Constitutive Activation of Akt and mTOR by Flt3 Internal Tandem Duplications Mediates Myeloid Leukemogenesis and Can Be Inhibited by Rapamycin.

Up to 30% of patients with acute myeloid leukemia (AML) harbor internal tandem duplications (ITD) within the FLT3 gene, a tyrosine kinase receptor. The signalling pathways and downstream targets are only partly understood. We demonstrated that the presence of Flt3-ITD constitutively activated Akt (PKB), a key serine-threonine kinase within the PI3-kinase pathway. The constitutive activation of Akt induced phosphorylation and transcriptional inhibition of the transcription factor Foxo3a. Also, forced Foxo3a activity reversed Flt3-ITD-mediated growth properties. Using a novel Akt inhibitor, we showed that Flt3-ITD-mediated Foxo3a phosphorylation was dependent on Akt. Conditional Akt activation targeted to the cell membrane conferred cytokine-independent survival, proliferation and transformation of myeloid progenitor cells. Induction of Akt activity in vivo promoted the development of a myeloproliferative disease in a syngeneic mouse model. Importantly, phosphorylation of Akt was frequently found in primary myeloid blasts, supporting that this signalling pathway may be important in the development of AML. Furthermore, activation of Akt constitutively activated the downstream kinase mammalian target of rapamycin (mTOR). mTOR has several important biological functions, notably in nutrient sensing and protein translation. Treatment with rapamycin, a specific inhibitor of mTOR, inhibited mTOR kinase activity at 1nM in all examined cell systems. Rapamycin caused cell cycle arrest, apoptosis and completely inhibited transformation in FLT3-ITD-positive cells. These effects were most pronounced in the low nanomolar range, which are therapeutic concentrations of rapamycin-treated patients. The biological advantage in Akt-expressing cells was fully reversed by treatment with rapamycin. In all examined primary blasts from AML patients, mTOR was inhibited by rapamycin. In short term proliferation assays, twelve out of 18 (67%) examined showed marked sensitivity with an IC50 below 10nM rapamycin, while one third were found to be more resistant.

Taken together, increased survival, proliferation and leukemic transformation by Flt3-ITD is mediated by Akt and mTOR, and can be inhibited by rapamycin. We conclude that rapamycin may be an attractive substance for the treatment of patients with AML.