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1.
Int J Mol Sci ; 22(14)2021 Jul 09.
Artigo em Inglês | MEDLINE | ID: mdl-34299013

RESUMO

Mucosal CD4+ T lymphocytes display a potent opioid-mediated analgesic activity in interleukin (IL)-10 knockout mouse model of inflammatory bowel diseases (IBD). Considering that endogenous opioids may also exhibit anti-inflammatory activities in the periphery, we examined the consequences of a peripheral opioid receptor blockade by naloxone-methiodide, a general opioid receptor antagonist unable to cross the blood-brain barrier, on the development of piroxicam-accelerated colitis in IL-10-deficient (IL-10-/-) mice. Here, we show that IL-10-deficient mice treated with piroxicam exhibited significant alterations of the intestinal barrier function, including permeability, inflammation-related bioactive lipid mediators, and mucosal CD4+ T lymphocyte subsets. Opioid receptor antagonization in the periphery had virtually no effect on colitis severity but significantly worsened epithelial cell apoptosis and intestinal permeability. Thus, although the endogenous opioid tone is not sufficient to reduce the severity of colitis significantly, it substantially contributes to the protection of the physical integrity of the epithelial barrier.


Assuntos
Colite/metabolismo , Interleucina-10/genética , Mucosa Intestinal/efeitos dos fármacos , Naloxona/análogos & derivados , Antagonistas de Entorpecentes/administração & dosagem , Piroxicam/farmacologia , Receptores Opioides/metabolismo , Animais , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios não Esteroides/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/genética , Linfócitos T CD4-Positivos/efeitos dos fármacos , Colite/induzido quimicamente , Colite/genética , Colite/patologia , Citocinas/genética , Citocinas/metabolismo , Células Epiteliais/efeitos dos fármacos , Inflamação/genética , Inflamação/metabolismo , Inflamação/patologia , Interleucina-10/metabolismo , Mucosa Intestinal/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Naloxona/farmacologia , Permeabilidade/efeitos dos fármacos , Compostos de Amônio Quaternário/farmacologia , Índice de Gravidade de Doença
2.
Int J Mol Sci ; 22(13)2021 Jun 29.
Artigo em Inglês | MEDLINE | ID: mdl-34209797

RESUMO

Macrophages play critical roles in both innate and adaptive immunity and are known for their high plasticity in response to various external signals. Macrophages are involved in regulating systematic iron homeostasis and they sequester iron by phagocytotic activity, which triggers M1 macrophage polarization and typically exerts antitumor effects. We previously developed a novel cryo-thermal therapy that can induce the mass release of tumor antigens and damage-associated molecular patterns (DAMPs), promoting M1 macrophage polarization. However, that study did not examine whether iron released after cryo-thermal therapy induced M1 macrophage polarization; this question still needed to be addressed. We hypothesized that cryo-thermal therapy would cause the release of a large quantity of iron to augment M1 macrophage polarization due to the disruption of tumor cells and blood vessels, which would further enhance antitumor immunity. In this study, we investigated iron released in primary tumors, the level of iron in splenic macrophages after cryo-thermal therapy and the effect of iron on macrophage polarization and CD4+ T cell differentiation in metastatic 4T1 murine mammary carcinoma. We found that a large amount of iron was released after cryo-thermal therapy and could be taken up by splenic macrophages, which further promoted M1 macrophage polarization by inhibiting ERK phosphorylation. Moreover, iron promoted DC maturation, which was possibly mediated by iron-induced M1 macrophages. In addition, iron-induced M1 macrophages and mature DCs promoted the differentiation of CD4+ T cells into the CD4 cytolytic T lymphocytes (CTL) subset and inhibited differentiation into Th2 and Th17 cells. This study explains the role of iron in cryo-thermal therapy-induced antitumor immunity from a new perspective.


Assuntos
Linfócitos T CD4-Positivos/efeitos dos fármacos , Crioterapia/efeitos adversos , Ferro/metabolismo , Ferro/farmacologia , Ativação de Macrófagos/efeitos dos fármacos , Linfócitos T Citotóxicos/efeitos dos fármacos , Animais , Linfócitos T CD4-Positivos/fisiologia , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/imunologia , Polaridade Celular/efeitos dos fármacos , Células Cultivadas , Feminino , Quelantes de Ferro/farmacologia , Ativação Linfocitária/efeitos dos fármacos , Ativação de Macrófagos/fisiologia , Macrófagos/efeitos dos fármacos , Macrófagos/fisiologia , Camundongos , Camundongos Endogâmicos BALB C , Células RAW 264.7 , Linfócitos T Citotóxicos/fisiologia
3.
Front Immunol ; 12: 568789, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34149679

