Degradation of rat hepatic cytochrome P-450 heme by 3,5-dicarbethoxy-2, 6-dimethyl-4-ethyl-1,4-dihydropyridine to irreversibly bound protein adducts

Abstract

Administration of 3,5-dicarbethoxy-2,6-dimethyl-4-ethyl-1,4-dihydropyridine (DDEP) (a structural analog of the dihydropyridine Ca 2+ antagonists) to untreated, phenobarbital-, or dexamethasone-pretreated rats results in time-dependent losses of hepatic cytochrome P-450 content. Functional markers for various cytochrome P-450 isozymes have permitted the identification of P-450 h, P-450 PB-1/k, and P-450 p as the isozymes inactivated preferentially by the drug. DDEP-mediated cytochrome P-450 destruction may be reproduced in vitro, is most prominent after pretreatment of rats with dexamethasone, pregnenolone 16α-carbonitrile or phenobarbital, and is blocked by triacetyloleandomycin. These findings together with the observation that DDEP markedly inactivates hepatic 2β- and 6β-testosterone hydroxylase and erythromycin N-demethylase tend to indict the steroid-inducible P-450 p isozyme as a key protagonist in this event. The precise mechanism of such DDEP-mediated P-450 p heme destruction is unclear, but involves prosthetic heme alkylation of the apocytochrome at its active site in what appears to be a novel mechanism-based “suicide” inactivation. Such inactivation appears to involve fragmentation of the heme to reactive metabolites that irreversibly bind to the protein, but the chemical structure of the heme-protein adducts is yet to be established. Intriguingly, such DDEP-mediated P-450 p destruction in vivo also results in accelerated loss of immunochemically detectable apocytochrome P-450 p. It remains to be determined whether or not this loss is due to enhanced proteolysis triggered by the structural modification of the apocytochrome.