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Effect of cimetidine on hepatic cytochromes P450 1A and P450 2C in male rats Law, Eva Yuk Wa


Cimetidine, an H₂-antagonist, is an inhibitor of hepatic cytochromes P450 in humans and male rats, both in vivo and in vitro. Results of early in vitro studies with rat hepatic microsomes showed that cimetidine, at concentrations in the millimolar range, inhibits many cytochrome P450-mediated enzyme activities and that the inhibition appears to be reversible. Thus, it has been thought that cimetidine is a general and reversible inhibitor of the cytochromes P450. However, these early in vitro studies were performed at cimetidine concentrations that were 100-1000-fold higher than the serum cimetidine concentrations present in rats and humans. As well, there is indirect evidence from in vivo studies in both species to suggest that cimetidine does not inhibit certain cytochromes P450. In male rats, in vivo administration of cimetidine results in inhibition of hepatic CYP2C11 and some unidentified cytochrome(s) P450, but not CYP2A1, CYP2B1/2 or CYP3A1/2. In addition, preincubation of rat microsomes in vitro with cimetidine and NADPH prior to the intiation of substrate oxidation increases the potency of its inhibition of CYP2C11 and results in a pattern of inhibition similar to that observed in vivo. These observations indicate that cimetidine exerts its inhibitory effect in vivo by acting as a catalysis-dependent inhibitor. It has been of interest to identify other rat cytochromes P450 that are affected by cimetidine and there is indirect evidence from the literature to suggest that cimetidine is an inhibitor of rat hepatic CYP1A1/2. The primary objective of the present study was to investigate the effect of cimetidine on other hepatic cytochromes P450 in male rats, in particular CYP1A1/2. The effects of in vivo cimetidine on methoxyresorufin and ethoxyresorufin O-dealkylase (MROD and EROD) activities were examined in microsomes from ß-napthoflavone (BNF)-induced and uninduced rats that had been administered cimetidine (150 mg/kg) or saline 90 min before decapitation. Phenacetin inhibition and immunoinhibition studies were conducted to determine the contribution of CYP1A and CYP2C enzymes to these activities. The effects of in vitro cimetidine on these activities were investigated in the absence or presence of a preincubation step. In vivo cimetidine had no effect on MROD or EROD activity in microsomes from BNF-induced rats. In microsomes from uninduced rats, in vivo cimetidine inhibited EROD activity, but appeared not to have an effect on MROD activity. Results of the phenacetin inhibition and antibody inhibition studies suggest that, in microsomes from BNF-induced rats, these activities are mainly due to CYP1A1 (with some contribution from CYP1A2) and that, in microsomes from uninduced rats, they are largely mediated by a CYP2C enzyme(s) other than CYP2C11. The addition of cimetidine in vitro to microsomes from BNF-induced or uninduced rats immediately prior to the initation of substrate oxidation resulted in inhibition of these activities with IC₅₀ values ranging from 0.25 - 4 mM. Preincubation of rat liver microsomes with cimetidine in the presence of NADPH for 15 min prior to the initiation of substrate oxidation had no effect on inhibition of MROD or EROD activity in BNF-induced rats. However, under the same conditions, preincubation enhanced the potency of the inhibition of EROD activity in uninduced rats by cimetidine 16-fold, but had no effect on MROD activity. These results suggest that, in male rats, cimetidine does not inhibit CYP1A1 and has little or no effect on CYP1A2, but does inhibit another CYP2C enzyme(s) in addition to CYP2C11. As in the case of CYP2C11, the inhibition of the CYP2C enzyme(s) responsible for EROD activity in uninduced rats appears to be mediated by a catalysis-dependent mechanism. In humans, cimetidine inhibits the hepatic clearance of theophylline. Because CYP1A2 has been implicated as being responsible for at least part of the metabolism of theophylline in human microsomes, it is possible that cimetidine is an inhibitor of human CYP1A2. Another objective of the present study was to determine whether cimetidine was a catalysis dependentinhibitor of human hepatic CYP1A2. EROD activity was used an indicator of human CYP1A2. Preincubation of human liver microsomes with cimetidine in the presence of NADPH prior to the initiation of substrate oxidation had no effect on inhibition of EROD activity. Although results of this preliminary experiment suggest that cimetidine has no catalysis-dependent effect on human CYP1A2, further studies are required to investigate the possible effects of cimetidine on this enzyme. Based on the observation from the present investigation that cimetidine inhibits another CYP2C enzyme(s) (possibly CYP2C6) in addition to CYP2C11 in male rats and the fact that human and rat cytochromes P450 are immunochemically related, it is hypothesized that cimetidine also inhibits one or more of the CYP2C enzymes in humans. Future studies with human cytochromes P450 should investigate the possible effects of cimetidine on CYP2C9 and CYP2C19.

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