RESUMO

Dysregulation of immune response was observed in COVID-19 patients. Thymosin alpha 1 (Tα1) is used in the management of COVID-19, because it is known to restore the homeostasis of the immune system during infections and cancers. We aim to observe the longitudinal changes in T lymphocyte subsets and to evaluate the efficacy of Tα1 for COVID-19. A retrospective study was conducted in 275 COVID-19 patients admitted to Shanghai public health clinical center. The clinical and laboratory characteristics between patients with different T lymphocyte phenotypes and those who were and were not treated with Tα1 were compared. Among the 275 patients, 137 (49.8%) were males, and the median age was 51 years [interquartile range (IQR): 37-64]. A total of 126 patients received Tα1 therapy and 149 patients did not. There were 158 (57.5%) patients with normal baseline CD4 counts (median:631/µL, IQR: 501~762) and 117 patients (42.5%) with decreased baseline CD4 counts (median:271/µL, IQR: 201~335). In those with decreased baseline CD4 counts, more patients were older (p<0.001), presented as critically ill (p=0.032) and had hypertension (p=0.008) compared with those with normal CD4 counts. There was no statistical difference in the duration of virus shedding in the upper respiratory tract between the two groups (p=0.214). In both the normal (14 vs 11, p=0.028) and the decreased baseline CD4 counts group (15 vs 11, p=0.008), duration of virus clearance in the patients with Tα1 therapy was significantly longer than that in those without Tα1 therapy. There was no significant difference in the increase of CD4+ (286 vs 326, p=0.851) and CD8+ T cell (154 vs 170, p=0.842) counts in the recovery period between the two groups with or without Tα1 therapy. Multivariate linear regression analysis showed that severity of illness (p<0.001) and Tα1 therapy (p=0.001) were associated with virus clearance. In conclusion, reduction of CD4+ T and CD8+ T cell counts were observed in COVID-19 patients. Tα1 may have no benefit on restoring CD4+ and CD8+ T cell counts or on the virus clearance. The use of Tα1 for COVID-19 need to be more fully investigated.


Assuntos
Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/efeitos dos fármacos , COVID-19/tratamento farmacológico , Timalfasina/uso terapêutico , Adjuvantes Imunológicos/farmacologia , Adulto , COVID-19/imunologia , China , Feminino , Humanos , Contagem de Linfócitos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , SARS-CoV-2
4.
Mol Cell Biol ; 41(9): e0008521, 2021 08 24.
Artigo em Inglês | MEDLINE | ID: mdl-34124936

RESUMO

Immune health requires innate and adaptive immune cells to engage precisely balanced pro- and anti-inflammatory forces. We employ the concept of chemical immunophenotypes to classify small molecules functionally or mechanistically according to their patterns of effects on primary innate and adaptive immune cells. The high-specificity, low-toxicity cyclin-dependent kinase 8 (CDK8) inhibitor 16-didehydro-cortistatin A (DCA) exerts a distinct tolerogenic profile in both innate and adaptive immune cells. DCA promotes regulatory T cells (Treg) and Th2 differentiation while inhibiting Th1 and Th17 differentiation in both murine and human cells. This unique chemical immunophenotype led to mechanistic studies showing that DCA promotes Treg differentiation in part by regulating a previously undescribed CDK8-GATA3-FOXP3 pathway that regulates early pathways of Foxp3 expression. These results highlight previously unappreciated links between Treg and Th2 differentiation and extend our understanding of the transcription factors that regulate Treg differentiation and their temporal sequencing. These findings have significant implications for future mechanistic and translational studies of CDK8 and CDK8 inhibitors.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Quinase 8 Dependente de Ciclina/antagonistas & inibidores , Fatores de Transcrição Forkhead/metabolismo , Fator de Transcrição GATA3/metabolismo , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Tolerância Imunológica/efeitos dos fármacos , Imunofenotipagem , Isoquinolinas/farmacologia , Adolescente , Adulto , Animais , Linfócitos T CD4-Positivos/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Quinase 8 Dependente de Ciclina/metabolismo , Humanos , Imunidade Inata/efeitos dos fármacos , Camundongos Endogâmicos BALB C , Pessoa de Meia-Idade , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-jun/metabolismo , Fatores de Transcrição STAT/metabolismo , Transdução de Sinais/efeitos dos fármacos , Adulto Jovem
5.
Int J Cancer ; 149(6): 1313-1321, 2021 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-34019700

RESUMO

CIGB-552 is a synthetic peptide that interacts with COMMD1 and upregulates its protein levels. The objectives of this phase I study were safety, pharmacokinetic profile, evaluation of the lymphocytes CD4+ and CD8+ and preliminary activity in patients with advanced tumors. A 3 + 3 dose-escalation design with seven dose levels was implemented. Patients were included until a grade 3 related adverse event occurred and the maximum tolerated dose was reached. The patients received subcutaneous administration of CIGB-552 three times per week for 2 weeks. Single-dose plasma pharmacokinetics was characterized at two dose levels, and tumor responses were classified by RECIST 1.1. Twenty-four patients received CIGB-552. Dose-limiting toxicity was associated with a transient grade 3 pruritic maculopapular rash at a dose of 7.0 mg. The maximum tolerated dose was defined as 4.7 mg. Ten patients were assessable for immunological status. Seven patients had significant changes in the ratio CD4/CD8 in response to CIGB-552 treatment; three patients did not modify the immunological status. Stable disease was observed in five patients, including two metastatic soft sarcomas. We conclude that CIGB-552 at dose 4.7 mg was well tolerated with no significant adverse events and appeared to provide some clinical benefits.


