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1.
Cancer Sci ; 110(9): 2846-2855, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31325200

RESUMO

DNA markers for pancreatic ductal adenocarcinoma (PDAC) are urgently needed for detection of minimally invasive disease. The epigenetic relevance of the cysteine dioxygenase 1 gene (CDO1) has been never investigated in PDAC. Three studies, including cellular experiments, tissue validation, and pilot testing for pancreatic cytology, were carried out. Promoter DNA methylation value (MV) of CDO1 was quantified by quantitative methylation-specific PCR. CDO1 expression was consistent with its promoter DNA methylation in 7 PDAC cell lines. In 160 retrospectively collected primary PDAC tumor tissues, MV was significantly higher compared to the corresponding noncancerous pancreas (area under the receiver operating characteristic curve [AUC] = 0.97, P < .0001), and CDO1 hypermethylation was highly specific to PDAC tumor tissues. CDO1 hypermethylation group (MV over 19) was significantly associated with diverse prognostic factors in PDAC. Surprisingly, it was significantly higher in prospectively collected PDAC cytology samples (n = 37), including both pancreatic juice (n = 12) and endoscopic ultrasound-fine needle aspiration (EUS-FNA) cytology (n = 25) compared to pancreatic benign diseases (AUC = 0.96, P < .0001). Detection of PDAC was confirmed by DNA testing in 35 of 37 patients (95% sensitivity); thus, it was more sensitive than cytology (33%) or EUS-FNA cytology (88%). Promoter DNA methylation of CDO1 is extremely specific for PDAC tumors, and accumulates with PDAC tumor progression. It could be a definitive diagnostic marker of PDAC in pancreatic juice or EUS-FNA cytology.


Assuntos
Biomarcadores Tumorais/genética , Carcinoma Ductal Pancreático/diagnóstico , Cisteína Dioxigenase/genética , Metilação de DNA , Neoplasias Pancreáticas/diagnóstico , Idoso , Carcinoma Ductal Pancreático/patologia , Linhagem Celular Tumoral , Cisteína Dioxigenase/metabolismo , Progressão da Doença , Aspiração por Agulha Fina Guiada por Ultrassom Endoscópico , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Pâncreas/patologia , Suco Pancreático/metabolismo , Neoplasias Pancreáticas/patologia , Projetos Piloto , Prognóstico , Regiões Promotoras Genéticas , Estudos Prospectivos , Estudos Retrospectivos , Sensibilidade e Especificidade
2.
Artigo em Inglês | MEDLINE | ID: mdl-31176866

RESUMO

Cysteine oxygenase (CDO) is a mononuclear nonhemoglobin enzyme that catalyzes the production of taurine through the cysteine (Cys) pathway and plays a key role in the biosynthesis of taurine in mammals. However, the function of CDOs in bony fish remains poorly understood. In this study, we cloned CDO genes (CaCDO1 and CaCDO2) from Carassius auratus. The cDNA sequences of both CaCDO1 and CaCDO2 encoded putative proteins with 201 amino acids, which included structural features typical of the CDO protein family. Multiple sequence alignment and phylogenetic analysis showed that CaCDO1 and CaCDO2 shared high sequence identities and similarities with C. carpio homologs. Quantitative real-time polymerase chain reaction (qRT-PCR) results revealed that CaCDO1 and CaCDO2 were both broadly expressed in all selected tissues and developmental stages in C. auratus but had differing mRNA levels. In addition, compared to those of the taurine-free group, the in vivo mRNA expression levels of both CaCDO1 and CaCDO2 significantly decreased with increasing dietary taurine levels from 1.0 to 9.0 g/kg. Furthermore, in vitro taurine treatments showed similar inhibitory effects on the expression of CaCDO1 and CaCDO2 in the intestines of C. auratus. Our results also showed that the mRNA expression of CaCDO2 in the intestines was higher than that of CaCDO1 in response to in vivo and in vitro taurine supplementation. Overall, these data may provide new insights into the regulation of fish CDO expression and provide valuable knowledge for improving dietary formulas in aquaculture.


Assuntos
Cisteína Dioxigenase/genética , Cisteína Dioxigenase/metabolismo , Proteínas de Peixes/genética , Proteínas de Peixes/metabolismo , Carpa Dourada/genética , Carpa Dourada/metabolismo , Taurina/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , DNA Complementar/genética , Regulação da Expressão Gênica no Desenvolvimento , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Carpa Dourada/crescimento & desenvolvimento , Isoenzimas/genética , Isoenzimas/metabolismo , Filogenia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Taurina/farmacologia , Distribuição Tecidual
3.
Anticancer Res ; 39(5): 2289-2298, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31092420

RESUMO

BACKGROUND/AIM: We previously identified that promoter DNA methylation of cysteine dioxygenase type 1 (CDO1) and homeobox only protein homeobox (HOPX) were both cancer specific, and have a clinical potential as prognostic biomarkers in breast cancer (BC). The present study compared the differential prognostic relevance of methylation status of the CDO1 and HOPX genes in BC. MATERIALS AND METHODS: Methylation levels (TaqMethVs) were quantified in 7 BC cell lines and 133 BC patients by TaqMan methylation-specific PCR and functional traits were explored for CDO1. RESULTS: TaqMethVs were associated between CDO1 and HOPX (r2=0.072, p=0.002). Multivariate Cox proportional hazards model could identify CDO1 hypermethylation as well as Ki-67 as independent prognostic factors related to disease-specific survival (p=0.016, p<0.001). Overexpression of CDO1 decreased the anchorage-independent growth capacity in BC cell lines. CONCLUSION: CDO1 is a definite tumor suppressor gene, while its prognostic relevance was more than expected in the context of its functional relevance.


