Altering DNA base excision repair: Use of nuclear and mitochondrial targeted N-methylpurine DNA glycosylase (MPG) to sensitize astroglia to chemotherapeutic agents

Mitochondrial DNA (mtDNA) is a sensitive target of anti-cancer DNA alkylating agents, and previous studies have revealed a strong correlation between cell type-specific mtDNA repair capacity and cell type-specific resistance in the central nervous system (CNS) to oxidation and alkylation treatment. An assessment of two primary cell types indicated that sensitivity in the CNS to DNA damaging agents may be attributable to an imbalance in mitochondrial base excision repair (BER). This study utilized a primary astrocyte model to investigate mtDNA repair modulation as a potential tool for sensitizing astrocytic tumors to genotoxic anti-cancer agents. An adenoviral-based expression system was employed to target high levels of N-methylpurine DNA glycosylase (MPG) to either the mitochondria or nucleus of primary astrocytes to test the hypothesis that an imbalance in mitochondrial BER would result in increased alkylation sensitivity in astrocytes. These studies revealed that increasing MPG activity dramatically increased BER kinetics in the mitochondria and nucleus. Modulating MPG activity in the mitochondria appeared to have little effect on alkylation sensitivity in primary astrocytes. However, increased nuclear MPG activity was lethal in astrocytes treated with methylnitrosourea (MNU). Caspase-3 cleavage was not detected in these transduced astrocytes, and both TUNEL and Annexin-V assays indicated that these alkylation sensitive astrocytes do not undergo a typical programmed cell death in response to MNU. An assessment of the steady-state levels of key apoptotic proteins suggests that the apoptotic machinery of astrocytes is adapted more for cell survival, thus astrocytes are less vulnerable to pro-apoptotic stimuli compared to other CNS cell types exhibiting greater sensitivity to chemotherapeutic agents. However, targeting this MPG-mediated alkylation sensitivity to astrocytic cells in combination with alkylating drugs and radiation might be a useful strategy for improving the prognosis for patients with astrocytoma.