Abstract
OBJECTIVES: Aromatase is an enzyme that catalyzes the conversion of androgens to estrogens. While inhibition of aromatase is a useful approach for treating breast cancer, it may also have toxicological consequences due to its endocrine disrupting/modulating effect. In this study, sensitivity and performance of two in vitro assays -a cell free and a cell-based- for evaluating aromatase activity were investigated by testing known aromatase inhibitors and partial validation of the methods was performed. Advantages and disadvantages of these methods are also discussed. MATERIALS AND METHODS: Aromatase activity was evaluated via two in vitro models; direct measurement with a cell-free assay using a fluorescent substrate and recombinant human enzyme and indirect evaluation with a cell-based assay where cell proliferation was determined in estrogen receptor positive human breast cancer cells (MCF-7 BUS) in the absence of estrogen and the presence of testosterone. RESULTS: In the cell-free direct measurement assay, reference compounds ketoconazole and aminoglutethimide have been shown to inhibit the aromatase enzyme with half-maximal inhibitory concentration (IC(50)) values concordant with literature. In cell-based indirect measurement assay, only ketoconazole dose-dependently inhibited cell proliferation with 3.47 x 10(-7) M IC(50). Inter-assay and intra-assay reproducibility of both methods was found to be within acceptable deviation levels. CONCLUSION: Both methods can be successfully applied. However, to evaluate the potential aromatase activity of the novel compounds in vitro, it seems better to perform both the cell-based and the cell-free assays that allows low-moderate biotransformation and eliminate cytotoxicity potential, respectively.