Research IndicatorsGraph generated 13 March 2017 using data from PubMed using criteria.
Mouse over the terms for more detail; many indicate links which you can click for dedicated pages about the topic. Tag cloud generated 13 March, 2017 using data from PubMed, MeSH and CancerIndex
Specific Cancers (6)
Data table showing topics related to specific cancers and associated disorders. Scope includes mutations and abnormal protein expression.
Note: list is not exhaustive. Number of papers are based on searches of PubMed (click on topic title for arbitrary criteria used).
OMIM, Johns Hopkin University
Referenced article focusing on the relationship between phenotype and genotype.
International Cancer Genome Consortium.
Summary of gene and mutations by cancer type from ICGC
Cancer Genome Anatomy Project, NCI
COSMIC, Sanger Institute
Somatic mutation information and related details
GEO Profiles, NCBI
Search the gene expression profiles from curated DataSets in the Gene Expression Omnibus (GEO) repository.
Latest Publications: NOX1 (cancer-related)
Bauer GCentral Signaling Elements of Intercellular Reactive Oxygen/Nitrogen Species-dependent Induction of Apoptosis in Malignant Cells.
Anticancer Res. 2017; 37(2):499-513 [PubMed
] Related Publications
Intercellular reactive oxygen/reactive nitrogen species-(ROS/RNS)-dependent induction of apoptosis in malignant cells is discussed as a potential control step during oncogenesis. In previous studies, the mechanism of intercellular apoptosis-inducing signaling was mainly established through the use of specific inhibitors and scavengers. Here, a detailed analysis was carried out based on small interfering ribonucleic acid (siRNA)-mediated knockdown of central players of intercellular ROS/RNS signaling and of the mitochondrial and the FAS receptor-dependent pathway of apoptosis. The data show that transforming growth factor β1, transforming growth factor β receptor, NADPH oxidase-1 (NOX1), NOX1 organizer, and NOX1 activator control the HOCl and the NO/peroxynitrite signaling pathways. Dual oxidase-1 (DUOX1) is specifically involved in HOCl signaling, and NO synthase in NO/peroxynitrite signaling. Both pathways utilize intracellular signal transduction through protein kinase C zeta, sphingomyelinase and central elements of the mitochondrial pathway of apoptosis, whereas the FAS receptor and FAS ligand do not seem to play a role.
Ito K, Ota A, Ono T, et al.Inhibition of Nox1 induces apoptosis by attenuating the AKT signaling pathway in oral squamous cell carcinoma cell lines.
Oncol Rep. 2016; 36(5):2991-2998 [PubMed
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NADPH oxidases, also known as the Nox family, are major sources of reactive oxygen species generation that regulate redox-sensitive signaling pathways. Recent studies have implicated the Nox family in cancer development and progression. However, the involvement of its members in the development of oral squamous cell carcinoma (OSCC) remains to be elucidated. To clarify this issue, we first analyzed mRNA expression of Nox/Duox family members (Nox1, Nox2, Nox3, Nox4, Nox5, Duox1 and Duox2) in five OSCC cell lines. Nox1 and Nox4 mRNAs were highly expressed in four OSCC cell lines. Western blot analysis revealed that the protein expression level of Nox1 was higher than that of Nox4 in the OSCC cell lines. In addition, knockdown of Nox1, but not Nox4, significantly suppressed cell viability and induced apoptosis in the HSC-2 and HSC-3 cells. We also found that a specific AKT inhibitor, perifosine, dose-dependently suppressed OSCC cell growth. Notably, Nox1 knockdown significantly attenuated the phosphorylation level of AKT. Furthermore, both Nox1 knockdown and perifosine treatment markedly enhanced the cisplatin-induced cytotoxic effect. Taken together, our results highlight that the Nox1/AKT signaling pathway plays an important role in cell survival in OSCC cells.
Reactive oxygen species (ROS) are an important endogenous source of DNA damage and oxidative stress in all cell types. Deficiency in tuberin resulted in increased oxidative DNA damage in renal cells. In this study, the role of tuberin in the regulating of ROS and NADPH oxidases was investigated. Formation of ROS and activity of NADPH oxidases were significantly higher in mouse embryonic fibroblasts and in primary culture of rat renal proximal tubular epithelial tuberin-deficient cells compared to wild-type cells. In addition, expression of NADPH oxidase (Nox)1, Nox2, and Nox4 (Nox isoforms) was higher in mouse embryonic fibroblasts and renal proximal tubular epithelial tuberin-deficient cells compared to wild-type cells. Furthermore, activity levels of NADPH oxidases and protein expression of all Nox isoforms were higher in the renal cortex of rat deficient in tuberin. However, treatment of tuberin-deficient cells with rapamycin showed significant decrease in protein expression of all Nox. Significant increase in protein kinase C βII expression was detected in tuberin-deficient cells, whereas inhibition of protein kinase C βII by bisindolylmaleimide I resulted in decreased protein expression of all Nox isoforms. In addition, treatment of mice deficient in tuberin with rapamycin resulted in significant decrease in all Nox protein expression. Moreover, protein and mRNA expression of all Nox were highly expressed in tumor kidney tissue of patients with tuberous sclerosis complex compared to control kidney tissue of normal subjects. These data provide the first evidence that tuberin plays a novel role in regulating ROS generation, NADPH oxidase activity, and Nox expression that may potentially be involved in development of kidney tumor in patients with tuberous sclerosis complex.
Pekarčíková L, Knopfová L, Beneš P, Šmarda Jc-Myb regulates NOX1/p38 to control survival of colorectal carcinoma cells.
