RT Journal Article
SR Electronic
T1 Ref-1/APE1 inhibition with novel small molecules blocks ocular neovascularization
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 296590
DO 10.1101/296590
A1 Sardar Pasha Sheik Pran Babu
A1 Kamakshi Sishtla
A1 Rania S. Sulaiman
A1 Bomina Park
A1 Melissa L. Fishel
A1 James H. Wikel
A1 Mark R. Kelley
A1 Timothy W. Corson
YR 2018
UL http://biorxiv.org/content/early/2018/04/06/296590.abstract
AB Ocular neovascular diseases like wet age-related macular degeneration are a major cause of blindness. Novel therapies are greatly needed for these diseases. One appealing antiangiogenic target is reduction-oxidation factor 1-apurinic/apyrimidinic endonuclease 1 (Ref-1/APE1). This protein can act as a redox-sensitive transcriptional activator for NF-κB and other pro-angiogenic transcription factors. An existing inhibitor of Ref-1’s function, APX3330, previously showed antiangiogenic effects. Here, we developed improved APX3330 derivatives and assessed their antiangiogenic activity. We synthesized APX2009 and APX2014 and demonstrated enhanced inhibition of Ref-1 function in a DNA-binding assay compared to APX3330. Both compounds were antiproliferative against human retinal microvascular endothelial cells (HRECs; GI50 APX2009: 1.1 µM, APX2014: 110 nM) and macaque choroidal endothelial cells (Rf/6a; GI50 APX2009: 26 µM, APX2014: 5.0 µM). Both compounds significantly reduced the ability of HRECs and Rf/6a cells to form tubes at mid nanomolar concentrations compared to control, and both significantly inhibited HREC and Rf/6a cell migration in a scratch wound assay. Ex vivo, both APX2009 and APX2014 inhibited choroidal sprouting at low micromolar and high nanomolar concentrations respectively. In the laser-induced choroidal neovascularization mouse model, intraperitoneal APX2009 treatment significantly decreased lesion volume by 4-fold compared to vehicle (p < 0.0001, ANOVA with Dunnett’s post hoc tests), without obvious intraocular or systemic toxicity. Thus, Ref-1 inhibition with APX2009 and APX2014 blocks ocular angiogenesis in vitro and ex vivo, and APX2009 is an effective systemic therapy for CNV in vivo, establishing Ref-1 inhibition as a promising therapeutic approach for ocular neovascularization. Visual AbstractAMDage-related macular degenerationANOVAanalysis of varianceAP-1activator protein 1APE1apurinic/apyrimidinic endonuclease 1APX2009(E)-N,N-diethyl-2-((3-methoxy-1,4-dioxo-1,4-dihydronaphthalen-2-yl)methylene)pentanamideAPX2014(E)-N-methoxy-2-((3-methoxy-1,4-dioxo-1,4-dihydronaphthalen-2-yl)methylene)pentanamideAPX3330(2E)-2-[(4,5-dimethoxy-2-methyl-3,6-dioxo-1,4-cyclohexadien-1-yl)methylene]-undecanoic acidATCCAmerican Type Culture CollectionBSAbovine serum albuminCNVchoroidal neovascularizationDMEMDulbecco’s modified Eagle’s mediumDMSOdimethyl sulfoxideEBM-2endothelial basal medium 2EGM-2endothelial growth medium 2EMSAelectrophoretic mobility shift assayFDAFood and Drug AdministrationFAfluorescein angiographyGCLganglion cell layerGI50median growth inhibitory concentrationGS-IB4isolectin B4 from Griffonia simplicifoliaHIF-1αhypoxia inducible factor 1αHRECshuman retinal microvascular endothelial cellsINLinner nuclear layerL-CNVlaser-induced choroidal neovascularizationNF-κBnuclear factor κ light-chain-enhancer of activated B cellsOCToptical coherence tomographyONLouter nuclear layerPBSphosphate buffered salinePDRproliferative diabetic retinopathyPKTpropylene glycol, Kolliphor HS15, Tween 80Ref-1reduction-oxidation factor 1ROPretinopathy of prematurityRP2Drecommended Phase II doseRPEretinal pigment epitheliumSTAT3signal transducer and activator of transcription 3VEGFvascular endothelial growth factorVLDLRvery low density lipoprotein receptor