Reactivation of DEP-1 by Prx-1 overexpression attenuates FLT3 ITD–dependent transformation. FLT3 ITD expressing 32D cells, harboring either nontargeting shRNA (not labeled for better clarity in panel C) or DEP-1 targeting shRNA (as indicated), were engineered by retroviral transduction to overexpress Prx-1, or were transduced with control vector. (A) Colony formation in methylcellulose of the 4 different cell pools was assessed. Left panel: example image. Right panel: quantification of colonies > 0.4 mm. (B) ROS formation and DEP-1 activity were assessed as described for Figures 2A and 1C, respectively (representative experiments of 3 with consistent results). (C) C3H/HeJ mice (10 mice per group) were injected with 2 × 10⁶ cells each of the indicated 32D cell pools. Survival was monitored and is displayed as Kaplan-Meier plot. Statistical significance was tested using Gehan-Breslow statistics for the survival curves; posthoc comparisons were made with the Holm-Sidak method for all pairwise multiple comparisons. DEP-1 knockdown efficiencies were assessed at day 11 as described in “Methods,” and are shown as qRT-PCR and immunoblotting data in the bottom panels. Note also equal DEP-1 expression in the cell pools harboring control shRNA. Survival of mice injected with cells harboring FLT3 ITD and Prx-1 was significantly prolonged (15.4 ± 0.3 days; P < .001) compared with mice injected with the corresponding cells additionally harboring DEP-1 shRNA (11.9 ± 0.3 days), with cells harboring FLT3 ITD but without Prx-1 12.5 ± 0.4 days), and with cells harboring FLT3 ITD and DEP-1 shRNA only (12.5 ± 0.2 days). There was no significant difference in the survival curves of mice injected with the latter 3 cell types.