Inhibitory Monoclonal Antibody to Human Cytochrome P450 2B6

Yang, Taoxi; Krausz, Kristopher W.; Shou, Magang; Yang, Shen K.; Buters, Jeroen; Gonzalez, Frank J.; Gelboin, Harry V.

doi:10.1016/s0006-2952(98)00018-5

Cited by 40 publications

(24 citation statements)

References 21 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…This percentage is higher than previously reported values (43, 3 24, 2 30% 1 ), and similar with 70, 9 85, 1 80, 7 or 90% 39 . The variation may also be due to other as yet unknown mutations or due to environmental factors.…”

Section: Discussionsupporting

confidence: 87%

“…4 Further characterization of CYP2B6 revealed higher mRNA and protein levels than previously observed. [5][6][7] Meanwhile, a number of structurally diverse classes of frequently used drugs have been recognized as CYP2B6 substrates: cyclophosphamide (CPA), 8 7-ethoxy-4-trifluoromethylcoumarin, 9 bupropion, 10 ifosphamide, 8,11,12 nicotine, 13 lidocaine, 14 S-mephenytoin, 15 verapamil, 14 amitriptyline, 4 diazepam, 16 7-benzyloxyresorufin, 16 nevirapine, 4 S-mephobarbital, 17 artemisinin, 18 and propofol 19 .…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Role of polymorphic human CYP2B6 in cyclophosphamide bioactivation

et al. 2003

View full text Add to dashboard Cite

The role of polymorphic CYP2B6 in cyclophosphamide (CPA) bioactivation was investigated in human liver microsomes. A total of 67 human liver specimens were first genotyped with respect to the CYP2B6*5 and CYP2B6*6 variant alleles. CYP2B6 apoprotein levels in 55 liver microsomal preparations were assessed by immunoblotting. 4-Hydroxy-CPA and hydroxy-bupropion were quantified by using HPLC and LC-MS, respectively. 7-Ethoxy-4-trifluoromethyl coumarin O-deethylase activity was measured fluorometrically. The frequencies of CYP2B6*5 and CYP2B6*6 mutant alleles were 9.0 and 16.4%, respectively. CYP2B6 protein expression was detected in 80% of the samples, with a large variation (0.003-2.234, arbitrary units). There was a high correlation between CYP2B6 apoprotein content and CPA 4-hydroxylation (n ¼ 55, r ¼ 0.81, Po0.0001). When based on the CYP2B6 apoprotein levels, the *6 carriers had significantly higher CPA 4-hydroxylation (Po0.05). CPA 4-hydroxylation also correlated significantly with other CYP2B6-specific reactions (n ¼ 20, Po0.0001). V max and K m for CPA 4-hydroxylation in recombinant CYP2B6 enzyme were 338 nmol/min/nmol enzyme and 1.4 mM, respectively. CYP2B6 showed much higher in vitro intrinsic clearance than previously observed in recombinant CYP2C19 and CYP2C9 variants in yeast expression system. Our results demonstrate that the polymorphic CYP2B6 is a major enzyme in the bioactivation of CPA. Moreover, we identified a strong impact of CYP2B6*6 on CPA 4-hydroxylation.

show abstract

Section: Discussionsupporting

confidence: 87%

Section: Introductionmentioning

confidence: 99%

Role of polymorphic human CYP2B6 in cyclophosphamide bioactivation

et al. 2003

View full text Add to dashboard Cite

show abstract

“…This should include enough low and high antibody concentrations to establish the titration curve. From the inhibition curve, one can assess the potency of the antibody from the slope and the relative contribution of this particular P450 isoform from maximal inhibition (Guengerich, 1988;Wang and Lu, 1997;Yang et al, 1998;Granvil et al, 2002). If there is only one P450 involved in the metabolism, greater than 90% inhibition is expected with pooled and individual human liver microsomes.…”

