Figure 2.

VE-821–mediated ATR inhibition attenuates cell cycle transit and replication fork progression in combination with HU and Gem (but not cytarabine [Ara-C]) in AML cell lines. (A) Unsynchronized populations of U937 cells were treated with 10 nM Gem, 100 µM HU, or 100 nM Ara-C (or vehicle control) in the presence of 1 µM VE-821 or DMSO control. Cell cycle profiles were determined by propidium iodide staining (in 2 × 10⁵ cells) immediately pretreatment (0 hours) and at 8-hour intervals posttreatment, as indicated. (B) MV4-11 cells were preincubated with 5 nM Gem, 15 µM HU, 50 nM Ara-C, or vehicle control for 1 hour, then pulse labeled with 25 mM 5-iodo-2′-deoxyuridine (IdU) for 40 minutes, followed by 250 mM 5-chloro-2′-deoxyuridine (CldU) for 40 minutes (with maintenance of exposure to drug throughout). Treatment with 1 µM VE-821 or DMSO control was applied simultaneously with CldU. Data points represent ratios of CldU:IdU length in individual replication tracts in the presence (open circles) or absence (solid circles) of VE-821 from 3 independent experiments. Solid lines indicate mean ratios. P values were calculated using the Mann-Whitney U test. Images below charts are representative images of individual replication tracts with IdU in green and CldU in red. Scale bars represent 10 µm. See supplemental Figure 4.