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Quantification of Plasma and Urine Thymidine and 2'-Deoxyuridine by LC-MS/MS for the Pharmacodynamic Evaluation of Erythrocyte Encapsulated Thymidine Phosphorylase in Patients with Mitochondrial Neurogastrointestinal Encephalomyopathy.

Kipper, K; Hecht, M; Antunes, NJ; Fairbanks, LD; Levene, M; Kalkan Uçar, S; Schaefer, A; Blakely, EL; Bax, BE (2020) Quantification of Plasma and Urine Thymidine and 2'-Deoxyuridine by LC-MS/MS for the Pharmacodynamic Evaluation of Erythrocyte Encapsulated Thymidine Phosphorylase in Patients with Mitochondrial Neurogastrointestinal Encephalomyopathy. J Clin Med, 9 (3). p. 788. ISSN 2077-0383 https://doi.org/10.3390/jcm9030788
SGUL Authors: Bax, Bridget Elizabeth

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Abstract

Mitochondrial neurogastrointestinal encephalomyopathy (MNGIE) is an ultra-rare disorder caused by mutations in TYMP, leading to a deficiency in thymidine phosphorylase and a subsequent systemic accumulation of thymidine and 2'-deoxyuridine. Erythrocyte-encapsulated thymidine phosphorylase (EE-TP) is under clinical development as an enzyme replacement therapy for MNGIE. Bioanalytical methods were developed according to regulatory guidelines for the quantification of thymidine and 2'-deoxyuridine in plasma and urine using liquid chromatography-tandem mass spectrometry (LC-MS/MS) for supporting the pharmacodynamic evaluation of EE-TP. Samples were deproteinized with 5% perchloric acid (v/v) and the supernatants analyzed using a Hypercarb column (30 × 2.1 mm, 3 µm), with mobile phases of 0.1% formic acid in methanol and 0.1% formic acid in deionized water. Detection was conducted using an ion-spray interface running in positive mode. Isotopically labelled thymidine and 2'-deoxyuridine were used as internal standards. Calibration curves for both metabolites showed linearity (r > 0.99) in the concentration ranges of 10-10,000 ng/mL for plasma, and 1-50 µg/mL for urine, with method analytical performances within the acceptable criteria for quality control samples. The plasma method was successfully applied to the diagnosis of two patients with MNGIE and the quantification of plasma metabolites in three patients treated with EE-TP.

Item Type: Article
Additional Information: © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
Keywords: 2’-deoxyuridine, EE-TP, LC–MS/MS, erythrocyte encapsulated thymidine phosphorylase, method validation, mitochondrial neurogastrointestinal encephalomyopathy, thymidine
SGUL Research Institute / Research Centre: Academic Structure > Molecular and Clinical Sciences Research Institute (MCS)
Journal or Publication Title: J Clin Med
ISSN: 2077-0383
Language: eng
Dates:
DateEvent
13 March 2020Published
11 March 2020Accepted
Publisher License: Creative Commons: Attribution 4.0
Projects:
Project IDFunderFunder ID
MR/K025406/1Medical Research Councilhttp://dx.doi.org/10.13039/501100000265
2016/22506-1São Paulo Research FoundationUNSPECIFIED
PubMed ID: 32183169
Go to PubMed abstract
URI: http://openaccess.sgul.ac.uk/id/eprint/111807
Publisher's version: https://doi.org/10.3390/jcm9030788

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