NF-kappa B is activated in response to temozolomide in an AKT-dependent manner and confers protection against the growth suppressive effect of the drug

Background: Most DNA-damaging chemotherapeutic agents activate the transcription factor nuclear factor kB (NF-kB). However, NF-kB activation can either protect from or contribute to the growth suppressive effects of the agent. We previously showed that the DNA-methylating drug temozolomide (TMZ) activates AKT, a positive modulator of NF-kB, in a mismatch repair (MMR) system-dependent manner. Here we investigated whether NF-kB is activated by TMZ and whether AKT is involved in this molecular event. We also evaluated the functional consequence of inhibiting NF-kB on tumor cell response to TMZ. Methods: AKT phosphorylation, NF-kB transcriptional activity, IkB-alpha degradation, NF-kB2/p52 generation, and RelA and NF-kB2/p52 nuclear translocation were investigated in TMZ-treated MMR-deficient (HCT116, 293TL alpha(-)) and/or MMR-proficient (HCT116/3-6, 293TL alpha(+), M10) cells. AKT involvement in TMZ-induced activation of NF-kB was addressed in HCT116/3-6 and M10 cells transiently transfected with AKT1-targeting siRNA or using the isogenic MMR-proficient cell lines pUSE2 and KD12, expressing wild type or kinase-dead mutant AKT1. The effects of inhibiting NF-kB on sensitivity to TMZ were investigated in HCT116/3-6 and M10 cells using the NF-kB inhibitor NEMO-binding domain (NBD) peptide or an anti-RelA siRNA.