Transcriptional down-regulation of Ataxia-Telangiectasia Mutated protein (ATM) radiosensitizes human prostate cancer cells

Our laboratory recently reported that pre-treatment with the protein kinase C activator 12- O-tetradecanoylphorbol 12-acetate (TPA) enables an apoptotic response in radiationresistant LNCaP human prostate cancer cells via activation of the enzyme ceramide synthase (CS) and de novo synthesis of the sphingolipid ceramide (1). A mechanism for the TPA effect was not, however, provided. Here, we show that TPA functions to decrease the cellular level of the Ataxia Telangiectasia-Mutated (ATM) protein, known to repress CS activation by radiation (2), via inhibition of ATM gene transcription. Gel-shift analysis demonstrated a significant reduction in binding of the Sp-1 transcription factor to the ATM promoter, and quantitative RT-PCR showed a 50% reduction of ATM mRNA between 8-16h of TPA treatment of LNCaP and CWR22-Rv1 prostate cancer cells. Exposure of TPA-treated cells to radiation did not further decrease the cellular ATM levels, but increased CS activity and apoptosis, inhibitable by the specific CS inhibitor, fumonisin B1. The specificity of ATM inactivation towards the enhancement of CS activation by radiation was demonstrated by treatment of LNCaP, CWR22-Rv1, PC3 and DU-145 human prostate cells with antisense-ATM oligonucleotides (ODN), which markedly reduced cellular ATM levels enhancing radiation-induced CS activation and apoptosis, for doses as low as 1Gy. These data suggest that the CS pathway is a generic mode of radiation-induced apoptosis in human prostate cancer cells, regulated by a suppressive function of ATM kinase and that pharmacologic inactivation of ATM may be of clinical relevance.