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1.
Chem Commun (Camb) ; 56(20): 3089-3092, 2020 Mar 10.
Artigo em Inglês | MEDLINE | ID: mdl-32052805

RESUMO

Reaction of FeIII(O2˙-)(TPP) with 2,3-dimethylindole at -40 °C gives the ring-opened, dioxygenated N-(2-acetyl-phenyl)-acetamide product. The reaction was monitored in situ by low-temperature UV-vis and 1H NMR spectroscopies. This work demonstrates that a discrete iron(iii)(superoxo) porphyrin is competent to carry out indole oxidation, as proposed for the tryptophan and indoleamine 2,3-dioxygenases.


Assuntos
Compostos Férricos/química , Indolamina-Pirrol 2,3,-Dioxigenase/química , Indóis/química , Metaloporfirinas/química , Superóxidos/química , Triptofano Oxigenase/química , Compostos Férricos/metabolismo , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Indóis/metabolismo , Metaloporfirinas/metabolismo , Estrutura Molecular , Oxirredução , Superóxidos/metabolismo , Triptofano Oxigenase/metabolismo
2.
Cancer Immunol Immunother ; 69(1): 57-67, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31802183

RESUMO

Tumors can utilize a diverse repertoire of immunosuppressive mechanisms to evade attack by the immune system. Despite promising success with blockade of immune checkpoints like PD-1 the majority of patients does not respond to current immunotherapies. The degradation of tryptophan into immunosuppressive kynurenine is an important immunosuppressive pathway. Recent attempts to target the key enzymes of this pathway-IDO1 and TDO2-have so far failed to show therapeutic benefit in the clinic, potentially caused by insufficient target engagement. We, therefore, sought to add an alternative, highly efficient approach to block the degradation of tryptophan by inhibiting the expression of IDO1 and TDO2 using locked nucleic acid (LNA)-modified antisense oligonucleotides (ASOs). We show that LNA-modified ASOs can profoundly inhibit the expression of IDO1 and TDO2 in cancer cells in vitro without using a transfection reagent with IC50 values in the sub-micromolar range. We furthermore measured kynurenine production by ASO-treated cancer cells in vitro and observed potently reduced kynurenine levels. Accordingly, inhibiting IDO1 expression in cancer cells in an in vitro system leads to increased proliferation of activated T cells in coculture. We furthermore show that combined treatment of cancer cells in vitro with IDO1-specific ASOs and small molecule inhibitors can reduce the production of kynurenine by cancer cells in a synergistic manner. In conclusion, we propose that a combination of LNA-modified ASOs and small molecule inhibitors should be considered as a strategy for efficient blockade of the degradation of tryptophan into kynurenine in cancer immunotherapy.


Assuntos
Antineoplásicos/farmacologia , Indolamina-Pirrol 2,3,-Dioxigenase/antagonistas & inibidores , Neoplasias/terapia , Oligonucleotídeos Antissenso/farmacologia , Triptofano Oxigenase/antagonistas & inibidores , Antineoplásicos/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Técnicas de Cocultura , Ensaios de Seleção de Medicamentos Antitumorais , Sinergismo Farmacológico , Humanos , Imunoterapia/métodos , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Concentração Inibidora 50 , Cinurenina/imunologia , Cinurenina/metabolismo , Ativação Linfocitária/efeitos dos fármacos , Neoplasias/imunologia , Oligonucleotídeos/administração & dosagem , Oligonucleotídeos/química , Oligonucleotídeos Antissenso/química , Linfócitos T/imunologia , Triptofano/imunologia , Triptofano/metabolismo , Triptofano Oxigenase/metabolismo
3.
Eur J Med Chem ; 184: 111750, 2019 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-31610376

RESUMO

Developing small molecules occupying the heme-binding site using computational approaches remains a challenging task because it is difficult to characterize heme-ligand interaction in heme-containing protein. Indoleamine 2,3-dioxygenase 1 (IDO1) is an intracellular heme-containing dioxygenase which is associated with the immunosuppressive effects in cancer. With IDO1 as an example, herein we report a combined virtual screening (VS) strategy including high-specificity heme-binding group (HmBG)-based pharmacophore screening and cascade molecular docking to identify novel IDO1 inhibitors. A total of four hit compounds were obtained and showed proper binding with the heme iron coordinating site. Further structural optimization led to a promising compound S18-3, which exerted potent anti-tumor efficacy in BALB/c mice bearing established CT26 tumors by activating the host's immune system. These results suggest that S18-3 merits further study to assess its potential for the intervention of cancer. Furthermore, our study also unveils a novel in silico-based strategy for identifying potential regulators for hemeproteins within short timeframe.