Assuntos
Neoplasias da Mama/genética , Cisteína Dioxigenase/genética , Metilação de DNA/genética , Proteínas de Homeodomínio/genética , Proteínas Supressoras de Tumor/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Intervalo Livre de Doença , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Antígeno Ki-67/genética , Pessoa de Meia-Idade , Prognóstico , Regiões Promotoras Genéticas , Modelos de Riscos Proporcionais
4.
PLoS One ; 14(1): e0211108, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30677088

RESUMO

Although small bowel cancer (SBC) is extremely rare, its prognosis is poor, and molecular mechanism of the SBC development remains unclear. The aim of our study is to elucidate whether DNA methylation of the promoter region of the cancer-specific methylation gene, cysteine dioxygenase 1 (CDO1), contributes to the carcinogenic process in SBC. The study group comprised patients with 53 patients with SBC, 107 colorectal cancer (CRC), and other rare tumors of the small intestine such as 4 malignant lymphomas, 2 leiomyosarcomas, and 9 gastrointestinal stromal tumors. We analyzed the extent of methylation in each tissue using quantitative TaqMan methylation-specific PCR for CDO1. Significantly higher CDO1 methylation was observed in cancer tissues compared with non-cancerous mucosa of the small intestine (ROC = 0.96). Among the various clinicopathological factors, positive correlation of CDO1 methylation with tumor diameter was observed (R = 0.31, p = 0.03), and the CDO1 methylation level was a possible prognostic factor for relapse-free survival (p = 0.09). Compared with CRC, SBC had a significantly poorer prognosis (p = 0.007) and displayed a significantly higher CDO1 methylation level (p < 0.0001). Intriguingly, especially in pStage I/II, there were robust prognostic difference between SBC and CRC (p = 0.08 / p < 0.0001), which may reflect CDO1 methylation status (p = 0.02 / p = 0.001). Among small bowel tumors, CDO1 methylation in SBC was higher in order of malignant lymphoma, cancer, and leiomyosarcoma/GIST (p = 0.002) by ANOVA. The CDO1 gene shows extremely cancer-specific hypermethylation, and it can be a prognostic marker in SBC.


Assuntos
Neoplasias Colorretais , Cisteína Dioxigenase , Metilação de DNA , DNA de Neoplasias , Leiomiossarcoma , Linfoma , Proteínas de Neoplasias , Idoso , Linhagem Celular Tumoral , Neoplasias Colorretais/enzimologia , Neoplasias Colorretais/genética , Neoplasias Colorretais/mortalidade , Neoplasias Colorretais/patologia , Cisteína Dioxigenase/genética , Cisteína Dioxigenase/metabolismo , DNA de Neoplasias/genética , DNA de Neoplasias/metabolismo , Intervalo Livre de Doença , Feminino , Tumores do Estroma Gastrointestinal/enzimologia , Tumores do Estroma Gastrointestinal/genética , Tumores do Estroma Gastrointestinal/mortalidade , Tumores do Estroma Gastrointestinal/patologia , Humanos , Leiomiossarcoma/enzimologia , Leiomiossarcoma/genética , Leiomiossarcoma/mortalidade , Leiomiossarcoma/patologia , Linfoma/enzimologia , Linfoma/genética , Linfoma/mortalidade , Linfoma/patologia , Masculino , Pessoa de Meia-Idade , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Estudos Retrospectivos , Taxa de Sobrevida
5.
Enzyme Microb Technol ; 120: 61-68, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30396400

RESUMO

Thiol dioxygenases are non-heme mononuclear-iron proteins and belong to the cupin superfamily. In 2014, mercaptosuccinate dioxygenase (Msdo) of Variovorax paradoxus B4 was identified as another bacterial cysteine dioxygenase (Cdo) homolog catalyzing the conversion of mercaptosuccinate (MS) into succinate and sulfite. To gain further insights into potentially important amino acid residues for enzyme activity, seven enzyme variants were generated and analyzed. (i) Three variants comprised the substitution of one conserved histidine residue each by leucine, either supposed to be mandatory for coordination of the Fe(II) cofactor (H93 and H95) or to be important for substrate positioning within the active site (H163). The corresponding enzyme variants were completely inactive confirming their essential roles for enzyme activity. (ii) Mutation C100S resulted as well in an inactive enzyme demonstrating its importance for either stability or activity of the protein. (iii) For eukaryotic Cdo, a hydrogen bond network for substrate positioning was postulated, and the corresponding amino acids are basically present in Msdo. Albeit the MsdoQ64A mutation exhibited an increased Km of 0.29 mM when compared to the wildtype with 0.06 mM, it did not significantly affect the specific activity. (iv) The variant MsdoR66A showed only very low activity even when high amounts of enzyme were applied indicating that this residue might be important for catalysis. (v) No strong effect had the mutation Y165F for which a specific enzyme activity of 10.22 µmol min-1 mg-1 protein and a Km value of 0.06 mM with high similarity to those of the wildtype enzyme were obtained. This residue corresponds to Y157 of human Cdo, which is part of the catalytic triad and is supposed to be involved in substrate positioning. Apparently, another residue could fulfill this role in Msdo, since the loss of Y165 did not have a strong effect.


