matadoras Naturais/metabolismo , Masculino , Camundongos , Camundongos Knockout , Muromegalovirus/imunologia , Análise de Componente Principal , RNA-Seq , Fatores de Transcrição STAT/genética , Transdução de Sinais/genética , Transdução de Sinais/imunologia
20.
Int J Mol Sci ; 22(7)2021 Mar 28.
Artigo em Inglês | MEDLINE | ID: mdl-33800561

RESUMO

Glioblastoma (GBM) is the most common and aggressive primary malignant brain tumor in adults. Natural Killer (NK) cells are potent cytotoxic effector cells against tumor cells inducing GBM cells; therefore, NK cell based- immunotherapy might be a promising target in GBM. T cell immunoglobulin mucin family member 3 (TIM3), a receptor expressed on NK cells, has been suggested as a marker of dysfunctional NK cells. We established TIM3 knockout in NK cells, using the clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein 9 (Cas9). Electroporating of TIM3 exon 2- or exon 5-targeting guide RNA- Cas9 protein complexes (RNPs) inhibited TIM3 expression on NK cells with varying efficacy. T7 endonuclease I mutation detection assays showed that both RNPs disrupted the intended genome sites. The expression of other checkpoint receptors, i.e., programmed cell death 1 (PD1), Lymphocyte-activation gene 3 (LAG3), T cell immunoreceptor with Ig and ITIM domains (TIGIT), and TACTILE (CD96) were unchanged on the TIM3 knockout NK cells. Real time cell growth assays revealed that TIM3 knockout enhanced NK cell-mediated growth inhibition of GBM cells. These results demonstrated that TIM3 knockout enhanced human NK cell mediated cytotoxicity on GBM cells. Future, CRISPR-Cas9 mediated TIM3 knockout in NK cells may prove to be a promising immunotherapeutic alternative in patient with GBM.


Assuntos
Neoplasias Encefálicas/genética , Sistemas CRISPR-Cas , Regulação Neoplásica da Expressão Gênica , Glioma/genética , Receptor Celular 2 do Vírus da Hepatite A/genética , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/terapia , Linhagem Celular Tumoral , Técnicas de Inativação de Genes , Genoma Humano , Glioma/metabolismo , Glioma/terapia , Humanos , Imunoterapia/métodos , Estimativa de Kaplan-Meier , Células Matadoras Naturais/metabolismo , Ligantes , Análise de Sequência com Séries de Oligonucleotídeos , RNA Guia/metabolismo , Transgenes
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