In vitro inhibition of cytidine triphosphate synthetase activity by cyclopentenyl cytosine in paediatric acute lymphocytic leukaemia

Cytidine triphosphate (CTP) synthetase is a key enzyme for the synthesis of cytosine (deoxy)ribonucleotides, catalysing the conversion of uridine trip hosphate (UTP) into cm, and has a high activity in several malignancies. In this preclinical study, the enzyme activity and mRNA expression of the enz yme and (deoxy)ribonucleotide concentrations were analysed in leukaemic cel ls of 57 children suffering from acute lymphocytic leukaemia (ALL). In addi tion, in vitro experiments were performed with the CTP synthetase inhibitor cyclopentenyl cytosine (CPEC). A significantly higher activity of CTP synt hetase (6.5 +/- 3.9 nmol CTP/mg/h) was detected in ALL cells than in lympho cytes of healthy controls (1.8 +/- 0.9 nmol CTP/mg/h, P < 0.001) that was i ndependent of white blood cell (WBC) count, blast percentage, age, gender o r type of ALL. The enzyme activity was not correlated with the CTP syntheta se mRNA expression. The activity of CTP synthetase in ALL cells compared wi th non-malignant CD34(+) bone marrow controls (5.6 +/- 2.4 nmol CTP/mg/h) w as not statistically different. In vitro treatment of ALL cells with CPEC i nduced a dose-dependent decrease of the CTP concentration. The lowest conce ntration of CPEC (0.63 mu M) induced a depletion of CTP of 41 +/- 20% and a depletion of dCTP of 27 +/- 2.1%. The degree of CTP depletion of ALL cells after treatment with CPEC was positively correlated with the activity of C TP synthetase. The inhibition of CTP synthetase in situ was confirmed by fl ux studies using radiolabelled uridine, From these results, it can be expec ted that CPEC has a cytostatic effect on lymphoblasts of children with ALL.