Assuntos
Aminoácidos/química , Comamonadaceae/enzimologia , Cisteína Dioxigenase/metabolismo , Dioxigenases/metabolismo , Mutação , Tiomalatos/metabolismo , Sequência de Aminoácidos , Aminoácidos/genética , Catálise , Domínio Catalítico , Cisteína Dioxigenase/química , Cisteína Dioxigenase/genética , Dioxigenases/química , Dioxigenases/genética , Mutagênese Sítio-Dirigida , Homologia de Sequência , Especificidade por Substrato
6.
PLoS One ; 13(5): e0194785, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29746493

RESUMO

BACKGROUND: Progression of colorectal cancer (CRC) has been explained by genomic abnormalities along with the adenoma-carcinoma sequence theory (ACS). The aim of our study is to elucidate whether the promoter DNA methylation of the cancer-specific methylation gene, cysteine dioxygenase 1 (CDO1), contributes to the carcinogenic process in CRC. METHODS: The study group comprised 107 patients with CRC who underwent surgical resection and 90 adenomas treated with endoscopic resection in the Kitasato University Hospital in 2000. We analyzed the extent of methylation in each tissue using quantitative TaqMan methylation-specific PCR for CDO1. RESULTS: The methylation level increased along with the ACS process (p < 0.0001), and statistically significant differences were found between normal-appearing mucosa (NAM) and low-grade adenoma (p < 0.0001), and between low-grade adenoma and high-grade adenoma (p = 0.01), but not between high-grade adenoma and cancer with no liver metastasis. Furthermore, primary CRC cancers with liver metastasis harbored significantly higher methylation of CDO1 than those without liver metastasis (p = 0.02). As a result, the area under the curve by CDO1 promoter methylation was 0.96, 0.80, and 0.67 to discriminate cancer from NAM, low-grade adenoma from NAM, and low-grade adenoma from high-grade adenoma, respectively. CONCLUSIONS: CDO1 methylation accumulates during the ACS process, and consistently contributes to CRC progression.


Assuntos
Adenocarcinoma/diagnóstico , Adenoma/diagnóstico , Neoplasias Colorretais/diagnóstico , Cisteína Dioxigenase/genética , Metilação de DNA , Regiões Promotoras Genéticas , Adenocarcinoma/enzimologia , Adenocarcinoma/genética , Adenoma/enzimologia , Adenoma/genética , Adulto , Idoso , Biomarcadores Tumorais , Neoplasias Colorretais/enzimologia , Neoplasias Colorretais/genética , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Prognóstico
7.
PLoS One ; 12(11): e0188178, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-29161283

RESUMO

BACKGROUND: Aberrant promoter DNA methylation of the cysteine dioxygenase 1 (CDO1) gene is found in various human cancers and is associated with clinical outcome. In this study, we assessed for the first time the clinicopathological significance of CDO1 methylation in primary gallbladder cancer (GBC) in comparison with non-malignant gallbladder disease. METHODS: CDO1 DNA methylation was quantified using quantitative TaqMan methylation specific PCR (Q-MSP) in 99 primary GBC patients together with the 78 corresponding non-tumor tissues and 26 benign gallbladder disease (including 7 patients with xanthogranulomatous cholecystitis) who underwent surgical resection between 1986 and 2014. RESULTS: The average CDO1 TaqMeth value of primary GBCs was 23.5±26. These values were significantly higher than those of corresponding non-tumor tissues (average 8±13, p < .0001) and diseased gallbladder tissues from patients with benign gallbladder diseases (average 0.98±1.6, p < .0001). CDO1 hypermethylation is also found in xanthogranulomatous cholecystitis. Using a cut-off value of 17.7, GBC cases with CDO1 hypermethylation (n = 47) showed significantly poorer prognosis than those with CDO1 hypomethylation (n = 52) (p = 0.0023). Multivariate Cox proportional hazards analysis identified that CDO1 hypermethylation was an independent prognostic factor. Notably, CDO1 hypermethylation showed prognostic relevance, especially in stage II GBC, in which it is highly anticipated to work as a predictive marker for candidates of adjuvant therapy. CONCLUSIONS: Promoter NA methylation of CDO1 was demonstrated for the first time to be a cancer-associated methylation in primary GBC, and it has the potential to be a prognostic biomarker of GBC for high-risk patients with stage II GBC.


Assuntos
Biomarcadores Tumorais/genética , Cisteína Dioxigenase/genética , Metilação de DNA/genética , Doenças da Vesícula Biliar/genética , Neoplasias da Vesícula Biliar/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Colecistite/complicações , Colecistite/genética , Colecistite/patologia , Feminino , Doenças da Vesícula Biliar/patologia , Neoplasias da Vesícula Biliar/complicações , Neoplasias da Vesícula Biliar/patologia , Humanos , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Prognóstico , Regiões Promotoras Genéticas , Xantomatose/complicações , Xantomatose/genética , Xantomatose/patologia
8.
Neoplasia ; 19(12): 1022-1032, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-29144989

