The development and prevalidation of an in vitro mutagenicity assay based on MutaMouse primary hepatocytes, Part II: Assay performance for the identification of mutagenic chemicals
- DOI
- Language of the publication
- English
- Date
- 2019-02-03
- Type
- Article
- Author(s)
- Cox, Julie A.
- Zwart, Edwin P.
- Luijten, Mirjam
- White, Paul A.
- Publisher
- Wiley
Abstract
As demonstrated in Part I, cultured MutaMouse primary hepatocytes (PHs) are suitable cells for use in an in vitro gene mutation assay due to their metabolic competence, their “normal” phenotype, and the presence of the MutaMouse transgene for reliable mutation scoring. The performance of these cells in an in vitro gene mutation assay is evaluated in this study, Part II. A panel of 13 mutagenic and nonmutagenic compounds was selected to investigate the performance of the MutaMouse PH in vitro gene mutation assay. The nine mutagens represent a range of classes of chemicals and include mutagens that are both direct-acting and requiring metabolic activation. All the mutagens tested, except for ICR 191, elicited significant, concentration-dependent increases in mutant frequency (MF) ranging from 2.6- to 14.4-fold over the control. None of the four nonmutagens, including two misleading, or “false,” positives (i.e., tertiary butylhydroquinone [TBHQ] and eugenol), yielded any significant increases in MF. The benchmark dose covariate approach facilitated ranking of the positive chemicals from most (i.e., 3-nitrobenzanthrone [3-NBA], benzo[a]pyrene [BaP], and aflatoxin B1 [AFB1]) to least (i.e., N-ethyl-N-nitrosourea [ENU]) potent. Overall, the results of this preliminary validation study suggest that this assay may serve as a complimentary tool alongside the standard genotoxicity test battery. This study, alongside Part I, illustrates the promise of MutaMouse PHs for use in an in vitro gene mutation assay, particularly for chemicals requiring metabolic activation.
Plain language summary
Risk assessments require the evaluation of a substance’s ability to cause genetic damage (i.e., genetic toxicity). Genetic toxicity tests using cultured cells (i.e., in vitro tests) are required in many jurisdictions, including Canada, for regulatory assessments of new substances. Existing and commonly used in vitro assays are not always reliable. Indeed, they have several known drawbacks, including inability to metabolically process chemicals. Additionally, there is a world-wide trend to reduce animal testing by developing and deploying novel in vitro genetic toxicity tests. In the companion to this paper (i.e., Part I), the isolation and characterization of liver cells (i.e., hepatocytes) from the MutaMouse, a mouse strain wherein mutations can be easily quantified, were described in detail. That work demonstrated that MutaMouse hepatocytes behave like normal liver cells, and are capable of metabolizing chemicals. In this study (i.e., Part II), the performance of an in vitro genetic toxicity assessment test based on MutaMouse hepatocytes was evaluated. The hepatocytes were exposed to 9 chemicals known to cause mutations, and 4 chemicals that do not cause mutations. These known genotoxicants represented different chemical classes with different modes of action. The performance of this assay exceeds the performance of other common in vitro assays; moreover, without the addition of rodent liver preparations (i.e., induced rat liver S9) to assist with the metabolism of chemicals tested in vitro. Taken together, the results of Part I and Part II demonstrate that a genetic toxicity assay based on cultured MutaMouse hepatocytes constitutes a promising tool for the reliable, cost-effective assessment of genotoxic hazard.
Subject
- Health,
- Health and safety