Effects of insulin and pathway inhibitors on the PI3K-Akt-mTOR phosphorylation profile in acute myeloid leukemia cells

Abstract

The phosphatidylinositol 3-kinase (PI3K)-Akt-mechanistic target of rapamycin (mTOR) pathway is constitutively activated in human acute myeloid leukemia (AML) cells and is regarded as a possible therapeutic target. Insulin is an agonist of this pathway and a growth factor for AML cells. We characterized the effect of insulin on the phosphorylation of 10 mediators in the main track of the PI3K-Akt-mTOR pathway in AML cells from 76 consecutive patients. The overall results showed that insulin significantly increased the phosphorylation of all investigated mediators. However, insulin effects on the pathway activation profile varied among patients, and increased phosphorylation in all mediators was observed only in a minority of patients; in other patients, insulin had divergent effects. Global gene expression profiling and proteomic/phosphoproteomic comparisons suggested that AML cells from these two patient subsets differed with regard to AML cell differentiation, transcriptional regulation, RNA metabolism, and cellular metabolism. Strong insulin-induced phosphorylation was associated with weakened antiproliferative effects of metabolic inhibitors. PI3K, Akt, and mTOR inhibitors also caused divergent effects on the overall pathway phosphorylation profile in the presence of insulin, although PI3K and Akt inhibition caused a general reduction in Akt pT308 and 4EBP1 pT36/pT45 phosphorylation. For Akt inhibition, the phosphorylation of upstream mediators was generally increased or unaltered. In contrast, mTOR inhibition reduced mTOR pS2448 and S6 pS244 phosphorylation but increased Akt pT308 phosphorylation. In conclusion, the effects of both insulin and PI3K-Akt-mTOR inhibitors differ between AML patient subsets, and differences in insulin responsiveness are associated with differential susceptibility to metabolic targeting.

The PI3K-Akt-mTOR Signaling Pathway in Human Acute Myeloid Leukemia (AML) Cells

“…Furthermore, the AML heterogeneity is also reflected by the different effects of PI3K-Akt-mTOR inhibition, and as also demonstrated by our research group, the effects of these inhibitors seem to vary considerably among AML patients [70,79,80,140]. No clear mutational profile or other pathological process related to the disease, has so far been detected to predict response to treatment, although these features must be evaluated in larger in vitro and in vivo studies.…”

“…Dysregulation of the PI3K-Akt-mTOR signaling pathway subsequent to oncogene activating mutations, oncogene amplification, upstream activation of RTKs, or inactivation of tumor suppressor genes has been demonstrated in many human malignancies, including AML [79,80]. The PI3K-Akt-mTOR pathway is central for hematopoietic cells, and regulates crucial functions such as proliferation, differentiation, and survival.…”

“…The PI3K-Akt-mTOR pathway is central for hematopoietic cells, and regulates crucial functions such as proliferation, differentiation, and survival. This pathway seems to be constitutively activated in 60% of AML patients, and this activation seems to be associated with decreased overall survival [79][80][81][82][83]. Mutations in membrane bound-proteins, such as RTKs or GTPases, are major causes of dysregulated PI3K-Akt-mTOR signaling and are observed in 55% of AML cases [13,66].…”

“…AML patients harbor leukemia cells that display heterogeneous constitutive PI3K-Akt-mTOR activation profiles [79,95]. These dissimilarities are part of more complex phenotypic variations, including transcriptional regulation, cross signaling, and cell communication and may be influenced by growth factor stimulation or various inhibitors of pathway activation [70,79,80]. This is probably again reflected in the heterogeneity between leukemic cells with regard to energy metabolism, amino acid metabolism and arachidonic acid metabolism, supported by the finding that modulation of arachidonic acid metabolism alters the activation of mTOR and its downstream mediators [80].…”

The PI3K/AKT Pathway Inhibitor ISC-4 Induces Apoptosis and Inhibits Growth of Leukemia in Preclinical Models of Acute Myeloid Leukemia

“…Multiple studies implicating the PI3K/AKT pathway in AML have been reported previously (50)(51)(52). Activation of this pathway, as identified by phosphorylation of the Ser473 residue on Akt, has been noted in ∼ 70% of human AML (8,33).…”