ReviewEnhancement of arabinocytosine (AraC) toxicity to AML cells by a differentiation agent combination

•Exposure of AML cells with DNA damage to differentiation agents increases cell death.•Vitamin D analogs and plant-derived antioxidants can produce synergistic cytotoxicity.•VDR expression is up-regulated by AraC, and further enhanced by 1-D2/CA combination.

Arabinocytosine (AraC, also known as cytarabine) is one of the mainstays of AML therapy, but like other DNA damaging therapeutic agents it is rarely curative by itself. There is an emerging realization that the therapeutic outcomes may be improved by combining AraC with other compounds. Here we report that the addition of a differentiating agent combination immediately following AraC damage to AML blasts, selectively increases the cell kill. The experiments were performed using cultured cells from established cell lines of AML (HL60 and U937). The cells were exposed to 100 nM AraC, a concentration which produced approximately 25-50% cell kill, followed by a combination of 100 nM 1alpha-hydroxyvitamin D2 (1-D2) and 10 μM carnosic acid (CA), which together can serve as a powerful differentiating agent combination for AML cells, but are not toxic alone. AraC-induced cell death, measured by annexin V/propidium iodide, was significantly (p < 0.01) increased by the 1-D2/CA combination in both cell lines, but not by 1-D2 or CA alone. The enhancement of cell death occurred by both apoptosis and necrosis, was associated with increased DNA damage and with higher levels of DNA damage response (DDR) activated marker Chk1, but the expression of p27, a cell cycle inhibitor protein, was not enhanced by 1-D2/CA. The principal finding is that a vitamin D analog 1-D2 combined with a plant-derived antioxidant CA can markedly augment the cytotoxic action of AraC, an anti-leukemia therapeutic agent.

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