Sex differences in the GSK3ß-mediated survival of adherent leukemic progenitors.

Therapeutic resistance of acute myeloid leukemia stem cells, enriched in the CD34(+)38(-)123(+) progenitor population, is supported by extrinsic factors such as the bone marrow niche. Here, we report that when adherent onto fibronectin or osteoblast components, CD34(+)38(-)123(+) progenitors survive through an integrin-dependent activation of glycogen synthase kinase 3ß (GSK3ß) by serine 9-dephosphorylation. Strikingly, GSK3ß-mediated survival was restricted to leukemic progenitors from female patients. GSK3ß inhibition restored sensitivity to etoposide, and impaired the clonogenic capacities of adherent leukemic progenitors from female patients. In leukemic progenitors from female but not male patients, the scaffolding protein RACK1, activated downstream of α(5)ß(1)-integrin engagement, was specifically upregulated and controlled GSK3ß activation through the phosphatase protein phosphatase 2A (PP2A). In a mirrored manner, survival of adherent progenitors (CD34(+)38(-)) from male but not female healthy donors was partially dependent on this pathway. We conclude that the GSK3ß-dependent survival pathway might be sex-specific in normal immature population and flip-flopped upon leukemogenesis. Taken together, our results strengthen GSK3ß as a promising target for leukemic stem cell therapy and reveal gender differences as a new parameter in anti-leukemia therapy.

The level of AKT phosphorylation on threonine 308 but not on serine 473 is associated with high-risk cytogenetics and predicts poor overall survival in acute myeloid leukaemia.

The phosphoinositide 3-kinase/Akt pathway is an important signalling pathway governing cell survival and proliferation in acute myeloid leukaemia (AML). As full activation of Akt requires phosphorylation on both threonine 308 (Thr308) and serine 473 (Ser473) residues, we studied the level of phosphorylation on the both sites in 58 AML samples by flow cytometry. The ratio of the mean fluorescence intensity of Thr308 and Ser473 represented a continuum ranging from 0.3 to 5.6 and from 0.4 to 2.87, respectively. There were no significant correlations between age, gender, French-American-British classification, leukocytosis, FLT3-ITD and Akt phosphorylation. However, the level of phosphorylation on Thr308, but not on Ser473, was significantly correlated with high-risk karyotype. Thr308(high) patients had significantly shorter overall survival (11 vs 47 months; P=0.01), event-free survival (9 vs 26 months; P=0.005) and relapse-free survival (10 months vs not reached; P=0.02) than Thr308(low) patients. Neither screening for AKT1 E17K mutation nor changes in the level of PTEN expression and phosphorylation could be linked to increased phosphorylation on Thr308 in high-risk cytogenetic AML cells. However, PP2A activity was significantly reduced in high-risk samples compared with intermediate-risk samples. Moreover, the specific Akt inhibitor, Akti-1/2, inhibited cell proliferation and clonogenic properties, and induced apoptosis in AML cells with high-risk cytogenetics, suggesting that Akt may represent a therapeutic target in high-risk AML.

PI-103, a dual inhibitor of Class IA phosphatidylinositide 3-kinase and mTOR, has antileukemic activity in AML.

The phosphatidylinositol 3-kinase (PI3K)/Akt and mammalian target of rapamycin complex 1 (mTORC1) signaling pathways are frequently activated in acute myelogenous leukemia (AML). mTORC1 inhibition with RAD001 induces PI3K/Akt activation and both pathways are activated independently, providing a rationale for dual inhibition of both pathways. PI-103 is a new potent PI3K/Akt and mTOR inhibitor. In human leukemic cell lines and in primary blast cells from AML patients, PI-103 inhibited constitutive and growth factor-induced PI3K/Akt and mTORC1 activation. PI-103 was essentially cytostatic for cell lines and induced cell cycle arrest in the G1 phase. In blast cells, PI-103 inhibited leukemic proliferation, the clonogenicity of leukemic progenitors and induced mitochondrial apoptosis, especially in the compartment containing leukemic stem cells. In contrast, apoptosis was not induced with RAD001 and IC87114 association, which specifically inhibits mTORC1 and p110delta activity, respectively. PI-103 had additive proapoptotic effects with etoposide in blast cells and in immature leukemic cells. Interestingly, PI-103 did not induce apoptosis in normal CD34(+) cells and had moderate effects on their clonogenic and proliferative properties. Here, we demonstrate that multitargeted therapy against PI3K/Akt and mTOR with PI-103 may be of therapeutic value in AML.