Assuntos
Antineoplásicos/administração & dosagem , Peptídeos Penetradores de Células/administração & dosagem , NF-kappa B/efeitos dos fármacos , Neoplasias/tratamento farmacológico , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adulto , Antineoplásicos/efeitos adversos , Antineoplásicos/farmacocinética , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/efeitos dos fármacos , Peptídeos Penetradores de Células/efeitos adversos , Peptídeos Penetradores de Células/farmacocinética , Relação Dose-Resposta a Droga , Esquema de Medicação , Feminino , Humanos , Injeções Subcutâneas , Masculino , Dose Máxima Tolerável , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Neoplasias/metabolismo , Neoplasias/patologia , Projetos de Pesquisa , Resultado do Tratamento
6.
Nat Commun ; 12(1): 2606, 2021 05 10.
Artigo em Inglês | MEDLINE | ID: mdl-33972557

RESUMO

Understanding resistance mechanisms to targeted therapies and immune checkpoint blockade in mutant KRAS lung cancers is critical to developing novel combination therapies and improving patient survival. Here, we show that MEK inhibition enhanced PD-L1 expression while PD-L1 blockade upregulated MAPK signaling in mutant KRAS lung tumors. Combined MEK inhibition with anti-PD-L1 synergistically reduced lung tumor growth and metastasis, but tumors eventually developed resistance to sustained combinatorial therapy. Multi-platform profiling revealed that resistant lung tumors have increased infiltration of Th17 cells, which secrete IL-17 and IL-22 cytokines to promote lung cancer cell invasiveness and MEK inhibitor resistance. Antibody depletion of IL-17A in combination with MEK inhibition and PD-L1 blockade markedly reduced therapy-resistance in vivo. Clinically, increased expression of Th17-associated genes in patients treated with PD-1 blockade predicted poorer overall survival and response in melanoma and predicated poorer response to anti-PD1 in NSCLC patients. Here we show a triple combinatorial therapeutic strategy to overcome resistance to combined MEK inhibitor and PD-L1 blockade.


Assuntos
Antineoplásicos Imunológicos/farmacologia , Antígeno B7-H1/metabolismo , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Neoplasias Pulmonares/tratamento farmacológico , Quinases de Proteína Quinase Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Proto-Oncogênicas p21(ras)/genética , Células Th17/metabolismo , Proteína Supressora de Tumor p53/genética , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Antígeno B7-H1/imunologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/mortalidade , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/imunologia , Sinergismo Farmacológico , Feminino , Humanos , Inibidores de Checkpoint Imunológico/imunologia , Imuno-Histoquímica , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/genética , Masculino , Camundongos , Camundongos Knockout , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Invasividade Neoplásica/genética , Invasividade Neoplásica/imunologia , Metástase Neoplásica , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Células Th17/imunologia , Proteína Supressora de Tumor p53/metabolismo
7.
Nat Commun ; 12(1): 2475, 2021 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-33931637

RESUMO

An innovative approach to eliminate HIV-1-infected cells emerging out of latency, the major hurdle to HIV-1 cure, is to pharmacologically reactivate viral expression and concomitantly trigger intracellular pro-apoptotic pathways in order to selectively induce cell death (ICD) of infected cells, without reliance on the extracellular immune system. In this work, we demonstrate the effect of DDX3 inhibitors on selectively inducing cell death in latent HIV-1-infected cell lines, primary CD4+ T cells and in CD4+ T cells from cART-suppressed people living with HIV-1 (PLWHIV). We used single-cell FISH-Flow technology to characterise the contribution of viral RNA to inducing cell death. The pharmacological targeting of DDX3 induced HIV-1 RNA expression, resulting in phosphorylation of IRF3 and upregulation of IFNß. DDX3 inhibition also resulted in the downregulation of BIRC5, critical to cell survival during HIV-1 infection, and selectively induced apoptosis in viral RNA-expressing CD4+ T cells but not bystander cells. DDX3 inhibitor treatment of CD4+ T cells from PLWHIV resulted in an approximately 50% reduction of the inducible latent HIV-1 reservoir by quantitation of HIV-1 RNA, by FISH-Flow, RT-qPCR and TILDA. This study provides proof of concept for pharmacological reversal of latency coupled to induction of apoptosis towards the elimination of the inducible reservoir.


Assuntos
Apoptose/efeitos dos fármacos , Azepinas/farmacologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , RNA Helicases DEAD-box/metabolismo , Infecções por HIV/imunologia , HIV-1/metabolismo , Imidazóis/farmacologia , Latência Viral/efeitos dos fármacos , Replicação Viral/efeitos dos fármacos , Antirretrovirais/farmacologia , Apoptose/genética , Azepinas/química , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD4-Positivos/virologia , Morte Celular/efeitos dos fármacos , Morte Celular/genética , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , RNA Helicases DEAD-box/antagonistas & inibidores , RNA Helicases DEAD-box/química , Inibidores Enzimáticos/farmacologia , Infecções por HIV/genética , Infecções por HIV/metabolismo , Infecções por HIV/virologia , HIV-1/efeitos dos fármacos , HIV-1/genética , Humanos , Imidazóis/química , Hibridização in Situ Fluorescente , Fator Regulador 3 de Interferon/metabolismo , Interferon beta/metabolismo , Células Jurkat , Simulação de Acoplamento Molecular , RNA Viral/metabolismo , Análise de Célula Única , Survivina/metabolismo , Ativação Viral/efeitos dos fármacos , Replicação Viral/genética
8.
Nat Commun ; 12(1): 2866, 2021 05 17.
Artigo em Inglês | MEDLINE | ID: mdl-34001890

RESUMO

Unlike HIV infection, which progresses to AIDS absent suppressive anti-retroviral therapy, nonpathogenic infections in natural hosts, such African green monkeys, are characterized by a lack of gut microbial translocation and robust secondary lymphoid natural killer cell responses resulting in an absence of chronic inflammation and limited SIV dissemination in lymph node B-cell follicles. Here we report, using the pathogenic model of antiretroviral therapy-treated, SIV-infected rhesus macaques that sequential interleukin-21 and interferon alpha therapy generate terminally differentiated blood natural killer cells (NKG2a/clowCD16+) with potent human leukocyte antigen-E-restricted activity in response to SIV envelope peptides. This is in contrast to control macaques, where less differentiated, interferon gamma-producing natural killer cells predominate. The frequency and activity of terminally differentiated NKG2a/clowCD16+ natural killer cells correlates with a reduction of replication-competent SIV in lymph node during antiretroviral therapy and time to viral rebound following analytical treatment interruption. These data demonstrate that African green monkey-like natural killer cell differentiation profiles can be rescued in rhesus macaques to promote viral clearance in tissues.