RESUMO

BACKGROUND: Ferroptosis is a recently discovered form of iron-dependent nonapoptotic cell death. It is characterized by loss of the activity of the lipid repair enzyme, glutathione peroxidase 4 (GPX4), and accumulation of lethal reactive lipid oxygen species. However, we still know relatively little about ferroptosis and its molecular mechanism in gastric cancer (GC) cells. Here, we demonstrate that erastin, a classic inducer of ferroptosis, induces this form of cell death in GC cells and that cysteine dioxygenase 1 (CDO1) plays an important role in this process. METHODS: We performed quantitative real-time polymerase chain reaction, Western blotting, cell viability assay, reactive oxygen species (ROS) assay, glutathione assay, lipid peroxidation assay, RNAi and gene transfection, immunofluorescent staining, dual-luciferase reporter assay, transmission electron microscopy, and chromatin immunoprecipitation assay to study the regulation of ferroptosis in GC cells. Mouse xenograft assay was used to figure out the mechanism in vivo. RESULTS: Silencing CDO1 inhibited erastin-induced ferroptosis in GC cells both in vitro and in vivo. Suppression of CDO1 restored cellular GSH levels, prevented ROS generation, and reduced malondialdehyde, one of the end products of lipid peroxidation. In addition, silencing COO1 maintained mitochondrial morphologic stability in erastin-treated cells. Mechanistically, c-Myb transcriptionally regulated CDO1, and inhibition of CDO1 expression upregulated GPX4 expression. CONCLUSIONS: Our findings give a better understanding of ferroptosis and its molecular mechanism in GC cells, gaining insight into ferroptosis-mediated cancer treatment.


Assuntos
Cisteína Dioxigenase/metabolismo , Piperazinas/metabolismo , Neoplasias Gástricas/metabolismo , Animais , Biomarcadores , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cisteína Dioxigenase/genética , Modelos Animais de Doenças , Heme/metabolismo , Xenoenxertos , Humanos , Ferro/metabolismo , Camundongos , Modelos Biológicos , Proteínas Proto-Oncogênicas c-myb/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Neoplasias Gástricas/patologia
9.
Int J Exp Pathol ; 98(2): 52-66, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-28439920

RESUMO

Amino acid metabolism is a significant metabolic activity in humans, especially of sulphur-containing amino acids, methionine and cysteine (Cys). Cys is cytotoxic and neurotoxic in nature; hence, mammalian cells maintain a constant intracellular level of Cys. Metabolism of Cys is mainly regulated by two thiol dioxygenases: cysteine dioxygenase (CDO) and 2-aminoethanethiol dioxygenase (ADO). CDO and ADO are the only human thiol dioxygenases reported with a role in Cys metabolism and localized to mitochondria. This metabolic pathway is important in various human disorders, as it is responsible for the synthesis of antioxidant glutathione and is also for the synthesis of hypotaurine and taurine. CDO is the most extensively studied protein, whose high-resolution crystallographic structures have been solved. As compared to CDO, ADO is less studied, even though it has a key role in cysteamine metabolism. To further understand ADO's structure and function, the three-dimensional structures have been predicted from I-TASSER and SWISS-MODEL servers and validated with PROCHECK software. Structural superimposition approach using iPBA web server further confirmed near-identical structures (including active sites) for the predicted protein models of ADO as compared to CDO. In addition, protein-protein interaction and their association in patho-physiology are crucial in understanding protein functions. Both ADO and CDO interacting partner profiles have been presented using STRING database. In this study, we have predicted a 3D model structure for ADO and summarized the biological roles and the pathological consequences which are associated with the altered expression and functioning of ADO and CDO in case of cancer, neurodegenerative disorders and other human diseases.


Assuntos
Cisteína Dioxigenase/metabolismo , Cisteína/metabolismo , Animais , Carotenoides/genética , Carotenoides/metabolismo , Cisteína Dioxigenase/química , Cisteína Dioxigenase/genética , Dioxigenases/genética , Dioxigenases/metabolismo , Glutationa/metabolismo , Humanos , Fígado/enzimologia , Metionina/metabolismo , Modelos Moleculares , Oxigenases/genética , Oxigenases/metabolismo , Compostos de Sulfidrila/metabolismo , Taurina/análogos & derivados , Taurina/metabolismo
10.
Gastric Cancer ; 20(5): 784-792, 2017 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-28243814

RESUMO

BACKGROUND: Minimal residual disease of the peritoneum is challenging for early cancer detection in gastric cancer (GC). Utility of PCR amplification of cancer-derived DNA has been considered feasible due to its molecular stability, however such markers have never been available in GC clinics. We recently discovered cancer-specific methylation of CDO1 gene in GC, and investigated the clinical potential to detect the minimal residual disease. METHODS: One hundred and two GC patients were investigated for peritoneal fluid cytology test (CY), and detection level of the promoter DNA methylation of CDO1 gene was assessed by quantitative methylation specific PCR (Q-MSP) in the sediments (DNA CY). RESULTS: (1) CY1 was pathologically confirmed in 8 cases, while DNA CY1 was detected in 18 cases. All 8 CY1 were DNA CY1. (2) DNA CY1 was recognized in 14.3, 25.0, 20.0, and 42.9%, in macroscopic Type II, small type III, large type III, and type IV, respectively, while it was not recognized in Type 0/I/V. (3) DNA CY1 was prognostic relevance in gastric cancer (p = 0.0004), and its significance was robust among Type III/IV gastric cancer (p = 0.006 for overall survival and p = 0.0006 for peritoneal recurrence free survival). (4) The peritoneal recurrence was hardly seen in GC patients with potent perioperative chemotherapy among those with DNA CY1. CONCLUSIONS: DNA CY1 detected by Q-MSP for CDO1 gene promoter DNA methylation has a great potential to detect minimal residual disease of the peritoneum in GC clinics as a novel DNA marker.


