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Alzheimer's disease (AD) is a neurodegenerative disease, and mild cognitive impairment (MCI) is considered a transitional stage between healthy aging and dementia. Early detection of MCI can help slow down the progression of AD. At present, there are few studies exploring the characteristics of abnormal dynamic brain activity in AD. This article uses a method called Leading Eigenvector Dynamics Analysis (LEiDA) to study resting-state functional magnetic resonance imaging (rs-fMRI) data of AD, MCI, and cognitively normal (CN) participants. By identifying repetitive states of phase coherence, inter group differences in brain dynamic activity indicators are examined. And the neurobehavioral scales were used to assess the relationship between abnormal dynamic activities and cognitive function. The results showed that in the indicators of occurrence probability and lifetime, the globally synchronized state of the patient group decreased. The activity state of the limbic regions significantly detected the difference between AD and the other two groups. Compared to CN, AD and MCI have varying degrees of increase in default and visual regions activity states. In addition, in the analysis related to the cognitive scales, it was found that individuals with poorer cognitive abilities were less active in the globally synchronized state, and more active in limbic regions activity state and visual regions activity state. Taken together, these findings reveal abnormal dynamic activity of resting-state networks in patients with AD and MCI, provide new insights into the dynamic analysis of brain networks, and contribute to a deeper understanding of abnormal spatial dynamic patterns in AD patients.
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BACKGROUND: Tandem C2 domains, nuclear (TC2N) is a C2 domain-containing protein that belongs to the carboxyl-terminal type (C-type) tandem C2 protein family, and acts as an oncogenic driver in several cancers. Previously, we preliminarily reported that TC2N mediates the PI3K-Akt signaling pathway to inhibit tumor growth of breast cancer (BC) cells. Beyond that, its precise biological functions and detailed molecular mechanisms in BC development and progression are not fully understood. METHODS: Tumor tissues of 212 BC patients were subjected to tissue microarray and further assessed the associations of TC2N expression with pathological parameters and FASN expression. The protein levels of TC2N and FASN in cell lines and tumor specimens were monitored by qRT-PCR, WB, immunofluorescence and immunohistochemistry. In vitro cell assays, in vivo nude mice model was used to assess the effect of TC2N ectopic expression on tumor metastasis and stemness of breast cancer cells. The downstream signaling pathway or target molecule of TC2N was mined using a combination of transcriptomics, proteomics and lipidomics, and the underlying mechanism was explored by WB and co-IP assays. RESULTS: Here, we found that the expression of TC2N remarkedly silenced in metastatic and poorly differentiated tumors. Function-wide, TC2N strongly inhibits tumor metastasis and stem-like properties of BC via inhibition of fatty acid synthesis. Mechanism-wise, TC2N blocks neddylated PTEN-mediated FASN stabilization by a dual mechanism. The C2B domain is crucial for nuclear localization of TC2N, further consolidating the TRIM21-mediated ubiquitylation and degradation of FASN by competing with neddylated PTEN for binding to FASN in nucleus. On the other hand, cytoplasmic TC2N interacts with import proteins, thereby restraining nuclear import of PTEN to decrease neddylated PTEN level. CONCLUSIONS: Altogether, we demonstrate a previously unidentified role and mechanism of TC2N in regulation of lipid metabolism and PTEN neddylation, providing a potential therapeutic target for anti-cancer.
