Elevated levels of PIP3 set off the recruitment of phos phatidylinositide dependent serine threonine kinase 1 and Akt on the cytoplasmic membrane in which PDK1 phosphorylates Akt on threonine 308. An addi tional phosphorylation on serine 473 is needed to fully activate Akt. Phosphorylation on threonine 308 naturally precedes phosphorylation on serine 473 but phosphorylation on serine 473 appears to be independent of PDK1. Despite the fact that several kinases, this kind of as integrin linked kinase, DNA dependent protein kinase, as well as the mTOR Rictor complicated happen to be pro posed to function as so known as PDK two, the identity of your serine 473 kinase is still controversial, There is accumulated evidence that LY294002 interferes with all the activation of Akt by inhibiting its upstream regulator PI3K.
In contrast, membrane tar geted alkylphosphocholines like ErPC3 interfere with membrane composition therefore affecting the recruit ment of Akt to the plasma membrane that is a prere quisite for its activation by PDK1, To the basis of this mechanism of action, ErPC3 and associated com pounds would even be successful in cells exactly where the substantial activity of Akt is caused by a constitutively active PI3K that is not inhibited by article source LY294002. In our hands, remedy with LY294002 resulted within a quick and constant downregulation of p Akt amounts while in the really LY294002 sensitive LNCaP cells. ErPC3 treatment also diminished p Akt ranges in LNCaP cells to a considerable sum. The reduce in p Akt was accom panied through the induction of cell PF-4929113 death by the two com pounds. This suggests that in LNCaP cells the constitutive activation of your survival kinase Akt takes place downstream of an overactive PI3K that’s inhibited by both, the PI3K inhibitor LY294002 as well as Akt inhibitor ErPC3.
In PC3 cells howerver, only ErPC3 reduced p Akt and induced cell death to a substantial sum when concentrations below 50 ?M were utilised. This sug gests that the higher p Akt levels in PC3 cells count on a LY294002 insensitive but ErPC3 delicate mechanism. Hence, PC3 cells may perhaps express a mutant PI3K that is insensitive to inhibition by LY294002. Alternatively, Akt activation in PC3 cells may perhaps happen independently from PI3K, e. g. by aberrant activation of Akt activating kinases or by loss or inactivation of p Akt phosphatases. There is certainly accumulated evidence that constitutive acti vation from the PI3K Akt pathway interferes using the cyto toxic action of ionizing radiation. However, it truly is regarded from earlier investigations that the antineo plastic efficacy of ErPC3 is elevated in human tumor cells when the drug is mixed with genotoxic agents like cytarabine, idarubicine or etoposide, or with ioniz ing radiation, respectively, Therefore, in the last set of in vitro experiments, we analyzed no matter whether treat ment using the Akt inhibitor ErPC3 would boost the quick time antineoplastic results of ionizing radiation within the prostate cancer cell lines.