Assuntos
Cisteína Dioxigenase/genética , Citodiagnóstico/métodos , Metilação de DNA , Neoplasias Gástricas/diagnóstico , Idoso , Líquido Ascítico/citologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Prognóstico , Regiões Promotoras Genéticas , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologia
11.
Dis Esophagus ; 30(3): 1-9, 2017 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-28184414

RESUMO

Methylation of cysteine dioxygenase type 1 (CDO1) gene, a tumor suppressor gene, has been studied in various cancers; however, there is no information regarding Barrett esophagus cancer. In this study, the clinical significance of CDO1 methylation in Barrett esophagus adenocarcinoma (BEA) was clarified. CDO1 gene promoter methylation was analyzed for DNA from the patient's specimens using quantitative methylation-specific polymerase chain reaction. Thirty-eight BEA patients who underwent resection were identified between 2000 and 2014. Hypermethylation of CDO1 gene was demonstrated to be frequently recognized even at early stage in BEA by quantitative methylation-specific polymerase chain reaction. In BEA, there is a robust prognostic difference between stage I and stage II/III/IV with regard to 5-year relapse-free survival (P = 0.0016) and 5-year overall survival (P = 0.0024), and the tumor size separated by 7 cm was also a prognostic factor. There was significant difference in CDO1 gene methylation according to the tumor size (P = 0.036). BEA patients with CDO1 gene methylation were shown marginally significantly poorer prognosis (P = 0.054) than otherwise patients. In conclusion, higher CDO1 gene methylation was seen in BEA at earlier stage than in squamous cell carcinoma, and it may account for aggressive phenotype of BEA.


Assuntos
Adenocarcinoma/genética , Esôfago de Barrett/genética , Cisteína Dioxigenase/genética , Metilação de DNA/genética , Neoplasias Esofágicas/genética , Predisposição Genética para Doença/epidemiologia , Adenocarcinoma/mortalidade , Adenocarcinoma/patologia , Adenocarcinoma/cirurgia , Adulto , Idoso , Idoso de 80 Anos ou mais , Esôfago de Barrett/patologia , Esôfago de Barrett/cirurgia , Transformação Celular Neoplásica/genética , Estudos de Coortes , Intervalo Livre de Doença , Neoplasias Esofágicas/mortalidade , Neoplasias Esofágicas/patologia , Neoplasias Esofágicas/cirurgia , Esofagectomia/métodos , Esofagectomia/mortalidade , Esofagoscopia/métodos , Feminino , Humanos , Japão , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase/métodos , Prognóstico , Modelos de Riscos Proporcionais , Estudos Retrospectivos , Medição de Risco , Estatísticas não Paramétricas , Análise de Sobrevida
12.
Dis Esophagus ; 30(2): 1-9, 2017 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-27629777

RESUMO

We have demonstrated that CDO1 methylation is frequently found in various cancers, including esophageal squamous cell carcinoma (ESCC), but its clinical relevance has remained elusive. CDO1 methylation was investigated in 169 ESCC patients who underwent esophagectomy between 1996 and 2007. CDO1 methylation was assessed by Q-MSP (quantitative methylation specific PCR), and its clinical significance, including its relationship to prognosis, was analyzed. (i) The median TaqMeth value of CDO1 methylation was 9.4, ranging from 0 to 279.5. CDO1 methylation was significantly different between cStage I and cStage II/III (P = 0.02). (ii) On the log-rank plot, the optimal cut-off value was determined to be 8.9; ESCC patients with high CDO1 methylation showed a significantly worse prognosis than those with low CDO1 methylation (P = 0.02). (iii) A multivariate Cox proportional hazards model identified only CDO1 hypermethylation as an independent prognostic factor (HR 2.00, CI 1.09-3.78, P = 0.03). (iv) CDO1 hypermethylation stratified ESCC patients' prognosis in cStage II/III for both neoadjuvant chemo(radio)therapy (NAC)-positive and NAC-negative cases. Moreover, the CDO1 methylation level was significantly lower in cases with Grade 2/3 than in those with Grade 0/1 (P = 0.02) among cStage II/III ESCC patients with NAC. Promoter DNA hypermethylation of CDO1 could be an independent prognostic factor in ESCC; it may also reflect NAC eradication of tumor cells in the primary tumors.


Assuntos
Carcinoma de Células Escamosas/genética , Cisteína Dioxigenase/genética , Metilação de DNA/genética , Neoplasias Esofágicas/genética , Esôfago/patologia , Regiões Promotoras Genéticas/genética , Idoso , Carcinoma de Células Escamosas/patologia , Carcinoma de Células Escamosas/terapia , Quimiorradioterapia Adjuvante , Neoplasias Esofágicas/patologia , Neoplasias Esofágicas/terapia , Carcinoma de Células Escamosas do Esôfago , Esofagectomia , Esôfago/cirurgia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Terapia Neoadjuvante/métodos , Gradação de Tumores , Estadiamento de Neoplasias , Reação em Cadeia da Polimerase/métodos , Prognóstico , Modelos de Riscos Proporcionais , Estudos Retrospectivos
13.
Cell Mol Life Sci ; 74(8): 1391-1412, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-27844098

