Characterisation and validation of an in vitro transactivation assay based on the 22Rv1/MMTV_GR-KO cell line to detect human androgen receptor agonists and antagonists

Park, Yooheon; Jung, Da Woon; Milcamps, Anne; Takeyoshi, Masahiro; Jacobs, Miriam N.; Houck, Keith A.; Ono, Atsushi; Bovee, Toine F.H.; Browne, Patience; Delrue, Nathalie; Kang, Yun Sook; Lee, Hee-Seok

doi:10.1016/j.fct.2021.112206

Cited by 4 publications

(2 citation statements)

References 21 publications

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“…Successfully validated in vitro methods with a clear regulatory purpose may become test guidelines (TG) at OECD. The AR-CALUX ® method was recently added to the existing TG458, joining the AR-EcoScreen™ already present in the TG458 and simultaneously including the 22Rv1/MMTV GR-KO TA which was also recently validated ( Park et al , 2021 ). This TG, now comprising 3 mechanistically similar ARTA methods, was recently adopted and published ( OECD, 2020b ).…”

Section: Discussionmentioning

confidence: 99%

“…This article describes the conduct and results of an international ring trial of an AR-CALUX ® transactivation assay that has recently been included in the updated TG458 of Androgen Receptor TransActivation assays (ARTA) (Androgen Receptor TransActivation Assays for Detection of Androgenic Agonist and Antagonist Activity of Chemicals using Stably Transfected Cell Lines; OECD, 2020b ). The original Test Guideline from 2016 described the AR-EcoScreen™ assay and the 2020 update include a further 2 newly validated assays: the ARTA described here and another mechanistically similar assay (the 22Rv1/MMTV-GR-KO assay) ( Milcamps et al , 2020 ; Park et al , 2021 ). Both ARTA assays contain reporter genes, which are activated by the Androgen Receptor (AR) when bound to a ligand, ie, a chemical with androgen receptor affinity.…”

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confidence: 99%

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Reliability of the AR-CALUX®In Vitro Method Used to Detect Chemicals with (Anti)Androgen Activity: Results of an International Ring Trial

Milcamps

Liška

Langezaal

et al. 2021

Toxicological Sciences

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The AR-CALUX® in vitro method is a reporter gene-based transactivation method where endocrine active chemicals with androgenic or anti-androgenic potential can be detected. Its primary purpose is for screening chemicals for further prioritisation and providing mechanistic (endocrine mode of action) information, as defined by the OECD Conceptual Framework for the testing and assessment of endocrine disrupting chemicals. This paper describes the conduct and results of an international ring trial with three EU-NETVAL laboratories and the test method developer. It was organised by EURL ECVAM to validate the method by testing 46 chemicals. A very good reproducibility within and between laboratories was concluded (94.7 – 100% and 100% concordance of classification) with low within and between laboratory variability (less than 2.5% CV on EC50 values). Moreover, the variability is within the range of other validated, mechanistically similar methods. In comparison to the AR-reference list compiled by ICCVAM, an almost 100% concordance of classifications was obtained. This method allows the detection of agonist and antagonist properties of a chemical. A specificity control test was developed during the validation study and added to the antagonist assay rendering the assay more specific. A comparison is made with the mechanistically similar methods AR-EcoScreen™ and 22Rv1/MMTV GR-KO TA. The AR-CALUX® method was approved for inclusion in the recently updated OECD test guideline TG458 which incorporates all three methods.

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Reliability of the AR-CALUX®In Vitro Method Used to Detect Chemicals with (Anti)Androgen Activity: Results of an International Ring Trial

Milcamps

Liška

Langezaal

et al. 2021

Toxicological Sciences

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Avoiding a reproducibility crisis in regulatory toxicology—on the fundamental role of ring trials

Jacobs,

Hoffmann,

Hollnagel

et al. 2024

Arch Toxicol

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The ongoing transition from chemical hazard and risk assessment based on animal studies to assessment relying mostly on non-animal data, requires a multitude of novel experimental methods, and this means that guidance on the validation and standardisation of test methods intended for international applicability and acceptance, needs to be updated. These so-called new approach methodologies (NAMs) must be applicable to the chemical regulatory domain and provide reliable data which are relevant to hazard and risk assessment. Confidence in and use of NAMs will depend on their reliability and relevance, and both are thoroughly assessed by validation. Validation is, however, a time- and resource-demanding process. As updates on validation guidance are conducted, the valuable components must be kept: Reliable data are and will remain fundamental. In 2016, the scientific community was made aware of the general crisis in scientific reproducibility—validated methods must not fall into this. In this commentary, we emphasize the central importance of ring trials in the validation of experimental methods. Ring trials are sometimes considered to be a major hold-up with little value added to the validation. Here, we clarify that ring trials are indispensable to demonstrate the robustness and reproducibility of a new method. Further, that methods do fail in method transfer and ring trials due to different stumbling blocks, but these provide learnings to ensure the robustness of new methods. At the same time, we identify what it would take to perform ring trials more efficiently, and how ring trials fit into the much-needed update to the guidance on the validation of NAMs.

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Azole pesticide products and their hepatic metabolites cause endocrine disrupting potential by suppressing the homo-dimerization of human estrogen receptor alpha

Jung

Jeong

Lee

2023

Environmental Pollution

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Characterisation and validation of an in vitro transactivation assay based on the 22Rv1/MMTV_GR-KO cell line to detect human androgen receptor agonists and antagonists

Cited by 4 publications

References 21 publications

Reliability of the AR-CALUX®In Vitro Method Used to Detect Chemicals with (Anti)Androgen Activity: Results of an International Ring Trial

Reliability of the AR-CALUX®In Vitro Method Used to Detect Chemicals with (Anti)Androgen Activity: Results of an International Ring Trial

Avoiding a reproducibility crisis in regulatory toxicology—on the fundamental role of ring trials

Azole pesticide products and their hepatic metabolites cause endocrine disrupting potential by suppressing the homo-dimerization of human estrogen receptor alpha

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