Silencing of ETV6/RUNX1 abrogates PI3K/AKT/mTOR signaling and impairs reconstitution of leukemia in xenografts

Abstract

The ETV6/RUNX1 (E/R) gene fusion is generated by the t(12;21) and found in approximately 25% of childhood B-cell precursor acute lymphoblastic leukemia. In contrast to the overwhelming evidence that E/R is critical for the initiation of leukemia, its relevance for the maintenance of overt disease is less clear. To investigate this issue, we suppressed the endogenous E/R fusion protein with lentivirally transduced short hairpin RNA in the leukemia cell lines REH and AT-2, and found a distinct reduction of proliferation and cell survival. In line with the observed concurrent inactivation of the phosphoinositide 3-kinase (PI3K)/AKT/ mammalian target of rapamycin (mTOR) pathway, pharmacological inhibition diminished the phosphorylation of AKT and ribosomal protein S6, and significantly increased the apoptosis rate in E/R-positive leukemias. Moreover, PI3K/mTOR inhibitors sensitized glucocorticoid-resistant REH cells to prednisolone, an observation of potential relevance for improving treatment of drug-resistant relapses. Of note, knockdown of the E/R fusion gene also severely impaired the repopulation capacity of REH cells in non-obese deficient/severe combined immunodeficient mice. Collectively, these data demonstrate that the E/R fusion protein activates the PI3K/AKT/mTOR pathway and is indispensible for disease maintenance. Importantly, these results provide a first rationale and justification for targeting the fusion gene and the PI3K/AKT/mTOR pathway therapeutically.

STAT3 mediates oncogenic addiction to TEL-AML1 in t(12;21) acute lymphoblastic leukemia

“…A recent report used shRNA-mediated silencing of TEL-AML1 in human leukemic cell lines to demonstrate cell cycle inhibition and impaired engraftment in a xenotransplant mouse model. 22 In line with this observation, we show that silencing TEL-AML1 expression with shRNA targeting the 59 region of TEL, still present in the fusion gene transcript, causes a block in G1/S cell cycle transition in Reh cells (supplemental Figure 8). Moreover, this is accompanied by reduced levels of Tyr705 STAT3 phosphorylation ( Figure 2B).…”

“…18 Analysis of the function of TEL-AML1 in human leukemic cell lines has demonstrated that its continued expression is required for survival, proliferation, and leukemia engraftment. [20][21][22] However, there is only limited information on the signaling pathways deregulated by the fusion in these cells. [22][23][24][25] To address this question, we have used small hairpin (sh)RNA-mediated silencing to demonstrate that TEL-AML1 is responsible for inducing signal transducer and activator of transcription 3 (STAT3) activation in human t(12;21) leukemia cells.…”

“…[20][21][22] However, there is only limited information on the signaling pathways deregulated by the fusion in these cells. [22][23][24][25] To address this question, we have used small hairpin (sh)RNA-mediated silencing to demonstrate that TEL-AML1 is responsible for inducing signal transducer and activator of transcription 3 (STAT3) activation in human t(12;21) leukemia cells. This activation is mediated via RAC1, results in transcriptional induction of MYC and is critical for the survival of TEL-AML1…”

Targeting the PI3K/Akt/mTOR signaling pathway in B-precursor acute lymphoblastic leukemia and its therapeutic potential

“…8 mTOR is a serine/threonine kinase, downstream of Akt, that controls cell proliferation and survival. mTOR is the catalytic subunit of two distinct multi-protein complexes, referred to as mTORC1 and mTORC2.…”

Phospho-Inositol-3-Kinase Activity and Dysregulation in Pediatric Leukemia and Lymphoma

“…One study found that the protein product of this fusion gene regulates PI3K-AKT-mTOR signaling, as evidenced by the fact that knocking it down with shRNA resulted in reduced AKT and ribosomal protein S6 (rpS6) phosphorylationtwo mTOR-mediated events-while at the same time increasing apoptosis. Furthermore, pharmacologic inhibition of the PI3K-AKT-mTOR pathway with either the PI3K inhibitor LY294002 or the dual PI3K/mTOR inhibitor PI-103 induced apoptosis in ALL cells and sensitized glucocorticoid-resistant leukemia cells to the apoptotic effect of prednisolone [77].…”