Assuntos
Antineoplásicos/farmacologia , Neoplasias do Colo/tratamento farmacológico , Hidrazinas/farmacologia , Indolamina-Pirrol 2,3,-Dioxigenase/antagonistas & inibidores , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Proliferação de Células/efeitos dos fármacos , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos , Ensaios de Seleção de Medicamentos Antitumorais , Células HeLa , Humanos , Hidrazinas/síntese química , Hidrazinas/química , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Estrutura Molecular , Relação Estrutura-Atividade , Células Tumorais Cultivadas
4.
Cancer Immunol Immunother ; 68(10): 1661-1669, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31606777

RESUMO

BACKGROUND: We previously found that PD-L1 expression is increased on tumor cells following vaccination treatments that lead to increased tumor-specific T cells that secrete IFNγ. Indoleamine 2,3-dioxygenase (IDO) is another IFNγ inducible gene that has potent immunosuppressive effects. There have been reports of IDO expression in prostate cancer; however, it is unknown whether IDO expression might similarly increase in prostate tumors following T-cell-based immunotherapy. METHODS: Blood samples from normal male blood donors (n = 12) and patients with different stages of prostate cancer (n = 89), including patients with metastatic, castration-resistant prostate cancer treated with a DNA vaccine and/or pembrolizumab, were evaluated for IDO activity by kynurenine and tryptophan levels. Metastatic tissue biopsies obtained pre- and post-treatments were evaluated for IDO expression. IDO suppression of vaccine-induced T-cell function was assessed by ELISPOT. RESULTS: Overall, IDO activity was increased in patients with more advanced prostate cancer. This activity, and IDO expression as detected immunohistochemically, increased following treatment with either a DNA vaccine encoding the prostatic acid phosphatase (PAP) tumor antigen or PD-1 blockade with pembrolizumab. Increased IDO activity after treatment was associated with the absence of clinical effect, as assessed by lack of PSA decline following treatment. Increased antigen-specific T-cell response, as measured by IFNγ release, to the vaccine target antigen was detected following in vitro stimulation of peripheral blood cells with 1-methyltryptophan. CONCLUSIONS: These findings suggest that IDO expression is a mechanism of immune evasion used by prostate cancer and that future clinical trials using T-cell-based immune strategies might best include IDO inhibition.


Assuntos
Vacinas Anticâncer/administração & dosagem , Imunoterapia , Indolamina-Pirrol 2,3,-Dioxigenase/antagonistas & inibidores , Recidiva Local de Neoplasia/tratamento farmacológico , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Neoplasias da Próstata/tratamento farmacológico , Linfócitos T/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/metabolismo , Seguimentos , Regulação Neoplásica da Expressão Gênica , Humanos , Indolamina-Pirrol 2,3,-Dioxigenase/imunologia , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Masculino , Pessoa de Meia-Idade , Terapia de Alvo Molecular , Recidiva Local de Neoplasia/imunologia , Recidiva Local de Neoplasia/metabolismo , Recidiva Local de Neoplasia/patologia , Prognóstico , Receptor de Morte Celular Programada 1/imunologia , Receptor de Morte Celular Programada 1/metabolismo , Neoplasias da Próstata/imunologia , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Linfócitos T/efeitos dos fármacos
5.
Nat Commun ; 10(1): 4766, 2019 10 18.
Artigo em Inglês | MEDLINE | ID: mdl-31628327

RESUMO

Trisomy 21 (T21) causes Down syndrome (DS), affecting immune and neurological function by ill-defined mechanisms. Here we report a large metabolomics study of plasma and cerebrospinal fluid, showing in independent cohorts that people with DS produce elevated levels of kynurenine and quinolinic acid, two tryptophan catabolites with potent immunosuppressive and neurotoxic properties, respectively. Immune cells of people with DS overexpress IDO1, the rate-limiting enzyme in the kynurenine pathway (KP) and a known interferon (IFN)-stimulated gene. Furthermore, the levels of IFN-inducible cytokines positively correlate with KP dysregulation. Using metabolic tracing assays, we show that overexpression of IFN receptors encoded on chromosome 21 contribute to enhanced IFN stimulation, thereby causing IDO1 overexpression and kynurenine overproduction in cells with T21. Finally, a mouse model of DS carrying triplication of IFN receptors exhibits KP dysregulation. Together, our results reveal a mechanism by which T21 could drive immunosuppression and neurotoxicity in DS.


Assuntos
Cromossomos Humanos Par 21/genética , Síndrome de Down/genética , Cinurenina/metabolismo , Receptores de Interferon/genética , Trissomia , Animais , Vias Biossintéticas/genética , Linhagem Celular , Citocinas/metabolismo , Síndrome de Down/metabolismo , Expressão Gênica , Humanos , Indolamina-Pirrol 2,3,-Dioxigenase/genética , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Metabolômica/métodos , Camundongos Endogâmicos C57BL , Ácido Quinolínico/metabolismo , Receptores de Interferon/metabolismo
6.
Cancer Sci ; 110(11): 3434-3441, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31509303

