Buick Julie K, Rowan-Carroll Andrea, Gagné Rémi, Williams Andrew, Chen Renxiang, Li Heng-Hong, Fornace Albert J, Chao Christy, Engelward Bevin P, Frötschl Roland, Ellinger-Ziegelbauer Heidrun, Pettit Syril D, Aubrecht Jiri, Yauk Carole L
Environmental Health Science and Research Bureau, Health Canada, Ottawa, ON, Canada.
Department of Oncology, Lombardi Comprehensive Cancer Center, Georgetown University Medical Center, Washington, DC, United States.
Front Toxicol. 2022 Sep 23;4:991590. doi: 10.3389/ftox.2022.991590. eCollection 2022.
Genotoxicity testing relies on the detection of gene mutations and chromosome damage and has been used in the genetic safety assessment of drugs and chemicals for decades. However, the results of standard genotoxicity tests are often difficult to interpret due to lack of mode of action information. The TGx-DDI transcriptomic biomarker provides mechanistic information on the DNA damage-inducing (DDI) capability of chemicals to aid in the interpretation of positive genotoxicity data. The CometChip assay was developed to assess DNA strand breaks in a higher-throughput format. We paired the TGx-DDI biomarker with the CometChip assay in TK6 cells to evaluate three model agents: nitrofurantoin (NIT), metronidazole (MTZ), and novobiocin (NOV). TGx-DDI was analyzed by two independent labs and technologies (nCounter and TempO-Seq). Although these anti-infective drugs are, or have been, used in human and/or veterinary medicine, the standard genotoxicity testing battery showed significant genetic safety findings. Specifically, NIT is a mutagen and causes chromosome damage, and MTZ and NOV cause chromosome damage in conventional tests. Herein, the TGx-DDI biomarker classified NIT and MTZ as non-DDI at all concentrations tested, suggesting that NIT's mutagenic activity is bacterial specific and that the observed chromosome damage by MTZ might be a consequence of test conditions. In contrast, NOV was classified as DDI at the second highest concentration tested, which is in line with the fact that NOV is a bacterial DNA-gyrase inhibitor that also affects topoisomerase II at high concentrations. The lack of DNA damage for NIT and MTZ was confirmed by the CometChip results, which were negative for all three drugs except at overtly cytotoxic concentrations. This case study demonstrates the utility of combining the TGx-DDI biomarker and CometChip to resolve conflicting genotoxicity data and provides further validation to support the reproducibility of the biomarker.
遗传毒性测试依赖于基因突变和染色体损伤的检测，几十年来一直用于药物和化学品的遗传安全性评估。然而，由于缺乏作用方式信息，标准遗传毒性测试的结果往往难以解释。TGx-DDI转录组学生物标志物提供了关于化学物质诱导DNA损伤（DDI）能力的机制信息，以帮助解释阳性遗传毒性数据。彗星芯片分析方法的开发是为了以更高通量的形式评估DNA链断裂。我们将TGx-DDI生物标志物与彗星芯片分析方法在TK6细胞中配对，以评估三种模型药物：呋喃妥因（NIT）、甲硝唑（MTZ）和新生霉素（NOV）。TGx-DDI由两个独立的实验室和技术（nCounter和TempO-Seq）进行分析。尽管这些抗感染药物现在或曾经用于人类和/或兽医学，但标准遗传毒性测试组合显示出显著的遗传安全性结果。具体而言，NIT是一种诱变剂，可导致染色体损伤，MTZ和NOV在传统测试中可导致染色体损伤。在此，TGx-DDI生物标志物在所有测试浓度下将NIT和MTZ分类为非DDI，这表明NIT的诱变活性具有细菌特异性，并且MTZ观察到的染色体损伤可能是测试条件的结果。相比之下，NOV在第二高测试浓度下被分类为DDI，这与NOV是一种细菌DNA促旋酶抑制剂且在高浓度下也影响拓扑异构酶II这一事实相符。彗星芯片结果证实了NIT和MTZ不存在DNA损伤，除了在明显的细胞毒性浓度下，这三种药物的结果均为阴性。本案例研究证明了将TGx-DDI生物标志物和彗星芯片相结合以解决相互矛盾的遗传毒性数据的实用性，并为支持该生物标志物的可重复性提供了进一步验证。
