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Quantification of the Uncertainties in Extrapolating From In Vitro Androgen Receptor Antagonism to In Vivo
Leon E Gray1, Johnathan R Furr2, Christy S Lambright1
1Reproductive and Developmental Toxicology Branch, PHITD, Office of Research and Development, U.S. Environmental Protection Agency (U.S. EPA), Research Triangle Park, North Carolina 27711.
Abstract:
Multiple molecular initiating events exist that disrupt male sexual differentiation in utero including androgen receptor (AR) antagonism and inhibition of synthesis, and metabolism of fetal testosterone. Disruption of androgen signaling by AR antagonists in utero reduces anogenital distance (AGD) and induces malformations in F1 male rat offspring. We are developing a quantitative network of adverse outcome pathways that includes multiple molecular initiating events and key events linking anti-AR activities to permanent reproductive abnormalities. Here, our objective was to determine how accurately the EC50s for AR antagonism in vitro or ED50s for reduced tissue growth in the Hershberger assay (HA) (key events in the adverse outcome pathway) predict the ED50s for reduced AGD in male rats exposed in utero to AR antagonists. This effort included in-house data and published studies from the last 60 years on AR antagonism in vitro and in vivo effects in the HA and on AGD after in utero exposure. In total, more than 250 studies were selected and included in the analysis with data from about 60 potentially antiandrogenic chemicals. The ability to predict ED50s for key events and adverse developmental effects from the in vitro EC50s displays considerable uncertainty with R2 values for HA and AGD of < 6%. In contrast, there is considerably less uncertainty in extrapolating from the ED50s in the HA to the ED50s for AGD (R2 value of about 85%). In summary, the current results suggest that the key events measured in the HA can be extrapolated with reasonable certainty to predict the ED50s for the adverse in utero effects of antiandrogenic chemicals on male rat offspring.
Insights
The Hershberger assay (HA) reliably predicts in utero antiandrogen effects on male rat offspring, unlike in vitro androgen receptor (AR) antagonism tests. This finding aids in assessing reproductive risks from endocrine-disrupting chemicals.
Area of Science:
- Reproductive toxicology
- Endocrine disruption
- Adverse Outcome Pathways (AOPs)
Background:
- Male sexual differentiation is vulnerable to in utero endocrine disruptors, particularly antiandrogens.
- Androgen receptor (AR) antagonism and altered testosterone metabolism disrupt fetal development, leading to reduced anogenital distance (AGD) and malformations.
- Adverse Outcome Pathways (AOPs) provide a framework for understanding chemical-induced toxicity from molecular initiating events to adverse outcomes.
Purpose of the Study:
- To evaluate the predictive accuracy of in vitro AR antagonism (EC50s) and Hershberger assay (HA) in vivo data (ED50s) for predicting reduced AGD in male rats exposed in utero to AR antagonists.
- To quantify the uncertainty in extrapolating key event ED50s to developmental toxicity ED50s.
Main Methods:
- Compiled data from over 250 studies spanning 60 years, including in vitro AR antagonism, in vivo HA data, and in utero AGD effects for approximately 60 antiandrogenic chemicals.
- Analyzed the correlation (R2 values) between in vitro EC50s, HA ED50s, and AGD ED50s to assess predictive power.
Main Results:
- In vitro AR antagonism (EC50s) showed considerable uncertainty in predicting HA (R2 < 6%) and AGD (R2 < 6%) ED50s.
- The Hershberger assay (HA) ED50s demonstrated strong predictive power for AGD ED50s (R2 ≈ 85%) after in utero exposure to AR antagonists.
- Extrapolation from HA key events to in utero effects showed significantly less uncertainty compared to in vitro data.
Conclusions:
- The Hershberger assay serves as a reliable predictor of in utero antiandrogenic effects on male rat offspring's AGD.
- In vitro androgen receptor antagonism assays have limited predictive value for in utero developmental toxicity.
- The HA's key events are valuable for predicting adverse outcomes of antiandrogenic chemicals during male sexual differentiation.