RESUMO

Immune checkpoint inhibitors have improved the prognosis of advanced melanoma. Although anti-programmed death ligand-1 (PD-L1) is a well-studied biomarker for response to anti-programmed death-1 PD-1 therapy in melanoma, its clinical relevance remains unclear. It has been established that the high expression of indoleamine 2,3-dioxygenase (IDO) is correlated to a response to anti-CTLA-4 treatment in melanoma. However, it is still unknown whether the IDO expression is associated with response to anti-PD-1 therapy in advanced melanoma. In addition, acral and mucosal melanomas, which comprise a great proportion of all melanomas in Asians, are genetically different subtypes from cutaneous melanomas; however, they have not been independently analyzed due to their low frequency in Western countries. To evaluate the association of IDO and PD-L1 expression with response to anti-PD-1 antibody in acral and mucosal melanoma patients, we analyzed 32 Japanese patients with acral and mucosal melanomas treated with anti-PD-1 antibody from the perspective of IDO and PD-L1 expression levels by immunohistochemistry (IHC). Multivariate Cox regression models showed that the low expression of IDO in tumors was associated with poor progression-free survival (HR = 0.33, 95% CI = 0.13-0.81, P = 0.016), whereas PD-L1 expression on tumors was not associated with progression-free survival. Significantly lower expression of IDO in tumors was found in non-responders compared to responders. Assessment of the IDO expression could be useful for the identification of suitable candidates for anti-PD-1 therapy among acral and mucosal melanomas patients. Further validation study is needed to estimate the clinical utility of our findings.


Assuntos
Antígeno CTLA-4/antagonistas & inibidores , Imunoterapia , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Melanoma/enzimologia , Receptor de Morte Celular Programada 1/metabolismo , Neoplasias Cutâneas/enzimologia , Idoso , Grupo com Ancestrais do Continente Asiático , Feminino , Humanos , Japão , Estimativa de Kaplan-Meier , Masculino , Melanoma/mortalidade , Melanoma/patologia , Melanoma/terapia , Pessoa de Meia-Idade , Mutação , Intervalo Livre de Progressão , Modelos de Riscos Proporcionais , Proteínas Proto-Oncogênicas B-raf/genética , Neoplasias Cutâneas/mortalidade , Neoplasias Cutâneas/patologia , Neoplasias Cutâneas/terapia
7.
Immunology ; 158(4): 353-361, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31557322

RESUMO

Reagents that activate the signaling adaptor stimulator of interferon genes (STING) suppress experimentally induced autoimmunity in murine models of multiple sclerosis and arthritis. In this study, we evaluated STING agonists as potential reagents to inhibit spontaneous autoimmune type I diabetes (T1D) onset in non-obese diabetic (NOD) female mice. Treatments with DNA nanoparticles (DNPs), which activate STING when cargo DNA is sensed, delayed T1D onset and reduced T1D incidence when administered before T1D onset. DNP treatment elevated indoleamine 2,3 dioxygenase (IDO) activity, which regulates T-cell immunity, in spleen, pancreatic lymph nodes and pancreas of NOD mice. Therapeutic responses to DNPs were partially reversed by inhibiting IDO and DNP treatment synergized with insulin therapy to further delay T1D onset and reduce T1D incidence. Treating pre-diabetic NOD mice with cyclic guanyl-adenyl dinucleotide (cGAMP) to activate STING directly delayed T1D onset and stimulated interferon-αß (IFN-αß), while treatment with cyclic diguanyl nucleotide (cdiGMP) did not delay T1D onset or induce IFN-αß in NOD mice. DNA sequence analyses revealed that NOD mice possess a STING polymorphism that may explain differential responses to cGAMP and cdiGMP. In summary, STING agonists attenuate T1D progression and DNPs enhance therapeutic responses to insulin therapy.


Assuntos
DNA/uso terapêutico , Diabetes Mellitus Tipo 1/tratamento farmacológico , Insulina/uso terapêutico , Proteínas de Membrana/agonistas , Nanopartículas/uso terapêutico , Linfócitos T/imunologia , Animais , GMP Cíclico/análogos & derivados , GMP Cíclico/metabolismo , DNA/química , Modelos Animais de Doenças , Sinergismo Farmacológico , Feminino , Humanos , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Proteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos NOD , Nanopartículas/química , Nucleotídeos Cíclicos/metabolismo , Polimorfismo Genético , Regulação para Cima
8.
Int J Med Sci ; 16(8): 1107-1115, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31523173

RESUMO

Astragalus membranaceus has been shown to possess anti-inflammation and antitumor properties. Several studies have indicated that extracts of Astragalus membranaceus (PG2) have growth inhibitory effects on tumor. However, the effect of PG2 on enhancing the chemotherapy, modulating tumor immune escape and their mechanism of action is unknown and need further investigation. Connexin (Cx) 43 is ubiquitous in cells and involved in facilitating the passage of chemotherapeutic drugs to bystander tumor cells. The indoleamine 2, 3-dioxygenase (IDO) depletes tryptophan, reduces the active T cell number and destroys immune surveillance. Herein, we provide evidence that the treatment of PG2 induced Cx43 expression, decreases IDO expression and enhances the distribution of chemotherapeutic drug. However, the effects of combination therapy (PG2 plus cisplatin) in animal models significantly retarded tumor growth and prolonged the survival. We believe that the information provided in this study may aid in the design of future therapy of PG2, suggest suitable combinations with chemotherapies.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Astragalus propinquus/química , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Extratos Vegetais/farmacologia , Animais , Antineoplásicos Fitogênicos/administração & dosagem , Antineoplásicos Fitogênicos/farmacologia , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Cisplatino/administração & dosagem , Conexina 43/metabolismo , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/farmacologia , Humanos , Indolamina-Pirrol 2,3,-Dioxigenase/antagonistas & inibidores , Camundongos Endogâmicos C57BL , Extratos Vegetais/administração & dosagem , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Linfócitos T/patologia , Ensaios Antitumorais Modelo de Xenoenxerto
9.
Phytomedicine ; 63: 153055, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31377585

