Ectopic expression of Myc, but not myr-AKT, bypasses the need for Notch activation and generates lymphomas resistant to Notch inhibition. Lethally irradiated LC recipient mice were reconstituted with 5FU-treated donor LCR BM cells after transduction with retroviral supernatants expressing empty vector (N = 8), myr-AKT (N = 7), or Myc (N = 7). (A) Kaplan-Meier curve showing fraction of mice without T-ALL after BMT. P values (log-rank test) were <.0001 (vector vs myr-AKT), .0012 (vector vs Myc), and not significant (n.s.) (myr-AKT vs Myc). (B) Flow cytometric analysis of CD4 and CD8 expression of vector, myr-AKT, and Myc-transduced thymic lymphomas. (C) Western blot analysis of vector, myr-AKT, and Myc-transduced T-ALLs (for antibodies and blotting procedures, see “Methods”). ICN1, staining observed with an antibody against the V1744 epitope, which only recognizes the γ-secretase cleaved ICN1; Notch1 NTM, staining observed with an antibody against the C-terminal region of Notch1, which recognizes the furin-processed NTM; p-AKT, staining observed with an antibody specific for AKT phosphorylated on S473; p-S6K, staining observed with an antibody specific for S6K phosphorylated on Thr389; t-AKT, total AKT; t-S6K, total S6K. The arrow indicates ICN1 likely resulting from 5′ rearrangements or other mutations as described in “Results.” The multiple bands within the brackets likely result from combined PEST mutations and 5′ rearrangements or other mutations as described in “Results.” (D) Primary T-ALL cell lines derived from vector, myr-AKT, and Myc T-ALLs were cultured ex vivo for 3 weeks. These cell lines were then treated with GSIs (1 μM JC-19) for 6 days and counted on day 6. Fold expansion was calculated based on the initial day 0 cell counts. DMSO, dimethyl sulfoxide.