A novel biosensor for prediction of global hypomethylation by decitabine and azacytadine in leukaemic cells

May, J E; Allainguillaume, Joel; Page, K; Stratford-Smith, C; Aruomaren, A; Alloush, H; Salisbury, V; Smith, A; Ruddock, M; Reid, C; Lamont, J; Anderson, E

Abstract

Decitabine (DAC) and azacytidine (AZA) are cytidine ana- logues and hypomethylating agents used in the manage- ment of patients with myelodysplastic syndrome (MDS), where the main goal is to prevent transformation to acute myeloid leukaemia (AML), which has poor prognosis. Increasingly DAC and AZA are being considered for MDS/ AML patients who may be ineligible for more cytotoxic induction regimens. Trials have shown AZA to be clinically favourable, with DAC being used in cases of resistance to AZA. Cellular uptake of DAC and AZA is via the mem- brane transporter hENT1, and subsequent phosphoryla- tion to the hypomethylating metabolite decitabine triphos- phate (DAC-TP) leads to DNA incorporation, covalent binding to and inactivation of DNA methyltransferases (DNMTs). Subsequent cell replication cycles result in DNA hypomethylation and potential re-expression of silenced tu- mour suppressor genes. This study aimed to assess the fea- sibility of rapidly detecting DAC-TP to predict subsequent hypomethylation using a previously validated biosensor sensitive to cytosine arabinoside (ara-C) (1). Leukaemic cell lines (KG-1a, HL-60, K562) were exposed to physi- ologically-relevant doses of DAC/AZA to assess whether uptake and metabolism to DAC-TP during the initial post- dosing period would predict subsequent cytotoxicity and hypomethylation. Following daily dosing over a 72-hour period HL-60 cells were most susceptible to DAC-induced cytotoxicity. High-resolution melting analysis concurrently performed on extracted DNA following 96 hour incuba- tion indicated hypomethylation at these doses, concurring with biosensor results from the initial post-dosing period. Studies are on-going in our laboratory to directly con rm DAC-TP levels in AML patient blasts and compare with global hypomethylation to de ne the intracellular drug ki- netics and further dissect the role of parent and metabo- lites in DAC/AZA-mediated toxicity. Our goal is to predict global hypomethylation and ultimately patient response to these agents using this rapid and cheap biosensor, reducing unnecessary costly treatment for patients (~$80,000pa per patient) where resistance emerges.

• Journal Article Type: Article
• Acceptance Date: Jul 15, 2015
• Publication Date: Nov 6, 2015
• Deposit Date: Apr 25, 2017
• Journal: Mutagenesis
• Print ISSN: 0267-8357
• Electronic ISSN: 1464-3804
• Publisher: Oxford University Press (OUP)
• Peer Reviewed: Peer Reviewed
• Volume: 30
• Issue: 6
• Pages: 856
• DOI: https://doi.org/10.1093/mutage/gev074
• Keywords: novel biosensor, prediction, global hypomethylation, decitabine, azacytadine, leukaemic cells
• Public URL: https://uwe-repository.worktribe.com/output/803563
• Publisher URL: https://academic.oup.com/mutage
• Contract Date: Apr 25, 2017