RESUMO

BACKGROUND: Indoleamine 2,3-dioxygenase 1 (IDO1), an important intracellular rate-limiting enzyme in the development of Hepatic fibrosis (HF), and has been proposed as a hallmark of HF. Danshensu (DSS) is a major bioactive component that isolated from a edible traditional Chinese medicinal herb Salviae Miltiorrhizae Radix et Rhizoma (Danshen), while, the anti-HF mode and mechanism of action of DSS have not been fully elucidated. METHODS: Carbon tetrachloride (CCl4)-induced rat HF model and TGF-ß1-induced hepatic stellate cell (HSC) model were employed to assess the in vivo and in vitro anti-HF effects of DSS. HSC-T6 cells stably expressing IDO1, a constitutively active IDO1 mutant, was used to determine the role of JAK2-STAT3 signaling in the DSS's anti-HF effects. RESULTS: We found that intragastric administration of DSS potently reduced fibrosis, inhibited IDO1 expression and STAT3 activity both in vitro and in vivo. Using molecular docking and molecular dynamics analysis, DSS was identified as a novel IDO1 inhibitor. Mechanistic studies indicated that DSS inhibited JAK2-STAT3 signaling, it reduced IDO1 expression, STAT3 phosphorylation and STAT3 nuclear localization. More importantly, overexpression of IDO1 diminished DSS's anti-HF effects. CONCLUSION: Our findings provide a pharmacological justification for the clinical use of DSS in treating HF, and suggest that DSS has the potential to be developed as a modern alternative and/or complimentary agent for HF treatment and prevention.


Assuntos
Indolamina-Pirrol 2,3,-Dioxigenase/antagonistas & inibidores , Janus Quinase 2/metabolismo , Lactatos/farmacologia , Cirrose Hepática Experimental/tratamento farmacológico , Fator de Transcrição STAT3/metabolismo , Animais , Tetracloreto de Carbono/toxicidade , Linhagem Celular , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Células Estreladas do Fígado/efeitos dos fármacos , Células Estreladas do Fígado/metabolismo , Células Estreladas do Fígado/patologia , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Janus Quinase 2/antagonistas & inibidores , Lactatos/química , Cirrose Hepática Experimental/induzido quimicamente , Cirrose Hepática Experimental/metabolismo , Masculino , Simulação de Acoplamento Molecular , Ratos Sprague-Dawley , Fator de Transcrição STAT3/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacos , Fator de Crescimento Transformador beta1/metabolismo , Fator de Crescimento Transformador beta1/farmacologia
10.
Eur J Med Chem ; 182: 111629, 2019 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-31445231

RESUMO

Targeting indoleamine 2,3-dioxygenase 1 (IDO1) has been identified as an attractive approach for the development of cancer immunotherapy. In this study, a series of phosphonamidate ester containing compounds were designed, synthesized and evaluated for their inhibitory activities against IDO1. Among them, compounds 16, 17, and 26 with good IDO1 inhibitory (HeLa IDO1 IC50 = 10-21 nM, hIDO1 IC50 = 78-121 nM) activities were selected for further investigation and showed good physicochemical properties. Furthermore, based on comparable PK profile and excellent IDO2/TDO inhibitory potency, representative compound 16 was selected for further bio-evaluation and characterized with good efficacy in suppressing lung metastasis (77% inhibition rate) of Lewis cells in vivo. Thus, compound 16 could be a potential and efficacious agent for further evaluation.


Assuntos
Antineoplásicos/farmacologia , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Descoberta de Drogas , Inibidores Enzimáticos/farmacologia , Indolamina-Pirrol 2,3,-Dioxigenase/antagonistas & inibidores , Neoplasias Pulmonares/tratamento farmacológico , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Células CACO-2 , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Células HeLa , Humanos , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Estrutura Molecular , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Ratos , Relação Estrutura-Atividade
11.
Folia Biol (Praha) ; 65(2): 101-108, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31464185

RESUMO

IgA nephropathy (IgAN) is the most common glomerulonephritis worldwide. Current studies have shown that the Th17/Treg immune balance may be involved in the occurrence of IgAN, but the exact mechanism is still unclear. Indoleamine 2,3-dioxygenase (IDO) is an enzyme that catalyses degradation of tryptophan (Trp) through the kynurenine (Kyn) pathway; it can control inflammation and immune response by inducing Trp starvation. IDO may be a key molecule in regulating the Th17/Treg immune balance. However, it is not clear whether IDO is involved in the IgAN disease occurrence by regulating the Th17/Treg immune balance. In this study, an IgAN mouse model was established. The mice were intraperitoneally inoculated with IDO inhibitor 1-MT or agonist ISS-ODN to observe whether the IDO signalling pathway participates in the occurrence and development of IgAN by regulating the Th17/Treg immune balance. The results showed that IDO inhibitor 1-MT significantly increased renal injury and glomerular IgA accumulation and up-regulated Th17/Treg and Th17-related cytokine expression in IgAN mice, while ISS-ODN significantly decreased renal injury and glomerular IgA accumulation, down-regulated Th17/Treg expression and inhibited Th17-related cytokine expression in IgAN mice. In conclusion, IDO was involved in the occurrence and progress of IgAN by regulating the Th17/ Treg balance.