Cell Signal. 2016; 28(8):924-36 [PubMed
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The c-Myb transcription factor is important for maintenance of immature cells of many tissues including colon epithelium. Overexpression of c-Myb occurring in colorectal carcinomas (CRC) as well as in other cancers often marks poor prognosis. However, the molecular mechanism explaining how c-Myb contributes to progression of CRC has not been fully elucidated. To address this point, we investigated the way how c-Myb affects sensitivity of CRC cells to anticancer drugs. Using CRC cell lines expressing exogenous c-myb we show that c-Myb protects CRC cells from the cisplatin-, oxaliplatin-, and doxorubicin-induced apoptosis, elevates reactive oxygen species via up-regulation of NOX1, and sustains the pro-survival p38 MAPK pathway. Using pharmacological inhibitors and gene silencing of p38 and NOX1 we found that these proteins are essential for the protective effect of c-Myb and that NOX1 acts upstream of p38 activation. In addition, our result suggests that transcription of NOX1 is directly controlled by c-Myb and these genes are strongly co-expressed in human tumor tissue of CRC patients. The novel c-Myb/NOX1/p38 signaling axis that protects CRC cells from chemotherapy described in this study could provide a new base for design of future therapies of CRC.
BACKGROUND: Although matrix metalloproteinase (MMP)-7 expression is correlated with increased metastatic potential in human colon cancer cells, the underlying molecular mechanism of invasive phenotype remains unknown. In the current study, we investigated the regulatory effects of membrane NADPH oxidase (NOX) and AMP activated protein kinase (AMPK) on MMP-7 expression and invasive phenotype change in colon cancer cells.
METHODS: Production of superoxide anion was measured by lucigenin chemiluminescence assay using whole cells and protein extracts (NADPH oxidase activity), and intracellular reactive oxygen species (ROS) by fluorescence microscopy using 2',7'-dichlorofluorescein diacetate (DCF-DA). Quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting were used to measure mRNA and protein levels, respectively. siRNA transfection was used to assess involvement of genes in cancer invasion, which were identified by Matrigel transwell invasion assay. Luciferase reporter assay was performed to identify transcription factors linked to gene expression.
RESULTS: Under basal conditions, less invasive human colon cancer cells (HT29 and Caco-2) showed low MMP-7 expression but high NOX1 expression and AMPK phosphorylation. Treatment of HT29 and Caco-2 cells with 12-O-tetradecanoylphorbol-13-acetate (TPA) induced an invasive phenotype response along with corresponding increases in ROS production and NOX2 and MMP-7 expression as well as reduced AMPK phosphorylation, which resemble basal conditions of highly invasive human colon cancer cells (SW620 and HCT116). In addition, inverse regulation between AMPK phosphorylation and NOX2 and MMP-7 expression was observed in HT29 cells treated with different concentrations of exogenous hydrogen peroxide. TPA-induced invasive phenotype in HT29 cells was abolished by treatment with Vit. E, DPI, apocynin, and NOX2 siRNA but not NOX1 siRNA, indicating NOX2-derived ROS production induced an invasive phenotype. TPA-induced induction of MMP-7 expression was suppressed by AP-1, NF-κB, and MAPK (ERK, p38, and JNK) inhibitors, whereas TPA-induced expression of NOX2 and its regulators, p47phox and p67phox, was blocked by p38 and NF-κB inhibitors.
CONCLUSIONS: Molecular switch from NOX1 to NOX2 in colon cancer cells induces ROS production and subsequently enhances MMP-7 expression by deactivating AMPK, which otherwise inhibits stimulus-induced autoregulation of ROS and NOX2 gene expression.
Guo S, Chen XThe human Nox4: gene, structure, physiological function and pathological significance.
J Drug Target. 2015; 23(10):888-96 [PubMed
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Increased generation of reactive oxygen species (ROS) has been implicated in the pathogenesis of a variety of diseases such as cardiovascular diseases and cancer. NADPH oxidase (Nox), a multicomponent enzyme, has been identified as one of the key sources of ROS. Nox4, one of the seven members of Nox family (Nox1, Nox2, Nox3, Nox4, Nox5, Duox1 and Duox2), has been extensively investigated in recent years. Its unique structures result in the constitutive generation of hydrogen peroxide (H2O2) as the main product. As a key oxygen sensor, Nox4-derived H2O2 plays diverse roles in cell proliferation, migration and death. Increased expression of Nox4 in cancer has been observed, which participates in metastasis, angiogenesis and apoptosis. Expression of Nox4 in endothelial cells actively mediated endothelial activation, dysfunction and injury, which contributes to the development of atherosclerosis, hypertension, cardiac hypertrophy and among others. This article explores the experimental studies related to the gene, structure, physiological function and pathological significance of Nox4. As Nox4 might serve as a potential target for the therapy of cardiovascular diseases and cancer, the Nox4 inhibitor is also discussed in this article.
Elatrech I, Marzaioli V, Boukemara H, et al.Escherichia coli LF82 differentially regulates ROS production and mucin expression in intestinal epithelial T84 cells: implication of NOX1.
Inflamm Bowel Dis. 2015; 21(5):1018-26 [PubMed
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BACKGROUND: Increased reactive oxygen species (ROS) production is associated with inflamed ileal lesions in Crohn's disease colonized by pathogenic adherent-invasive Escherichia coli LF82. We investigated whether such ileal bacteria can modulate ROS production by epithelial cells, thus impacting on inflammation and mucin expression.
METHODS: Ileal bacteria from patients with Crohn's disease were incubated with cultured epithelial T84 cells, and ROS production was assayed using the luminol-amplified chemiluminescence method. The gentamicin protection assay was used for bacterial invasion of T84 cell. The expression of NADPH oxidase (NOX) subunits, mucin, and IL-8 was analyzed by quantitative real-time PCR and Western blots. Involvement of NOX and ROS was analyzed using diphenyleneiodonium (DPI) and N-acetylcysteine (NAC).
