Abstract 4365: A Bhas 42 cell transformation assay sensitive to Ames-negative and Ames-discordant carcinogens: Its performance for the prediction of chemical carcinogenicity

Abstract

Abstract Introduction: A cell transformation assay (CTA) can detect the carcinogenicity of chemicals, but is classified into a category different from a genotoxicity assay which has been used for the prediction of chemical carcinogenicity. CTA detects non-genotoxic carcinogens as well as genotoxic ones. Most of non-genotoxic carcinogens are considered to be tumor-promoters and many of them have been detected as promoters in a two-stage BALB/c 3T3 CTA, which mimics a in vivo two-stage carcinogenesis test. The Bhas 42 cells were established from the BALB/c 3T3 cells through transfection of v-Ha-ras gene and regarded as initiated cells in the two-stage carcinogenesis theory. Using the Bhas 42 cells, a short-term CTA was developed. Bhas 42 CTA is superior to conventional CTAs in cost and labor performance. It consists of an initiation assay and a promotion assay to detect initiating activity and promoting activity, respectively. We applied this short-term CTA to 92 chemicals to characterize the assay and evaluate its performance for the prediction of chemical carcinogenicity. Methods: In the initiation assay, the Bhas 42 cells were seeded at a density of 2000 cells/mL onto 6-well microplates (Day 0), treated with test chemicals from Day 1, when the cells were sparse, to Day 4, and thereafter cultured in normal medium until Day 21. In the promotion assay, the cells were seeded at a density of 7000 cells/mL, treated with chemicals from Day 4, when the cells were sub-confluent, to Day 14, and maintained in normal medium until Day 21. The cultures were fixed and stained with Giemsa's solution. The transformed foci were judged on the basis of morphological characteristics. A statistical analysis for the number of transformed foci per well was performed by the multiple comparison using the Dunnett method. Results and Discussion: Our results of Bhas 42 CTA were compared with existing genotoxicity data. Bhas 42 CTA could detect Ames-negative and Ames-discordant chemicals, and the promotion assay detected most of them, confirming that the Bhas 42 cells act as initiated cells in the CTA. The concordance, sensitivity, specificity, positive predictivity, negative predictivity, false positive and false negative were calculated from the assay results for known carcinogens and non-carcinogens. Among these performance characteristics, the specificity and positive predictivity were high and exceeded 80%. The other performance characteristics were equivalent to those of genotoxicity tests. From these results, we considered that the accuracy of prediction for chemical carcinogenicity would be improved by introducing Bhas 42 CTA into the battery of in vitro assays. Acknowledgement: This work was supported by the New Energy and Industrial Technology Development Organization/the Ministry of Economy, Trade and Industry of Japan. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4365.