Received for publication January 3, 2008.
Revised March 27, 2008.
Accepted for publication April 14, 2008.

AKT is Activated in an ATR-dependent Manner in Response to Temozolomide and Confers Protection against Drug-induced Cell Growth Inhibition

Abstract

The PI3K/AKT pathway is activated frequently in human cancer and has been implicated in tumor cell proliferation, survival and chemoresistance. In this study we addressed the role of AKT in cellular responses to the therapeutic methylating agent temozolomide (TMZ), and investigated the possible link between TMZ-induced modulation of AKT function and activation of ATR- and ATM-dependent signalling pathways. We found that clinically relevant concentrations of TMZ caused activation of endogenous AKT in lymphoblastoid cells, and in colon and breast cancer cells, and that this molecular event required a functional mismatch repair system. Transfection of a dominant-negative kinase-dead form of AKT1 into breast cancer cells abrogated TMZ-induced activation of endogenous AKT and markedly enhanced cell sensitivity to the drug. Similarly, exposure of the MMR-proficient cell lines to the AKT inhibitor SH-5 impaired AKT phosphorylation in response to TMZ and significantly increased cell chemosensitivity. Furthermore, siRNA-mediated reduction of AKT1 expression in colon cancer cells potentiated the growth inhibitory effects of TMZ. Inhibition of ATM expression in colon cancer cells by siRNA did not impair TMZ-induced activation of AKT, whereas siRNA-mediated inhibition of ATR prevented AKT activation in response to the drug and increased cell chemosensitivity. These results strongly support the hypothesis that clinical benefit could be obtained by combining TMZ with inhibitors of the AKT pathway. Moreover, they provide the first evidence of a novel function of ATR as an upstream activator of AKT in response to DNA damage induced by O⁶-guanine methylating agents.

Key words: Mechanisms of cell killing/apoptosis, Resistance