Akt activation suppresses Chk2-mediated, methylating agent-induced G2 arrest and protects from temozolomide-induced mitotic catastrophe and cellular senescence

Pharmacologic inhibition of the DNA signal transducers Chk1 and p38 blocks G(2) arrest and sensitizes glioblastoma cells to chemotherapeutic methylating agent-induced cytotoxicity. Because Akt pathway activation has been suggested to also block G(2) arrest induced by DNA-damaging agents and because glioma cells frequently have high levels of Akt activation, we examined the contribution of the Akt pathway to methylating agent-induced G(2) arrest and toxicity. U87MG human glioma cells containing an inducible Akt expression construct were incubated with inducing agent or vehicle, after which the cells were exposed to temozolomide and assayed for activation of the components of the G(2) arrest pathway and survival. Temozolomide-treated control cells activated the DNA damage signal transducers Chk1, Chk2, and p38, leading to Cdc25C and Cdc2 inactivation, prolonged G(2) arrest, and loss of clonagenicity by a combination of senescence and mitotic catastrophe. Temozolomide-treated cells induced to overexpress Akt, however, exhibited significantly less drug-induced Cdc25C/Cdc2 inactivation and less G(2) arrest. Akt-mediated suppression of G(2) arrest was associated not with alterations in Chk1 or p38 activation but rather with suppression of Chk2 activation and reduced recruitment of Chk2 to sites of damage in chromatin. Unlike bypass of the G(2) checkpoint induced by pharmacologic inhibitors of Chk1 or p38, however, Akt-induced bypass of G(2) arrest suppressed, rather than enhanced, temozolomide-induced senescence and mitotic catastrophe. These results show that whereas Akt activation suppresses temozolomide-induced Chk2 activation and G(2) arrest, the overriding effect is protection from temozolomide-induced cytotoxicity. The Akt pathway therefore represents a new target for the sensitization of gliomas to chemotherapeutic methylating agents such as temozolomide.