3-AMINOBENZAMIDE AND OR O-6-BENZYLGUANINE EVALUATED AS AN ADJUVANT TOTEMOZOLOMIDE OR BCNU TREATMENT IN CELL-LINES OF VARIABLE MISMATCH REPAIR STATUS AND O-6-ALKYLGUANINE-DNA ALKYLTRANSFERASE ACTIVITY/

O-6-benzylguanine (O-6-BG) and 3-aminobenzamide (3-AB) inhibit the DNA repair proteins O-6-alkylguanine-DNA alkyltransferase (AGT) and poly( ADP-ribose) polymerase (PARP) respectively. The effect of O-6-BG and/o r 3-AB on temozolomide and 1, 3-bis(2-chloroethyl)-nitrosourea (BCNU) cytotoxicity, was assessed in seven human tumour cell lines: six with an AGT activity of >80 fmol mg(-1) protein (Mer(+)) and one with an AG T activity of <3 fmol mg(-1) protein (Mer(-)). Three of the Mer(+) cel l lines (LS174T, DLD1 and HCT116) were considered to exhibit resistanc e to methylation by a mismatch repair deficiency (MMR(-)), each being known to exhibit microsatellite instability, and DLD1 and HCT116 havin g well-characterised defects in DNA mismatch binding. Potentiation was defined as the ratio between an IC50 achieved without and with a part icular inhibitor treatment. Temozolomide or BCNU cytotoxicity was not potentiated by either inhibitor in the Mer(-) cell line. Preincubation with O-6-BG (100 mu M for 1 h) was found to potentiate the cytotoxici ty of temozolomide by 1.35- to 1.57-fold in Mer(+)/MMR(+) cells, but h ad no significant effect in Mer(+)/MMR(-) cells. In comparison, O-6-BG pretreatment enhanced BCNU cytotoxicity by 1.94- to 2.57-fold in all Mer(+) cell lines. Post-incubation with 3-AB (2 mM, 48 h) potentiated temozolomide by 1.35- to 1.59-fold in Mer(+)/MMR(+) cells, and when co mbined with O-6-BG pretreatment produced an effect which was at least additive, enhancing cytotoxicity by 1.97- to 2.16-fold. 3-AB treatment also produced marked potentiation (2.20- to 3.12-fold) of temozolomid e cytotoxicity in Mer(+)/MMR(-) cells. In contrast, 3-AB produced marg inal potentiation of BCNU cytotoxicity in only three cell lines (1.19- to 1.35-fold), and did not enhance the cytotoxicity of BCNU with O-6- BG treatment in any cell line. These data suggest that the combination of an AGT and PARP inhibitor may have a therapeutic role in potentiat ing temozolomide activity, but that the inhibition of poly(ADP-ribosyl )ation has little effect on the cytotoxicity of BCNU.