Assuntos
Neoplasias da Mama , Animais , Camundongos , Humanos , Feminino , Neoplasias da Mama/patologia , Camundongos Nus , Fosfatidilinositol 3-Quinases/metabolismo , Transdução de Sinais , Ácidos Graxos , Linhagem Celular Tumoral , Proteínas Proto-Oncogênicas c-akt/metabolismo , PTEN Fosfo-Hidrolase/genética , Proliferação de Células , Regulação Neoplásica da Expressão GênicaRESUMO
Hepatocellular carcinoma (HCC), one of the most prevalent types of cancer worldwide, has an exceedingly poor prognosis. Tandem C2 domain nuclear protein (TC2N) has been implicated in tumorigenesis and serves as an oncogene or tumor suppressor in different types of cancer. Here, we explore the possible regulatory activities and molecular mechanisms of TC2N in HCC progression. However, TC2N expression was significantly upregulated in HCC tissues and hepatoma cell lines, and this upregulation was positively correlated with tumor progression in HCC patients. The ectopic overexpression of TC2N accelerated the proliferation, migration, and invasion of HCC cells, whereas its knockdown showed the opposite effects. Bioinformatics analysis showed that TC2N participates in the regulation of the Wnt/ß-catenin signaling pathway. Mechanistically, TC2N activated the Wnt/ß-catenin signaling pathway by regulating the expression levels of ß-catenin and its downstream targets CyclinD1, MMP7, c-Myc, c-Jun, AXIN2, and glutamine synthase. Furthermore, the deletion of ß-catenin effectively neutralized the regulation of TC2N in HCC proliferation and metastasis. Overall, this study showed that TC2N promotes HCC proliferation and metastasis by activating the Wnt/ß-catenin signaling pathway, indicating that TC2N might be a potential molecular target for the treatment of HCC.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Humanos , Carcinoma Hepatocelular/metabolismo , Neoplasias Hepáticas/metabolismo , beta Catenina/metabolismo , Via de Sinalização Wnt/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Regulação Neoplásica da Expressão GênicaRESUMO
An I2-DMSO-mediated multicomponent [3+1+2] cascade annulation reaction using aryl methyl ketones, enaminones, and benzo[d]isoxazol-3-amine as substrates has been developed. This metal-free reaction involved the transannulation of benzo[d]isoxazol-3-amines with the formation of two C-N bonds and a C-C bond in one pot. Notably, a pyrimidine ring with a 1,4-dicarbonyl scaffold could efficiently transform into a pyrimido[4,5-d]pyridazine skeleton. The phenolic hydroxyl group of the target product could undergo further modification with pharmaceuticals, demonstrating the utility of this method.
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Colorectal cancer (CRC) is the third most common cancer in men and the second most common cancer in women worldwide. CRC is the second leading cause of cancer-related deaths. Although some progress in the treatment of CRC has been achieved, the molecular mechanism of CRC is still unclear. In this study, alcohol dehydrogenase 1C(ADH1C) was first identified as a target gene closely associated with the development of CRC by the comprehensive application of transcriptomics, proteomics, metabonomics and in silico analysis. The ADH1C mRNA and protein expression in CRC cell lines and tumor tissues was lower than that in normal intestinal epithelial cell lines and healthy tissues. Overexpression of ADH1C inhibited the growth, migration, invasion and colony formation of CRC cell lines and prevented the growth of xenograft tumors in nude mice. The inhibitory effects of ADH1C on CRC cells in vitro were exerted by reducing the expression of PHGDH/PSAT1 and the serine level. This inhibition could be partially reversed by adding serine to the culture medium. These results showed that ADH1C is a potential drug target in CRC.
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Álcool Desidrogenase , Neoplasias Colorretais , Álcool Desidrogenase/genética , Álcool Desidrogenase/metabolismo , Animais , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células/genética , Neoplasias Colorretais/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Redes e Vias Metabólicas , Camundongos , Camundongos Nus , RNA Mensageiro/metabolismo , Serina/genética , Serina/metabolismoRESUMO
Glioblastoma multiforme (GBM) is the most malignant and lethal primary brain tumor in adults accounting for about 50% of all gliomas. The only treatment available for GBM is the drug temozolomide, which unfortunately has frequent drug resistance issue. By analyzing the hub genes of GBM via weighted gene co-expression network analysis (WGCNA) of the cancer genome atlas (TCGA) dataset, and using the connectivity map (CMAP) platform for drug repurposing, we found that multiple azole compounds had potential anti-GBM activity. When their anti-GBM activity was examined, however, only three benzimidazole compounds, i.e. flubendazole, mebendazole and fenbendazole, potently and dose-dependently inhibited proliferation of U87 and U251 cells with IC50 values below 0.26 µM. Benzimidazoles (0.125-0.5 µM) dose-dependently suppressed DNA synthesis, cell migration and invasion, and regulated the expression of key epithelial-mesenchymal transition (EMT) markers in U87 and U251 cells. Benzimidazoles treatment also dose-dependently induced the GBM cell cycle arrest at the G2/M phase via the P53/P21/cyclin B1 pathway. Furthermore, the drugs triggered pyroptosis of GBM cells through the NF-κB/NLRP3/GSDMD pathway, and might also concurrently induced mitochondria-dependent apoptosis. In a nude mouse U87 cell xenograft model, administration of flubendazole (12.5, 25, and 50 mg · kg-1 · d-1, i.p, for 3 weeks) dose-dependently suppressed the tumor growth without obvious adverse effects. Taken together, our results demonstrated that benzimidazoles might be promising candidates for the treatment of GBM.
