PT  - JOURNAL ARTICLE
AU  - Sally A Coulthard
AU  - Sarah McGarrity
AU  - Kalvin Sahota
AU  - Philip Berry
AU  - Christopher P.F. Redfern
TI  - Three Faces of Mercaptopurine Cytotoxicity in vitro: Methylation, Nucleotide Homeostasis and Deoxythioguanosine in DNA
AID  - 10.1124/dmd.118.081844
DP  - 2018 Jan 01
TA  - Drug Metabolism and Disposition
PG  - dmd.118.081844
4099  - http://dmd.aspetjournals.org/content/early/2018/06/08/dmd.118.081844.short
4100  - http://dmd.aspetjournals.org/content/early/2018/06/08/dmd.118.081844.full
AB  - Mercaptopurine (MP) is a cytotoxic thiopurine important for the treatment of cancer and autoimmune diseases. MP and other thiopurine drugs undergo extensive intracellular metabolism but the mechanisms of action are poorly characterised. In particular, it is unknown how different metabolites contribute to cytotoxicity and incorporation of thiopurine bases into DNA. The aim of this study was to ask whether cytotoxicity results from the incorporation of thioguanosine nucleotides into DNA, an alternative thiopurine metabolite, or a combination of factors. Therefore, we measured the cytotoxicity, metabolism, and incorporation of thioguanosine into DNA in response to MP or MP metabolites. Thiopurine metabolites varied in cytotoxicity, with methyl-thioinosine- mono-phosphate and thioguanosine-tri-phosphate the most toxic, and the methyl-thioguanosine nucleotides the least. We show, using liquid chromatography-tandem mass spectrometry, how different metabolites may perturb biochemical pathways, particularly disrupting guanosine nucleotide homeostasis which may contribute to the mechanism of action of thiopurines. Although there was no correlation between metabolite cytotoxicity and the levels of 6-methylthioinosine-mono-phosphate or thioguanosine incorporation into DNA as individual factors, a combined analysis suggested that these factors together had a major influence on cytotoxicity. This study emphasises the importance of enzymes of nucleotide homeostasis, methylation and demethylation in thiopurine effects. These results will facilitate the development of dynamic biochemical models of thiopurine biochemistry which will improve our understanding of mechanisms of action in relevant target tissues.