The novel antiangiogenic VJ115 inhibits the NADH oxidase ENOX1 and cytoskeleton-remodeling proteins
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Targeting tumor vasculature represents a rational strategy for controlling cancer. (Z)-(+/−)-2-(1-benzylindol-3-ylmethylene)-1-azabicyclo[2.2.2]octan-3-ol (denoted VJ115) is a novel chemical entity that inhibits the enzyme ENOX1, a NADH oxidase. Genetic and small molecule inhibition of ENOX1 inhibits endothelial cell tubule formation and tumor-mediated neo-angiogenesis. Inhibition of ENOX1 radiosensitizes tumor vasculature, a consequence of enhanced apoptosis. However, the molecular mechanisms underlying these observations are not well understood. Herein, we mechanistically link ENOX1-mediated regulation of cellular NADH concentrations with proteomics profiling of endothelial cell protein expression following exposure to VJ115. Pathway Studios network analysis of potential effector molecules identified by the proteomics profiling indicated that a VJ115 exposure capable of altering intracellular NADH concentrations impacted proteins involved in cytoskeletal reorganization. The analysis was validated using RT-PCR and immunoblotting of selected proteins. RNAi knockdown of ENOX1 was shown to suppress expression of stathmin and lamin A/C, proteins identified by the proteomics analysis to be suppressed upon VJ115 exposure. These data support the hypothesis that VJ115 inhibition of ENOX1 can impact expression of proteins involved in cytoskeletal reorganization and support a hypothesis in which ENOX1 activity links elevated cellular NADH concentrations with cytoskeletal reorganization and angiogenesis.
KeywordsENOX1 Angiogenesis Proteomics 2D-DIGE NADH
Supported in part by grants from U.S. National Institutes of Health/National Cancer Institute grants RO1CA140409, T32CA093240, Vanderbilt-Ingram Cancer Center grant P30 CA68485, and the Vanderbilt Academic Venture Capital fund for proteomics. We are thankful to Drs. V.J. Sonar and Y.T.R. Reddy for the synthesis of VJ115.
Conflict of interest statement
The authors declare that they have no conflict of interest
- 6.Venkateswaran A SK, Melville DB, Levic DS, Rybski WM, Walsh AJ, Skala MC, Crooks PA, Freeman ML, Knapik EW. The NADH oxidase Enox1 is required for vasculogenesis in Zebrafish. In Preparation.Google Scholar
- 14.Avi-Dor Y, Olson JM, Doherty MD, Kaplan NO (1962) Fluorescence of pyridine nucleotides in mitochondria. J Biol Chem 237:2377–2383Google Scholar
- 25.Lee SH, Dominguez R (2010) Regulation of actin cytoskeleton dynamics in cells. Mol Cells 29:311–25.Google Scholar
- 34.Muriel O, Echarri A, Hellriegel C, Pavon DM, Beccari L, Del Pozo MA (2011) Phosphorylated filamin A regulates actin-linked caveolae dynamics. J Cell Sci 124:2763–76.Google Scholar