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Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Benzimidazóis/farmacologia , Neoplasias do Sistema Nervoso Central/tratamento farmacológico , Glioblastoma/tratamento farmacológico , Antineoplásicos/química , Benzimidazóis/química , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Neoplasias do Sistema Nervoso Central/metabolismo , Neoplasias do Sistema Nervoso Central/patologia , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Glioblastoma/metabolismo , Glioblastoma/patologia , Humanos , Estrutura Molecular , Relação Estrutura-Atividade , Células Tumorais CultivadasRESUMO
Glioblastoma (GBM), a malignant brain tumor, is a world-wide health problem because of its poor prognosis and high rates of recurrence and mortality. Apolipoprotein C1 (APOC1) is the smallest of apolipoproteins, implicated in many diseases. Recent studies have shown that APOC1 promotes tumorigenesis and development of several types of cancer. In this study we investigated the role of APOC1 in GBM tumorigenesis. Using in silico assays we showed that APOC1 was highly expressed in GBM tissues and its expression was closely related to GBM progression. We showed that APOC1 protein expression was markedly increased in four GBM cell lines (U251, U138, A172 and U87) compared to the normal brain glia cell lines (HEB, HA1800). In U251 cells, overexpression of APOC1 promoted cell proliferation, migration, invasion and colony information, which was reversed by APOC1 knockdown. APOC1 knockdown also markedly inhibited the growth of GBM xenografts in the ventricle of nude mice. We further demonstrated that APOC1 reduced ferroptosis by inhibiting KEAP1, promoting nuclear translocation of NRF2 and increasing expression of HO-1 and NQO1 in GBM cells. APOC1 also induced ferroptosis resistance by increasing cystathionine beta-synthase (CBS) expression, which promoted trans-sulfuration and increased GSH synthesis, ultimately leading to an increase in glutathione peroxidase-4 (GPX4). Thus, APOC1 plays a key role in GBM tumorigenesis, conferring resistance to ferroptosis, and may be a promising therapeutic target for GBM.
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Apolipoproteína C-I , Ferroptose , Glioblastoma , Proteína 1 Associada a ECH Semelhante a Kelch , Fator 2 Relacionado a NF-E2 , Animais , Humanos , Camundongos , Apolipoproteína C-I/metabolismo , Carcinogênese/metabolismo , Linhagem Celular Tumoral , Transformação Celular Neoplásica , Cistationina beta-Sintase/metabolismo , Regulação Neoplásica da Expressão Gênica , Glioblastoma/metabolismo , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Camundongos Nus , Fator 2 Relacionado a NF-E2/metabolismoRESUMO
Glioblastoma (GBM) is the most common and lethal primary brain tumor in adults, but there is no effective drug available for GBM. Avasimibe is a potent inhibitor of acyl-coenzyme A: cholesterol acyltransferase-1 (ACAT-1), which was used to treat atherosclerosis. Experimental evidence and bioinformatics have shown that avasimibe has anticancer activity. In this study we investigated the anticancer effects of avasimibe on human glioblastoma cells and the underlying mechanisms. Our results showed that avasimibe dose-dependently inhibited the proliferation of U251 and U87 human glioblastoma cells with IC50 values of 20.29 and 28.27 µM, respectively, at 48 h. Avasimibe (7.5, 15, 30 µM) decreased the DNA synthesis, and inhibited the colony formation of the tumor cells. Treatment of avasimibe also dose-dependently increased the apoptotic rate of tumor cells, decreased the mitochondrial membrane potential, induced the activity of caspase-3/7, and increased the protein expression of cleaved caspase-9, cleaved PARP and Bax in U251 and U87 cells. RNA-sequencing analyses revealed that avasimibe suppressed the expression of CDK2, cyclin E1, CDK4, cyclin D, CDK1, cyclin B1, Aurora A, and PLK1, while induced the expression of p53, p21, p27, and GADD45A, which was validated by Western blot analysis. These results demonstrated that avasimibe induced mitochondria-dependent apoptosis in glioblastoma cells, which was associated with arresting the cell cycle at G0/G1 phase and G2/M phase by regulating the p53/p21 pathway, p53/GADD45A and Aurora A/PLK1 signaling pathways. In U87 xenograft nude mice model, administration of avasimibe (15, 30 mg·kg-1·d-1, ip, for 18 days) dose-dependently inhibit the tumor growth. Taken together, our results demonstrated that avasimibe might be a promising chemotherapy drug in the treatment of GBM.