Assuntos
Glomerulonefrite por IGA/enzimologia , Glomerulonefrite por IGA/imunologia , Imunidade , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Linfócitos T Reguladores/imunologia , Células Th17/imunologia , Animais , Citocinas/metabolismo , Feminino , Rim/lesões , Rim/metabolismo , Rim/patologia , Camundongos Endogâmicos BALB C , Transdução de Sinais
12.
J Enzyme Inhib Med Chem ; 34(1): 1481-1488, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31423846

RESUMO

Indoleamine 2,3-dioxygenase 1 (IDO1), a tryptophan catabolising enzyme, is known as a tumour cell survival factor that causes immune escape in several types of cancer. Flavonoids of Sophora flavescens have a variety of biological benefits for humans; however, cancer immunotherapy effect has not been fully investigated. The flavonoids (1-6) isolated from S. flavescens showed IDO1 inhibitory activities (IC50 4.3-31.4 µM). The representative flavonoids (4-6) of S. flavescens were determined to be non-competitive inhibitors of IDO1 by kinetic analyses. Their binding affinity to IDO1 was confirmed using thermal stability and surface plasmon resonance (SPR) assays. The molecular docking analysis and mutagenesis assay revealed the structural details of the interactions between the flavonoids (1-6) and IDO1. These results suggest that the flavonoids (1-6) of S. flavescens, especially kushenol E (6), as IDO1 inhibitors might be useful in the development of immunotherapeutic agents against cancers.


Assuntos
Antineoplásicos/farmacologia , Inibidores Enzimáticos/farmacologia , Flavonoides/farmacologia , Indolamina-Pirrol 2,3,-Dioxigenase/antagonistas & inibidores , Sophora/química , Antineoplásicos/química , Antineoplásicos/isolamento & purificação , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/química , Inibidores Enzimáticos/isolamento & purificação , Flavonoides/química , Flavonoides/isolamento & purificação , Células HeLa , Humanos , Indolamina-Pirrol 2,3,-Dioxigenase/genética , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Modelos Moleculares , Estrutura Molecular , Mutagênese Sítio-Dirigida , Relação Estrutura-Atividade , Células Tumorais Cultivadas
13.
PLoS Pathog ; 15(8): e1007955, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31461509

RESUMO

Most intracellular pathogens that reside in a vacuole prevent transit of their compartment to lysosomal organelles. Effector mechanisms induced by the pro-inflammatory cytokine Interferon-gamma (IFNγ) can promote the delivery of pathogen-occupied vacuoles to lysosomes for proteolytic degradation and are therefore important for host defense against intracellular pathogens. The bacterial pathogen Coxiella burnetii is unique in that, transport to the lysosome is essential for replication. The bacterium modulates membrane traffic to create a specialized autophagolysosomal compartment called the Coxiella-containing vacuole (CCV). Importantly, IFNγ signaling inhibits intracellular replication of C. burnetii, raising the question of which IFNγ-activated mechanisms restrict replication of a lysosome-adapted pathogen. To address this question, siRNA was used to silence a panel of IFNγ-induced genes in HeLa cells to identify genes required for restriction of C. burnetii intracellular replication. This screen demonstrated that Indoleamine 2,3-dioxygenase 1 (IDO1) contributes to IFNγ-mediated restriction of C. burnetii. IDO1 is an enzyme that catabolizes cellular tryptophan to kynurenine metabolites thereby reducing tryptophan availability in cells. Cells deficient in IDO1 function were more permissive for C. burnetii replication when treated with IFNγ, and supplementing IFNγ-treated cells with tryptophan enhanced intracellular replication. Additionally, ectopic expression of IDO1 in host cells was sufficient to restrict replication of C. burnetii in the absence of IFNγ signaling. Using differentiated THP1 macrophage-like cells it was determined that IFNγ-activation resulted in IDO1 production, and that supplementation of IFNγ-activated THP1 cells with tryptophan enhanced C. burnetii replication. Thus, this study identifies IDO1 production as a key cell-autonomous defense mechanism that limits infection by C. burnetii, which suggests that peptides derived from hydrolysis of proteins in the CCV do not provide an adequate supply of tryptophan for bacterial replication.


