Quantification of deoxythioguanosine in human DNA with LC-MS/MS, a marker for thiopurine therapy optimisation.

Autor: Carlsson B; Department of Clinical Pharmacology and Department of Biomedical and Clinical Sciences, Linköping University, Linköping, Sweden. bjorn.carlsson@regionostergotland.se., Karlsson L; Department of Clinical Pharmacology and Department of Biomedical and Clinical Sciences, Linköping University, Linköping, Sweden., Ärlemalm A; Department of Clinical Pharmacology and Department of Biomedical and Clinical Sciences, Linköping University, Linköping, Sweden.; Department of Biomedical and Clinical Sciences, Linköping University, Linköping, Sweden., Sund S; Department of Clinical Pharmacology and Department of Biomedical and Clinical Sciences, Linköping University, Linköping, Sweden., Appell ML; Department of Biomedical and Clinical Sciences, Linköping University, Linköping, Sweden.
Jazyk: angličtina
Zdroj: Analytical and bioanalytical chemistry [Anal Bioanal Chem] 2024 Dec; Vol. 416 (29), pp. 6711-6723. Date of Electronic Publication: 2024 Oct 13.
DOI: 10.1007/s00216-024-05581-6
Abstrakt: In the treatment of diseases such as acute childhood leukaemia (ALL) and inflammatory bowel disease (IBD), the thiopurines azathioprine, 6-mercaptopurine, and 6-thioguanine are used. Thiopurines are antimetabolites and immunomodulators used to maintain remission in patients. They are all prodrugs and must be converted into the competing antimetabolites thioguanosine triphosphate and deoxythioguanosine triphosphate for final incorporation into RNA or DNA. The current therapeutic drug monitoring (TDM) method measures the sum of the formed metabolites in the sample, after acidic hydrolysis at high temperature. In this work, the goal is to measure these drugs closer to their pharmacological endpoints, once incorporated into DNA. After extracting DNA from whole blood, followed by DNA hydrolysis, 2'-deoxythioguanosine (dTG) and the complementary natural nucleobase 2'-deoxycytidine (dC) were measured. Chromatographic separation on a HSS T3 column followed by mass spectrometric detection was performed in multi-reaction monitoring (MRM) mode on a Xevo TQ-XS with ESI in positive mode, within 5 min. The concentration range for dTG was 0.04-5 nmol/L, and for dC, 0.1-12.5 µmol/L. The lower limit of detection was determined to a concentration of 0.003 nmol/L for dTG and 0.019 µmol/L for dC. The intra- and inter-assay imprecision for the quality controls ranged between 3.0 and 5.1% and between 8.4 and 10.9%, respectively. Sample stability for up to 4 years is shown. In summary, a sensitive method to quantify the thiopurines incorporated into DNA as dTG has been developed and will be used in further clinical studies for a better understanding of the mode of action of the thiopurines and the use of this method in TDM.
Competing Interests: Declarations. Ethics approval: The study was approved by the Ethics Commission of Linköping University (diary number EPN 2018/47-31) and was conducted in accordance with the Declaration of Helsinki. All subjects who provided blood samples signed informed consent. Conflict of interest: The authors declare no competing interests.
(© 2024. The Author(s).)
Databáze: MEDLINE
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