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Acetamidas/farmacologia , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Glioblastoma/tratamento farmacológico , Sulfonamidas/farmacologia , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Reposicionamento de Medicamentos , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Camundongos Endogâmicos BALB C , Mitocôndrias/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacosRESUMO
Triple-negative breast cancer (TNBC) is characterized by low expression of human epidermal growth factor receptor-2 (HER2), estrogen receptor (ER), and progesterone receptor (PR), which is the most aggressive subtype with poor outcome among breast cancers. The underlying mechanisms of TNBC remain unclear and there is a lack of biomarkers. In this study we conducted an in silico assay and found that FOXC1 was highly expressed in ER-/PR-/HER2- breast cancers, which was confirmed by qRT-PCR, immunohistochemistry, and Western blot analysis. FOXC1 was more highly expressed in TNBCs than the other breast cancers. Kaplan-Meier plotter revealed that expression of FOXC1 was associated with overall survival (OS) of patients with breast cancers. Expression of FOXC1 was reversely associated with level of H3K27me3, which was methylated by EZH2. In MCF-7 and T47D cells, inhibition of EZH2 by DZNeP or GSK343 concentration- and time-dependently increased expression of FOXC1. Finally, we demonstrated that the expression of FOXC1 was associated with resistance of doxorubicin treatment of breast cancer cells. In conclusion, these results suggest that FOXC1 may be a potential biomarker or drug target for TNBCs, and that downregulation of FOXC1 could have therapeutic value in treatment of TNBCs.
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Biomarcadores Tumorais/metabolismo , Proteína Potenciadora do Homólogo 2 de Zeste/metabolismo , Fatores de Transcrição Forkhead/metabolismo , Histonas/metabolismo , Neoplasias de Mama Triplo Negativas/metabolismo , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/fisiologia , Humanos , MetilaçãoRESUMO
Emerging evidences have revealed tumor-specific gene methylation is considered to be a promising non-invasive biomarker for many different types of cancers. This study was determined whether TMEM196 gene hypermethylation and downregulation are considered to be promising biomarkers for early diagnosis and prognosis in lung cancer. Methylation status was detected with methylation-specific PCR. Kaplan-Meier survival curves and Cox regression analysis were used to determine the significance of prognosis. TMEM196 gene was hypermethylated in 68.1% (64/94) of lung cancer tissues, 52.8% (67/127) of plasma and 55.2% (79/143) of sputum samples, but unmethylated (0/50) in normal tissues. TMEM196 methylation in plasma and sputum samples was significantly correlated with that in the corresponding paired tumor tissues (r = 0.750, r = 0.880, P < 0.001). TMEM196 aberrant methylation in cancer tissues, plasma and sputum DNA was significantly associated with age and pathological type (P < 0.05). TMEM196 high methylation could robustly distinguish lung cancer patients (AUC = 0.905) from normal subjects and patients with TMEM196 high methylation have a significantly poorer survival than those with low level from The Cancer Genome Atlas (Wilcoxon P < 0.001). Multivariate models showed TMEM196 methylation is an independent prognostic marker in lung cancer. Furthermore, the overall survival of patients with low TMEM196 expression was significantly poorer than that of TMEM196-high patients (P < 0.001, log-rank test). Low TMEM196 expression in tumor tissues was found to predict poorer survival (HR = 3.007; 95%CI, 1.918-4.714). Our study provided new insights into the clinical importance and potential use of TMEM196 methylation and expression as novel early diagnostic and prognostic biomarkers for human lung cancers.