Assuntos
Antirretrovirais/farmacologia , Interferon gama/farmacologia , Interleucinas/farmacologia , Células Matadoras Naturais/efeitos dos fármacos , Síndrome de Imunodeficiência Adquirida dos Símios/tratamento farmacológico , Vírus da Imunodeficiência Símia/efeitos dos fármacos , Animais , Antivirais/farmacologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/virologia , Feminino , Células Matadoras Naturais/virologia , Ativação Linfocitária/efeitos dos fármacos , Macaca mulatta , Síndrome de Imunodeficiência Adquirida dos Símios/virologia , Vírus da Imunodeficiência Símia/fisiologia , Carga Viral/efeitos dos fármacos , Viremia/sangue , Viremia/tratamento farmacológico
9.
J Virol ; 95(12)2021 05 24.
Artigo em Inglês | MEDLINE | ID: mdl-33827940

RESUMO

The antiapoptotic protein BCL2 inhibits death of HIV-infected cells. Previously, we showed that the BCL2 inhibitor venetoclax selectively kills acutely HIV-infected cells and reduces HIV DNA in latently infected CD4 T cells ex vivo after reactivation with anti-CD3/anti-CD28. However, there is a need to identify a combination therapy with venetoclax and a clinically relevant latency reversal agent. Ixazomib is an oral proteasome inhibitor which we have shown reactivates latent HIV and predisposes reactivated cells to cell death. Here, we determined that the combination of venetoclax and ixazomib kills more latently HIV-infected cells and leads to greater reduction in HIV replication than either treatment alone in vitro in a T cell model. However, combination treatment of ex vivo CD4 T cells from antiretroviral therapy (ART)-suppressed, HIV-positive participants resulted in unanticipated and unacceptable nonspecific toxicity in primary cells. Therefore, while we show proof of concept that multiple agents can enhance selective killing of HIV-infected cells, the combination of venetoclax and ixazomib has unacceptable toxicity in primary cells, and so further investigation is needed to identify a clinically relevant latency reversal agent to combine with venetoclax as a novel strategy to reduce the size of the HIV reservoir.IMPORTANCE A cure for HIV would require eliminating cells that contain the virus in a latent form from the body. Current antiretroviral medications are unable to rid the body of latently infected cells. Here, we show that a combination of investigational agents-ixazomib plus venetoclax-which reactivate latent virus and predispose infected cells to apoptosis may reduce latent virus in a T cell model, but at the expense of nonspecific toxicity in primary cells.


Assuntos
Fármacos Anti-HIV/farmacologia , Compostos de Boro/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Glicina/análogos & derivados , HIV-1/efeitos dos fármacos , Sulfonamidas/farmacologia , Fármacos Anti-HIV/toxicidade , Apoptose/efeitos dos fármacos , Compostos de Boro/toxicidade , Compostos Bicíclicos Heterocíclicos com Pontes/toxicidade , Linfócitos T CD4-Positivos/virologia , Linhagem Celular , Linhagem Celular Tumoral , Células Cultivadas , Quimioterapia Combinada , Glicina/farmacologia , Glicina/toxicidade , HIV-1/fisiologia , Humanos , Células Jurkat , Provírus/efeitos dos fármacos , Sulfonamidas/toxicidade , Resposta a Proteínas não Dobradas , Ativação Viral , Latência Viral , Replicação Viral/efeitos dos fármacos
10.
Nat Commun ; 12(1): 1980, 2021 03 31.
Artigo em Inglês | MEDLINE | ID: mdl-33790300

RESUMO

The majority of patients with systemic lupus erythematosus (SLE) have high expression of type I IFN-stimulated genes. Mitochondrial abnormalities have also been reported, but the contribution of type I IFN exposure to these changes is unknown. Here, we show downregulation of mitochondria-derived genes and mitochondria-associated metabolic pathways in IFN-High patients from transcriptomic analysis of CD4+ and CD8+ T cells. CD8+ T cells from these patients have enlarged mitochondria and lower spare respiratory capacity associated with increased cell death upon rechallenge with TCR stimulation. These mitochondrial abnormalities can be phenocopied by exposing CD8+ T cells from healthy volunteers to type I IFN and TCR stimulation. Mechanistically these 'SLE-like' conditions increase CD8+ T cell NAD+ consumption resulting in impaired mitochondrial respiration and reduced cell viability, both of which can be rectified by NAD+ supplementation. Our data suggest that type I IFN exposure contributes to SLE pathogenesis by promoting CD8+ T cell death via metabolic rewiring.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Perfilação da Expressão Gênica/métodos , Interferon Tipo I/imunologia , Lúpus Eritematoso Sistêmico/imunologia , Adulto , Idoso , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/metabolismo , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Feminino , Humanos , Interferon Tipo I/metabolismo , Interferon Tipo I/farmacologia , Lúpus Eritematoso Sistêmico/genética , Lúpus Eritematoso Sistêmico/metabolismo , Ativação Linfocitária/efeitos dos fármacos , Ativação Linfocitária/genética , Ativação Linfocitária/imunologia , Redes e Vias Metabólicas/genética , Pessoa de Meia-Idade , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Adulto Jovem
11.
J Immunol ; 206(9): 2015-2028, 2021 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-33820855

