CTL-killing of LSCs in vivo depends on leukemia load. (A) Experimental design. 2 × 10⁴ H8 LSCs from H8 CML mice were FACS-purified 20 d after transplantation and were transferred i.v. into irradiated (6.5 Gy) secondary BL/6 recipients. 18 h later, mice were either left untreated (Ø, n = 4) or treated with 3 × 10⁶ FACS-purified p14 CTLs i.v. (n = 4). 48 h later, BM was harvested and analyzed for BCR/ABL-GFP⁺ cells (B) and LSCs (C) by FACS. Pooled data from two independent experiments are shown. (D) Kaplan-Meier survival curves resulting from H8 CML mice either left untreated (Ø, n = 5) or treated with 5 × 10⁶ p14 effector CTLs (p14, n = 5) 7 d after transplantation. (E–I) H8 CML mice 18 d after transplantation were either left untreated (Ø, n = 4) or treated with 3 × 10⁶ FACS-purified p14 effector CTLs (p14, n = 5). 2 d later, (E) the frequencies of LSCs in lin⁻BCR/ABL-GFP⁺ cells, the numbers of total BM cells (F), the numbers of LSCs in the BM (G), the frequencies of LSK cells in lin⁻GFP⁻ cells (H), and the numbers of LSK cells in the BM (I) were analyzed. One representative plot of 4–5 is shown for each group. (J) Kaplan-Meier survival curves resulting from H8 CML mice either left untreated (Ø, n = 9) or treated with 5 × 10⁶ p14 effector CTLs (p14, n = 10) 19 d after transplantation. Data are displayed as mean ± SEM. Statistics: Student’s t test (B, C, F, G, and I), log-rank test (D and J). *, P < 0.05; **, P < 0.005. LSCs, leukemia stem cells; LSK, lin⁻Sca-1⁺c-kit^hi.