High-throughput screening (HTPS) assays to detect inhibitors of thyroperoxidase (TPO), the enzymatic catalyst for thyroid hormone (TH) synthesis, are not currently available. Herein, we describe the development of a HTPS TPO inhibition assay. Rat thyroid microsomes and a fluorescent peroxidase substrate, Amplex UltraRed (AUR), were employed in an end-point assay for comparison to the existing kinetic guaiacol (GUA) oxidation assay. Following optimization of assay metrics, including Z′, dynamic range, and activity, using methimazole (MMI), the assay was tested with a 21-chemical training set. The potency of MMI-induced TPO inhibition was greater with AUR compared to GUA. The dynamic range and Z′ score with MMI were as follows: 127-fold and 0.62 for the GUA assay, 18-fold and 0.86 for the 96-well AUR assay, and 11.5-fold and 0.93 for the 384-well AUR assay. The 384-well AUR assay drastically reduced animal use, requiring one-tenth of the rat thyroid microsomal protein needed for the GUA 96-well format assay. Fourteen chemicals inhibited TPO, with a relative potency ranking of MMI > ethylene thiourea > 6-propylthiouracil > 2,2′,4,4′-tetrahydroxy-benzophenone > 2-mercaptobenzothiazole > 3-amino-1,2,4-triazole > genistein > 4-propoxyphenol > sulfamethazine > daidzein > 4-nonylphenol > triclosan > iopanoic acid > resorcinol. These data demonstrate the capacity of this assay to detect diverse TPO inhibitors. Seven chemicals acted as negatives: 2-hydroxy-4-methoxybenzophenone, dibutylphthalate, diethylhexylphthalate, diethylphthalate, 3,5-dimethylpyrazole-1-methanol, methyl 2-methyl-benzoate, and sodium perchlorate. This assay could be used to screen large numbers of chemicals as an integral component of a tiered TH-disruptor screening approach.

Katie B. Paul

Joan M. Hedge

Daniel M. Rotroff

Michael W. Hornung

Kevin M. Crofton

Steven O. Simmons

Chemical Research in Toxicology

毒理学化学研究发表关于毒理学广泛主题的文章、快速报告、化学概况、评论、观点、致编辑的信和 TOXWATCH，这些主题为化学和分子理解以及基于结构预测生物学结果的能力提供信息和流程。该杂志报道的活动的首要目标是提供促进人类安全和生态系统保护的智能解决方案所需的知识和创新方法。该杂志强调对现象学观察的毒性机制的洞察力。它坚持严格的化学、物理和数学标准来表征和应用现代技术。

Chemical Research in Toxicology is an academic journal focused on the field of toxicology, publishing research articles, rapid reports, chemical profiles, reviews, viewpoints, letters to the editor, and TOXWATCH on a wide range of topics in toxicology. The journal aims to provide the knowledge and innovative approaches necessary for promoting human safety and protecting ecosystems, emphasizing a deep understanding of toxic mechanisms. It adheres to rigorous standards in chemistry, physics, and mathematics to characterize and apply modern technologies.

《Chemical Research in Toxicology》是一本专注于毒理学领域的学术期刊，发表关于毒理学广泛主题的研究文章、快速报告、化学概况、评论、观点、致编辑的信件以及 TOXWATCH。该期刊旨在提供促进人类安全和生态系统保护所需的知识和创新方法，强调对毒性机制的深入理解。它坚持严格的化学、物理和数学标准，以表征和应用现代技术。

Development of a Thyroperoxidase Inhibition Assay for High-Throughput Screening

The development of a HTPS TPO inhibition assay that was used to screen large numbers of chemicals as an integral component of a tiered TH-disruptor screening approach and demonstrated the capacity of this assay to detect diverse TPO inhibitors.

Category	Quartile
医学	3
医学, 毒理学	2
医学, 药物化学	3
医学, 化学综合	3

Development of a Thyroperoxidase Inhibition Assay for High-Throughput Screening

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