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Resistance to 2-Chloro-2'-deoxyadenosine of the Human B-cell Leukemia Cell Line EHEB¹

Laboratory of Physiological Chemistry, Christian de Duve Institute of Cellular Pathology, B-1200 Brussels, Belgium [S. C., E. V. D. N., C. S., A. D., G. V. d. B., F. B.]; Department of Hematology, Cliniques universitaires Saint-Luc, Université catholique de Louvain, B-1200 Brussels, Belgium [E. V. D. N., A. F.]; and Tumoral Metabolism Research Unit, C. H. Jolimont-Lobbes, B-7100 La Louvière, Belgium [J-F. R.]

The effects of 2-chloro-2'-deoxyadenosine (CdA, cladribine), an adenosine deaminase-resistant analogue toxic for both proliferating and resting lymphoid cells, were investigated in the human leukemia cell line EHEB, which was derived from a patient with B-cell chronic lymphocytic leukemia. These cells were found to be less sensitive to CdA than B-cell chronic lymphocytic leukemia lymphocytes (25-fold) and other human lymphoblastic cell lines (10–1000-fold). Phosphorylation of CdA by deoxycytidine kinase and intracellular accumulation of 2-chloro-2'-deoxyadenosine triphosphate (CdATP) were similar in EHEB cells and in other CdA-sensitive cell lines. In contrast, the inhibitory effect of CdA on ribonucleotide reductase activity, which was investigated in situ by the conversion of cytidine into deoxyribonucleotides and its incorporation into DNA, was much less pronounced in EHEB cells than in other human lymphoblastic cells. Accordingly, concentrations of deoxynucleoside triphosphates did not decrease and even tended to rise. Unexpectedly, incorporation of thymidine and deoxycytidine into DNA was increased severalfold after a 24-h incubation with CdA. CdA also increased the activities of deoxycytidine kinase and thymidine kinase approximately 4-fold. Analysis of the cell cycle by flow cytometry showed that after 24 h, CdA provoked an increase in the proportion of cells in S phase, synthesizing DNA. We conclude that the EHEB cell line is resistant to the cytotoxic action of CdA not only because of a lack of inhibition of ribonucleotide reduction but also because CdA, in contrast with its known effects, provokes in this cell line an increase in the proportion of cells replicating their DNA. Unraveling of the mechanism of this effect may shed light on clinical resistance to CdA.

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