Cancer Therapy: Preclinical Effects of Selective Checkpoint Kinase 1 Inhibition on Cytarabine Cytotoxicity in Acute Myelogenous Leukemia Cells In Vitro

Purpose: Previous studies have shown that the replication checkpoint, which involves the kinases ataxia telangiectasiamutated andRad3 related (ATR) andChk1, contributes to cytarabine resistance in cell lines. In the present study, we examinedwhether this checkpoint is activated in clinical acutemyelogenous leukemia (AML) during cytarabine infusion in vivo and then assessed the impact of combining cytarabine with the recently described Chk1 inhibitor SCH 900776 in vitro. Experimental design: AML marrow aspirates harvested before and during cytarabine infusion were examined by immunoblotting. HumanAML lines treatedwith cytarabine in the absence or presence of SCH 900776 were assayed for checkpoint activation by immunoblotting, nucleotide incorporation into DNA, and flow cytometry. Long-term effects in AML lines, clinical AML isolates, and normal myeloid progenitors were assayed using clonogenic assays. Results: Immunoblotting revealed increased Chk1 phosphorylation, a marker of checkpoint activation, inmore thanhalf of Chk1-containing AMLs after 48hours of cytarabine infusion. In humanAML lines, SCH 900776 not only disrupted cytarabine-induced Chk1 activation and S-phase arrest but also markedly increased cytarabine-induced apoptosis. Clonogenic assays demonstrated that SCH 900776 enhanced the antiproliferative effects of cytarabine in AML cell lines and clinical AML samples at concentrations that had negligible impact on normal myeloid progenitors. Conclusions: These results not only provide evidence for cytarabine-induced S-phase checkpoint activation in AML in the clinical setting, but also show that a selective Chk1 inhibitor can overcome the S-phase checkpoint and enhance the cytotoxicity of cytarabine. Accordingly, further investigation of the cytarabine/SCH 900776 combination in AML appears warranted. Clin Cancer Res; 18(19);