RESULTS: Among different bacteria tested, only LF82 induced an increase of ROS production by T84 cells in a dose-dependent manner. This response was inhibited by DPI and NAC. Heat- or ethanol-attenuated LF82 bacteria and the mutant LF82ΔFimA, which does not express pili type 1 and poorly adheres to epithelial cells, did not induce the oxidative response. The LF82-induced oxidative response coincides with its invasion in T84 cells, and both processes were inhibited by DPI. Also, we observed an increased expression of NOX1 and NOXO1 in response to LF82 bacteria versus the mutant LF82ΔFimA. Furthermore, LF82 inhibited mucin gene expression (MUC2 and MUC5AC) in T84 cells while increasing the chemotactic IL-8 expression, both in a DPI-sensitive manner.
CONCLUSIONS: Adherent-invasive E. coli LF82 induced ROS production by intestinal NADPH oxidase and altered mucin and IL-8 expression, leading to perpetuation of inflammatory lesions in Crohn's disease.
Wang F, Lin F, Zhang P, et al.Thioredoxin-1 inhibitor, 1-methylpropyl 2-imidazolyl disulfide, inhibits the growth, migration and invasion of colorectal cancer cell lines.
Oncol Rep. 2015; 33(2):967-73 [PubMed
] Related Publications
1-Methylpropyl 2-imidazolyl disulfide (PX-12) has been proposed as an inhibitor of thioredoxin-1 (Trx-1) with antitumor activity. However, the antitumor activity of the Trx-1 redox signaling inhibitor PX-12 on colorectal cancer is still obscure. In the present study, we showed that PX-12 inhibited the growth of colorectal cancer DLD-1 and SW620 cells in a dose- and time-dependent manner. Further analysis demonstrated that PX-12 reduced cell colony formation and induced a G2/M phase arrest of the cell cycle. In addition, PX-12 treatment induced apoptosis, as observed by the increased number of Annexin V-positive cells and increased activation of caspase-3. Notably, a low dose of PX-12 inhibited colorectal cancer cell migration and invasion. Treatment of cancer cells with PX-12 reduced NOX1, CDH17 and S100A4 mRNA expression, and increased KLF17 mRNA expression. Moreover, PX-12 decreased S100A4 protein expression in the colorectal cancer cells. Collectively, the present study demonstrates the antitumor effects and therapeutic potential of PX-12 in colorectal cancer.
Cancers exhibit abnormal molecular signatures associated with disease initiation and progression. Molecular signatures could improve cancer screening, detection, drug development and selection of appropriate drug therapies for individual patients. Typically only very small amounts of tissue are available from patients for analysis and biopsy samples exhibit broad heterogeneity that cannot be captured using a single marker. This report details application of an in-house custom designed GenomeLab System multiplex gene expression assay, the hCellMarkerPlex, to assess predictive gene signatures of normal, adenomatous polyp and carcinoma colon tissue using archived tissue bank material. The hCellMarkerPlex incorporates twenty-one gene markers: epithelial (EZR, KRT18, NOX1, SLC9A2), proliferation (PCNA, CCND1, MS4A12), differentiation (B4GANLT2, CDX1, CDX2), apoptotic (CASP3, NOX1, NTN1), fibroblast (FSP1, COL1A1), structural (ACTG2, CNN1, DES), gene transcription (HDAC1), stem cell (LGR5), endothelial (VWF) and mucin production (MUC2). Gene signatures distinguished normal, adenomatous polyp and carcinoma. Individual gene targets significantly contributing to molecular tissue types, classifier genes, were further characterised using real-time PCR, in-situ hybridisation and immunohistochemistry revealing aberrant epithelial expression of MS4A12, LGR5 CDX2, NOX1 and SLC9A2 prior to development of carcinoma. Identified gene signatures identify aberrant epithelial expression of genes prior to cancer development using in-house custom designed gene expression multiplex assays. This approach may be used to assist in objective classification of disease initiation, staging, progression and therapeutic responses using biopsy material.
A role for intracellular ROS production has been recently implicated in the pathogenesis and progression of a wide variety of neoplasias. ROS sources, such as NAD(P)H oxidase (Nox) complexes, are frequently activated in AML (acute myeloid leukemia) blasts and strongly contribute to their proliferation, survival, and drug resistance. Myelodysplastic syndromes (MDS) comprise a heterogeneous group of disorders characterized by ineffective hematopoiesis, with an increased propensity to develop AML. The molecular basis for MDS progression is unknown, but a key element in MDS disease progression is the genomic instability. NADPH oxidases are now recognized to have specific subcellular localizations, this targeting to specific compartments for localized ROS production. Local Nox-dependent ROS production in the nucleus may contribute to the regulation of redox-dependent cell growth, differentiation, senescence, DNA damage, and apoptosis. We observed that Nox1, 2, and 4 isoforms and p22phox and Rac1 subunits are expressed in MDS/AML cell lines and MDS samples, also in the nuclear fractions. Interestingly, Nox4 interacts with ERK and Akt1 within nuclear speckle domain, suggesting that Nox4 could be involved in regulating gene expression and splicing factor activity. These data contribute to the elucidation of the molecular mechanisms used by nuclear ROS to drive MDS evolution to AML.
Expression of membrane-associated NADPH oxidase (NOX1) represents a characteristic feature of malignant cells. NOX1-derived extracellular superoxide anions are the basis for autocrine stimulation of proliferation, but also drive the HOCl and the NO/peroxynitrite signaling pathways. This may cause the elimination of transformed cells. Tumor cells express membrane-associated catalase that efficiently protects the cells against apoptosis-inducing reactive oxygen species (ROS) signaling. Membrane-associated superoxide dismutase (SOD) plays a co-modulatory protective role that is functionally interrelated with the protective effect mediated by catalase. Due to the co-localization of NOX1, catalase and SOD on the outer membrane of tumor cells, specific inhibition of membrane-associated SOD causes superoxide anion-dependent inhibition of catalase. This establishes a strong apoptotic signaling through the NO/peroxynitrite pathway. In parallel, it causes a drastic decrease in the concentration of proliferation-stimulating H2O2. Knowledge of the biochemical network on the surface of tumor cells should, therefore, allow development of specific novel strategies for tumor therapy, based on the specific features of tumor cell-specific extracellular ROS interactions.