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Biomarcadores Tumorais/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Metilação de DNA , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/genética , Proteínas de Membrana/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Estudos de Casos e Controles , Feminino , Seguimentos , Humanos , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , Prognóstico , Regiões Promotoras Genéticas , Taxa de SobrevidaRESUMO
Bisphenol A (BPA) impairs male fertility by acting as an endocrine disruptor. However, the mechanisms by which BPA cause reproductive toxicity are not fully elucidated. Here, we explored the role of XAF1, a novel pro-apoptosis molecule, in BPA-induced abnormal spermatogenesis and the transcriptional regulation mechanism of BPA-induced XAF1. BPA exposure detrimentally impacted spermatogenesis by inducing excessive germ cell apoptosis. XAF1 was upregulated in germ cells after BPA exposure, which was involved in the apoptosis pathway. In addition, the expression levels of XIAP and XAF1 were inversely correlated after BPA exposure. Knockdown of XAF1 expression partially inhibited the apoptosis of GC-2 cells, suppressed the activation of caspase 3 and improved the BPA-induced XIAP expression. Moreover, IFNß expression levels were significantly upregulated after BPA exposure both in vitro and in vivo, and these levels were positively related to the expression of XAF1. Furthermore, IFNß knockdown reduced the expression of XAF1 and increased the expression of XIAP in BPA-treated GC-2 cells. Together, these data indicated that BPA triggers male germ cell apoptosis in mice via the IFNß-XAF1-XIAP pathway, which may contribute to BPA-induced testis toxicity.
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Apoptose/efeitos dos fármacos , Compostos Benzidrílicos/toxicidade , Proteínas F-Box/efeitos dos fármacos , Células Germinativas/efeitos dos fármacos , Proteínas Inibidoras de Apoptose/efeitos dos fármacos , Interferon beta/efeitos dos fármacos , Fenóis/toxicidade , Transdução de Sinais/efeitos dos fármacos , Proteínas Adaptadoras de Transdução de Sinal , Animais , Proteínas Reguladoras de Apoptose , Linhagem Celular , Proteínas F-Box/genética , Expressão Gênica/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Proteínas Inibidoras de Apoptose/genética , Interferon beta/genética , Masculino , Camundongos , Espermatogênese/efeitos dos fármacos , Testículo/patologia , Regulação para Cima/efeitos dos fármacosRESUMO
BACKGROUND: Different histological subtypes of non-small cell lung cancer (NSCLC) show different molecular characteristics and responses to therapeutic strategy. Identification of specific gene, clarification of its special roles and molecular mechanisms are crucial for developing new therapeutic approach for particular subtype patients. METHODS: Surgical specimens of 540 NSCLC patients were recruited. Immunohistochemistry was used to detect SOX30 expression, and correlations with clinical parameters were analyzed. Functional experiments and gene ontology analysis were performed to investigate roles of SOX30. Network analysis, TOP/FOP-Flash assays, luciferase reporter assays and ChIP-PCR assays were performed to determine the mechanism. Survival analyses were calculated by Kaplan-Meier and Cox regression. Recovery experiment was investigated the importance of the target of SOX30. RESULTS: SOX30 expression is closely associated with histological types of NSCLC, and metastasis of adenocarcinoma (ADC) patients but not of squamous cell carcinoma (SCC) patients. SOX30 strongly inhibits cancer cell migration and invasion in ADC cell lines, whrereas not affects cell migration and invasion in SCC cell lines. The genes associated with SOX30 preferentially enrich in metastasis process and Wnt-signaling in only ADC patients. Consistently, SOX30 is negatively associated with the expression of Wnt-signaling and metastasis-related gene CTNNB1 (ß-catenin) in ADC, but not in SCC. At the molecular level, SOX30 represses Wnt-signaling by directly transcriptional inhibition of CTNNB1 in ADC, and also not in SCC. In the clinical, SOX30 is a favorable and independent prognostic factor in ADC patients, whereas is an unfavorable and independent prognostic factor in SCC patients. Moreover, SOX30 expression is a double face early-stage prognostic biomarker in ADC and SCC patients. In addition, forcible restoration of CTNNB1 indeed can inhibit the anti-metastatic role of SOX30 in ADC patients. CONCLUSIONS: In early-stage ADC patients, elevated SOX30 expression inhibits tumor-metastasis by directly binding to CTNNB1 promoter resulting in a favorable prognosis of these patients. However, in early-stage SCC patients, SOX30 has no inhibitory role on tumor-metastasis due to not binding to CTNNB1 promoter leading to an unfavorable prognosis of the patients. This study highlights a special role and prognostic value of SOX30 in ADC, providing a novel therapeutic target for particular subtype NSCLC patients.