RESUMO

Hydrogen sulfide (H2S) has profound biological effects within living organisms and is now increasingly being considered alongside other gaseous signalling molecules, such as nitric oxide (NO) and carbon monoxide (CO). Conventional use of pharmacological and molecular approaches has spawned a rapidly growing research field that has identified H2S as playing a functional role in cell-signalling and post-translational modifications. Recently, a number of laboratories have reported the use of siRNA methodologies and genetic mouse models to mimic the loss of function of genes involved in the biosynthesis and degradation of H2S within tissues. Studies utilising these systems are revealing new insights into the biology of H2S within the cardiovascular system, inflammatory disease, and in cell signalling. In light of this work, the current review will describe recent advances in H2S research made possible by the use of molecular approaches and genetic mouse models with perturbed capacities to generate or detoxify physiological levels of H2S gas within tissues.


Assuntos
Vias Biossintéticas , Sulfeto de Hidrogênio/metabolismo , Animais , Carbono-Oxigênio Liases/genética , Carbono-Oxigênio Liases/metabolismo , Cistationina beta-Sintase/genética , Cistationina beta-Sintase/metabolismo , Cisteína Dioxigenase/genética , Cisteína Dioxigenase/metabolismo , Dioxigenases/genética , Dioxigenases/metabolismo , Regulação da Expressão Gênica , Técnicas de Inativação de Genes/métodos , Humanos , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Quinona Redutases/genética , Quinona Redutases/metabolismo , Transdução de Sinais , Sulfurtransferases/genética , Sulfurtransferases/metabolismo
14.
Epigenetics ; 11(12): 871-880, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27689475

RESUMO

Molecular biomarkers may facilitate the distinction between aggressive and clinically insignificant prostate cancer (PCa), thereby potentially aiding individualized treatment. We analyzed cysteine dioxygenase 1 (CDO1) promoter methylation and mRNA expression in order to evaluate its potential as prognostic biomarker. CDO1 methylation and mRNA expression were determined in cell lines and formalin-fixed paraffin-embedded prostatectomy specimens from a first cohort of 300 PCa patients using methylation-specific qPCR and qRT-PCR. Univariate and multivariate Cox proportional hazards and Kaplan-Meier analyses were performed to evaluate biochemical recurrence (BCR)-free survival. Results were confirmed in an independent second cohort comprising 498 PCa cases. Methylation and mRNA expression data from the second cohort were generated by The Cancer Genome Atlas (TCGA) Research Network by means of Infinium HumanMethylation450 BeadChip and RNASeq. CDO1 was hypermethylated in PCa compared to normal adjacent tissues and benign prostatic hyperplasia (P < 0.001) and was associated with reduced gene expression (ρ = -0.91, P = 0.005). Using two different methodologies for methylation quantification, high CDO1 methylation as continuous variable was associated with BCR in univariate analysis (first cohort: HR = 1.02, P = 0.002, 95% CI [1.01-1.03]; second cohort: HR = 1.02, P = 0.032, 95% CI [1.00-1.03]) but failed to reach statistical significance in multivariate analysis. CDO1 promoter methylation is involved in gene regulation and is a potential prognostic biomarker for BCR-free survival in PCa patients following radical prostatectomy. Further studies are needed to validate CDO1 methylation assays and to evaluate the clinical utility of CDO1 methylation for the management of PCa.


Assuntos
Biomarcadores Tumorais/genética , Cisteína Dioxigenase/genética , Neoplasias da Próstata/genética , Adulto , Idoso , Biomarcadores Tumorais/biossíntese , Cisteína Dioxigenase/biossíntese , Metilação de DNA/genética , Intervalo Livre de Doença , Regulação Neoplásica da Expressão Gênica , Inativação Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Prognóstico , Regiões Promotoras Genéticas/genética , Prostatectomia , Neoplasias da Próstata/patologia , Neoplasias da Próstata/cirurgia
15.
J Mol Biol ; 428(20): 3999-4012, 2016 10 09.
Artigo em Inglês | MEDLINE | ID: mdl-27477048

RESUMO

In mammals, the non-heme iron enzyme cysteine dioxygenase (CDO) helps regulate Cys levels through converting Cys to Cys sulfinic acid. Its activity is in part modulated by the formation of a Cys93-Tyr157 crosslink that increases its catalytic efficiency over 10-fold. Here, 21 high-resolution mammalian CDO structures are used to gain insight into how the Cys-Tyr crosslink promotes activity and how select competitive inhibitors bind. Crystal structures of crosslink-deficient C93A and Y157F variants reveal similar ~1.0-Å shifts in the side chain of residue 157, and both variant structures have a new chloride ion coordinating the active site iron. Cys binding is also different from wild-type CDO, and no Cys-persulfenate forms in the C93A or Y157F active sites at pH6.2 or 8.0. We conclude that the crosslink enhances activity by positioning the Tyr157 hydroxyl to enable proper Cys binding, proper oxygen binding, and optimal chemistry. In addition, structures are presented for homocysteine (Hcy), D-Cys, thiosulfate, and azide bound as competitive inhibitors. The observed binding modes of Hcy and D-Cys clarify why they are not substrates, and the binding of azide shows that in contrast to what has been proposed, it does not bind in these crystals as a superoxide mimic.


