SEM, RS4,11 and Jurkat cells showed a decrease of the phosphorylated as well as the total form of 4EBP 1 after an incubation of 4 and 24 h. In contrast, MOLT4 cells displayed an increase of the phosphorylated form at these points of time. PDA 66 does not inhibit kinase activity of recombinant GSK3B as distinct as SB 216763 The effect of PDA 66 on the GSK3B enzyme activity selleck kinase inhibitor was determined by incubation with the specific substrate pGS2, PDA 66 or SB 216763 and ATP. The following addition of Kinase Glo reagent converts the remaining ATP into a lu minescence signal which correlates with enzyme inhibition. SB 216763 demonstrated a stable inhibition of GSK3B at concentrations from 0. 1 to 5 uM which was statistically sig nificant at 5 uM. Compared to this PDA 66 showed a less pronounced inhibition of enzyme activity at concentration from 0.

1 to 1 uM which were not significant. Discussion The prognosis of ALL in adult patients is still poor and requires further research for new therapeutic ap proaches. In this study we could demonstrate for the first time a pronounced antiproliferative effect of the novel arylindolylmaleimide PDA 66 on different B and T ALL cell lines. We investigated the influence of PDA 66 on ALL cells in respect of proliferation, metabolic activ ity, morphology, apoptosis, cell cycle arrest, and activa tion of PI3K Akt and Wnt B catenin signaling pathways. Furthermore, the effect on kinase activity of GSK3B was determined. PDA 66 was recently synthesized and described as an analogue to SB 216763, which is a known GSK3B inhibi tor.

The inhibition of this kinase has been extensively examined in various neoplastic cells types and demon strated an attenuated proliferation in malignant cells. Investigating the influence of PDA 66 on the en zyme activity of human recombinant GSK3B we found a minor inhibition in vitro which was much less distinct and not significant compared to our results obtained with SB 216763. While the basic molecular structure of SB 216763 and PDA 66 is the same, both compounds differ in their substitution patterns. In comparison to SB 216763, PDA 66 is characterized by an unprotected 2 methylindole group and a methylated maleimide group. Additionally, the 2,4 dichloro substitution pattern is replaced with a 4 acetyl group in PDA 66. These structural changes are supposed to be key in the reduced capacity of PDA 66 to inhibit GSK3B.

The influence of PDA 66 on GSK3B activity including other key enzymes of Wnt B catenin and PI3K Akt sig naling pathways was also investigated by western blot. Affirming the results obtained by kinase activity selleckchem assay, we found no enhanced activation of the Wnt B catenin pathway. Considering the role of GSK3B in Wnt signal ing an increase of B catenin would have been expected when inhibiting GSK3B. Furthermore, no effect on the protein expression of GSK3B and no distinct activation of Akt were detectable.