Despite a series of persuasive experimental evidence for the involvement of eyes absent protein tyrosine phosphatases in various human cancers, no small-molecule inhibitor has been reported so far. We have identified seven novel inhibitors of eyes absent homologue 2 (Eya2) with IC(50) values ranging from 1 to 70 μm by the virtual screening with docking simulations and enzyme inhibition assay. Atomic charges of the active-site Mg(2+) ion complex are calculated to enhance the accuracy of docking simulations. The newly discovered inhibitors are structurally diverse and have various chelating groups for the Mg(2+) ion. The interactions with the amino acid residues responsible for the stabilizations of the inhibitors in the active site of Eya2 are addressed in detail.
"EYA is a metallo-enzyme , , . However the inhibitors described here are not metal-chelators, unlike compounds identified in a recently reported in silico screen performed using the isolated catalytic domain of EYA2 . While metal chelators can be high affinity inhibitors of metalloenzymes, as therapeutics they present challenges such as depletion of physiologically needed metal ions and inhibition of non-targeted metalloenzymes. "
[Show abstract][Hide abstract] ABSTRACT: Eyes Absents (EYA) are multifunctional proteins best known for their role in organogenesis. There is accumulating evidence that overexpression of EYAs in breast and ovarian cancers, and in malignant peripheral nerve sheath tumors, correlates with tumor growth and increased metastasis. The EYA protein is both a transcriptional activator and a tyrosine phosphatase, and the tyrosine phosphatase activity promotes single cell motility of mammary epithelial cells. Since EYAs are expressed in vascular endothelial cells and cell motility is a critical feature of angiogenesis we investigated the role of EYAs in this process. Using RNA interference techniques we show that EYA3 depletion in human umbilical vein endothelial cells inhibits transwell migration as well as Matrigel-induced tube formation. To specifically query the role of the EYA tyrosine phosphatase activity we employed a chemical biology approach. Through an experimental screen the uricosuric agents Benzbromarone and Benzarone were found to be potent EYA inhibitors, and Benzarone in particular exhibited selectivity towards EYA versus a representative classical protein tyrosine phosphatase, PTP1B. These compounds inhibit the motility of mammary epithelial cells over-expressing EYA2 as well as the motility of endothelial cells. Furthermore, they attenuate tubulogenesis in matrigel and sprouting angiogenesis in the ex vivo aortic ring assay in a dose-dependent fashion. The anti-angiogenic effect of the inhibitors was also demonstrated in vivo, as treatment of zebrafish embryos led to significant and dose-dependent defects in the developing vasculature. Taken together our results demonstrate that the EYA tyrosine phosphatase activity is pro-angiogenic and that Benzbromarone and Benzarone are attractive candidates for repurposing as drugs for the treatment of cancer metastasis, tumor angiogenesis, and vasculopathies.
PLoS ONE 04/2012; 7(4):e34806. DOI:10.1371/journal.pone.0034806 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: INTRODUCTION: Rhodanine-based compounds have been associated with numerous biological activities. After many years of research in drug discovery, they have gained a reputation as being pan assay interference compounds (PAINS) and frequent hitters in screening campaigns. Rhodanine-based compounds are also aggregators that can non-specifically interact with target proteins as well as Michael acceptors and interfere photometrically in biological assays due to their color. AREAS COVERED: The authors review the recently reported biological activities of rhodanine-based compounds. Furthermore, the article provides details of their synthesis and occurrence in compound libraries through high-throughput screening (HTS) and virtual high-throughput screening (VHTS). Additionally, the authors provide the reader with possible mechanisms of non-specific target modulation, analysis of the crystal structures of enzyme-rhodanine complexes and a comparison of rhodanine and thiazolidine-2,4-dione moieties. EXPERT OPINION: The biological activity of compounds possessing a rhodanine moiety should be considered very critically despite the convincing data obtained in biological assays. In addition to the lack of selectivity, unusual structure-activity relationship profiles and safety and specificity problems mean that rhodanines are generally not optimizable.
Expert Opinion on Drug Discovery 05/2012; 7(7):549-60. DOI:10.1517/17460441.2012.688743 · 3.54 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Although Eyes absent protein tyrosine phosphatases proved to be involved in various human cancers by a series of persuasive experimental evidence, only a very few number of small-molecule inhibitors have been reported so far. We have been able to identify 29 novel inhibitors of Eyes absent homologue 2 (Eya2) by means of a structure-based de novo design with the two known inhibitor scaffolds that contain a proper chelating group for the active-site Mg(2+) ion. Because these newly found inhibitors were screened for having desirable physicochemical properties as a drug candidate and exhibited a moderate inhibitory activity with IC(50) values ranging from 6 to 50μM, they deserve consideration for further investigation to develop new anticancer medicines. Structural features relevant to the stabilization of the identified inhibitors in the active site of Eya2 phosphatase are discussed in detail.
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