Recent studies suggest that gentian violet (GV) may have anticancer activity by inhibiting for instance NADPH oxidases (Nox genes) whose overexpression is linked to tumor progression. Nox1 overexpression has been shown to inhibit transcriptional activity of the oncosuppressor p53, impairing tumor cell response to anticancer drugs. The tumor suppressor p53 is a transcription factor that, upon cellular stress, is activated to induce target genes involved in tumor cell growth inhibition and apoptosis. Thus, its activation is important for efficient tumor eradication. In this study, we examined the effect of GV on wild-type (wt) p53 activity in cancer cells. We found that GV was able to overcome the inhibitory effect of the NADPH oxidase Nox1 on p53 transcriptional activity. For the first time we show that GV was able to directly induce p53/DNA binding and transcriptional activity. In vitro, GV markedly induced cancer cell death and apoptotic marker PARP cleavage in wtp53-carrying cells. GV-induced cell death was partly inhibited in cells deprived of p53, suggesting that the anticancer activity of GV may partly depend on p53 activation. GV is US Food and Drug Administration approved for human use and may, therefore, have therapeutic potential in the management of cancer through p53 activation.
Li Q, Fu GB, Zheng JT, et al.NADPH oxidase subunit p22(phox)-mediated reactive oxygen species contribute to angiogenesis and tumor growth through AKT and ERK1/2 signaling pathways in prostate cancer.
Biochim Biophys Acta. 2013; 1833(12):3375-85 [PubMed
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Excessive generation of reactive oxygen species (ROS) in cancer cells is associated with cancer development, but the underlying mechanisms and therapeutic significance remain elusive. In this study, we reported that levels of ROS and p22(phox) expression are greatly increased in human prostate cancer tissues, and knockdown of p22(phox) by specific small interfering RNA (siRNA) decreased ROS levels in prostate cancer cells. We also showed that stable downregulation of p22(phox) in prostate cancer cells inhibited cell proliferation and colony formation, which was mediated by AKT and extracellular signal-regulated kinase (ERK)1/2 signaling pathways and their downstream molecules hypoxia-inducible factor 1α (HIF-1α) and vascular endothelial growth factor (VEGF). The NADPH oxidase subunit NOX1 was also elevated in prostate cancer cells, and was involved in activation of AKT/ERK/HIF-1/VEGF pathway and regulation of cell proliferation. Knockdown of p22(phox) resulted in inhibition of tumor angiogenesis and tumor growth in nude mice. These findings reveal a new function of p22(phox) in tumor angiogenesis and tumor growth, and suggest that p22(phox) is a potential novel target for prostate cancer treatment.
BACKGROUND: Breast cancer, a heterogeneous disease has been broadly classified into oestrogen receptor positive (ER+) or oestrogen receptor negative (ER-) tumour types. Each of these tumours is dependent on specific signalling pathways for their progression. While high levels of survivin, an anti-apoptotic protein, increases aggressive behaviour in ER- breast tumours, oxidative stress (OS) promotes the progression of ER+ breast tumours. Mechanisms and molecular targets by which OS promotes tumourigenesis remain poorly understood.
RESULTS: DETA-NONOate, a nitric oxide (NO)-donor induces OS in breast cancer cell lines by early re-localisation and downregulation of cellular survivin. Using in vivo models of HMLE(HRAS) xenografts and E2-induced breast tumours in ACI rats, we demonstrate that high OS downregulates survivin during initiation of tumourigenesis. Overexpression of survivin in HMLE(HRAS) cells led to a significant delay in tumour initiation and tumour volume in nude mice. This inverse relationship between survivin and OS was also observed in ER+ human breast tumours. We also demonstrate an upregulation of NADPH oxidase-1 (NOX1) and its activating protein p67, which are novel markers of OS in E2-induced tumours in ACI rats and as well as in ER+ human breast tumours.
CONCLUSION: Our data, therefore, suggest that downregulation of survivin could be an important early event by which OS initiates breast tumour formation.
Doroshow JH, Gaur S, Markel S, et al.Effects of iodonium-class flavin dehydrogenase inhibitors on growth, reactive oxygen production, cell cycle progression, NADPH oxidase 1 levels, and gene expression in human colon cancer cells and xenografts.
Free Radic Biol Med. 2013; 57:162-75 [PubMed
] Free Access to Full Article Related Publications
Iodonium-class flavoprotein dehydrogenase inhibitors have been demonstrated to possess antiproliferative potential and to inhibit reactive oxygen production in human tumor cells, although the mechanism(s) that explains the relationship between altered cell growth and the generation of reactive oxygen species (ROS) remains an area of active investigation. Because of the ability of these compounds to inhibit the activity of flavoprotein-containing epithelial NADPH oxidases, we chose to examine the effects of several iodonium-class flavoprotein inhibitors on human colon cancer cell lines that express high, functional levels of a single such oxidase (NADPH oxidase 1, or Nox1). We found that diphenyleneiodonium (DPI), di-2-thienyliodonium (DTI), and iodonium diphenyl inhibited the growth of Caco2, HT-29, and LS-174T colon cancer cells at concentrations (10-250nM for DPI, 0.5-2.5μM for DTI, and 155nM to 10μM for iodonium diphenyl) substantially lower than needed for DU145 human prostate cancer cells, which do not possess functional NADPH oxidase activity. Drug treatment was associated with decreased H2O2 production and diminished intracellular ROS levels, lasting up to 24h, after short-term (1-h) exposure to the iodonium analogs. Decreased tumor cell proliferation was caused, in part, by a profound block in cell cycle progression at the G1/S interface in both LS-174T and HT-29 cells exposed to either DPI or DTI; and the G1 block was produced, for LS-174T cells, by upregulation of p27 and a drug concentration-related decrease in the expression of cyclins D1, A, and E that was partially prevented by exogenous H2O2. Not only did DPI and DTI decrease intracellular ROS, they both also significantly decreased the mRNA expression levels of Nox1, potentially contributing to the prolonged reduction in tumor cell reactive oxygen levels. We also found that DPI and DTI significantly decreased the growth of both HT-29 and LS-174T human tumor xenografts, at dose levels that produced peak plasma concentrations similar to those utilized for our in vitro experiments. These findings suggest that iodonium analogs have therapeutic potential for NADPH oxidase-containing human colon cancers in vivo and that at least part of their antineoplastic mechanism of action may be related to targeting Nox1.
