Inhibition of O6-Alkylguanine DNA-Alkyltransferase or Poly(ADP-ribose) Polymerase Increases Susceptibility of Leukemic Cells to Apoptosis Induced by Temozolomide

Abstract

High levels of expression of the DNA repair enzyme O6-alkylguanine DNA-alkyltransferase (OGAT) (EC 2.1.1.63) account for tumor cell resistance to methylating agents. Previous studies suggested that methylating triazenes might have a potential role for the treatment of acute leukemias with low levels of OGAT. In the current study, we transduced the human OGAT cDNA in OGAT-deficient leukemia cell clones. OGAT-transduced cells were more resistant than their OGAT-deficient counterparts to apoptosis triggered by the methylating triazene temozolomide (TZM), as indicated by the results of flow cytometry, terminal deoxynucleotidyl transferase assay, and analysis of DNA fragmentation. Depletion of OGAT activity by O6-benzylguanine increased leukemia cell sensitivity to TZM-mediated apoptosis. Moreover, combined treatment of cells with TZM and benzamide, an inhibitor of the poly(ADP-ribose) polymerase (EC 2.4.2.30), increased the apoptosis induced by the methylating agent. These results demonstrate for the first time that methyl adducts at the O6 position of guanine, which are specifically removed by OGAT, are the principal DNA lesions responsible for the induction of apoptosis on treatment of leukemic cells with the methylating triazene TZM. This study also supports the possible use of TZM for the treatment of acute leukemias and suggests new strategies to increase the susceptibility of tumor cells to methylating triazenes in the clinic.