title: Characterization of NADPH oxidase 5 expression in human tumors and tumor cell lines with a novel mouse monoclonal antibody.
abstract: Reactive oxygen species generated by NADPH oxidase 5 (Nox5) have been implicated in physiological and pathophysiological signaling pathways, including cancer development and progression. However, because immunological tools are lacking, knowledge of the role of Nox5 in tumor biology has been limited; the expression of Nox5 protein across tumors and normal tissues is essentially unknown. Here, we report the characterization and use of a mouse monoclonal antibody against a recombinant Nox5 protein (bp 600-746) for expression profiling of Nox5 in human tumors by tissue microarray analysis. Using our novel antibody, we also report the detection of endogenous Nox5 protein in human UACC-257 melanoma cells. Immunofluorescence, confocal microscopy, and immunohistochemical techniques were employed to demonstrate Nox5 localization throughout UACC-257 cells, with perinuclear enhancement. Tissue microarray analysis revealed, for the first time, substantial Nox5 overexpression in several human cancers, including those of prostate, breast, colon, lung, brain, and ovary, as well as in malignant melanoma and non-Hodgkin lymphoma; expression in most nonmalignant tissues was negative to weak. This validated mouse monoclonal antibody will promote further exploration of the functional significance of Nox5 in human pathophysiology, including tumor cell growth and proliferation.
authors.affiliation: Laboratory of Molecular Pharmacology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Smitha
authors.initials: S
authors.lastname: Antony
authors.affiliation: Laboratory of Molecular Pharmacology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Yongzhong
authors.initials: Y
authors.lastname: Wu
authors.affiliation: Laboratory of Pathology, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Stephen M
authors.initials: SM
authors.lastname: Hewitt
authors.affiliation: Pathology/Histotechnology Laboratory, SAIC Frederick, Inc./Frederick National Laboratory for Cancer Research, National Institutes of Health, Frederick, MD 21702, USA.
authors.forename: Miriam R
authors.initials: MR
authors.lastname: Anver
authors.affiliation: Pathology/Histotechnology Laboratory, SAIC Frederick, Inc./Frederick National Laboratory for Cancer Research, National Institutes of Health, Frederick, MD 21702, USA.
authors.forename: Donna
authors.initials: D
authors.lastname: Butcher
authors.affiliation: Laboratory of Molecular Pharmacology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Guojian
authors.initials: G
authors.lastname: Jiang
authors.affiliation: Laboratory of Molecular Pharmacology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Jennifer L
authors.initials: JL
authors.lastname: Meitzler
authors.affiliation: Division of Cancer Treatment and Diagnosis, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Han
authors.initials: H
authors.lastname: Liu
authors.affiliation: Laboratory of Molecular Pharmacology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Agnes
authors.initials: A
authors.lastname: Juhasz
authors.affiliation: Laboratory of Molecular Pharmacology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Jiamo
authors.initials: J
authors.lastname: Lu
authors.affiliation: Division of Cancer Treatment and Diagnosis, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Krishnendu K
authors.initials: KK
authors.lastname: Roy
authors.affiliation: Laboratory of Molecular Pharmacology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA; Division of Cancer Treatment and Diagnosis, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA. Electronic address: doroshoj@mail.NIH.gov.
authors.forename: James H
authors.initials: JH
authors.lastname: Doroshow
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal; D014408:Biomarkers, Tumor / Q000032:analysis*; D015153:Blotting, Western; D045744:Cell Line, Tumor; D005260:Female; D005455:Fluorescent Antibody Technique; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D008565:Membrane Proteins / Q000032:analysis / Q000096:biosynthesis*; D051379:Mice; D018613:Microscopy, Confocal; D000074664:NADPH Oxidase 5; D019255:NADPH Oxidases / Q000032:analysis / Q000096:biosynthesis*; D009369:Neoplasms / Q000201:enzymology*; D060888:Real-Time Polymerase Chain Reaction; D046888:Tissue Array Analysis
chemical_list: D000911:Antibodies, Monoclonal; D014408:Biomarkers, Tumor; D008565:Membrane Proteins; D000074664:NADPH Oxidase 5; D019255:NADPH Oxidases; C431571:NOX5 protein, human
keywords: Duox; Free radicals; Melanoma; Mouse monoclonal Nox5 antibody; NADPH oxidase; NADPH oxidase 5; Nox; ROS; Reactive oxygen species; Superoxide; Tissue microarray; UACC-257 cells; dual oxidase; reactive oxygen species