Assuntos
Coxiella burnetii/patogenicidade , Interações Hospedeiro-Patógeno , Indolamina-Pirrol 2,3,-Dioxigenase/antagonistas & inibidores , Lisossomos/virologia , Febre Q/prevenção & controle , RNA Interferente Pequeno/genética , Replicação Viral/genética , Coxiella burnetii/efeitos dos fármacos , Células HeLa , Humanos , Indolamina-Pirrol 2,3,-Dioxigenase/genética , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Interferon gama/farmacologia , Macrófagos/metabolismo , Macrófagos/patologia , Macrófagos/virologia , Febre Q/genética , Febre Q/virologia , Triptofano/metabolismo
14.
Int J Mol Sci ; 20(13)2019 Jun 28.
Artigo em Inglês | MEDLINE | ID: mdl-31261636

RESUMO

Protein-ligand docking is a widely used method to generate solutions for the binding of a small molecule with its target in a short amount of time. However, these methods provide identification of physically sound protein-ligand complexes without a complete view of the binding process dynamics, which has been recognized to be a major discriminant in binding affinity and ligand selectivity. In this paper, a novel piece of open-source software to approach this problem is presented, called GPathFinder. It is built as an extension of the modular GaudiMM platform and is able to simulate ligand diffusion pathways at atomistic level. The method has been benchmarked on a set of 20 systems whose ligand-binding routes were studied by other computational tools or suggested from experimental "snapshots". In all of this set, GPathFinder identifies those channels that were already reported in the literature. Interestingly, the low-energy pathways in some cases indicate novel possible binding routes. To show the usefulness of GPathFinder, the analysis of three case systems is reported. We believe that GPathFinder is a software solution with a good balance between accuracy and computational cost, and represents a step forward in extending protein-ligand docking capacities, with implications in several fields such as drug or enzyme design.


Assuntos
Simulação de Acoplamento Molecular/métodos , Software , Algoritmos , Aquaporinas/química , Aquaporinas/metabolismo , Proteínas Arqueais/química , Proteínas Arqueais/metabolismo , Sítios de Ligação , Citocromo P-450 CYP2C19/química , Citocromo P-450 CYP2C19/metabolismo , Humanos , Indolamina-Pirrol 2,3,-Dioxigenase/química , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Ligantes , Ligação Proteica
15.
Gastroenterology ; 157(4): 1093-1108.e11, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31325428

RESUMO

BACKGROUND & AIMS: Inflammation, injury, and infection up-regulate expression of the tryptophan metabolizing enzyme indoleamine 2,3-dioxygenase 1 (IDO1) in the intestinal epithelium. We studied the effects of cell-specific IDO1 expression in the epithelium at baseline and during intestinal inflammation in mice. METHODS: We generated transgenic mice that overexpress fluorescence-tagged IDO1 in the intestinal epithelium under control of the villin promoter (IDO1-TG). We generated intestinal epithelial spheroids from mice with full-length Ido1 (controls), disruption of Ido1 (knockout mice), and IDO1-TG and analyzed them for stem cell and differentiation markers by real-time polymerase chain reaction, immunoblotting, and immunofluorescence. Some mice were gavaged with enteropathogenic Escherichia coli (E2348/69) to induce infectious ileitis, and ileum contents were quantified by polymerase chain reaction. Separate sets of mice were given dextran sodium sulfate or 2,4,6-trinitrobenzenesulfonic acid to induce colitis; intestinal tissues were analyzed by histology. We utilized published data sets GSE75214 and GDS2642 of RNA expression data from ilea of healthy individuals undergoing screening colonoscopies (controls) and patients with Crohn's disease. RESULTS: Histologic analysis of small intestine tissues from IDO1-TG mice revealed increases in secretory cells. Enteroids derived from IDO1-TG intestine had increased markers of stem, goblet, Paneth, enteroendocrine, and tuft cells, compared with control enteroids, with a concomitant decrease in markers of absorptive cells. IDO1 interacted non-enzymatically with the aryl hydrocarbon receptor to inhibit activation of NOTCH1. Intestinal mucus layers from IDO1-TG mice were 2-fold thicker than mucus layers from control mice, with increased proportions of Akkermansia muciniphila and Mucispirillum schaedleri. Compared to controls, IDO1-TG mice demonstrated an 85% reduction in ileal bacteria (P = .03) when challenged with enteropathogenic E coli, and were protected from immune infiltration, crypt dropout, and ulcers following administration of dextran sodium sulfate or 2,4,6-trinitrobenzenesulfonic acid. In ilea of Crohn's disease patients, increased expression of IDO1 correlated with increased levels of MUC2, LYZ1, and aryl hydrocarbon receptor, but reduced levels of SLC2A5. CONCLUSIONS: In mice, expression of IDO1 in the intestinal epithelial promotes secretory cell differentiation and mucus production; levels of IDO1 are positively correlated with secretory cell markers in ilea of healthy individuals and Crohn's disease patients. We propose that IDO1 contributes to intestinal homeostasis.