Assuntos
Cisteína Dioxigenase/química , Cisteína Dioxigenase/metabolismo , Inibidores Enzimáticos/metabolismo , Animais , Cristalografia por Raios X , Cisteína/genética , Cisteína/metabolismo , Cisteína Dioxigenase/genética , Mamíferos , Modelos Moleculares , Proteínas Mutantes/química , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Ligação Proteica , Conformação Proteica , Tirosina/genética , Tirosina/metabolismo
16.
Clin Cancer Res ; 22(13): 3361-71, 2016 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-26842235

RESUMO

PURPOSE: Lung cancer remains as the leading cause of cancer-related death worldwide, mainly due to late diagnosis. Cytology is the gold-standard method for lung cancer diagnosis in minimally invasive respiratory samples, despite its low sensitivity. We aimed to identify epigenetic biomarkers with clinical utility for cancer diagnosis in minimally/noninvasive specimens to improve accuracy of current technologies. EXPERIMENTAL DESIGN: The identification of novel epigenetic biomarkers in stage I lung tumors was accomplished using an integrative genome-wide restrictive analysis of two different large public databases. DNA methylation levels for the selected biomarkers were validated by pyrosequencing in paraffin-embedded tissues and minimally invasive and noninvasive respiratory samples in independent cohorts. RESULTS: We identified nine cancer-specific hypermethylated genes in early-stage lung primary tumors. Four of these genes presented consistent CpG island hypermethylation compared with nonmalignant lung and were associated with transcriptional silencing. A diagnostic signature was built using multivariate logistic regression model based on the combination of four genes: BCAT1, CDO1, TRIM58, and ZNF177 Clinical diagnostic value was also validated in multiple independent cohorts and yielded a remarkable diagnostic accuracy in all cohorts tested. Calibrated and cross-validated epigenetic model predicts with high accuracy the probability to detect cancer in minimally and noninvasive samples. We demonstrated that this epigenetic signature achieved higher diagnostic efficacy in bronchial fluids as compared with conventional cytology for lung cancer diagnosis. CONCLUSIONS: Minimally invasive epigenetic biomarkers have emerged as promising tools for cancer diagnosis. The herein obtained epigenetic model in combination with current diagnostic protocols may improve early diagnosis and outcome of lung cancer patients. Clin Cancer Res; 22(13); 3361-71. ©2016 AACR.


Assuntos
Biomarcadores Tumorais/genética , Cisteína Dioxigenase/genética , Metilação de DNA/genética , Proteínas de Ligação a DNA/genética , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/genética , Transaminases/genética , Proteínas com Motivo Tripartido/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Ilhas de CpG/genética , Detecção Precoce de Câncer/métodos , Epigênese Genética/genética , Feminino , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Regiões Promotoras Genéticas , Estudos Retrospectivos
17.
Sci Rep ; 6: 21231, 2016 Feb 16.
Artigo em Inglês | MEDLINE | ID: mdl-26880478

RESUMO

Animals have varied taurine biosynthesis capability, which was determined by activities of key enzymes including cysteine dioxygenase (CDO) and cysteine sulfinate decarboxylase (CSD). However, whether CDO and CSD are differentially regulated across species remains unexplored. In the present study, we examined the regulations of CDO and CSD in rainbow trout and Japanese flounder, the two fish species with high and low taurine biosynthesis ability respectively. Our results showed that the expression of CDO was lower in rainbow trout but more responsive to cysteine stimulation compared to that in Japanese flounder. On the other hand, both the expression and catalytic efficiency (k(cat)) of CSD were higher in rainbow trout than those of Japanese flounder. A three-residue substrate recognition motif in rainbow trout CSD with sequence of F126/S146/Y148 was identified to be responsible for high k(cat), while that with sequence of F88/N108/F110 in Japanese flounder led to low k(cat), as suggested by site-directed mutagenesis studies. In summary, our results determined new aspects of taurine biosynthesis regulation across species.


Assuntos
Linguado/metabolismo , Oncorhynchus mykiss/metabolismo , Taurina/biossíntese , Sequência de Aminoácidos , Animais , Carboxiliases/genética , Carboxiliases/metabolismo , Clonagem Molecular , Cisteína Dioxigenase/genética , Cisteína Dioxigenase/metabolismo , DNA Complementar , Ativação Enzimática , Linguado/genética , Expressão Gênica , Fígado/metabolismo , Oncorhynchus mykiss/genética , Análise de Sequência de DNA
18.
Sci Rep ; 6: 19296, 2016 Jan 14.
Artigo em Inglês | MEDLINE | ID: mdl-26763277

RESUMO

Mesenchymal stem cells (MSCs) are multipotent cells, which can give rise to variety of cell types, including adipocytes and osteoblasts. Previously, we have shown that cysteine dioxygenase type 1 (Cdo1) promoted adipogenesis of primary mouse bone marrow stromal cells (BMSCs) and 3T3-L1 pre-adipocytes via interaction with Pparγ. However, the role of Cdo1 in osteogenesis remains unclear. Here, we demonstrated that expression of Cdo1 was elevated during osteoblastic differentiation of BMSCs in vitro. Interestingly, knockdown of Cdo1 by siRNA led to an increased expression of osteogenic related genes, elevated alkaline phosphatase (ALP) activity, and enhanced mineralization. Overexpression of Cdo1 in BMSCs inversely suppressed the osteogenesis. Furthermore, we found that overexpression of Cdo1 impaired Wnt signaling and restricted the Wnt3a induced expression of osteogenic transcriptional factors, such as Runx2 and Dlx5. Collectively, our findings indicate Cdo1 suppresses osteogenic differentiation of BMSCs, through a potential mechanism which involves in Wnt signaling reduction concomitantly.


