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Dose–response relationship of temozolomide, determined by the Pig-a, comet, and micronucleus assay

  • Genotoxicity and Carcinogenicity
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Abstract

Temozolomide (TMZ), a monofunctional alkylating agent, was selected as a model compound to determine its quantitative genotoxic dose–response relationship in different tissues (blood, liver, and jejunum) and endpoints [Pig-a-, comet-, and micronucleus assay (MNT)] in male rats. TMZ was administered p.o. over 5 consecutive days (day 1–5), followed by a treatment-free period of 50 days (day 6–56) and a final administration prior to necropsy (day 57–59). TMZ showed a dose-dependent increase in DNA damage in all interrogated endpoints. A statistically significant increase in Pig-a mutant phenotypes was observed on day 44 starting at 7.5 mg/kg/day for mutant reticulocytes (for RETCD59−) and at 3.75 mg/kg/day for mutant red blood cells (RBCCD59−), respectively. In addition, a statistically significant increase in cytogenetic damage, as measured by micronucleated reticulocytes, was observed starting at 3.75 mg/kg/day on day 3 and 1.5 mg/kg/day on day 59. DNA strand breaks, as detected by the comet assay, showed a dose-dependent and statistically significant increase in liver, blood, and jejunum starting at doses of 3.75, 3.75, and 7.5 mg/kg/day, respectively. The dose–response relationships of the Pig-a, MNT, and comet data were analyzed for possible points of departure (PoD) using the benchmark-dose (BMD) software PROAST with different critical effect sizes (CES) (BMD0.1, BMD0.5, BMD1, and BMD1SD). Overall, PoD values show a high concordance between different tissues and endpoints, underlining the suitability of this experimental design to explore quantitative dose–response relationships in a variety of different tissues and endpoints, while minimizing animal use.

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Acknowledgements

We would like to thank Elmar Gocke for his valuable input in the design and set-up of the experiment and Leilei Tang for her contribution in performing the experiments.

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Authors and Affiliations

  1. Pharmaceutical Sciences, Roche Innovation Center Basel, F. Hoffmann La Roche Ltd., Grenzacherstrasse 124, 4070, Basel, Switzerland

    M. Guérard, G. Duran-Pacheco, J. Funk & A. Zeller

  2. Institute of Life Science, Swansea University Medical School, Singleton Park, Swansea, SA3 5DE, UK

    G. Johnson

  3. Litron Laboratories, 3500 Winton Place, Rochester, NY, USA

    S. Dertinger

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  1. M. Guérard
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  2. G. Johnson
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  3. S. Dertinger
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  4. G. Duran-Pacheco
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  5. J. Funk
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  6. A. Zeller
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Correspondence to M. Guérard.

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Guérard, M., Johnson, G., Dertinger, S. et al. Dose–response relationship of temozolomide, determined by the Pig-a, comet, and micronucleus assay. Arch Toxicol 91, 2443–2453 (2017). https://doi.org/10.1007/s00204-016-1923-4

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  • DOI: https://doi.org/10.1007/s00204-016-1923-4

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