High-Performance Affinity Beads for Identifying Anti-NF-κ B Drug Receptors

Hara, Masaki; Shimizu, Noriaki; Nishi, Takeyuki; Shima, Daisuke; Aizawa, Shin; Tanaka, Hirotoshi; Hatakeyama, Mamoru; Kawaguchi, Haruma; Handa, Hiroshi

doi:10.1016/s0076-6879(02)53038-x

Cited by 18 publications

(15 citation statements)

References 7 publications

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“…E3330 holds clinical therapeutic potential as a specific inhibitor of APE1 redox function [18]. The importance of this function is highlighted by results demonstrating that NF-κB-mediated gene expression is regulated by APE1 redox activity, without effects on IκBα degradation [19]. E3330 was also found to selectively inhibit growth/migration of human pancreatic cancer cells [20], suggesting that the APE1 redox function could represent a good candidate for inhibition of tumor invasion and metastasis.…”

Section: Introductionmentioning

confidence: 99%

Specific Inhibition of the Redox Activity of Ape1/Ref-1 by E3330 Blocks Tnf-Α-Induced Activation of Il-8 Production in Liver Cancer Cell Lines

et al. 2013

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APE1/Ref-1 is a main regulator of cellular response to oxidative stress via DNA-repair function and co-activating activity on the NF-κB transcription factor. APE1 is central in controlling the oxidative stress-based inflammatory processes through modulation of cytokines expression and its overexpression is responsible for the onset of chemoresistance in different tumors including hepatic cancer. We examined the functional role of APE1 overexpression during hepatic cell damage related to fatty acid accumulation and the role of the redox function of APE1 in the inflammatory process. HepG2 cells were stably transfected with functional and non-functional APE1 encoding plasmids and the protective effect of APE1 overexpression toward genotoxic compounds or FAs accumulation, was tested. JHH6 cells were stimulated with TNF-α in the presence or absence of E3330, an APE1 redox inhibitor. IL-8 promoter activity was assessed by a luciferase reporter assay, gene expression by Real-Time PCR and cytokines (IL-6, IL-8, IL-12) levels measured by ELISA. APE1 over-expression did not prevent cytotoxicity induced by lipid accumulation. E3330 treatment prevented the functional activation of NF-κB via the alteration of APE1 subcellular trafficking and reduced IL-6 and IL-8 expression induced by TNF-α and FAs accumulation through blockage of the redox-mediated activation of NF-κB. APE1 overexpression observed in hepatic cancer cells may reflect an adaptive response to cell damage and may be responsible for further cell resistance to chemotherapy and for the onset of inflammatory response. The efficacy of the inhibition of APE1 redox activity in blocking TNF-α and FAs induced inflammatory response opens new perspectives for treatment of inflammatory-based liver diseases.

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Section: Introductionmentioning

confidence: 99%

Specific Inhibition of the Redox Activity of Ape1/Ref-1 by E3330 Blocks Tnf-Α-Induced Activation of Il-8 Production in Liver Cancer Cell Lines

et al. 2013

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“…This molecule has previously been shown to block Ape1 function in studies with NFkB (35) and in hematopoietic cells (82) and is very specific for Ape1 with a high binding affinity and specificity (34, 65). In order to continue these studies, E3330, which is not commercially available, had to be synthesized using a new approach, which we developed and described in the methods section.…”

Section: Resultsmentioning

confidence: 99%

Role of the Multifunctional DNA Repair and Redox Signaling Protein Ape1/Ref-1 in Cancer and Endothelial Cells: Small-Molecule Inhibition of the Redox Function of Ape1

Luo

Delaplane

Jiang

et al. 2008

Antioxidants & Redox Signaling

148

263

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The DNA base excision repair pathway is responsible for the repair of DNA damage caused by oxidation/alkylation and protects cells against the effects of endogenous and exogenous agents. Removal of the damaged base by creates a baseless (AP) site. AP endonuclease1 (Ape1) acts upon this site to continue the BER pathway repair. Failure to repair baseless sites leads to DNA strand breaks and cytotoxicity. In addition to Ape1's repair role, it also functions as a major redox signaling factor to reduce and activate transcription factors such as AP1, p53, HIF-1α and others which control the expression of genes important for cell survival and cancer promotion and progression. Thus the Ape1 protein interacts with proteins involved in DNA repair, growth signaling pathways and pathways involved in tumor promotion and progression. While knockdown studies using siRNA have been informative in studying the role of Ape1 in both normal and cancer cells, knocking down Ape1 does not reveal the individual role of Ape1's redox or repair functions. The identification of small molecule inhibitors of specific Ape1 functions is critical for mechanistic studies and translational applications. Here we discuss small molecule inhibition of Ape1 redox and its effect on both cancer and endothelial cells.