Weyemi U, Redon CE, Parekh PR, et al.NADPH Oxidases NOXs and DUOXs as putative targets for cancer therapy.
Anticancer Agents Med Chem. 2013; 13(3):502-14 [PubMed
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Reactive oxygen species (ROS) form a class of molecules with both positive and negative impacts on cellular health. Negatively, ROS may react with cellular constituents including proteins, lipids, and DNA to generate an array of oxidative lesions. These lesions may compromise genome stability which is critical for long-term cellular homeostasis and healthy progeny. Paradoxically, ROS also function as strong signalling molecules that mediate various growth-related responses, so their presence is also essential for cellular metabolism. While ROS are generated in an unregulated manner by physical stresses such as exposure to ionizing radiation and biochemical malfunctions such as mitochondrial leakage, cells also contain the NADPH oxidases NOXs and DUOXs, which specifically generate ROS in a wide variety of tissues. While the NOXs/DUOXs may be involved in maintaining optimal cellular redox levels, there is also accumulating evidence that NADPH oxidases-derived ROS may elevate the risk for genomic instability and cancer. Cancer cells may produce high levels of ROS, and in some cases, the source of these ROS has been linked to NOX/DUOX deregulation as reported for prostate cancer (NOX1 and NOX5), melanoma and glioblastoma (NOX4) among others. In addition, recent studies reveal that targeting NADPH oxidases with NOXs inhibitors may impair tumor growth in vivo; indicating that these proteins may be useful targets in future clinical strategies to fight cancer. This review provides an overview of the current knowledge concerning these enzymes, their roles in cancer, and their potential as targets in future cancer therapies.
Elevated aerobic glycolysis in cancer cells (the Warburg effect) may be attributed to respiration injury or mitochondrial dysfunction, but the underlying mechanisms and therapeutic significance remain elusive. Here we report that induction of mitochondrial respiratory defect by tetracycline-controlled expression of a dominant negative form of DNA polymerase γ causes a metabolic shift from oxidative phosphorylation to glycolysis and increases ROS generation. We show that upregulation of NOX is critical to support the elevated glycolysis by providing additional NAD+. The upregulation of NOX is also consistently observed in cancer cells with compromised mitochondria due to the activation of oncogenic Ras or loss of p53, and in primary pancreatic cancer tissues. Suppression of NOX by chemical inhibition or genetic knockdown of gene expression selectively impacts cancer cells with mitochondrial dysfunction, leading to a decrease in cellular glycolysis, a loss of cell viability, and inhibition of cancer growth in vivo. Our study reveals a previously unrecognized function of NOX in cancer metabolism and suggests that NOX is a potential novel target for cancer treatment.
Liu F, Gomez Garcia AM, Meyskens FLNADPH oxidase 1 overexpression enhances invasion via matrix metalloproteinase-2 and epithelial-mesenchymal transition in melanoma cells.
J Invest Dermatol. 2012; 132(8):2033-41 [PubMed
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NADPH oxidase 1 (Nox1) is a member of the NADPH oxidase family that has not been well characterized in the melanocytic cell lineage. Here we demonstrated that Nox1 and Nox4 were detected in melanocytic lineage, with only Nox1 detected in normal human melanocytes and Nox4 in a subset of metastatic melanoma cell lines. The protein level and enzymatic activity of Nox1 was elevated in all melanoma cells as compared with normal melanocytes. Overexpression of GFP-Nox1 protein in Wm3211 primary melanoma cells increased invasion rate by 4- to 6-fold as measured by Matrigel invasion assay, whereas knocking down or inhibiting Nox1 decreased invasion by approximately 40-60% in Wm3211 and SK-Mel-28 cells. Matrix metalloproteinase-2 (MMP-2) was increased by Nox1 overexpression at the mRNA, protein, and activity levels, and decreased by Nox1 knockdown. MMP-2 promoter activity was also regulated by Nox1 knockdown. In addition, stable clones overexpressing Nox1 exhibited an epithelial-mesenchymal transition (EMT) as examined by cell morphology and EMT markers; knockdown or inhibiting Nox1 led to a reversal of EMT. Supplementing MMP-2 to culture media did not induce EMT, suggesting that EMT induction by Nox1 was not through MMP-2 upregulation. In summary, Nox1 was overexpressed in all melanoma cell lines examined, and enhanced cell invasion by MMP-2 upregulation and EMT induction.
