The mechanisms of Ara-C-induced apoptosis of resting B-chronic lymphocytic leukemia cells

Department of Hematology, Leiden University Medical Center, P.O. Box 9600 2300 RC Leiden, The Netherlands. j.f.vries@lumc.nl

BACKGROUND AND OBJECTIVES: Cytarabine (Ara-C) is commonly used for the treatment of acute leukemia. Incorporation of Ara-C into DNA is a key event in the mechanism of killing of proliferating leukemic cells. Previously, we demonstrated that Ara-C was cytotoxic to proliferating but not to resting (G(O)) malignant cells from patients with acute leukemia. In contrast, here we show unexpected apoptosis of G(O) B-chronic lymphocytic leukemia (CLL) cells by Ara-C in a dose-dependent manner. In this study we analyzed which cellular processes were involved in Ara-C-mediated killing of G(O)-B-CLL cells. DESIGN AND METHODS: Using primary B-CLL cells (>98% in G(O)), we examined the mechanisms of Ara-C-mediated apoptosis in resting G(O) cells. CFSE-based cytotoxicity assays combined with cell cycle analysis were used to perform a long-term analysis of Ara-C-mediated killing of B-CLL cells. The effects of Ara-C on DNA and RNA synthesis were studied using various 3H-incorporation experiments. RESULTS: Ara-C-mediated cell death of B-CLL cells showed the characteristics of normal apoptosis, such as phosphatidyl serine exposure and caspase activation. The mechanism of killing of quiescent B-CLL cells by Ara-C was shown not to be dependent on DNA replication. In contrast, CD40L-activated B-CLL cells showed S-phase-specific depletion of proliferating CLL cells. We demonstrated that Ara-C was converted into its active triphosphate by G(O)-B-CLL cells, coinciding with a 30% inhibition of RNA synthesis. INTERPRETATION AND CONCLUSIONS: In conclusion, our data indicate that Ara-C can induce apoptosis in resting G(O)-B-CLL cells using a mechanism independent of cell proliferation and DNA replication but associated with inhibition of RNA synthesis and downregulation of Mcl-1.

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