RESUMO

The cGAS-cyclic GMP-AMP (cGAMP)-stimulator of IFN genes (STING) pathway induces a powerful type I IFN (IFN-I) response and is a prime candidate for augmenting immunity in cancer immunotherapy and vaccines. IFN-I also has immune-regulatory functions manifested in several autoimmune diseases and is a first-line therapy for relapsing-remitting multiple sclerosis. However, it is only moderately effective and can induce adverse effects and neutralizing Abs in recipients. Targeting cGAMP in autoimmunity is unexplored and represents a challenge because of the intracellular location of its receptor, STING. We used microparticle (MP)-encapsulated cGAMP to increase cellular delivery, achieve dose sparing, and reduce potential toxicity. In the C57BL/6 experimental allergic encephalomyelitis (EAE) model, cGAMP encapsulated in MPs (cGAMP MPs) administered therapeutically protected mice from EAE in a STING-dependent fashion, whereas soluble cGAMP was ineffective. Protection was also observed in a relapsing-remitting model. Importantly, cGAMP MPs protected against EAE at the peak of disease and were more effective than rIFN-ß. Mechanistically, cGAMP MPs showed both IFN-I-dependent and -independent immunosuppressive effects. Furthermore, it induced the immunosuppressive cytokine IL-27 without requiring IFN-I. This augmented IL-10 expression through activated ERK and CREB. IL-27 and subsequent IL-10 were the most important cytokines to mitigate autoreactivity. Critically, cGAMP MPs promoted IFN-I as well as the immunoregulatory cytokines IL-27 and IL-10 in PBMCs from relapsing-remitting multiple sclerosis patients. Collectively, this study reveals a previously unappreciated immune-regulatory effect of cGAMP that can be harnessed to restrain T cell autoreactivity.


Assuntos
Micropartículas Derivadas de Células/imunologia , Encefalomielite Autoimune Experimental/imunologia , Interferon Tipo I/imunologia , Proteínas de Membrana/imunologia , Nucleotídeos Cíclicos/imunologia , Transdução de Sinais/imunologia , Animais , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Micropartículas Derivadas de Células/metabolismo , Células Cultivadas , Citocinas/imunologia , Citocinas/metabolismo , Encefalomielite Autoimune Experimental/metabolismo , Encefalomielite Autoimune Experimental/prevenção & controle , Feminino , Humanos , Interferon Tipo I/metabolismo , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/metabolismo , Proteínas de Membrana/agonistas , Proteínas de Membrana/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos , Camundongos Knockout , Nucleotídeos Cíclicos/administração & dosagem , Nucleotídeos Cíclicos/metabolismo , Transdução de Sinais/efeitos dos fármacos
12.
Eur J Med Chem ; 219: 113393, 2021 Jul 05.
Artigo em Inglês | MEDLINE | ID: mdl-33845236

RESUMO

ZAP-70 (zeta-chain associated protein kinase 70 kDa) signaling pathway and its functions have been involved in the development and adaptive immune signaling of T cell. It thus represents a promising target for autoimmune diseases. Although reversible ZAP-70 kinase domain inhibitors have been developed, they are either weak or nonselective. We report herein the structure-guided development of the first potent and covalent inhibitor of ZAP-70 kinase domain. In particular, compound 18 (RDN009) showed good selectivity for ZAP-70 over structurally related Syk, and displayed potent inhibitory effects on T cell proliferation, activation, and inflammatory cytokine production. A mass spectrometry analysis further confirmed the covalent linkage between the inhibitor and ZAP-70 protein at C346. Overall, the covalent inhibitor RDN009 represents a potent and selective probe of ZAP-70 for further development for treatment of autoimmune diseases.


Assuntos
Inibidores de Proteínas Quinases/química , Proteína-Tirosina Quinase ZAP-70/antagonistas & inibidores , Animais , Sítios de Ligação , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/metabolismo , Proliferação de Células/efeitos dos fármacos , Citocinas/análise , Citocinas/metabolismo , Avaliação Pré-Clínica de Medicamentos , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Simulação de Acoplamento Molecular , Inibidores de Proteínas Quinases/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Relação Estrutura-Atividade , Quinase Syk/antagonistas & inibidores , Quinase Syk/metabolismo , Proteína-Tirosina Quinase ZAP-70/metabolismo
13.
FEBS Lett ; 595(11): 1587-1603, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33792041

RESUMO

OX40 is a costimulatory molecule that belongs to the tumor necrosis factor receptor (TNFR) superfamily. OX40 agonist-based combinations are emerging as promising candidates for novel cancer immunotherapy. Clinical trials have shown that OX40 agonist antibodies could lead to better results in cancer patients. Using a hybridoma platform and three different types of immunization strategies, namely recombinant protein, DNA, and overexpressing cells, we identified a chimeric anti-OX40 antibody (mAb035-hIgG1 from DNA immunization) that shows excellent binding specificity, and slightly stronger activation of human memory CD4+ T cells and similar potent antitumor activity compared with BMS 986178, an anti-OX40 antibody currently being evaluated for the treatment of solid tumors. This paper further systematically investigates the antigen-specific immune response, the number of binders, epitope bins, and functional activities of antibodies among different immunization strategies. Interestingly, we found that different immunization strategies affect the biological activity of monoclonal antibodies.