Recent metabolic profiles of human prostate cancer tissues showed a significant increase in cysteine (Cys) and a significant decrease in reduced glutathione (GSH) during cancer progression from low- to high-grade Gleason scores. Cys is primarily localized extracellularly, whereas GSH is present mostly inside the cell. We hypothesized that extra- or intracellular redox state alterations differentially regulate cell invasion in PC3 prostate carcinoma cells versus PrEC normal prostate epithelial cells. Cells were exposed to media with calculated Cys/CySS redox potentials (E(h)CySS) ranging from -60 to -180mV. After 3h exposure to a reducing extracellular redox state (E(h)CySS=-180mV), matrix metalloprotease (MMP), gelatinase, and NADPH oxidase activities increased, correlating with increases in cell invasion, cell migration, and extracellular hydrogen peroxide levels in PC3 cells but not PrECs. Knockdown of NADPH oxidase or MMP with silencing RNAs during cultivation with E(h)CySS=-180mV medium significantly decreased PC3 cell invasion. Modulation of extra- and intracellular redox states by exposure of PC3 cells to Cys/CySS-free medium (approx E(h)CySS=-87mV) containing 500μMN-acetylcysteine resulted in a more reducing intracellular redox state and a significant decrease in cell invasive ability. The decrease in PC3 cell invasion induced by these conditions correlated with a decrease in MMP activity. Our studies demonstrated that an extracellular redox state that was more reducing than a physiologic microenvironment redox state increased PC3 cancer cell invasive ability, whereas an intracellular redox environmental that was more reducing than an intracellular physiologic redox state inhibited PC3 cell invasive ability.
Cao JY, Mansouri S, Frappier LChanges in the nasopharyngeal carcinoma nuclear proteome induced by the EBNA1 protein of Epstein-Barr virus reveal potential roles for EBNA1 in metastasis and oxidative stress responses.
J Virol. 2012; 86(1):382-94 [PubMed
] Free Access to Full Article Related Publications
Epstein-Barr virus (EBV) infection is causatively associated with a variety of human cancers, including nasopharyngeal carcinoma (NPC). The only viral nuclear protein expressed in NPC is EBNA1, which can alter cellular properties in ways that may promote oncogenesis. Here, we used 2-dimensional difference gel electrophoresis (2-D DiGE) to profile changes in the nuclear proteome that occur after stable expression of EBNA1 in the EBV-negative NPC cell line CNE2. We found that EBNA1 consistently altered the levels of a small percentage of the nuclear proteins. The identification of 19 of these proteins by mass spectrometry revealed that EBNA1 upregulated three proteins affecting metastatic potential (stathmin 1, maspin, and Nm23-H1) and several proteins in the oxidative stress response pathway, including the antioxidants superoxide dismutase 1 (SOD1) and peroxiredoxin 1 (Prx1). Western blot analysis verified that EBNA1 expression upregulated and EBNA1 silencing downregulated these proteins. In addition, transcripts for stathmin 1 were induced by EBNA1, whereas EBNA1 only affected Prx1 and SOD1 at the protein level. Further investigation of the EBNA1 effects on the redox pathway showed that long-term EBNA1 expression in NPC resulted in increased reactive oxygen species (ROS) and increased levels of the NADPH oxidases NOX1 and NOX2, known to generate ROS. In addition, EBNA1 depletion in EBV-positive cells decreased NOX2 and ROS. The results show multiple roles for EBNA1 in the oxidative stress response pathway and suggest mechanisms by which EBNA1 may promote NPC metastases.
Sattayakhom A, Ittiwat W, Stremmel W, Chamulitrat WRedox regulation of cytokeratin 18 protein by NADPH oxidase 1 in preneoplastic human epithelial cells.
J Cancer Res Clin Oncol. 2011; 137(11):1669-78 [PubMed
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INTRODUCTION: A catalytic subunit of NADPH oxidase 1 (Nox1) is implicated to be involved in neoplastic progression in human epithelial cancers. We had previously demonstrated that Nox1 overexpression of immortalized epithelial cells was able to induce the generation of progenitor cells that expressed fetal-type cytokeratins 8 and 18.
PURPOSE: We aimed to investigate the direct effects and underlying mechanisms of Nox1 on expression of cytokeratin 18 (CK18).
METHODS: Immortalized human epithelial GM16 cells with low CK18 were used in Nox1 overexpression experiments. NuB2 cells with high CK18 were used in Nox1 knockdown experiments. Protein expression of CK18, phosphorylated and ubiquitinated CK18 were analyzed by Western blot.
RESULTS: With no effects on the mRNA levels, CK18 protein was increased upon Nox1 overexpression and decreased upon Nox1 knockdown. Treatment with proteasome inhibitor MG132 prevented CK18 degradation and increased CK18 protein indicating translational regulation of CK18. Treatment for NuB2 cells with N-acetyl-L: -cysteine, diphenyleneiodonium, or apocynin decreased CK18 protein levels indicating its regulation involving reactive oxygen species and flavoprotein Nox. It has been known that phosphorylation of CK18 regulates CK18 turnover by ubiquination. Consistently, Nox1 modulated CK18 phosphorylation at ser52. Nox1 knockdown and treatment with diphenyleneiodonium accumulated the levels of ubiquinated CK18 enhancing degradation causing decreased CK18 protein.
CONCLUSION: We demonstrated that Nox1 was able to induce CK18 stabilization by inhibiting CK18 protein degradation in a phosphorylation-dependent manner. CK18 accumulation induced by Nox1 is consistent with the persistence of fetal-type CK18 protein in many epithelial carcinomas.
Hexavalent chromium [Cr(VI)] is a well-known human carcinogen associated with the incidence of lung cancer. Although overproduction of reactive oxygen species (ROS) has been suggested to play a major role in its carcinogenicity, the mechanisms of Cr(VI)-induced ROS production remain unclear. In this study, we investigated the role of NADPH oxidase (NOX), one of the major sources of cellular ROS, in Cr(VI)-induced oxidative stress and carcinogenesis. We found that short-term exposure to Cr(VI) (2μM) resulted in a rapid increase in ROS generation in Beas-2B cells, and concomitantly increased NOX activity and expression of NOX members (NOX1-3 and NOX5) and subunits (p22(phox), p47(phox), p40(phox), and p67(phox)). Cr(VI) also induced phosphorylation of p47(phox) and membrane translocation of p47(phox) and p67(phox), further confirming NOX activation. Knockdown of p47(phox) with a short hairpin RNA attenuated the ROS production induced by Cr(VI). Chronic exposure (up to 3 months) to low doses of Cr(VI) (0.125, 0.25, and 0.5μM) also promoted ROS generation and the expression of NOX subunits, such as p47(phox) and p67(phox), but inhibited the expression of main antioxidant enzymes, such as superoxidase dismutase (SOD) and glutathione peroxidase (GPx). Chronic Cr(VI) exposure resulted in transformation of Beas-2B cells, increasing cell proliferation, anchorage independent growth in soft agar, and forming aggressive tumors in nude mice. Stable knockdown of p47(phox) or overexpression of SOD1, SOD2, or catalase (CAT) eliminated Cr(VI)-induced malignant transformation. Our results suggest that NOX plays an important role in Cr(VI)-induced ROS generation and carcinogenesis.