Assuntos
Bactérias/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Diferenciação Celular , Microbioma Gastrointestinal , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Doenças Inflamatórias Intestinais/enzimologia , Doenças Inflamatórias Intestinais/microbiologia , Mucosa Intestinal/enzimologia , Mucosa Intestinal/microbiologia , Receptores de Hidrocarboneto Arílico/metabolismo , Receptores Notch/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Estudos de Casos e Controles , Linhagem Celular , Linhagem da Célula , Modelos Animais de Doenças , Células Epiteliais/enzimologia , Células Epiteliais/microbiologia , Células Epiteliais/patologia , Genótipo , Humanos , Indolamina-Pirrol 2,3,-Dioxigenase/deficiência , Indolamina-Pirrol 2,3,-Dioxigenase/genética , Doenças Inflamatórias Intestinais/genética , Doenças Inflamatórias Intestinais/patologia , Mucosa Intestinal/patologia , Camundongos Knockout , Fenótipo , Receptores de Hidrocarboneto Arílico/genética , Receptores Notch/genética , Via Secretória , Transdução de Sinais , Células-Tronco/enzimologia , Células-Tronco/microbiologia , Células-Tronco/patologia
16.
Mol Med Rep ; 20(3): 2954-2962, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31322231

RESUMO

A number of studies have linked abnormalities in the function of the serotonergic and noradrenergic systems to the pathophysiology of depression. It has been reported that selective serotonin reuptake inhibitors promote the expression of tryptophan hydroxylase (TPH), which is involved in the synthesis of serotonin. However, limited evidence of TPH alteration has been found in selective serotonin and noradrenaline reuptake inhibitors (SNRIs), and more key enzymes need to be investigated. The aim of the present study was to determine whether venlafaxine (VLX; a classical SNRI) regulates TPH and other key enzymes responsible for the synthesis and metabolism of monoaminergic transmitters in rats with chronic unpredictable stress (CUS). The present results suggested that CUS­exposed rats exhibited decreased locomotor activity in the open­field test and increased immobility time in the forced swim test, as compared with the controls. Pretreatment with VLX (20 mg/kg) significantly increased locomotor activity and reduced immobility time in the CUS­exposed rats. In addition, VLX (20 mg/kg) treatment prevented the CUS­induced reduction in tyrosine hydroxylase and TPH expression in the cortex and hippocampus. Furthermore, VLX alleviated the CUS­induced oxidative stress in the serum, cortex and hippocampus. However, VLX administration did not have an effect on indoleamine­2,3­dioxygenase overexpression in the hippocampus. It was therefore concluded that the regulation of abnormalities in the synthesis and metabolism of monoaminergic transmitters may be associated with the antidepressant effects of VLX, suggesting that multimodal pharmacological treatments can efficiently treat depression.


Assuntos
Antidepressivos de Segunda Geração/uso terapêutico , Monoaminas Biogênicas/metabolismo , Depressão/tratamento farmacológico , Triptofano Hidroxilase/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismo , Cloridrato de Venlafaxina/uso terapêutico , Animais , Antidepressivos de Segunda Geração/farmacologia , Comportamento Animal/efeitos dos fármacos , Depressão/etiologia , Depressão/fisiopatologia , Hipocampo/efeitos dos fármacos , Hipocampo/fisiopatologia , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Masculino , Monoaminoxidase/metabolismo , Ratos Sprague-Dawley , Estresse Fisiológico/efeitos dos fármacos , Cloridrato de Venlafaxina/farmacologia
17.
Transplant Proc ; 51(6): 2141-2145, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31307771

RESUMO

Indoleamine 2,3-dioxygenase (IDO) is an important enzyme in immune response regulation; cells that express IDO can suppress T-cell responses. Endothelial cells (ECs) play an important role in graft rejection. Therefore, we investigated the role of IDO expression by vascular ECs in immunoregulation. We demonstrated that interferon gamma can upregulate IDO expression in cultured ECs. Moreover, IDO induction by interferon gamma required IKKα activation, a part of the noncanonical NF-κB signaling pathway. In addition, IκB kinase α (IKKα) silencing resulted in significantly reduced IPO expression, demonstrating an essential role of NF-κB signaling pathway in IDO activation in vitro. These results may have an implication for regulating the immune response to alloantigens. The results obtained are important not only in understanding the role of ECs in the regulation of the transplantation immune response, but also in determining a potential therapeutic target for inhibiting allograft rejection.


Assuntos
Células Endoteliais/metabolismo , Rejeição de Enxerto/metabolismo , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Interferon gama/metabolismo , NF-kappa B/metabolismo , Animais , Regulação Enzimológica da Expressão Gênica , Rejeição de Enxerto/imunologia , Humanos , Quinase I-kappa B/metabolismo , Ratos , Transdução de Sinais/efeitos dos fármacos , Linfócitos T/imunologia
18.
J Exp Clin Cancer Res ; 38(1): 314, 2019 Jul 17.
Artigo em Inglês | MEDLINE | ID: mdl-31315643

