Inhibition of O6-methylguanine-DNA methyltransferase by glucose-conjugated inhibitors:: Comparison with nonconjugated inhibitors and effect on fotemustine and temozolomide-induced cell death

The DNA repair protein O-6-methylguanine-DNA methyltransferase (MGMT) is an important suicide enzyme involved in the defense against O-6-alkylating mutagens. It also plays a role in the resistance of tumors to anticancer drugs targeting the O-6-position of guanine, such as temozolomide and fotemustine. Several potent MGMT inhibitors have been developed sensitizing cells to O-6-alkylating agents. Aimed at targeting MGMT inhibitors to tumor cells, we synthesized MGMT inhibitory compounds conjugated with glucose to improve uptake in tumor cells. Here, we compared O-6-benzylguanine, O-6-2-fluoropyridinylmethylguanine (O(6)FPG), O-6-3-iodobenzylguanine, O-6-4-bromothenylguanine, and O-6-5-iodothenylguanine with the corresponding C8-linker beta-D-glucose derivatives. All glucose conjugated inhibitors were 3- to 5-fold less effective than the corresponding nonconjugated drugs as to MGMT inhibition that was measured in cell extracts ( in vitro) and cultivated HeLaS3 cells (in vivo). Except for O(6)FPG, IC50 values of the guanine derivatives applied in vitro and in vivo were correlated. A similar correlation was not obvious for the corresponding glucosides, indicating differences in cellular uptake. C8-alpha-D-glucosides were less effective than beta-glucosides. From the newly developed glucose-conjugated inhibitors tested, O-6-4-bromothenylguanine-C8-beta-D-glucoside (O(6)BTG-C8-betaGlu) was most potent in inhibiting MGMT both in vitro and in vivo. At a concentration of 0.1 muM, it inhibited cellular MGMT to completion. It was not toxic, even when applied chronically to cells at high dose ( up to 20 muM). O(6)BTG-C8-betaGlu strongly potentiated the killing effect of fotemustine and temozolomide, causing reversal from MGMT+ to MGMT - phenotype. Therefore, O(6)BTG-C8-betaGlu seems to be especially suitable for approaching MGMT inhibitor targeting in tumor therapy.