The DNA damage induced by, and cellular responses to, the C-1027 family of antitumor antibiotic radiomimetic enediynes
Kennedy, Daniel R.
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My thesis research involves the characterization of a novel pathway that cells activate in response to DNA double strand breaks. The activation of DNA repair and cell cycle checkpoint responses following the induction of DNA double strand breaks requires activation of the phosphatidylinositol 3-kinase-related kinase (PIKK) family member (ATM). The only observed exception occurs when cells are treated with the radiomimetic enediyne C-1027. The first chapter determined that the cellular response to C-1027 can utilize both ATM and the related PIKK ATR to activate cell cycle checkpoint responses. Since C-1027 induces DNA double strand breaks (DSBs), we wanted to determine what was different about C-1027 induced damage from that of ionizing radiation and other radiomimetics. The second chapter further examined C-1027-induced DNA damage by characterizing three bio-engineered C-1027 analogs, each containing a single modification in the enediyne chromophore for their ability to induce DNA damage and ATM-independent activation of cell cycle checkpoints. While all of the analogs were less potent in inducing DSBs, the least potent analog retained the ability to induce ATM-independent DNA damage responses while the other analogs did not. This observation led us to believe that select C-1027 family members induce a second type of DNA damage other than DSB. The last chapter examined the ability of C-1027 and its analogs to induce cellular DNA inter-strand crosslinks, based upon a previous observation of the parent compound under anaerobic conditions in cell-free systems. It was determined that both C-1027 and desmethyl robustly induced DNA ICLs under anaerobic cell-free and aerated cellular conditions. Furthermore, drug induced activation of ATM and ATR was dependent on the robust induction of DNA DSBs and ICLs respectively, as C-1027 robustly induced both lesions and both DNA damage responses with either ATM or ATR, desmethyl induced mostly ICLs and requires ATR, while deschloro induced mostly DSBs and required ATM.