Redox regulation of epidermal growth factor receptor (EGFR) signaling helps protect cells against oxidative stress. In this study, we investigated whether the cytotoxicity of an EGFR tyrosine kinase inhibitor, erlotinib (ERL), was mediated by induction of oxidative stress in human head and neck cancer (HNSCC) cells. ERL elicited cytotoxicity in vitro and in vivo while increasing a panel of oxidative stress parameters which were all reversible by the antioxidant N-acetyl cysteine. Knockdown of EGFR by using siRNA similarly increased these oxidative stress parameters. Overexpression of mitochondrial targeted catalase but not superoxide dismutase reversed ERL-induced cytotoxicity. Consistent with a general role for NADPH oxidase (NOX) enzymes in ERL-induced oxidative stress, ERL-induced cytotoxicity was reversed by diphenylene iodonium, a NOX complex inhibitor. ERL reduced the expression of NOX1, NOX2, and NOX5 but induced the expression of NOX4. Knockdown of NOX4 by using siRNA protected HNSCC cells from ERL-induced cytotoxicity and oxidative stress. Our findings support the concept that ERL-induced cytotoxicity is based on a specific mechanism of oxidative stress mediated by hydrogen peroxide production through NOX4 signaling.
Reactive oxygen species, ROS, are regulators of endothelial cell migration, proliferation and survival, events critically involved in angiogenesis. Different isoforms of ROS-generating NOX enzymes are expressed in the vasculature and provide distinct signaling cues through differential localization and activation. We show that mice deficient in NOX1, but not NOX2 or NOX4, have impaired angiogenesis. NOX1 expression and activity is increased in primary mouse and human endothelial cells upon angiogenic stimulation. NOX1 silencing decreases endothelial cell migration and tube-like structure formation, through the inhibition of PPARα, a regulator of NF-κB. Administration of a novel NOX-specific inhibitor reduced angiogenesis and tumor growth in vivo in a PPARα dependent manner. In conclusion, vascular NOX1 is a critical mediator of angiogenesis and an attractive target for anti-angiogenic therapies.
The incidence rates of urinary bladder cancer continue to rise yearly, and thus new therapeutic approaches and early diagnostic markers for bladder cancer are urgently needed. Thus, identifying the key mediators and molecular mechanisms responsible for the survival of bladder cancer has valuable implications for the development of therapy. In this study, the role of BLT2, a receptor for leukotriene B((4)) (LTB((4))) and 12(S)-hydroxyeicosatetraenoic acid (HETE), in the survival of bladder cancer 253J-BV cells was investigated. We found that the expression of BLT2 is highly elevated in bladder cancer cells. Also, we observed that blockade of BLT2 with an antagonist or BLT2 siRNA resulted in cell cycle arrest and apoptotic cell death, suggesting a role of BLT2 in the survival of human bladder cancer 253J-BV cells. Further experiments aimed at elucidating the mechanism by which BLT2 mediates survival revealed that enhanced level of reactive oxygen species (ROS) are generated via a BLT2-dependent up-regulation of NADPH oxidase members NOX1 and NOX4. Additionally, we observed that inhibition of ROS generation by either NOX1/4 siRNAs or treatment with an ROS-scavenging agent results in apoptotic cell death in 253J-BV bladder cancer cells. These results demonstrated that a 'BLT2-NOX1/4-ROS' cascade plays a role in the survival of this aggressive bladder cancer cells, thus pointing to BLT2 as a potential target for anti-bladder cancer therapy.
DNA damage is a well-known initiator of tumorigenesis. Studies have shown that most cancer cells rely on aerobic glycolysis for their bioenergetics. We sought to identify a molecular link between genomic mutations and metabolic alterations in neoplastic transformation. We took advantage of the intrinsic genomic instability arising in xeroderma pigmentosum C (XPC). The XPC protein plays a key role in recognizing DNA damage in nucleotide excision repair, and patients with XPC deficiency have increased incidence of skin cancer and other malignancies. In cultured human keratinocytes, we showed that lentivirus-mediated knockdown of XPC reduced mitochondrial oxidative phosphorylation and increased glycolysis, recapitulating cancer cell metabolism. Accumulation of unrepaired DNA following XPC silencing increased DNA-dependent protein kinase activity, which subsequently activated AKT1 and NADPH oxidase-1 (NOX1), resulting in ROS production and accumulation of specific deletions in mitochondrial DNA (mtDNA) over time. Subcutaneous injection of XPC-deficient keratinocytes into immunodeficient mice led to squamous cell carcinoma formation, demonstrating the tumorigenic potential of transduced cells. Conversely, simultaneous knockdown of either NOX1 or AKT1 blocked the neoplastic transformation induced by XPC silencing. Our results demonstrate that genomic instability resulting from XPC silencing results in activation of AKT1 and subsequently NOX1 to induce ROS generation, mtDNA deletions, and neoplastic transformation in human keratinocytes.
