Thyroid method 4c: Inhibition of THs sulfation using liquid chromatography

Topic: Endocrine disruption

Test Method Number:

TM2019-12 (EU)

Short Name of TM:

THSUL-LC

Year received:

2019

Responsible Organisation:

EURL ECVAM - European Union

General Comments:

Method Description

This method assesses the potential of test items to inhibit the metabolism and excretion of thyroid hormones (THs,T3 and T4).
Sulfation appears to be a primary step leading to the irreversible inactivation of TH and it is carried out by sulfotransferases (SULT); it increases the water solubility of TH, facilitating the excretion of sulfated TH via bile and urine. In humans, as sulfated TH can be hydrolysed to their precursor by the sulfatases present in tissues and gut microbiota, they are important as a reservoir for biological active iodothyronines.
This assay measures the inhibition of THs by liquid chromatography/mass spectrometry (LC/MS). The potential inhibition of the compounds towards sulfation of T3 and T4 is determined by measuring and comparing the thyroid hormones sulfate conjugates formation, after incubation with and without the test chemicals.

This method was selected with other methods to assess modulating effects on the thyroid system (OECD, 2014). Information...

This method was selected with other methods to assess modulating effects on the thyroid system (OECD, 2014). Information on these selected methods is available in TSAR as ‘Thyroid methods’.

Track Approval Status

Submission
Validation
Peer-review
Recommendation
Regulatory acceptance/Standards

Submission

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Date	Stage	Comment	Documents/Links
Jun 2017	Assessment finalised	Selection of the test method for the Thyroid Validation Study based on the OECD scoping document on 'in vitro and ex vivo methods for the identification of modulators of thyroid hormone signalling' and guidance from the Endocrine Disrupters Testing and Assessment Advisory Group considering scientific advances since the issuing of the OECD scoping document, human relevance, possibility for High Throughput screening and availability of method elements (such as reagents and detection method) to a wide range of future users.	OECD scoping document (OECD, 2014)OECD detailed review paper (OECD, 2012)OECD Endocrine Disrupters Testing and Assessment Advisory Group

Validation

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Date	Stage	Comment	Documents/Links
Sep 2019	Finalised	Preparatory work carried out by EURL ECVAM before the start of the validation study between November 2017 and September 2019. During this phase, intellectual property was addressed, legal agreements with method developer, test system supplier and EU-NETVAL laboratory were established and adequate training of the EU-NETVAL laboratory was ensured. An outline protocol describing the current status of the method, the quality controlled test system and reference and control items were made available to EU-NETVAL. OECD Good In Vitro Method Practices (GIVIMP, OECD, 2018) has been used as guidance.	OECD Good In Vitro Method Practices (GIVIMP, OECD, 2018)
Oct 2019	Finalised	Start of the EURL ECVAM coordinated validation study. EU-NETVAL facility 'Accelera Srl (Italy)' agreed to develop one method from available published methods and to assess the method.	EU-NETVAL
Feb 2021	Finalised	Final selection of the validation set chemicals to be used in the thyroid validation study PART 2. Selection was based on the advice provided by 15 experts in in vitro, in vivo and clinical thyroid disruption during an expert meeting in November 2019.
Jan 2022	Stopped	Completion of method definition and description prior to experimental assessment was stopped. The method would need further development.

Peer-review

Recommendation

Regulatory acceptance/Standards