In silico and in vitro human metabolism of IOX2, a performance-enhancing doping agent

IOX2 is a potent inhibitor of prolyl hydroxylase 2, a key enzyme in the regulation of hypoxia-inducible factor (HIF) and oxygen homeostasis. As such, it can be used to enhance athletic performance and is currently banned by the World Anti-Doping Agency (WADA). Detection of metabolites is critical to demonstrate drug use in doping. However, there is currently little data on IOX2 human metabolism. Our aim was to identify relevant biomarkers of IOX2 use in humans. For this purpose, IOX2 was incubated with 10-donor-pooled human hepatocytes for 3 h, incubates were analyzed by liquid chromatography-high-resolution tandem mass spectrometry (LC-HRMS/MS), and LC-HRMS/MS data were screened with Compound Discoverer (Thermo Scientific) for a comprehensive identification of IOX2 metabolites. Additionally, IOX2 human metabolites were predicted with GLORYx open-access software (University of Hamburg, Germany) to assist in the LC-HRMS/MS analysis and data mining. Thirteen metabolites were identified, oxidation at the quinolinyl group, O-glucuronidation, and combinations being predominant biotransformations. The results were consistent with previous animal studies and a single case of oral microdose administration. We suggest hydroxyquinolinyl-IOX2 as major biomarker of IOX2 use in biological samples, glucuronide hydrolysis being critical to increase IOX2 and hydroxyquinolinyl-IOX2 detectability in urine.

 

Comments:

It seems like you're describing a study or research related to the identification of biomarkers for IOX2, focusing on its metabolism in human hepatocytes. The detection and identification of metabolites are crucial in proving the use of IOX2 in doping cases.

Your study utilized liquid chromatography-high-resolution tandem mass spectrometry (LC-HRMS/MS) to analyze the metabolites produced when IOX2 was incubated with human hepatocytes. This method allows for a comprehensive identification of IOX2 metabolites. Additionally, you employed Compound Discoverer software for data analysis and GLORYx software for predicting IOX2 metabolites, aiding in the LC-HRMS/MS analysis and data interpretation.

Your findings highlighted thirteen identified metabolites, with predominant biotransformations involving oxidation at the quinolinyl group and O-glucuronidation. These results align with previous animal studies and a single instance of oral microdose administration.

Your suggestion of hydroxyquinolinyl-IOX2 as a major biomarker for detecting IOX2 use in biological samples, with emphasis on glucuronide hydrolysis to enhance detectability in urine, is significant. This information could potentially aid in developing more effective methods for detecting IOX2 use in anti-doping testing.

Is there anything specific you're looking to discuss or explore further regarding this research?

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Cat.No.	Product Name	Information
S2919	IOX2	IOX2 (JICL38) is a potent inhibitor of HIF-1α prolyl hydroxylase-2 (PHD2) with IC50 of 21 nM in a cell-free assay, >100-fold selectivity over JMJD2A, JMJD2C, JMJD2E, JMJD3, or the 2OG oxygenase FIH.

Related Targets

HIF