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Adenocarcinoma de Pulmão/metabolismo , Biomarcadores Tumorais/biossíntese , Neoplasias Pulmonares/metabolismo , Fatores de Transcrição SOX/biossíntese , Proteínas Supressoras de Tumor/biossíntese , Via de Sinalização Wnt/fisiologia , Células A549 , Adenocarcinoma de Pulmão/patologia , Idoso , Feminino , Humanos , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , PrognósticoRESUMO
The identification of the key genetic and epigenetic changes underlying lung carcinogenesis would aid effective early diagnosis and targeted therapies for lung cancer. In this study, we screened a novel hypermethylated gene ankyrin repeat domain 18B (ANKRD18B), to determine whether it is regulated by DNA methylation and clarify its biological and clinical implications in lung cancer. Methylation status and expression level were analyzed by methylation-specific PCR, bisulfite genomic sequencing, and quantitative reverse transcription-polymerase chain reaction (qRT-PCR). We detected ANKRD18B hypermethylation in 52 of 98 (53.1%) primary lung cancer tissues and in nine of 10 (90%) cell lines, whereas no methylation was seen in 10 normal lung tissue samples. ANKRD18B methylation was more frequently observed in patients with poor differentiation (P < 0.05). Notably, 62 pairs of samples from patients whose tumor tissue showed hypermethylation of ANKRD18B exhibited the same aberrant methylation in 72.7% and 69.7% of their corresponding plasma and sputum samples, respectively; whereas no hypermethylation of ANKRD18B was detected in the sputum and plasma from patients whose tumor sample lacked this alteration. In addition, ANKRD18B mRNA expression was significantly decreased or silenced in lung cancer tissues and cell lines associated with hypermethylation of the ANKRD18B region. Demethylation agent 5-aza-2'-deoxycytidine significantly increased ANKRD18B mRNA expression in lung cancer cell lines. Furthermore, overexpression of ANKRD18B suppressed lung cancer cell growth. These results suggest that the expression of ANKRD18B is regulated by CpG island hypermethylation in lung cancer. Our findings confirm the importance of the identification of new markers of epigenetic dysregulation in cancer.
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Metilação de DNA , Epigênese Genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/genética , Pulmão/patologia , Sequência de Bases , Linhagem Celular Tumoral , Proliferação de Células , Ilhas de CpG , Feminino , Humanos , Pulmão/metabolismo , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , RNA Mensageiro/genéticaRESUMO
Using genome-wide methylation screening, we found Aristaless-like homeobox-4 (ALX4) preferentially methylated in lung cancer. ALX4 is a putative transcription factor that belongs to the family of paired-class homeoproteins involved in epithelial development. However, the role of ALX4 in tumorigenesis remains largely unclear. Here, we analyzed its epigenetic regulation, biological functions and related molecular mechanisms in lung cancer. CpG island methylation and expression of ALX4 were evaluated by methylation-specific polymerase chain reaction (PCR), bisulfite genomic sequencing, reverse-transcription PCR and Western blotting. ALX4 functions were determined by cell viability, colony formation, flow cytometry and in vivo tumorigenicity assays. ALX4 hypermethylation was detected in 55% (54/98) of primary lung cancers compared to none (0/20) of the normal lung tissue samples tested (p < 0.01). ALX4 was readily expressed in normal lung tissues with an unmethylated status, but downregulated or silenced in 90% (9/10) of lung cancer cell lines with a hypermethylation status. Demethylation experiments further confirmed that loss of ALX4 expression was regulated by CpG island hypermethylation. Re-expression of ALX4 in lung cancer cell lines suppressed cell viability, colony formation and migration, whereas it induced apoptosis and G1/S arrest and restrained the tumorigenicity in nude mice. These effects were associated with upregulation of proapoptotic proteins caspase-7, -8 and -9, and downregulation of Bcl-2. On the other hand, knockdown of ALX4 expression by siRNA increased cell viability and proliferation, whereas it inhibited apoptosis and cell cycle arrest. In conclusion, our results suggest that ALX4 is a novel putative tumor suppressor with epigenetic silencing in lung carcinogenesis.
