In vitro and in vivo mammalian mutation assays support a nonmutagenic mechanism of carcinogenicity for hydrazine

Abstract

Hydrazine has been described as a mutagenic, probable human carcinogen. It is mutagenic in in vitro systems such as bacterial reverse mutation (Ames) tests and some yeast systems, as well as in in vivo systems with drosophila. It was shown to cause chromosome damage both in vitro and in vivo but was negative in some wellvalidated mammalian mutation systems such as CHO HPRT assays. Importantly, there is only one in vivo gene mutation test reported, which was negative. Our objective was to determine if hydrazine is mutagenic in mammalian test systems. Thus, we conducted an in vitro gene mutation test in Muta™Mouse lung epithelial cells (FE1 cell assay) and a regulatory-compliant in vivo Big Blue ® mouse test. Consistent with previous reports, an additional six-well Ames assay showed that hydrazine was mutagenic to bacteria. The FE1 cell assay was negative in conditions with and without metabolic activation when tested to cytotoxicity limits. In the Big Blue ® mouse study, female mice received dosages of hydrazine up to 10.9 mg/kg via drinking water for 28 days. This dose is comparable to a dose used in a carcinogenicity study where female mice had significant increases in hepatocellular adenoma at 11.5 mg/kg. There were no increases in mutant frequency in liver and lung, two tissues sensitive to the carcinogenic effects of hydrazine in mice. Our research shows that hydrazine is not mutagenic in mammalian cells either in vitro or in vivo, indicating mutagenicity may not play a role in the carcinogenicity of hydrazine.

Exploring the molecular and functional cellular response to hydrazine via transcriptomics and DNA repair mutant cell lines

“…These data illustrate how some genetic toxicology assays may give rise to positive (or in some cases, equivocal) in vitro genotoxicity tests, but then may result in a negative in vivo mutation test, referencing our previous work (Nicolette et al, 2020 ). Taking a step back, we should be clear that a chemical can be genotoxic but may not be an in vivo mutagen.…”

“…Standard assays used in determining genotoxicity and mutagenicity have been conducted with hydrazine. Our previous results confirmed the results of others showing that hydrazine is positive in the bacterial mutation (Ames) test, as well as in an vitro gene mutation yeast test (CERI, 2007 ; IARC, 1999 ; Nicolette et al, 2020 ). Additionally, hydrazine has been found to be positive in the chromosomal aberration test in Chinese hamster ovary (CHO) cells, but not in human lymphocytes (CERI, 2007 ; IARC, 1999 ).…”

“…Hydrazine is associated with the induction of DNA single‐strand breaks and alkali‐labile sites (comet assay). However, hydrazine failed to induce gene mutation in in vitro FE1 mouse cell and in vivo Big Blue mouse mutation studies (Nicolette et al, 2020 ). The lack of a hydrazine‐induced increase in mutation frequency measured in the organs for which carcinogenicity has been noted prompted us to explore additional mechanistic experimental work.…”

“…Despite the positive Ames test and DNA damage data, recent in vitro and in vivo mutation studies have been negative (Nicolette et al, 2020 ). We demonstrated that hydrazine did not increase mutations in vitro in Muta Mouse FE1 lung epithelial cells, which are derived from a tissue that has been shown to be sensitive to carcinogenic effects of hydrazine.…”

“…We demonstrated that hydrazine did not increase mutations in vitro in Muta Mouse FE1 lung epithelial cells, which are derived from a tissue that has been shown to be sensitive to carcinogenic effects of hydrazine. Additionally, hydrazine was not mutagenic in a good laboratory practice‐compliant (GLP) 28‐day Big Blue mouse study that explored mutation frequencies in both lung and liver at doses approximating those that induced tumors (Matsumoto et al, 2016 ; Nicolette et al, 2020 ).…”