Transformation by tyrosine kinase oncogenes (TKOs) in myeloid malignancies, including BCR-ABL in chronic myeloid leukemia, FLT3ITD in acute myeloid leukemia or JAK2V617F in myeloproliferative neoplasms, is associated with increased growth and cytoskeletal abnormalities. Using targeted approaches against components of the superoxide-producing NADPH-oxidases, including NADPH oxidase 2 (NOX2), NOX4 and the common p22(phox) subunit of NOX1-4, myeloid cells were found to display reduced cell growth and spontaneous migration. Consistent with a role of NOXs as regulators of membrane proximal signaling events in nonphagocytic cells, NOX2 and NOX4 were not involved in the excess production of intracellular reactive oxygen species and did not significantly increase oxygen consumption. All NOX family members are controlled in part through levels of the rate-limiting substrate NADPH, which was found to be significantly elevated in TKO-transformed cells. Also, reduced phosphorylation of the actin filament crosslinking protein myristoylated alanine-rich C-kinase substrate (MARCKS) in response to suppression of p22(phox) hints at a novel effector of NOX signaling. MARCKS was also found to be required for increased migration. Overall, these data suggest a model whereby NOX links metabolic NADPH production to cellular events that directly contribute to transformation.
Gianni D, Taulet N, DerMardirossian C, Bokoch GMc-Src-mediated phosphorylation of NoxA1 and Tks4 induces the reactive oxygen species (ROS)-dependent formation of functional invadopodia in human colon cancer cells.
Mol Biol Cell. 2010; 21(23):4287-98 [PubMed
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The NADPH oxidase family, consisting of Nox1-5 and Duox1-2, catalyzes the regulated formation of reactive oxygen species (ROS). Highly expressed in the colon, Nox1 needs the organizer subunit NoxO1 and the activator subunit NoxA1 for its activity. The tyrosine kinase c-Src is necessary for the formation of invadopodia, phosphotyrosine-rich structures which degrade the extracellular matrix (ECM). Many Src substrates are invadopodia components, including the novel Nox1 organizer Tks4 and Tks5 proteins. Nox1-dependent ROS generation is necessary for the maintenance of functional invadopodia in human colon cancer cells. However, the signals and the molecular machinery involved in the redox-dependent regulation of invadopodia formation remain unclear. Here, we show that the interaction of NoxA1 and Tks proteins is dependent on Src activity. Interestingly, the abolishment of Src-mediated phosphorylation of Tyr110 on NoxA1 and of Tyr508 on Tks4 blocks their binding and decreases Nox1-dependent ROS generation. The contemporary presence of Tks4 and NoxA1 unphosphorylable mutants blocks SrcYF-induced invadopodia formation and ECM degradation, while the overexpression of Tks4 and NoxA1 phosphomimetic mutants rescues this phenotype. Taken together, these results elucidate the role of c-Src activity on the formation of invadopodia and may provide insight into the mechanisms of tumor formation in colon cancers.
FaO rat hepatoma cells proliferate in the absence of serum through a mechanism that requires activation of the epidermal growth factor receptor (EGFR) pathway. The aim of this work was to analyze the molecular mechanisms that control EGFR activation in these and other liver tumor cells. Reactive oxygen species production is observed a short time after serum withdrawal in FaO cells, coincident with up-regulation of the NADPH oxidase NOX1. NOX1-targeted knockdown, the use of antioxidants, or pharmacological inhibition of NADPH oxidase attenuates autocrine growth, coincident with lower mRNA levels of EGFR and its ligand transforming growth factor-alpha (TGF-alpha) and a decrease in phosphorylation of EGFR. EGFR-targeted knockdown induces similar effects on cell growth and downstream signals to those observed in NOX1-depleted cells. Early NOX1 activation induces both a feedback-positive loop via an Src-ERK pathway that up-regulates its own levels, and a parallel signaling pathway through p38 MAPK and AKT resulting in EGFR and TGF-alpha up-regulation. Human hepatocellular carcinoma cell lines, but not non-tumoral hepatocytes, show autocrine growth upon serum withdrawal, which is also coincident with NOX1 up-regulation that mediates EGFR and TGF-alpha expression. The use of antioxidants, or pharmacological inhibition of NADPH oxidase, effectively attenuates autocrine growth in hepatocellular carcinoma cell lines. In summary, results presented in this study indicate that NOX1 might control autocrine cell growth of liver tumor cells through regulation of the EGFR pathway.
Reactive oxygen species (ROS) are known to be involved in many physiological and pathological processes. Initially ROS-producing NADPH oxidase (NOX) proteins were thought to be present in phagocytes. However, recent studies have demonstrated that NOX proteins are expressed in many other cell types and tissues. NOX family members' expression and function seems to vary from tissue to tissue. We determined the expression of the NOX family of proteins (NOX1-5) in normal breast tissue and breast tumors. Our study revealed that normal breast tissues express NOX1, 4 and 5 genes. Similar pattern of expression was revealed in a breast epithelial cell line. We found that NOX4 was overexpressed in the majority of breast cancer cell lines and primary breast tumors. NOX4 was also overexpressed in ovarian tumors. Overexpression of NOX4 in normal breast epithelial cells resulted in cellular senescence, resistance to apoptosis, and tumorigenic transformation. Overexpression of NOX4 in already transformed breast tumor cells also showed increased tumorigenicity. Strong evidence suggests that regulation of these processes occurs through NOX4 generation of ROS in the mitochondria. We demonstrate that the NOX4 protein contains a 73 amino acid long mitochondrial localization signal at the N-terminus that is capable of transporting a passenger protein GFP into the mitochondria. Treatment of NOX4 overexpressing cells with catalase resulted in decreased tumorigenic characteristics. Together, this study provides evidence for an oncogenic function for NOX4 protein localized to mitochondria and suggests that NOX4 is a novel source of ROS produced in the mitochondria. This study also identifies a possible treatment of NOX4-induced breast cancer by antioxidant treatment.