The apurinic/apyrimidinic endonuclease 1/redox factor‐1 (Ape‐1/Ref‐1) is a multi‐functional protein, involved in DNA repair and the activation of redox‐sensitive transcription factors. The Ape‐1/Ref‐1 redox domain acts as a cytoprotective element in normal endothelial cells, mitigating the deleterious effects of apoptotic stimuli through induction of survival signals. We explored the role of the Ape‐1/Ref‐1 redox domain in the maintenance of tumor‐associated endothelium, and of endothelial progenitor cells (EPCs), which contribute to tumor angiogenesis. We demonstrate that E3330, a small molecule inhibitor of the Ape‐1/Ref‐1 redox domain, blocks the in vitro growth of pancreatic cancer‐associated endothelial cells (PCECs) and EPCs, which is recapitulated by stable expression of a dominant‐negative redox domain mutant. Further, E3330 blocks the differentiation of bone marrow‐derived mesenchymal stem cells (BMSCs) into CD31⁺ endothelial progeny. Exposure of PCECs to E3330 results in a reduction of H‐ras expression and intracellular nitric oxide (NO) levels, as well as decreased DNA‐binding activity of the hypoxia‐inducible transcription factor, HIF‐1α. E3330 also reduces secreted and intracellular vascular endothelial growth factor expression by pancreatic cancer cells, while concomitantly downregulating the cognate receptor Flk‐1/KDR on PCECs. Inhibition of the Ape‐1/Ref‐1 redox domain with E3330 or comparable angiogenesis inhibitors might be a potent therapeutic strategy in solid tumors. J. Cell. Physiol. 219: 209–218, 2009. © 2008 Wiley‐Liss, Inc.