ATR dependent phosphorylation of Chk1 was not inhibited by CP466722, despite the fact that ATM dependent phosphorylation of Chk2 was blocked in these cells. Failure to inhibit aphidicolin induced Chk1 bcr-abl phosphorylation in cells lacking ATM presented a lot more certain evidence that CP466722 does not inhibit ATR kinase in cells. DNA PK is another PIKK member of the family that contributes to damage induced signaling and equally ATM and DNA PK may phosphorylate histone H2AX on Serine139 following IR. Phosphorylation of histone H2AX was assessed in wild form and A T cells since DNA PK phosphorylates this page in the lack of ATM kinase activity, to investigate potential effects of CP466722 on DNA PK. While H2AX phosphorylation following IR was restricted by CP466722 or KU55933 in wild type cells, these ATM inhibitors did not prevent IR induced H2AX phosphorylation in A T cells, demonstrating a lack of noticeable effects Caspase inhibitor on DNA PK. In response to growth factor stimulation, AKT is activated by phosphorylation of threonine 308 by the PI3K pathway and serine 473 by other PIKK household members. Human fibroblasts were serum starved for 24h before being activated with IGF I either in the presence or lack of CP466722, KU55933 or Wortmannin, to demonstrate that CP466722 wasn’t inhibiting PI3K or PIKK family unit members. Serum starvation resulted in an almost complete lack of AKT phosphorylation. These phosphorylation functions were strongly induced upon addition of IGF I to serum starved cells and, as expected, were strongly inhibited by the recognized PI3K inhibitor wortmannin. No inhibition was observed with CP466722 or KU55933 treatment. Taken together, these results show that CP466722 inhibits Cellular differentiation ATM kinase, but doesn’t affect the cellular activity of PI3K or PIKK household members. Abl and Src kinases were identified in the original in vitro screens as possible targets of CP466722. To handle whether CP466722 inhibits cellular Abl and Src kinases, we employed a mouse pre B cell product. In this technique, the BCR Abl fusion protein is constitutively energetic, driving autophosphorylation of deposit tyrosine 245 and phosphorylation of a goal CrkL on tyrosine 207. Src kinase undergoes intermolecular autophosphorylation of deposit tyrosine 416 on its activation loop to become fully activated. In cells expressing BCR Abl, SRC kinases are activated and increased levels of Src phosphorylation have been reported suggesting that Src is effective and undergoing autophosphorylation. As a control, CP466722 and KU55933 were shown to inhibit ATM kinase activity in the mouse pre B cells as shown by disruption of p53 phosphorylation and p53 stabilization in Akt1 inhibitor reaction to IR. The mouse pre B cells were treated with CP466722, KU55933 or Imatinib as a positive control, to determine perhaps the inhibitors affected Abl and Src kinase exercise. Autophosphorylation of BCR Abl, endogenous Abl, and Abl dependent phosphorylation of CrkL were all found in control mouse pre B cells, as expected.