Abstract

The exposure of cells to O6-methylguanine or O6-benzylguanine is known to reduce the enzymatic activity of O6-alkylguanine-DNA alkyltransferase, which leads to a sensitivity enhancement to chloroethylnitrosourea cytotoxic effects. The main disadvantage of the guanine derivatives is their low water solubility, which makes their formulation difficult for clinical use in humans. To overcome this problem, water-soluble O6-alkylguanine-DNA alkyltransferase inhibitors have been synthesized and their ability to increase the chloroethylnitrosourea potency in vitro and in vivo was evaluated. Four water-soluble molecules (O6-methyl-N-acetylguanosine; O6-methyl-N-acetyldeoxyguanosine; O6-benzyl-N-acetylguanosine, BNAG; and O6-benzyl-N-acetyldeoxyguanosine, BNADG) were tested for sensitivity of M4Beu cells to N'-(2-chloroethyl]-N[2-(methylsulfonyl)ethyl]-N'-nitrosourea (cystemustine) based on the colony-forming ability of M4Beu melanoma cells. The cell sensitivity to cystemustine was increased by benzylated derivative pretreatment but not with methylated derivative pretreatment. Furthermore, BNAG or BNADG pretreatment followed by cystemustine was less cytotoxic than BNAG or BNADG given simultaneously and followed 24 hr later by BNAG or BNADG. Comparative studies performed with O6-benzylguanine on the same model showed that this inhibitor was effective at lower concentrations than the corresponding guanosine or deoxyguanosine analogs. Preliminary pharmacological assays were carried out in nude mice bearing the M4Beu tumor to determine whether the BNAG-cystemustine combination has greater antitumor activity than cystemustine alone. Simultaneous i.p. injection of 200 mg/kg of BNAG and 15 mg/kg of cystemustine followed by an i.p. injection 4 hr later of 200 mg/kg of BNAG led to a significant enhancement of inhibition of tumor growth.