Abstract B63: Targeting cancer cells with a glucose-conjugated DNA repair inhibitor.

Abstract

Abstract Alkylating agents are important chemotherapeutic drugs used for the treatment of several types of cancers, including brain tumors, melanoma and lymphoma. These chemotherapeutic agents have a strong affinity towards oxygen atoms in DNA giving rise to the important genotoxic DNA lesions O6-methylguanine and O6-chloroethylguanine, which are responsible for the cytotoxic effects of several alkylating anticancer drugs (e.g. temozolomide and lomustine). The DNA repair protein O6-methylguanine-DNA methyltransferase (MGMT) is considered as an important player of drug resistance because it removes these DNA adducts from the DNA. The MGMT protein restores guanine in the DNA by a suicide repair reaction leading to irreversible inactivation and degradation of the protein. Therefore depletion of MGMT renders cells more susceptible to treatment with alkylating anticancer drugs. There has been numerous attempts to inactivate MGMT with different inhibitors like O6-benzylguanine (O6-BG) or O6-(4-bromothenyl) guanine (O6-BTG, lomeguatrib) in glioblastoma and malignant melanoma cells. These strategies however have failed thus far, mainly due to the severe side effects associated with the systemic sensitization of the patients to chemotherapeutic agents. The application of targeting strategies for MGMT inhibitors therefore should improve chemotherapy by selective inhibition of MGMT in the tumor. In our previous work we have shown that O6-BG and O6-BTG bound to glucose is efficient in inhibiting MGMT on enzyme level and in the cell, causing sensitization to DNA alkylating N-nitrosoureas (Kaina et. al., 2004; 2010). Here we show, using a quantitative MGMT assay, that intracellular uptake of O6-benzylguanine-C8-glucose (O6-BG-Glu) occurs by a transporter-dependent mechanism and not through passive diffusion. We observed differences in the transport kinetics between several cell lines. Most cancer cell lines are able to take up the inhibitor, whereas normal cells only show marginal uptake of the glucose-conjugated inhibitor. The uptake is probably not depended on the glucose transporter GLUT since blocking of GLUT does not abolishes MGMT inhibition. Uptake of the glucose conjugated inhibitors in cancer cells leads to an increased DNA damage response, induction of apoptosis and significant decrease in surviving fraction after treatment with alkylating anticancer drugs, pointing to the usefulness of the glucose-mediated MGMT inhibitor targeting strategy.