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Metilação de DNA , Proteínas de Ligação a DNA/genética , Epigênese Genética , Inativação Gênica , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Fatores de Transcrição/genética , Animais , Apoptose , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Western Blotting , Caspases/metabolismo , Adesão Celular , Ciclo Celular , Movimento Celular , Proliferação de Células , Células Cultivadas , Ilhas de CpG , Proteínas de Ligação a DNA/antagonistas & inibidores , Proteínas de Ligação a DNA/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Pulmão/metabolismo , Pulmão/patologia , Neoplasias Pulmonares/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Estadiamento de Neoplasias , Regiões Promotoras Genéticas , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/metabolismo , Ensaio Tumoral de Célula-TroncoRESUMO
The living hyena species (spotted, brown, striped and aardwolf) are remnants of a formerly diverse group of more than 80 fossil species, which peaked in diversity in the Late Miocene (about 7-8 Ma). The fossil history indicates an African origin, and morphological and ancient DNA data have confirmed that living spotted hyenas (Crocuta crocuta) of Africa were closely related to extinct Late Pleistocene cave hyenas from Europe and Asia. The current model used to explain the origins of Eurasian cave hyena populations invokes multiple migrations out of Africa between 3.5-0.35 Ma. We used mitochondrial DNA sequences from radiocarbon-dated Chinese Pleistocene hyena specimens to examine the origin of Asian populations, and temporally calibrate the evolutionary history of spotted hyenas. Our results support a far more recent evolutionary timescale (430-163 kya) and suggest that extinct and living spotted hyena populations originated from a widespread Eurasian population in the Late Pleistocene, which was only subsequently restricted to Africa. We developed statistical tests of the contrasting population models and their fit to the fossil record. Coalescent simulations and Bayes Factor analysis support the new radiocarbon-calibrated timescale and Eurasian origins model. The new Eurasian biogeographic scenario proposed for the hyena emphasizes the role of the vast steppe grasslands of Eurasia in contrast to models only involving Africa. The new methodology for combining genetic and geological data to test contrasting models of population history will be useful for a wide range of taxa where ancient and historic genetic data are available.
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Evolução Biológica , Hyaenidae/genética , Filogenia , Animais , Ásia , Teorema de Bayes , Citocromos b/genética , DNA Mitocondrial/genética , Europa (Continente) , Fósseis , Genética Populacional , Modelos Genéticos , Dados de Sequência Molecular , Análise de Sequência de DNARESUMO
Background: Non-small cell lung cancer (NSCLC), including the lung squamous cell carcinoma (LUSC) and lung adenocarcinoma (LUAD) subtypes, is a malignant tumor type with a poor 5-year survival rate. The identification of new powerful diagnostic biomarkers, prognostic biomarkers, and potential therapeutic targets in NSCLC is urgently required. Methods: The UCSC Xena, UALCAN, and GEO databases were used to screen and analyze differentially expressed genes, regulatory modes, and genetic/epigenetic alterations in NSCLC. The UCSC Xena database, GEO database, tissue microarray, and immunohistochemistry staining analyses were used to evaluate the diagnostic and prognostic values. Gain-of-function assays were performed to examine the roles. The ESTIMATE, TIMER, Linked Omics, STRING, and DAVID algorithms were used to analyze potential molecular mechanisms. Results: NR3C2 was identified as a potentially important molecule in NSCLC. NR3C2 is expressed at low levels in NSCLC, LUAD, and LUSC tissues, which is significantly related to the clinical indexes of these patients. Receiver operating characteristic curve analysis suggests that the altered NR3C2 expression patterns have diagnostic value in NSCLC, LUAD, and especially LUSC patients. Decreased NR3C2 expression levels can help predict poor prognosis in NSCLC and LUAD patients but not in LUSC patients. These results have been confirmed both with database analysis and real-world clinical samples on a tissue microarray. Copy number variation contributes to low NR3C2 expression levels in NSCLC and LUAD, while promoter DNA methylation is involved in its downregulation in LUSC. Two NR3C2 promoter methylation sites have high sensitivity and specificity for LUSC diagnosis with clinical application potential. NR3C2 may be a key participant in NSCLC development and progression and is closely associated with the tumor microenvironment and immune cell infiltration. NR3C2 co-expressed genes are involved in many cancer-related signaling pathways, further supporting a potentially significant role of NR3C2 in NSCLC. Conclusions: NR3C2 is a novel potential diagnostic and prognostic biomarker and therapeutic target in NSCLC.