Mechanism of Redox Regulation by Ape1/Ref-1

Ape1/Ref-1 的氧化还原调节机制

• 批准号: 7144509
• 负责人: Millie M Georgiadis
• 金额: $ 24.2万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-07-01 至 2011-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7144509
• 关键词: clinical research; neoplasm /cancer; neoplastic cell; oxidation reduction reaction; role; transcription factor

DESCRIPTION (provided by applicant): Apurinic/apyrimidinic endonuclease (Ape1), an essential Base Excision Repair (BER) enzyme, has been shown to have elevated or altered levels of expression in a number of cancers.(1-8) In addition to its DMA repair activity, Ape1 has a redox function that serves to reduce a number of transcription factors including AP-1 (Fos/Jun), HIF-1a, NF-kB, PAX, HLF, p53, and others (reviewed in 1). Despite the discovery of Apel's redox activity more than 10 years ago, there is currently no detailed mechanism to describe how Ape1 reduces these transcription factors and conflicting data in the literature regarding the role of cysteine 65, thought to be critical for the redox function. We propose to revisit the mechanism of Apel's redox activity and its role in cancer etiology with the specific goal of exploring this unique and as yet unexplored activity as a target for the development of cancer therapeutics in the future. Our ability to detail the mechanism of Apel's redox activity and its role in cancer cells will be aided greatly by the use of 3-[5-(2,3-dimethoxy-6-methyl-1,4- benzoquinoyl)]-2-nonyl-2-propionic acid, (E3330), a quinone derivative, that has been shown (1) to bind specifically to Ape1 in a direct binding assay with high affinity and (2) to inhibit the redox activity of Ape1 both in vitro and in cancer cell lines. As detailed in Specific Aims 1 and 2, we have proposed to use X-ray crystallography, site-directed mutagenesis, chemical cross-linking, and hydrogen/deuterium exchange experiments in order to identify the residues that are required for Apel's redox activity and sites of interactions between Ape1 and the transcription factors that it reduces. In Specific Aim 3, we propose to inhibit the redox activity of Ape1 with E3330 and thereby determine the role of Apel's redox activity on downstream targets in normal and cancer cell lines. Relevance: Apurinic/apyrimidinic endonuclease (Ape1) has been shown to have elevated or altered levels of expression in a number of cancers. This multifunctional enzyme is an attractive target for the development of chemotherapeutics. We propose here to elucidate the mechanism of the redox function of Ape1 and explore its role in tumor cell response following treatment with DNA damaging agents.

DESCRIPTION (provided by applicant): Apurinic/apyrimidinic endonuclease (Ape1), an essential Base Excision Repair (BER) enzyme, has been shown to have elevated or altered levels of expression in a number of cancers.(1-8) In addition to its DMA repair activity, Ape1 has a redox function that serves to reduce a number of transcription factors including AP-1 (Fos/Jun), HIF-1a, NF-KB，PAX，HLF，P53等（1次审查）。尽管十多年前发现了APEL的氧化还原活性，但目前尚无详细的机制来描述APE1如何减少有关半胱氨酸65作用的文献中这些转录因子和相互矛盾的数据，这对氧化还原功能至关重要。我们建议重新审视APEL的氧化还原活性的机制及其在癌症病因中的作用，其特定目标是探索这种独特而尚未开发的活动，以此作为未来癌症治疗剂发展的目标。我们有能力通过使用3- [5-（2,3-二甲氧基-6-甲基-1-甲基-1,4-苯甲酰烯酰基）的能力在很大程度上有助于帮助，[2,3-二甲基-6-甲基-1,4-苯甲酰基）] - 2-氯-2-丙酸），（E3330）（E3330），（E3330），与Quinone衍生物相结合，APE（1）与Quinone衍生物相结合，APE（1）均具有APE。 （2）在体外和癌细胞系中抑制APE1的氧化还原活性。正如特定目标1和2中所述，我们提议使用X射线晶体学，定点诱变，化学交联以及氢/氘交换实验，以识别APEL的氧化还原活性和相互作用位点所需的残留物以及APE1之间的相互作用和降低的转录因子之间的相互作用。在特定目标3中，我们建议用E3330抑制APE1的氧化还原活性，从而确定APEL的氧化还原活性在正常和癌细胞系中下游靶标的作用。相关性：已证明许多癌症中的表达水平升高或变化，肾上腺素/丙酰胺核酸内切酶（APE1）的表达水平升高或改变。这种多功能酶是化学治疗剂开发的有吸引力的靶标。我们在这里提议阐明APE1的氧化还原功能的机制，并探索其在用DNA损伤剂处理后在肿瘤细胞反应中的作用。