Assuntos
Anticorpos Monoclonais/farmacologia , Antineoplásicos Imunológicos/farmacologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Imunização/métodos , Receptores OX40/imunologia , Proteínas Recombinantes de Fusão/farmacologia , Animais , Anticorpos Monoclonais/biossíntese , Anticorpos Monoclonais/isolamento & purificação , Afinidade de Anticorpos , Especificidade de Anticorpos , Antineoplásicos Imunológicos/isolamento & purificação , Antineoplásicos Imunológicos/metabolismo , Bioensaio , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/imunologia , Células CHO , Cricetulus , Feminino , Adjuvante de Freund/administração & dosagem , Expressão Gênica , Genes Reporter , Células HEK293 , Humanos , Hibridomas/química , Hibridomas/imunologia , Fragmentos Fc das Imunoglobulinas/biossíntese , Fragmentos Fc das Imunoglobulinas/isolamento & purificação , Fragmentos Fc das Imunoglobulinas/farmacologia , Células Jurkat , Luciferases/genética , Luciferases/metabolismo , Ativação Linfocitária/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , NF-kappa B/genética , NF-kappa B/imunologia , Receptores OX40/antagonistas & inibidores , Receptores OX40/genética , Proteínas Recombinantes de Fusão/biossíntese , Proteínas Recombinantes de Fusão/isolamento & purificação
14.
Methods Mol Biol ; 2285: 191-200, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33928554

RESUMO

Flow cytometric evaluation of phosphorylation status of signal transduction molecules is a useful method to study T-cell signaling pathways. As mutations occurring in TCR complex molecules, common gamma chain family's cytokines, their receptors or molecules involved in these pathways can lead to severe immune system defects, the study of T-cell signal transduction can be applied to both basic and clinical/translational research areas. In the present chapter, we show two different protocols for the study of T- cell response to an antigen-like stimulus and to IL-2.


Assuntos
Linfócitos T CD4-Positivos/metabolismo , Citometria de Fluxo , Fosfoproteínas/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Transdução de Sinais , Animais , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Células Cultivadas , Humanos , Interleucina-2/farmacologia , Fosforilação , Receptores de Antígenos de Linfócitos T/agonistas , Projetos de Pesquisa , Transdução de Sinais/efeitos dos fármacos , Acetato de Tetradecanoilforbol/farmacologia , Fluxo de Trabalho
15.
PLoS Pathog ; 17(4): e1009141, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33826675

RESUMO

HIV persists during antiretroviral therapy (ART) as integrated proviruses in cells descended from a small fraction of the CD4+ T cells infected prior to the initiation of ART. To better understand what controls HIV persistence and the distribution of integration sites (IS), we compared about 15,000 and 54,000 IS from individuals pre-ART and on ART, respectively, with approximately 395,000 IS from PBMC infected in vitro. The distribution of IS in vivo is quite similar to the distribution in PBMC, but modified by selection against proviruses in expressed genes, by selection for proviruses integrated into one of 7 specific genes, and by clonal expansion. Clones in which a provirus integrated in an oncogene contributed to cell survival comprised only a small fraction of the clones persisting in on ART. Mechanisms that do not involve the provirus, or its location in the host genome, are more important in determining which clones expand and persist.


Assuntos
Antirretrovirais/uso terapêutico , Infecções por HIV/virologia , Leucócitos Mononucleares/virologia , Oncogenes/genética , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/virologia , DNA Viral/genética , Humanos , Oncogenes/imunologia , Provírus/genética , Replicação Viral/genética
16.
Chem Biol Interact ; 341: 109451, 2021 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-33798506

RESUMO

The pathogenesis of rheumatoid arthritis (RA) is characterized by synoviocyte hyperplasia and proinflammatory cytokine secretion, as well as the destruction of cartilage and bone. Glaucocalyxin A (GLA) is an alkaloid derived from a Chinese medicinal plant that exhibits anti-inflammatory, anti-tumor and neuroprotective properties. We investigated the effects of GLA on RA-fibroblast-like synoviocytes (FLS cells), and collagen-induced arthritis (CIA), and further explored the underlying mechanisms. GLA inhibited TNF-a-induced RA-FLS proliferation, increased apoptotic ratios and upregulated levels of caspase-3, cleaved PARP, and Bax. GLA also inhibited the expression of IL-10, IL-1ß, and IL-6 in vitro. Levels of p-STAT3 were downregulated in a dose-dependent manner. Over-expression of STAT3 partly neutralized the GLA-mediated elevation of caspase-3 and cleaved PARP levels as well as the downregulation of IL-10, IL-1B and IL-6 expression levels. This suggests that GLA inactivated the STAT3 pathway. Furthermore, the production of inflammatory cytokines in RA-FLS and a CIA rat model were inhibited effectively by GLA. Taken together, our data suggest that GLA is a potential long-term therapeutic agent for patients with RA.


