Advantages and Disadvantages of Two In Vitro Assays in Evaluating Aromatase Activity: “A Cell-Based and a Cell-Free Assay”
Makaleler > Tam Makale · 2021
Yazarlar: ELİF İNCE ERGÜÇ, Senem Ozcan-Sezer, Hande GÜRER ORHAN
Turkish Journal of Pharmaceutical Sciences, SCI Expanded İndekslerine Giren Dergi, Uluslararası, Scopus, 2021, cilt 19, ss 626-629
DOI: 10.4274/tjps.galenos.2021.85530
Özet
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.