Interlaboratory prevalidation of a new in vitro transcriptional activation assay for the screening of (anti-)androgenic activity of chemicals using the UALH-hAR cell line

We report an interlaboratory evaluation of a recently developed androgen receptor (AR) transactivation assay using the UALH-hAR reporter cell line that stably expresses the luciferase gene under the transcriptional control of androgen receptor elements (AREs) with no glucocorticoid receptor (GR) crosstalk. Herein, a two-step prev-alidation study involving three laboratories was conducted to assess performance criteria of the method such as transferability as well as robustness, sensitivity, and specificity. The first step consisted in the validation of the transfer of the cell line to participant laboratories through the testing of three reference chemicals: the AR agonist dihydrotestosterone, the AR antagonist hydroxyflutamide and the glucocorticoid dexamethasone. Secondly, a blinded study was conducted by screening a selection of ten chemicals, including four AR agonists, five AR antagonists, and one non-active chemical. All test compounds yielded the same activity profiles in all labora-tories. The logEC50 (agonist assay) or logIC50 (antagonist assay) were in the same range, with intra-laboratory coefficients of variation (CVs) of 0.1-3.4% and interlaboratory CVs of 1-4%, indicating very good within-and between-laboratory reproducibility. Our results were consistent with literature and regulatory data (OECD TG458). Overall, this interlaboratory study demonstrated that the UALH-hAR assay is transferable, produces reliable, accurate and specific (anti)androgenic activity of chemicals, and can be considered for further regu-latory validation.

Automated summary

In this study, a novel androgen receptor (AR) transactivation assay using the UALH-hAR reporter cell line was evaluated. A two-step prev-validation study involving three laboratories was conducted to assess the method's transferability, robustness, sensitivity, and specificity. The assay demonstrated good reproducibility and produced reliable, accurate, and specific (anti)androgenic activity of chemicals, making it suitable for regulatory validation.