Assuntos
Artrite Reumatoide/tratamento farmacológico , Diterpenos do Tipo Caurano/farmacologia , Fator de Transcrição STAT3/metabolismo , Animais , Anti-Inflamatórios não Esteroides/farmacologia , Apoptose/efeitos dos fármacos , Artrite Experimental/tratamento farmacológico , Artrite Experimental/patologia , Artrite Reumatoide/metabolismo , Artrite Reumatoide/patologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Citocinas/metabolismo , Humanos , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Inflamação/patologia , Masculino , Camundongos Endogâmicos DBA , Ratos Wistar , Fator de Transcrição STAT3/genética , Sinoviócitos/efeitos dos fármacos , Sinoviócitos/metabolismo , Sinoviócitos/patologia , Células Th17/efeitos dos fármacos , Células Th17/fisiologia , Fator de Necrose Tumoral alfa/farmacologia
17.
Nat Immunol ; 22(4): 423-433, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33767427

RESUMO

Individuals infected with human immunodeficiency virus type-1 (HIV-1) show metabolic alterations of CD4+ T cells through unclear mechanisms with undefined consequences. We analyzed the transcriptome of CD4+ T cells from patients with HIV-1 and revealed that the elevated oxidative phosphorylation (OXPHOS) pathway is associated with poor outcomes. Inhibition of OXPHOS by the US Food and Drug Administration-approved drug metformin, which targets mitochondrial respiratory chain complex-I, suppresses HIV-1 replication in human CD4+ T cells and humanized mice. In patients, HIV-1 peak viremia positively correlates with the expression of NLRX1, a mitochondrial innate immune receptor. Quantitative proteomics and metabolic analyses reveal that NLRX1 enhances OXPHOS and glycolysis during HIV-1-infection of CD4+ T cells to promote viral replication. At the mechanistic level, HIV infection induces the association of NLRX1 with the mitochondrial protein FASTKD5 to promote expression of mitochondrial respiratory complex components. This study uncovers the OXPHOS pathway in CD4+ T cells as a target for HIV-1 therapy.


Assuntos
Linfócitos T CD4-Positivos/virologia , Genômica , Infecções por HIV/virologia , HIV-1/crescimento & desenvolvimento , Metaboloma , Metabolômica , Fosforilação Oxidativa , Proteoma , Transcriptoma , Replicação Viral , Animais , Antivirais/farmacologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Modelos Animais de Doenças , Feminino , Perfilação da Expressão Gênica , Redes Reguladoras de Genes , Células HEK293 , Infecções por HIV/tratamento farmacológico , Infecções por HIV/imunologia , Infecções por HIV/metabolismo , HIV-1/efeitos dos fármacos , HIV-1/imunologia , HIV-1/metabolismo , Interações Hospedeiro-Patógeno , Humanos , Células Jurkat , Masculino , Metformina/farmacologia , Camundongos , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Fosforilação Oxidativa/efeitos dos fármacos , Proteômica , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Carga Viral , Replicação Viral/efeitos dos fármacos
18.
Int J Mol Sci ; 22(4)2021 Feb 16.
Artigo em Inglês | MEDLINE | ID: mdl-33669259

RESUMO

BACKGROUND: Pseurotins, a family of secondary metabolites of different fungi characterized by an unusual spirocyclic furanone-lactam core, are suggested to have different biological activities including the modulation of immune response. PURPOSE: Complex characterization of the effects of pseurotin D on human lymphocyte activation in order to understand the potential of pseurotin to modulate immune response in humans. METHODS: CD4+ and CD8+ T cells and CD19+ B cells isolated from human blood were activated by various activators simultaneously with pseurotin D treatment. The effects of pseurotin were tested on the basis of changes in cell viability, apoptosis, activation of signal transducers and activators of transcription (STAT) signaling pathways, production of tumor necrosis factor (TNF)-α by T cells, expression of activation markers CD69 and CD25 on T cells and Human Leukocyte Antigen-DR isotype (HLA-DR) on B cells, and the differentiation markers CD20, CD27, CD38, and immunoglobulin (Ig) D on B cells. RESULTS: Pseurotin D significantly inhibited the activation of both CD4+ and CD8+ human T cells complemented by the inhibition of TNF-α production without significant acute toxic effects. The Pseurotin D-mediated inhibition of T-cell activation was accompanied by the induction of the apoptosis of T cells. This corresponded with the inhibited phosphorylation of STAT3 and STAT5. In human B cells, pseurotin D did not significantly inhibit their activation; however, it affected their differentiation. CONCLUSIONS: Our results advance the current mechanistic understanding of the pseurotin-induced inhibition of lymphocytes and suggest pseurotins as new attractive chemotypes for future research in the context of immune-modulatory drugs.