Assuntos
Cisteína Dioxigenase/metabolismo , Células-Tronco Mesenquimais/metabolismo , Osteogênese , Via de Sinalização Wnt , Animais , Biomarcadores , Diferenciação Celular/genética , Cisteína Dioxigenase/genética , Regulação da Expressão Gênica , Inativação Gênica , Células-Tronco Mesenquimais/citologia , Camundongos , Osteoblastos/metabolismo , Fenótipo , Interferência de RNA , RNA Interferente Pequeno/genética
19.
PLoS One ; 11(1): e0144862, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26785325

RESUMO

Using pharmacological unmasking microarray, we identified promoter DNA methylation of cysteine dioxygenase 1 (CDO1) gene in human cancer. In this study, we assessed the clinicopathological significance of CDO1 methylation in primary breast cancer (BC) with no prior chemotherapy. The CDO1 DNA methylation was quantified by TaqMan methylation specific PCR (Q-MSP) in 7 BC cell lines and 172 primary BC patients with no prior chemotherapy. Promoter DNA of the CDO1 gene was hypermethylated in 6 BC cell lines except SK-BR3, and CDO1 gene expression was all silenced at mRNA level in the 7 BC cell lines. Quantification of CDO1 methylation was developed using Q-MSP, and assessed in primary BC. Among the clinicopathologic factors, CDO1 methylation level was not statistically significantly associated with any prognostic factors. The log-rank plot analysis elucidated that the higher methylation the tumors harbored, the poorer prognosis the patients exhibited. Using the median value of 58.0 as a cut-off one, disease specific survival in BC patients with CDO1 hypermethylation showed significantly poorer prognosis than those with hypomethylation (p = 0.004). Multivariate Cox proportional hazards model identified that CDO1 hypermethylation was prognostic factor as well as Ki-67 and hormone receptor status. The most intriguingly, CDO1 hypermethylation was of robust prognostic relevance in triple negative BC (p = 0.007). Promoter DNA methylation of CDO1 gene was robust prognostic indicator in primary BC patients with no prior chemotherapy. Prognostic relevance of the CDO1 promoter DNA methylation is worthy of being paid attention in triple negative BC cancer.


Assuntos
Neoplasias da Mama/diagnóstico , Neoplasias da Mama/genética , Cisteína Dioxigenase/genética , Metilação de DNA , Regiões Promotoras Genéticas , Adulto , Idoso , Biomarcadores Tumorais , Neoplasias da Mama/mortalidade , Linhagem Celular Tumoral , Epigênese Genética , Feminino , Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Prognóstico , Análise de Sequência de DNA , Transcrição Genética
20.
Appl Environ Microbiol ; 82(3): 910-21, 2016 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-26590284

RESUMO

Cysteine dioxygenases (Cdos), which catalyze the sulfoxidation of cysteine to cysteine sulfinic acid (CSA), have been extensively studied in eukaryotes because of their roles in several diseases. In contrast, only a few prokaryotic enzymes of this type have been investigated. In Ralstonia eutropha H16, two Cdo homologues (CdoA and CdoB) have been identified previously. In vivo studies showed that Escherichia coli cells expressing CdoA could convert 3-mercaptopropionate (3MP) to 3-sulfinopropionate (3SP), whereas no 3SP could be detected in cells expressing CdoB. The objective of this study was to confirm these findings and to study both enzymes in detail by performing an in vitro characterization. The proteins were heterologously expressed and purified to apparent homogeneity by immobilized metal chelate affinity chromatography (IMAC). Subsequent analysis of the enzyme activities revealed striking differences with regard to their substrate ranges and their specificities for the transition metal cofactor, e.g., CdoA catalyzed the sulfoxidation of 3MP to a 3-fold-greater extent than the sulfoxidation of cysteine, whereas CdoB converted only cysteine. Moreover, the dependency of the activities of the Cdos from R. eutropha H16 on the metal cofactor in the active center could be demonstrated. The importance of CdoA for the metabolism of the sulfur compounds 3,3'-thiodipropionic acid (TDP) and 3,3'-dithiodipropionic acid (DTDP) by further converting their degradation product, 3MP, was confirmed. Since 3MP can also function as a precursor for polythioester (PTE) synthesis in R. eutropha H16, deletion of cdoA might enable increased synthesis of PTEs.


Assuntos
Coenzimas/metabolismo , Cupriavidus necator/enzimologia , Cisteína Dioxigenase/genética , Cisteína Dioxigenase/metabolismo , Ácido 3-Mercaptopropiônico/metabolismo , Cromatografia de Afinidade , Coenzimas/química , Cupriavidus necator/genética , Cupriavidus necator/metabolismo , Cisteamina/farmacologia , Cisteína/análogos & derivados , Cisteína/metabolismo , Cisteína Dioxigenase/química , Cisteína Dioxigenase/isolamento & purificação , Cinética , Mercaptoetanol/farmacologia , Propionatos/metabolismo , Proteínas Recombinantes/metabolismo , Análise de Sequência de DNA , Especificidade por Substrato
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