Summary of Study ST002297

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR001471. The data can be accessed directly via it's Project DOI: 10.21228/M8KH70 This work is supported by NIH grant, U2C- DK119886.

See: https://www.metabolomicsworkbench.org/about/howtocite.php

This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.

Study ID	ST002297
Study Title	Comprehensive biotransformation analysis of phenylalanine-tyrosine metabolism reveals alternative routes of metabolite clearance in nitisinone-treated alkaptonuria (Urine metabolomic analysis)
Study Type	Urine metabolomic analysis (study 2 of 2)
Study Summary	Background: Metabolomic analyses in alkaptonuria (AKU) have recently revealed alternative pathways in phenylalanine-tyrosine (phe-tyr) metabolism from biotransformation of homo-gentisic acid (HGA), the active molecule in this disease. The aim of this research was to study the phe-tyr metabolic pathway and whether the metabolites upstream of HGA, increased in nitisinone-treated patients, also undergo phase 1 and 2 biotransformation reactions. Methods: Metabolomic analyses were performed on serum and urine from patients partaking in the SONIA 2 phase 3 international randomised-controlled trial of nitisinone in AKU (EudraCT no. 2013-001633-41). Serum and urine samples were taken from the same patients at baseline (pre-nitisinone) then at 24 and 48 months on nitisinone treatment (patients N = 47 serum; 53 urine) or no treatment (patients N = 45 serum; 50 urine). Targeted feature extraction was per-formed to specifically mine data for the entire complement of theoretically predicted phase 1 and 2 biotransformation products derived from phenylalanine, tyrosine, 4-hydroxyphenylpyruvic acid and 4-hydroxyphenyllactic acid, in addition to phenylalanine-derived metabolites with known increases in phenylketonuria. Results: In total, we ob-served 13 phase 1 and 2 biotransformation products from phenylalanine through to HGA. Each of these products were observed in urine and two were detected in serum. The derivatives of the metabolites upstream of HGA were markedly increased in urine of nitisinone-treated patients (fold change 1.2-16.2) and increases in 12 of these compounds were directly proportional to the degree of nitisinone-induced hypertyrosinaemia (correlation coefficient with serum tyrosine = 0.2-0.7). Increases in the urinary phenylalanine metabolites were also observed across consecutive visits in the treated group. Conclusions: Nitisinone treatment results in marked increases in a wider network of phe-tyr metabolites than shown before. This network comprises alternative biotransformation products from the major metabolites of this pathway, produced by reactions including hydration (phase 1) and bioconjugation (phase 2) of acetyl, methyl, acetylcysteine, glucuronide, glycine and sulfate groups. We propose that these alternative routes of phe-tyr metabolism, predominantly in urine, minimise tyrosinaemia as well as phenylalanaemia.
Institute	University of Liverpool Institute of Life Course & Medical Sciences
Department	Department of Musculoskeletal & Ageing Science
Last Name	Brendan
First Name	Norman
Address	William Henry Duncan Building, 6 West Derby Street, Liverpool, UK. L7 8TX
Email	bnorman@liverpool.ac.uk
Phone	+447809606497
Submit Date	2022-09-28
Num Groups	2
Total Subjects	103
Num Males	65
Num Females	38
Raw Data Available	Yes
Raw Data File Type(s)	d
Analysis Type Detail	LC-MS
Release Date	2022-10-14
Release Version	1

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Sample Preparation:

Sampleprep ID:	SP002389
Sampleprep Summary:	Prior to analysis, urine samples were thawed at room temperature before vortexing and centrifugation at 1500 ×g for 5 minutes. 150 µL of each urine sample was aliquoted into a 1 mL 96-well plate (Waters Corporation, Wilmslow, UK) and diluted with 450 µL of deionised water. Samples were mixed on a plate shaker (MTS 2/4m IKA) at 600 rpm for 10 min and sub-aliquoted into multiple replicate 96-well plates before storage at -80 °C ready for analysis. Prior to analysis, sample plates were thawed then agitated on a plate shaker (MTS 2/4m IKA) at 600 rpm for 10 min.

Sampleprep ID:

SP002389

Sampleprep Summary:

Prior to analysis, urine samples were thawed at room temperature before vortexing and centrifugation at 1500 ×g for 5 minutes. 150 µL of each urine sample was aliquoted into a 1 mL 96-well plate (Waters Corporation, Wilmslow, UK) and diluted with 450 µL of deionised water. Samples were mixed on a plate shaker (MTS 2/4m IKA) at 600 rpm for 10 min and sub-aliquoted into multiple replicate 96-well plates before storage at -80 °C ready for analysis. Prior to analysis, sample plates were thawed then agitated on a plate shaker (MTS 2/4m IKA) at 600 rpm for 10 min.