The in vitro BALB/c 3T3 cell transformation assay to screen the anti-transforming activity of chemical compounds

The in vitro transformation test on BALB/c 3T3 cells resembles tumor onset and progression in vivo and represents a useful and flexible model to screen the carcinogenic potential of both genotoxic and non genotoxic chemicals. It shows good predictability, high sensitivity and specificity and is time-saving and less expensive compared to animal studies (Sakai et al, 2007). Several improvements to the basic protocol were proposed aiming at increasing the cell transformation frequency, shortening the length of the assay and reducing the cytotoxic effects associated to chemical exposure (Matthews et al, 1993). The results obtained in our laboratory confirm the suitability of the improved protocol to discriminate carcinogenic compounds and support the use of BALB/c 3T3 cell transformation assay as a possible alternative to predict carcinogenic risk to humans (Mascolo et al, 2010). In order to identify compounds which could be able to interfere with the tumor development we set up an anti-transformation assay (Vaccari et al, 1999). Several molecules belonging to different chemical classes, as estrogen antagonists, polyphenolic compounds and retinoids, has been tested as promising tumor chemopreventives. The treatment has been performed according to different exposure schedules to better define the potential protective effects and the steps of the multistage carcinogenesis process they affect. The chemicals has been administered to BALB/c-3T3 cells :1) for 48 hours, before cell plating (pre-treatment); 2) for 48 hours, concurrently with the carcinogen (simultaneous treatment); 3) throughout the duration of the experiment, after the exposure to the carcinogen, at each medium change (chronic treatment). A cytoxicity assay was performed to estimate the corresponding cloning efficiency. The cell treatment with the carcinogen determines the induction of morphologically aberrant foci; the outcome of BALB/c 3T3 cell exposure to the potential anti-transforming chemicals is the reduction of the number of transformed foci/plate as well as the decrease of the transformation frequency. Results give evidence that the proposed in vitro test could be useful for the preclinical screening of promising chemopreventive agents.