Figure 2. Deletion of Pten compromises BCR-ABL1 and NRAS-driven leukemogenesis.

(a) The percentage of GFP⁺ cells were measured in BCR-ABL1 (left) or NRAS^G12D (right) transformed Pten^fl/fl pre-B cells transduced with inducible Cre-ER^T2-GFP or GFP empty vector (EV) control after induction of Cre with tamoxifen by flow cytometry. For each plot, P < 0.0001 was obtained by contingency table. (b) Comparison of colony forming ability of BCR-ABL1 (left) or NRAS^G12D (right) transformed Pten^fl/fl pre-B cells after Pten deletion. (c) Measuring the induction of cell cycle arrest following 24 h of tamoxifen induced Pten deletion in BCR-ABL1 (left) or NRAS^G12D (right) transformed Pten^fl/fl pre-B cells. The percentages of each phase of the cell cycle are indicated. (d) The prevalence of cellular senescence was measure using β-galactosidase assay after deletion of Pten in BCR-ABL1 (left) or NRAS^G12D (right) transformed Pten^fl/fl pre-B cells. The percentages of cells stained with blue dots are depicted on the panels. Results in (a–d) are representative of two independent experiments. Scale bars in (b) and (d) represent 1 mm and 15 μm respectively. P values (b–d) were calculated by Student’s t test. (e) NOD/SCID recipient mice injected with 10⁵ Pten^fl/fl BCR-ABL1-transformed pre-B ALL transduced with Cre-ER^T2 (left) were treated with tamoxifen (0.4 mg/mouse) or corn oil (vehicle) (n = 7 per group). In a parallel experiment, NOD/SCID recipient mice were injected with 10⁵ Pten^fl/fl NRAS^G12D transformed pre-B ALL cells transduced with Cre-ER^T2 or empty vector (EV) control (right). In the latter experiment all mice were treated with Tamoxifen (0.4 mg/mouse; n = 7 per group). P values were calculated by Mantel-Cox log-rank test. All cells were labeled with luciferase before injection. Treatments were started on day 5 after transplantation and continued for ten consecutive days. Error bars (a–b, d) represent S.D.