Ablation of PI3K blocks BCR-ABL leukemogenesis in mice, and a dual PI3K/mTOR inhibitor prevents expansion of human BCR-ABL+ leukemia cells
Michael G. Kharas, … , Kevan M. Shokat, David A. Fruman
Michael G. Kharas, … , Kevan M. Shokat, David A. Fruman
Published August 14, 2008
Citation Information: J Clin Invest. 2008;118(9):3038-3050. https://doi.org/10.1172/JCI33337.
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Research Article

Ablation of PI3K blocks BCR-ABL leukemogenesis in mice, and a dual PI3K/mTOR inhibitor prevents expansion of human BCR-ABL+ leukemia cells

Abstract

Some cases of pre–B cell acute lymphoblastic leukemia (pre–B-ALL) are caused by the Philadelphia (Ph) chromosome–encoded BCR-ABL oncogene, and these tend to have a poor prognosis. Inhibitors of the PI3K/AKT pathway reduce BCR-ABL–mediated transformation in vitro; however, the specific PI3K isoforms involved are poorly defined. Using a murine model of Ph+ pre–B-ALL, we found that deletion of both Pik3r1 and Pik3r2, genes encoding class IA PI3K regulatory isoforms, severely impaired transformation. BCR-ABL–dependent pre/pro-B cell lines could be established at low frequency from progenitors that lacked these genes, but the cells were smaller, proliferated more slowly, and failed to cause leukemia in vivo. These cell lines displayed nearly undetectable PI3K signaling function and were resistant to the PI3K inhibitor wortmannin. However, they maintained activation of mammalian target of rapamycin (mTOR) and were more sensitive to rapamycin. Treatment with rapamycin caused feedback activation of AKT in WT cell lines but not PI3K-deficient lines. A dual inhibitor of PI3K and mTOR, PI-103, was more effective than rapamycin at suppressing proliferation of mouse pre–B-ALL and human CD19+CD34+ Ph+ ALL leukemia cells treated with the ABL kinase inhibitor imatinib. Our findings provide mechanistic insights into PI3K dependency in oncogenic networks and provide a rationale for targeting class IA PI3K, alone or together with mTOR, in the treatment of Ph+ ALL.

Authors

Michael G. Kharas, Matthew R. Janes, Vanessa M. Scarfone, Michael B. Lilly, Zachary A. Knight, Kevan M. Shokat, David A. Fruman

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Figure 1

Decreased BCR-ABL–mediated (p190) colony transformation of both α-null and α/β-null progenitor B cells.

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Decreased BCR-ABL–mediated (p190) colony transformation of both α-null a...
(A) Plating of p190-infected whole BM (5 × 104 cells) from 3- to 6-week-old mice of the indicated genotypes, on M3630 (CFU–pre-B) medium. *P < 0.05 versus WT; #P < 0.01 versus β-null; †P < 0.001 versus β-null; 1-way ANOVA, n = 3–7 independent experiments; mean values ± SEM are shown. (B) Schematic flowchart depicting the clonogenic expansion and assessment of PI3K expression status in L-CFCs. CFU–pre-B colonies were scored at day 7 after infection, and single colonies were selected and transferred to liquid culture. Outgrowth of p190+ L-CFCs was then quantified (“expansion” was defined as reaching ≥1 × 106 cells), followed by immunoblot assessment of Pik3r1 deletion and p55γ upregulation. Table 1 displays percentages of colonies meeting requirements for expansion, deletion of Pik3r1, and absence of p55γ upregulation. (C) monitoring class IA PI3K subunit expression in expanded L-CFCs. Representative immunoblot shows multiple clones of each genotype with variable loss of p85α and p55γ upregulation. p85α-specific antibody was used to distinguish p85α from p85β. White boxes represent clones that failed to delete p85α or upregulated p55γ. Black boxes represent clones that were used for in vitro and/or in vivo assays. (D) Immunoblot for class IA PI3K catalytic subunits in established L-CFCs. Representative blot for 3 independent clones.