RESUMO

BACKGROUND: IDO1 (Indoleamine 2,3-dioxygenase 1) inhibits host anti-tumor immune response by exhausting tryptophan in tumor microenvironment, but the pathogenic mechanisms of IDO1 in gastric cancer (GC) cells need to be further explored. METHODS: The aim of this study was to use CCLE (Cancer Cell Line Encyclopedia) transcriptomic data of GC cell lines for WGCNA (Weighted Gene Co-expression Network Analysis) analysis, and explore the potential functions and mechanisms of IDO1 in GC progression in vitro and in vivo. RESULTS: The higher expression level of IDO1 was identified in 4 out of 7 GC cell lines. Increased IDO1 expression strongly promoted cell migration via its metabolite kynurenine and was associated with pathways of immune activation according to GSEA (Gene Set Enrichment Analysis). The functions of IDO1 were closely associated with extracellular matrix, collagen metabolic and catabolic process by WGCNA analysis. Among five hub genes (AXL, SGCE, COL12A1, ANTXR1, LOXL2), COL12A1 and LOXL2 were upregulated in GC tissues. IDO1 disclosed positive correlation with six collagen genes by coefficient matrix diagram. Knockdown of IDO1 decreased the expression of LOXL2, COL6A1, COL6A2 and COL12A1 in GC cells in both mRNA and protein levels. Of them, knockdown of COL12A1 inhibited cell migration more apparently than knockdown of others. IDO1 and COL12A1 revealed synergistic efficacy on promoting cell migration via a positive feedback sustained by MAPK pathway. This bioprocess was mediated by IDO1 metabolite kynurenine and integrin ß1. A popliteal lymph nodemetastasis model was established for verifying metastatic promotion of IDO1 and COL12A1 in GC. CONCLUSIONS: IDO1 and COL12A1 synergistically promoted GC metastasis. The novel findings suggested that both IDO1 and COL12A1 may be promising targets on anti-cancer treatment in GC.


Assuntos
Colágeno Tipo XII/metabolismo , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Cinurenina/metabolismo , Metástase Linfática/patologia , Neoplasias Gástricas/patologia , Animais , Linhagem Celular Tumoral , Movimento Celular , Colágeno Tipo XII/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Indolamina-Pirrol 2,3,-Dioxigenase/genética , Metástase Linfática/genética , Sistema de Sinalização das MAP Quinases , Camundongos , Transplante de Neoplasias , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Microambiente Tumoral , Regulação para Cima
19.
Cells ; 8(7)2019 07 12.
Artigo em Inglês | MEDLINE | ID: mdl-31336879

RESUMO

Galectin-3 regulates numerous biological processes in the gut. We investigated molecular mechanisms responsible for the Galectin-3-dependent regulation of colon inflammation and evaluated whether Galectin-3 may be used as biomarker for monitoring the progression of ulcerative colitis (UC). The differences in disease progression between dextran sodium sulphate-treated wild type and Galectin-3-deficient mice were investigated and confirmed in clinical settings, in 65 patients suffering from mild, moderate, and severe colitis. During the induction phase of colitis, Galectin-3 promoted interleukin-1ß-induced polarization of colonic macrophages towards inflammatory phenotype. In the recovery phase of colitis, Galectin-3 was required for the immunosuppressive function of regulatory dendritic cells (DCs). Regulatory DCs in Galectin-3:Toll-like receptor-4:Kynurenine-dependent manner promoted the expansion of colon-infiltrated T regulatory cells (Tregs) and suppressed Th1 and Th17 cell-driven colon inflammation. Concentration of Galectin-3 in serum and stool samples of UC patients negatively correlated with clinical, endoscopic, and histological parameters of colitis. The cutoff serum values of Galectin-3 that allowed the discrimination of mild from moderate and moderate from severe colitis were 954 pg/mL and 580 pg/mL, respectively. Fecal levels of Galectin-3 higher than 553.44 pg/mL indicated attenuation of UC. In summing up, Galectin-3 regulates the cross-talk between colon-infiltrating DCs and Tregs and represents a new biomarker for monitoring the progression of UC.


Assuntos
Colite Ulcerativa/patologia , Células Dendríticas/patologia , Galectina 3/fisiologia , Linfócitos T Reguladores/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Biomarcadores/metabolismo , Células Cultivadas , Progressão da Doença , Feminino , Humanos , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade
20.
Eur J Med Chem ; 179: 38-55, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31233921

RESUMO

Indoleamine 2, 3-dioxygenase 1 (IDO1) plays a key role in tryptophan catabolism which is an important mechanism in immune tolerance. The small molecule epacadostat is the most advanced IDO1 inhibitor, but its phase III trials as a single agent or in combinations with PD-1 antibody failed to show appreciable objective responses. To gain more insight on the antitumor efficacy of IDO1 inhibitors, we have designed a series of analogues of epacadostat by incorporating a cyclic aminosulfonamide moiety as the sidechain capping functionality. Compound 5a was found to display significant potency against recombinant hIDO1 and hIDO1 expressed HEK293 cancer cells. This compound has improved physico-chemical properties, acceptable PK parameters as well as optimal cardiac safety. Similar to epacadostat, 5a is ineffective as single agent in the CT-26 syngeneic xenograft model, however, the combination of 5a with PD-1 antibody showed both elevated tumor growth inhibition and prolonged median life span.


Assuntos
Aminas/farmacologia , Antineoplásicos/farmacologia , Desenho de Drogas , Inibidores Enzimáticos/farmacologia , Indolamina-Pirrol 2,3,-Dioxigenase/antagonistas & inibidores , Oxidiazóis/farmacologia , Aminas/síntese química , Aminas/química , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Feminino , Células HEK293 , Humanos , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Estrutura Molecular , Oxidiazóis/síntese química , Oxidiazóis/química , Ratos , Ratos Sprague-Dawley , Relação Estrutura-Atividade
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