Jaruga Pawel, Kant Melis, Luzadder Michael M, Lloyd R Stephen, Boldogh Istvan, Dizdaroglu Miral
Biomolecular Measurement Division, National Institute of Standards and Technology, Gaithersburg, Maryland 20899, United States.
Oregon Institute of Occupational Health Sciences, Oregon Health & Science University, Portland, Oregon 97239, United States.
Biochemistry. 2025 Apr 15;64(8):1788-1796. doi: 10.1021/acs.biochem.4c00419. Epub 2025 Apr 3.
DNA glycosylases of the base excision repair pathway have become clinically validated drug targets for the treatment of several diseases. Human OGG1 (hOGG1) is specific for the removal of the highly mutagenic 8-oxoguanine (8-oxo-Gua) and 2,6-diamino-4-hydroxy-5-formamidopyrimidine (FapyGua) from damaged DNA. To develop clinically approved drugs, various small-molecule inhibitors of hOGG1 have been developed to inhibit its glycosylase and lyase activities, with 4-(4-bromo-2-oxo-3H-benzimidazol-1-yl)--(4-iodophenyl)piperidine-1-carboxamide (TH5487) shown to be a potent inhibitor. The inhibition of hOGG1 by TH5487 has been shown to suppress cancer cell growth, pulmonary inflammation, and lung fibrosis and sensitize cancer cells to ionizing radiation, confirming hOGG1 as a target for pharmaceutical intervention. While the assays that identified TH5487 utilized an oligodeoxynucleotide with the target substrate being 8-hydroxyadenine mispaired with cytosine, measurements of TH5487-mediated inhibition of the release of 8-oxo-Gua and FapyGua have not been reported. In the present work, we investigated the inhibition of hOGG1 by TH5487 using genomic DNA with multiple lesions and gas chromatography-tandem mass spectrometry with isotope dilution to measure inhibition of hOGG1-catalyzed DNA base lesion removal from DNA. An oligodeoxynucleotide containing 8-oxo-Gua was also used to measure the half-maximal inhibitory concentration (IC), which is 0.800 μmol/L ± 0.061 μmol/L. We show that TH5487 efficiently inhibits the excision of both 8-oxo-Gua and FapyGua, and a minor substrate 4,6-diamino-5-formamidopyrimidine (FapyAde) from DNA with the IC values of 1.6 μmol/L, 3.1 μmol/L, and 3.1 μmol/L, respectively. The results suggest that the approach used in the present work may be applied for future studies of hOGG1 inhibition by TH5487 on cellular and animal disease models.
碱基切除修复途径中的DNA糖基化酶已成为治疗多种疾病的临床验证药物靶点。人类OGG1（hOGG1）特异性地从受损DNA中去除高度致突变的8-氧代鸟嘌呤（8-oxo-Gua）和2,6-二氨基-4-羟基-5-甲酰胺基嘧啶（FapyGua）。为开发临床批准的药物，已开发出多种hOGG1的小分子抑制剂来抑制其糖基化酶和裂解酶活性，其中4-(4-溴-2-氧代-3H-苯并咪唑-1-基)- (4-碘苯基)哌啶-1-甲酰胺（TH5487）被证明是一种有效的抑制剂。TH5487对hOGG1的抑制作用已被证明可抑制癌细胞生长、肺部炎症和肺纤维化，并使癌细胞对电离辐射敏感，证实hOGG1是药物干预的靶点。虽然鉴定TH5487的实验使用了一种寡脱氧核苷酸，其靶底物是与胞嘧啶错配的8-羟基腺嘌呤，但尚未报道TH5487介导的对8-oxo-Gua和FapyGua释放抑制的测量。在本研究中，我们使用具有多个损伤的基因组DNA和同位素稀释气相色谱-串联质谱法研究了TH5487对hOGG1的抑制作用，以测量hOGG1催化从DNA中去除DNA碱基损伤的抑制情况。还使用了含有8-oxo-Gua的寡脱氧核苷酸来测量半数最大抑制浓度（IC），其为0.800 μmol/L±0.061 μmol/L。我们表明，TH5487能有效抑制DNA中8-oxo-Gua、FapyGua以及次要底物4,6-二氨基-5-甲酰胺基嘧啶（FapyAde）的切除，其IC值分别为1.6 μmol/L、3.1 μmol/L和3.1 μmol/L。结果表明，本研究中使用的方法可能适用于未来在细胞和动物疾病模型上研究TH5487对hOGG1的抑制作用。