Assuntos
Antígenos CD19/metabolismo , Linfócitos B/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Fatores Imunológicos/farmacologia , Ativação Linfocitária/efeitos dos fármacos , Pirrolidinonas/farmacologia , Adolescente , Adulto , Apoptose/efeitos dos fármacos , Linfócitos B/efeitos dos fármacos , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Feminino , Voluntários Saudáveis , Humanos , Masculino , Pessoa de Meia-Idade , Transdução de Sinais/efeitos dos fármacos , Fator de Necrose Tumoral alfa/metabolismo , Adulto Jovem
19.
Cancer Med ; 10(7): 2509-2522, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33704935

RESUMO

Prostate cancer-associated fibroblasts (prostate CAFs) are essential components of the tumor microenvironment and can promote tumor progression through their immunosuppressive functions. MPSSS, a novel polysaccharide purified from Lentinus edodes, has been reported to have anti-tumor activity. MPSSS could also inhibit the immunosuppressive function of prostate CAFs, which has been demonstrated through that the secretome of MPSSS-treated prostate CAFs could inhibit the proliferation of T cells. However, how the secretome of MPSSS-treated prostate CAFs influence prostate cancer progression is still unclear. Interestingly, we found that the low molecular weight (3-100kD) secretome of prostate CAFs (lmwCAFS) could promote the growth of PC-3 cells, while that of MPSSS-treated prostate CAFs (MT-lmwCAFS) could inhibit their growth. We carried out comparative secretomic analysis of lmwCAFS and MT-lmwCAFS to identify functional molecules that inhibit the growth of PC-3 cells, and proteomic analysis of lmwCAFS-treated PC-3 cells and MT-lmwCAFS-treated PC-3 cells to investigate the underlying molecular mechanism. These analyses suggest that TGF-ß3 from MT-lmwCAFS may inhibit the growth of PC-3 cells. The validated experiments revealed that TGF-ß3 from MT-lmwCAFS activated p21 expression in PC-3 cells by regulating the FoxO pathway thereby inducing G0/G1 cell cycle arrest of PC-3 cells. Overall, our data demonstrated that MPSSS reversed the ability of prostate CAFs to suppress the cell viability of PC-3 cells, which might provide a potential therapeutic strategy to prevent prostate cancer progression.


Assuntos
Fibroblastos Associados a Câncer/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Fatores de Transcrição Forkhead/metabolismo , Polissacarídeos Fúngicos/farmacologia , Neoplasias da Próstata/patologia , Proteômica , Fator de Crescimento Transformador beta3/farmacologia , Actinas/metabolismo , Animais , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/imunologia , Fibroblastos Associados a Câncer/química , Fibroblastos Associados a Câncer/fisiologia , Pontos de Checagem do Ciclo Celular , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Progressão da Doença , Vesículas Extracelulares/química , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Peso Molecular , Células PC-3 , Neoplasias da Próstata/terapia , Cogumelos Shiitake/química , Microambiente Tumoral/imunologia
20.
BMC Cancer ; 21(1): 233, 2021 Mar 06.
Artigo em Inglês | MEDLINE | ID: mdl-33676448

RESUMO

BACKGROUND: It was demonstrated that multifunctional protein APE1 (Apurinic/apyrimidinic endonuclease 1) is closely related to tumor immune microenvironment in a number of investigations, Meanwhile, the abundance of tumor infiltrating lymphocytes (TILs) has been shown as a prognosis indicator in some researches. However, it remains unclear whether APE1 is involved in the process of TILs affecting the prognosis of patients. To this end, we investigated the associations between APE1 and TILs in non-small cell lung cancer (NSCLC) and explored whether APE1 would influence the associations of CD4+ T cells infiltration with the prognosis of patients. METHODS: Genome-wide expression datasets were obtained from the Gene Expression Omnibus (GEO) public database under accession number GSE68465, GSE30219, GSE31210 and GSE50081. MCPcounter and CIBERSORT analysis was conducted to evaluate the abundance of TILs in 1006 NSCLC patients of GEO database. Spearman correlation tests were used to evaluate correlations between abundance of various TILs and APE1 expression. RFS (recurrence free survival) was estimated using the Kaplan-Meier method and the Cox proportional-hazards model. The expression level of APE1 and tumor-infiltrating CD4+ T cells was evaluated by immunohistochemistry (IHC). RESULTS: The results showed that the abundance of CD4+ naïve T cells was negatively associated with the APE1 expression. CD4+ naïve T cells infiltration was a favorable prognostic factor for RFS, however, there was no effect of CD4+ T cells infiltration on RFS in patients with high APE1 expression. Subsequently, it was further confirmed that CD4+ T cells infiltration was negatively associated with the APE1 expression level in 108 NSCLC tissue samples; high CD4+ T cells infiltration was associated with longer RFS in low APE1 expression group but not in APE1 high expression group. CONCLUSION: These results suggested that APE1 may affect the relationship between CD4+ T cells infiltration and prognosis in NSCLC. This study provides new insights into predictors of outcome in patients with NSCLC, and suggests that combining immunotherapy and APE1-targeted therapy may be a promising treatment for NSCLC.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/mortalidade , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/metabolismo , Neoplasias Pulmonares/mortalidade , Recidiva Local de Neoplasia/epidemiologia , Antineoplásicos Imunológicos/farmacologia , Antineoplásicos Imunológicos/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Carcinoma Pulmonar de Células não Pequenas/imunologia , Carcinoma Pulmonar de Células não Pequenas/patologia , Carcinoma Pulmonar de Células não Pequenas/terapia , Quimioterapia Adjuvante/métodos , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/antagonistas & inibidores , Conjuntos de Dados como Assunto , Intervalo Livre de Doença , Feminino , Perfilação da Expressão Gênica , Humanos , Estimativa de Kaplan-Meier , Pulmão/imunologia , Pulmão/patologia , Pulmão/cirurgia , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/terapia , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Linfócitos do Interstício Tumoral/imunologia , Masculino , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/imunologia , Recidiva Local de Neoplasia/prevenção & controle , Análise de Sequência com Séries de Oligonucleotídeos , Pneumonectomia , Prognóstico , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/imunologia
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