A monoclonal antibody to rat liver cytochrome P450 IIC11 strongly and regiospecifically inhibits constitutive benzo[a]pyrene metabolism and DNA binding

The monoclonal antibody MAb 1‐68‐11, prepared to constitutive cytochrome P450 IIC11 (2c/RLM5) from male Sprague‐Dawley rat liver, was used to study the contribution of the class of cytochrome P450s epitopically related to P450 IIC11 to the regiospecific metabolism of benzo[a]pyrene (BP) and its binding to DNA. The effect of MAb 1‐68‐11 was determined on the conversion of BP to BP‐9,10‐dihydrodiol, BP‐7,8‐dihydrodiol, BP‐4,5‐dihydrodiol, BP phenols, and BP quinones, and on the P450‐dependent DNA binding catalyzed by P450 in microsomes from uninduced male and female Wistar and Sprague‐Dawley rat livers, as well as 3‐methylcholanthrene‐ and phenobarbital (PB)‐induced male Wistar rat livers. In liver microsomes from untreated male rats, MAb 1‐68‐11 inhibited BP‐9,10‐dihydrodiol formation by 80%; in liver microsomes from untreated female rats, the inhibition was 100%. BP‐7,8‐dihydrodiol formation was inhibited from 38 to 77% in microsomes from males and 50% in those from females. In microsomes from PB‐induced rats, inhibition of the 9,10‐dihydrodiol and the 7,8‐dihydrodiol was 90% and 73%, respectively, whereas BP‐4,5‐dihydrodiol formation was enhanced 80%. In microsomes from 3‐methylcholanthrene‐treated rats, no inhibition of MAb 1‐68‐11 was observed on either the metabolism of BP or its binding to DNA. In contrast, the binding of BP to DNA was completely inhibited by MAb 1‐68‐11 in microsomes from uninduced male Wistar rats and 70% in PB‐induced microsomes. ³²P‐postlabeling analysis showed that formation of the major stable adduct, BP diol epoxide bound at C‐10 to the 2‐amino of deoxyguanosine, was strongly inhibited in uninduced and PB‐induced microsomes. Formation of the major labile BP‐DNA adduct 7‐(benzo[a]pyren‐6‐yl) guanine (BP‐N7Gua) was inhibited about 60% in microsomes from untreated male Wistar rats. These results show that MAb 1‐68‐11 regiospecifically inhibits cytochrome P450 IIC11 and epitopically related P450s that metabolize BP at the 7,8 and 9,10 positions. MAb 1‐68‐11 also inhibits enzyme‐catalyzed binding of BP to DNA in the specific formation of BP‐N7Gua and adducts detected by the ³²P‐postlabeling technique.