Comparison of In Vitro and In Vivo Screening Models for Androgenic and Estrogenic Activities

Abstract

Identification of nuclear receptor-mediated endocrine activities is important in a variety of fields, ranging from pharmacological and clinical screening, to food and feed safety, toxicological monitoring, and risk assessment. Traditionally animal studies such as the Hershberger and Allen-Doisy tests are used for the assessment of androgenic and estrogenic potencies, respectively. To allow fast analysis of the activities of new chemicals, food additives, and pharmaceutical compounds, high-throughput screening strategies have been developed. Here, a panel of mainly steroidal compounds, screened in different in vitro assays, was compared with two human U2-OS cell line-based CALUX (Chemically Activated LUciferase eXpression) reporter gene assays for androgens (AR CALUX) and estrogens (ERalpha CALUX). Correlations found between the data of these two CALUX reporter gene assays and data obtained with other in vitro screening assays measuring receptor binding or reporter gene activation (CHO cell line-based) were good (correlation coefficients (r2) between 0.54 and 0.76; p < 0.0001). Good correlations were also found between the in vitro and in vivo data (correlation coefficient r2 = 0.46 for the AR CALUX vs. Hershberger assay and r2 = 0.87 for the ERalpha CALUX vs. Allen-Doisy assay). The variations in the results obtained with the reporter gene assays (CALUX vs. CHO cell line based) were relatively small, showing the robustness of these types of assays. Using hierarchical clustering, bioactivity relationships between compounds but also relationships between various bioassays were determined. The in vitro assays were found to be good predictors of in vivo androgenic or estrogenic activity of a range of compounds, allowing prescreen and/or possible reduction of animal studies.

High-Resolution Mass Spectrometric Identification and Quantification of Glucocorticoid Compounds in Various Wastewaters in The Netherlands

“…Human U2OS osteosarcoma cells, stably transfected with a GR-controlled luciferase reporter gene construct were cultured as described previously (27). GR CALUX bioassays were performed as described by Van der Linden et al (15).…”

In-Vitro Methods of Screening of Anti-Cancer Agent

“…[4] Mostly murine host systems are used for experimental tumor therapy because of the availability of in-bred lines at relatively low costs. In-vitro models to define the mechanisms of action of a given compound Once a compound has demonstrated robust cytotoxic activity against a panel of human cancer cell lines and deserves further investigation in in-vivo models, it is important to clarify its mechanism of action and to identify its exact molecular targets.…”

The ChemScreen project to design a pragmatic alternative approach to predict reproductive toxicity of chemicals

“…These assays comprise a panel of highly specific human CALUX ® reporter gene assays, which includes a selection out of about 50 different assays. This panel is based on the human U2-OS cell line which are modified to express reporter genes for separate pathways [10,11]. The high selectivity is reached through the use of reporter genes with synthetic promoters consisting of minimal promoter elements coupled to multimerised, highly selective response elements driving the expression of the reporter gene luciferase.…”

“…This mechanism-based approach of toxicity testing was one of the cornerstones of the ChemScreen program. In 2005 we already developed a panel of mechanism-based CALUX assays to assess hormonal activity of compounds [10], a panel which has shown to be highly predictive for such activities in experimental animals [11,12]. Some of these mechanistic assays also formed a part of the promising battery of tests used in the ReProTect [7].…”