1-chloro-N,N-diethyl-1,1-diphenyl-1-(phenylmethyl)phosphoramine

Endpoint summary

Additional information

Three in vitro tests (bacterial reverse mutation test, mammalian cell gene mutation test and mammalian chromosome abberation test) and one in vivo genotoxicity test (mammalian bone marrow micronucleus test) are available. The studies were performed according to standard OECD guidelines and in compliance with GLP. None of the tests showed evidence of genotoxicity.

Bacterial reverse mutation:

In a reverse gene mutation assay in bacteria, performed according to the OECD No.471 guideline and in compliance with GLP, GM102E diluted in DMSO was tested in S. typhimuriumT A 1535, TA 1537, TA 1538, TA 98 and TA 100 in the presence and the absence of mammalian metabolic activation (S9). Four known mutagens (Sodium azide; 2-Aminoanthracene; 2-nitrofluorene and 9-aminoacridine), dissolved in dimethylsulfoxide (except for sodium azide which was dissolved in sterile distilled water), were used to check the sensitivity of the test system.

The results of the two main tests showed that toxicity was observed in the presence of S9-mix in strains TA 1535 and TA 1538 at the top dose tested of 3.3 mg/plate. In the absence of S9-mix, toxicity was noted from 333 µg/plate in strain TA 1535, from 1 mg/plate in strain TA 1538 and at 3.3 mg/plate in strains 1537, TA 98 and TA100. The number of revertants for the vehicle and positive controls met the acceptance criteria. The study was therefore considered to be valid. The test item did not induce any noteworthy or biologically relevant increase in the number of revertants, in any of the other tested strains.  

Under the test conditions, GM102E did not show any mutagenic activity in the bacterial reverse mutation test using Salmonella typhimurium.

 

In vitro mammalian cell gene mutation:

In an in vitro mammalian cell mutation assay, performed according to the OECD No.476 and in compliance with the GLP, GM102E dissolved in water for injections was tested in the L5178Y Tk +/- mouse lymphoma cell line in the presence and the absence of mammalian metabolic activation (S9 mix).

The vehicle controls had acceptable mutant frequency values that were within the normal range for the L5178Y cell line at the TK +/- locus. The positive control items induced marked increases in the mutant frequency indicating the satisfactory performance of the test and of the activity of the metabolising system.

Under the experimental conditions of this study, the GM102E did not show any mutagenic activity in the mouse lymphoma assay, up to 2 mM in the presence of a rat metabolizing system (3-hour treatment) or up to 75 μg/mL (3-hour treatment) or 50 μg/mL (24-hour treatment) in the absence of a rat metabolizing system. In conclusion, the test item was considered not to be mutagenic to L5178Y cells under the conditions of the test.

 

In vitro mammalian chromosome aberration:

The potential of the test item, GM102E, to induce chromosome aberrations in cultured human lymphocytes was investigated in an in vitro mammalian chromosome aberration test, performed according to the OECD No.473 and in compliance with the GLP.

The frequencies of cells with structural chromosome aberrations for the vehicle and positive controls were as specified in acceptance criteria. The study was therefore considered to be valid.

Under the experimental conditions of this study, the GM102Edid not induce chromosome aberrations in cultured human lymphocytes, either in the presence or absence of a rat liver metabolizing system. In conclusion, the test item was considered not to be mutagenic to human lymphocytes under the conditions of the test.

 

In vivo mammalian micronucleus test:

A bone marrow micronucleus assay was conducted in Sprague-Dawley rats according to the micronucleus test in order to assess the in vivo genotoxicity of GM102E. The study was conducted to a standard OECD guideline and in compliance with good laboratory practices. Animals (15 males and females per test substance dose and negative control) were treated by gavage with the test substance at 50 mg/kg bw in deionized water in a single administration. The positive control, Mitomycin C, was administered intraperitoneally to 5 males and 5 female.

Bone marrow cells were harvested at 18-hour, 42 -hour and 66 -hour post-treatment. There is no significant difference between the micronucleus frequency in the treated group in comparison with the control group at any sampling time. The group of animals treated with the positive control, Mitomycin C, showed a statistically increased frequency of micronucleated cells in comparison with the control group. Under the conditions of the in vivo test, the registered substance showed no evidence of causing chromosome damage or bone marrow cell toxicity.

Justification for selection of genetic toxicity endpoint
Studies performed according to standard OECD guidelines and in compliance with GLP.

Short description of key information:
- Genetic toxicity in vitro: negative (bacterial reverse mutation test, mammalian cell gene mutation test and mammalian chromosome abberation test)
- Genetic toxicity in vivo: negative (mammalian bone marrow micronucleus test)

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

None of the available tests showed evidence of genotoxicity. GM102E is therefore considered to be non-genotoxic and does not require classification for genetic toxicity according to the classification criteria of EU Regulation 1272/2008 (CLP).