Murine oral carcinogenesis model

To establish the 4-nitroquinoline 1-oxide (4-NQO) model, female, immunocompetent C57BL/6J mice aged 8 weeks were purchased from Jackson Laboratories. Protocols for animal experiments were approved by the Institutional Animal Care and Use Committee (IACUC) under the reference #18042580. To induce the development of oral carcinomas, mice were administered the carcinogen 4-NQO (Tokyo Chemical Industry CO., LTD.) in drinking water once a week. Therefore, 4-NQO was dissolved in propylene glycol (Sigma-Aldrich) as stock solution (4 mg/mL), which was immediately diluted to a concentration of 0.1 mg/mL in the drinking water. 4-NQO was protected from light at all times. Mice were treated with 4-NQO for 16 weeks followed by the provision of normal drinking water.