BackgroundTargeted therapy approaches have been successfully introduced into the treatment of several cancers. The multikinase inhibitor Sorafenib has antitumor activity in solid tumors and its effects on acute lymphoblastic leukemia (ALL) cells are still unclear.MethodsALL cell lines (SEM, RS4;11 and Jurkat) were treated with Sorafenib alone or in combination with cytarabine, doxorubicin or RAD001. Cell count, apoptosis and necrosis rates, cell cycle distribution, protein phosphorylation and metabolic activity were determined.ResultsSorafenib inhibited the proliferation of ALL cells by cell cycle arrest accompanied by down-regulation of CyclinD3 and CDK4. Furthermore, Sorafenib initiated apoptosis by cleavage of caspases 3, 7 and PARP. Apoptosis and necrosis rates increased significantly with most pronounced effects after 96 h. Antiproliferative effects of Sorafenib were associated with a decreased phosphorylation of Akt (Ser473 and Thr308), FoxO3A (Thr32) and 4EBP-1 (Ser65 and Thr70) as early as 0.5 h after treatment. Synergistic effects were seen when Sorafenib was combined with other cytotoxic drugs or a mTOR inhibitor emphasizing the Sorafenib effect.ConclusionSorafenib displays significant antileukemic activity in vitro by inducing cell cycle arrest and apoptosis. Furthermore, it influences PI3K/Akt/mTOR signaling in ALL cells.

No abstract

Background: The phosphatidylinositol 3 Kinase (PI3K)/Akt signalling pathway is involved in the regulation of proliferation, apoptosis and angiogenesis. Dysregulation of PI3K/Akt has been described for several solid and hematological tumors. Its role in acute lymphoblastic leukemogenesis (ALL) is unkown. Here, we investigated whether the inhibition of PI3K/Akt kinases influences apoptosis, necrosis and cell proliferation. Patients and Methods: ALL and high grade NHL cell lines with different cytogenetics and phenotypes were used (SEM, RS4;11, REH, Jurkat, DOGKIT). Phosphorylationstatus of Akt (Ser473, Thr308) and FOXO3A (Thr32) were determined by western blot (WB). Cells were incubated for 96h with LY29004 (5μM, 12.5μM, 25μM; PI3K inhibitor) or sorafenib (0.73μM, 7.3μM). In order to detect synergistic effects with other cytotoxic drugs cells were treated with combinations of sorafenib and doxorubicine, cytosin-arabinoside, and mTOR inhibitor (RAD001), respecitvely. Cell number, apoptosis, necrosis, protein phosphorylation and metabolic activity were determined at 4h, 24h, 48h, 72h, and 96h by microscopy, flow cytometry, WB and WST-1 testing. Whole genome Affymetrix gene expression arrays (U133 Plus 2.0) were performed in order to detect differential gene expression compared to controls treated with DMSO (dimethylsulfoxid) only. Results: In all ALL cell lines phosphorylated Akt (pAKT) was detected. Levels of pAkt and pFOXO3A differed between cell lines significantly, with Jurkat and SEM demonstrating high activation levels. PI3K inhibition by LY29004 led to decreased proliferation in SEM cells with only slight increases in apoptosis and moderate increase in necrosis rates (20%). Sorafenib inhibited the proliferation of SEM, Jurkat and RS4;11 significantly, with most pronounced effects at 96h. Maximal apoptosis and necrosis rates increased and ranged from 11 to 39% and 53 to 84%, respectively. Metabolic activity decreased significantly already after 24h. As early as 0.5h after treatment complete disappearance (SEM, RS4;11) or marked decrease (Jurkat) in levels of pAkt and pFOXO occurred. Combination of sorafenib with conventional cytotoxic drugs failed to demonstrate synergistic effects. Sorafenib treatment induced differential gene expression in several genes e.g. genes involved in apoptosis such as BIMBAM, caspases and CDKs. Conclusions: Inhibition of the PI3K/Akt pathway seems to be a potential therapeutic target in ALL and high grade lymphoma cells. Sorafenib as a multikinase inhibitor approved for clincial application in solid tumors displayes significant antileukemic activity in vitro and might be a potential drug for a targed therapy approach in ALL.