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“…SG beads have several advantages over conventional affinity purification supports; their extremely large surface area results in a relatively high binding capacity, and their lack of pores facilitates the efficient removal of residual proteins. Thus, SG beads enable rapid and efficient purification of target proteins for a wide range of chemical compounds (17).…”

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confidence: 99%

Porphyrin Accumulation in Mitochondria Is Mediated by 2-Oxoglutarate Carrier

et al. 2006

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Heme (Fe-protoporphyrin IX), an endogenous porphyrin derivative, is an essential molecule in living aerobic organisms and plays a role in a variety of physiological processes such as oxygen transport, respiration, and signal transduction. For the biosynthesis of heme or the mitochondrial heme proteins, heme or its biosynthetic precursor porphyrin must be transported into mitochondria from cytosol. The mechanism of porphyrin accumulation in the mitochondrial inner membrane is unclear. In the present study, we analyzed the mechanism of mitochondrial translocation of porphyrin derivatives. We showed that palladium meso-tetra(4-carboxyphenyl)porphyrin (PdTCPP), a phosphorescent porphyrin derivative, accumulated in the mitochondria of several cell lines. Using affinity latex beads, we showed that 2-oxoglutarate carrier (OGC), the mitochondrial transporter of 2-oxoglutarate, bound to PdTCPP, and in vitro PdTCPP inhibited 2-oxoglutarate uptake into mitochondria in a competitive manner (K i ‫؍‬ 15 M). Interestingly, all types of porphyrin derivatives examined in this study competitively inhibited 2-oxoglutarate uptake into mitochondria, including protoporphyrin IX, coproporphyrin III, and hemin. Furthermore, mitochondrial accumulation of porphyrins was inhibited by 2-oxoglutarate or OGC inhibitor. These results suggested that porphyrin accumulation in mitochondria is mediated by OGC and that porphyrins are able to competitively inhibit 2-oxoglutarate uptake into mitochondria. This is the first report of a putative mechanism for accumulation of porphyrins in the mitochondrial inner membrane.Porphyrins consist of a tetrapyrrole ring structure and are most widely and efficiently used in energy metabolism. Heme (Fe-protoporphyrin IX), a porphyrin derivative, is a prosthetic molecule for several hemoproteins, and plays an essential role in various biological processes such as oxygen transport, respiration, and signal transduction. The biosynthesis of heme is a multistep process that starts with the condensation of glycineand succinyl-CoA to form 5-aminolevulinate (1). Heme is ultimately formed in the mitochondrial matrix space following translocation of the heme precursor, co-proporphyrinogen III, which is generated in cytosol, into mitochondria (1). Heme is then utilized in the mitochondrial matrix for the synthesis of a variety of heme proteins such as the cytochromes. Thus, it is necessary for heme biosynthesis and synthesis of the mitochondrial heme proteins that heme and porphyrin precursors must be transported into mitochondria from cytosol. However, the mechanism by which this mitochondrial accumulation occurs is unclear.Some metalloporphyrin derivatives are fluorescence-or phosphorescence-emitting molecules. These molecules undergo a shift from a low energy ground state to a high energy excited state upon exposure to UV light. Fluorescent or phosphorescent light is emitted as the molecules decay from their high energy state back down to the ground state. PdTCPP 3 (Fig. 1A) emits long-lived phosphorescence in respons...

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High-Performance Affinity Beads for Identifying Anti-NF-κ B Drug Receptors

Cited by 18 publications

References 7 publications

Specific Inhibition of the Redox Activity of Ape1/Ref-1 by E3330 Blocks Tnf-Α-Induced Activation of Il-8 Production in Liver Cancer Cell Lines

Specific Inhibition of the Redox Activity of Ape1/Ref-1 by E3330 Blocks Tnf-Α-Induced Activation of Il-8 Production in Liver Cancer Cell Lines

Role of the Multifunctional DNA Repair and Redox Signaling Protein Ape1/Ref-1 in Cancer and Endothelial Cells: Small-Molecule Inhibition of the Redox Function of Ape1

Porphyrin Accumulation in Mitochondria Is Mediated by 2-Oxoglutarate Carrier

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