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Metabolite complex formation of orphenadrine with cytochrome P450. Involvement of CYP2C11 and CYP3A isozymes
Abstract:
Expression and inhibition of cytochrome P450 (CYP) isozymes capable of forming an orphenadrine metabolite complex were studied in microsomes of untreated and inducer-treated male and female rats. High levels of complex-forming isozymes were found in microsomes of untreated male as compared to female rats. Treatment of male rats with several P450 inducers did not considerably increase the extent of in vitro complex formation. In female rats, however, phenobarbital or dexamethasone treatments led to pronounced induction. The isozyme specificity of complex formation was investigated by several approaches including: 1. inhibition by orphenadrine of isozyme-specific P450 activities, such as hydroxylation of testosterone, O-dealkylation of pentoxy-and ethoxyresorufin and complex formation with triacetyloleandomycin (TAO), 2. inhibition of orphenadrine complex formation by metyrapone, TAO, and cimetidine, and 3. correlation of complex levels with immunochemically, enzymatically, or spectroscopically determined amounts of P450 isozymes. Our data suggest that CYP2C11, a CYP3A isozyme and an unidentified P450 species are involved in complex formation with orphenadrine, but exclude the involvement of CYP1A1/2 and CYP2B1/2. The capability of CYP2C11 to form a metabolite complex with orphenadrine is strongly suggested for the following reasons: 1. Efficient inhibition of testosterone 2 alpha- and 16 alpha-hydroxylation by complex formation with orphenadrine in microsomes of untreated male rats, 2. high expression of orphenadrine-complexing isozymes in untreated male compared to female rats, 3. specific inhibition of in vitro complex formation by cimetidine, 4. suppression of complex-forming isozymes by 3-methylcholanthrene and beta-naphthoflavone, and 5. concomitant induction of complex-forming isozymes, immunodetectable CYP2C11, and testosterone 2 alpha-hydroxylase by stanozolol. That at least one, but not all, CYP3A isozymes is involved in complex formation is concluded from inhibition experiments with TAO that show that orphenadrine complexation can be significantly inhibited in microsomes of dexamethasone-treated, but not in microsomes of untreated rats. Furthermore, complex formation with TAO is not inhibited by orphenadrine in microsomes of phenobarbital (PB)-treated rats. In PB-treated female rats, a further unidentified complex-forming isozyme can be detected that is not inhibited by complex formation with TAO.
Insights
This study identifies cytochrome P450 (CYP) isozymes involved in orphenadrine metabolite complex formation, highlighting CYP2C11 and CYP3A in male rats. Sex differences in CYP expression influence these interactions.
Area of Science:
- Biochemistry
- Pharmacology
- Toxicology
Background:
- Cytochrome P450 (CYP) enzymes play crucial roles in drug metabolism and detoxification.
- Orphenadrine can form metabolite- সcomplexes with certain CYP isozymes, impacting drug interactions.
- Understanding sex-based differences in CYP activity is vital for personalized medicine.
Purpose of the Study:
- To investigate the specific cytochrome P450 (CYP) isozymes responsible for forming orphenadrine metabolite complexes.
- To explore the influence of sex and inducer treatments on CYP-orphenadrine complex formation.
- To elucidate the roles of CYP2C11 and CYP3A isozymes in these interactions.
Main Methods:
- Studied CYP isozyme activity in male and female rat liver microsomes.
- Utilized in vitro complex formation assays with orphenadrine.
- Employed isozyme-specific inhibition studies using various P450 inhibitors (e.g., cimetidine, TAO).
- Correlated complex levels with immunochemical and enzymatic quantification of CYP isozymes.
Main Results:
- Higher levels of orphenadrine-complexing isozymes were observed in male rats compared to females.
- Phenobarbital and dexamethasone treatments induced complex formation in female rats.
- CYP2C11 and at least one CYP3A isozyme were identified as key players in orphenadrine complex formation.
- CYP1A1/2 and CYP2B1/2 were excluded from this interaction.
Conclusions:
- CYP2C11 is strongly implicated in orphenadrine metabolite complex formation, particularly in male rats.
- Specific CYP3A isozymes contribute to orphenadrine complexation, with induction observed in females.
- Sex-dependent expression and inducibility of CYP isozymes significantly influence orphenadrine metabolism and potential drug interactions.