Section: Current Approaches Used In P450 Identificationmentioning

confidence: 99%

“…Because of the demand of resources and expertise, not every pharmaceutical company can produce their own monoclonal antibodies. With the exception of a limited number of publications (Guengerich, 1988;Wang and Lu, 1997;Yang et al, 1998;Granvil et al, 2002), most of the antibody inhibition studies used only a single antibody concentration to demonstrate inhibition of the reaction of interest, perhaps due to limited supply of antibodies. If the inhibition at this selected antibody concentration is greater than 90%, one can conclude that the particular P450 isoform is the major enzyme responsible for the metabolism.…”

Section: Current Approaches Used In P450 Identificationmentioning

confidence: 99%

“…Using dmd.aspetjournals.org inhibitory monoclonal antibodies, Gelboin et al (1999) reported very large variability in the contribution of individual P450 enzyme to the metabolism of various drugs. For example, CYP2D6 contributes to 10 to 70% of bufuralol 1Ј-hydroxylase activity in different liver samples (Gelboin et al, 1997) while CYP2B6 contributes to 8 to 42% of phenanthrene metabolism in all liver microsomal preparations examined (Yang et al, 1998). Thus, when conducting P450 in vitro reaction phenotyping, it is desirable to use pooled human liver microsomes to obtain a composite picture on the P450 enzymes responsible for the metabolism of compound of interest.…”

Section: Variable Cytochrome P450 Contribution and The Hidden Factormentioning

confidence: 99%

See 1 more Smart Citation

Cytochrome P450 In Vitro Reaction Phenotyping: A Re-evaluation of Approaches Used for P450 Isoform Identification

Lu¹,

Wang²,

Lin³

2003

Drug Metab Dispos

View full text Add to dashboard Cite

ABSTRACT:Marker substrates, chemical inhibitors, and inhibitory antibodies are important tools for the identification of cytochrome P450 (P450) isoform responsible for the metabolism of therapeutic agents in vitro. In view of the versatile and nonspecific nature of P450 enzymes, many of the marker substrates and chemical inhibitors used for P450 in vitro reaction phenotyping are isoform selective but not specific. Recently, the use of marker substrate and chemical inhibitors in CYP2D6 in vitro reaction phenotyping was questioned by Granvil et al. (2002). In comparison of a panel of 15 recombinant P450 enzymes, they found that in addition to CYP2D6, CYP1A1 is also capable of catalyzing the formation of 4-hydroxylated metabolite of debrisoquine and that the intrinsic clearance of debrisoquine by CYP2D6-mediated 4-hydroxylation is only about twice that by CYP1A1. In their study, they have also demonstrated that quinidine inhibits both CYP2D6-and CYP1A1-mediated debrisoquine 4-hydroxylation. In view of these important findings, we have reevaluated various approaches used to identify P450 isoform(s) responsible for the metabolism of therapeutic agents. While acknowledging the value of inhibitory antibodies in P450-phenotyping studies, it is our opinion that in well conducted in vitro experiments, isoform-selective chemical inhibitors can also provide valuable and reliable information. Hopefully, future efforts may produce even better P450 isoform-selective marker substrates and inhibitors.For most drugs, biotransformation is the major route of elimination, and oxidative metabolism by P450 1 enzymes is a common metabolic pathway (Rendic, 2002). Therefore, it is important to assess the relative contribution of metabolic pathways to the overall elimination processes and to identify the P450 isoforms responsible for oxidative reactions. In vitro P450 phenotyping has proven to be very successful in predicting the potential of drug interactions and the polymorphic impact on drug disposition (Lin and Lu, 1998;Dahl, 2002). As a result, pharmaceutical companies routinely include in vitro reaction P450 phenotyping in the drug candidate selection process.Ever since the pioneering work of Smith and coworkers (Mahgoub et al., 1977;Sloan et al., 1978), the 4-hydroxylation of debrisoquine has been recognized as a marker reaction of human CYP2D6. Urinary metabolic ratio defined as debrisoquine to 4-hydroxydebrisoquine measured following an oral dose of debrisoquine is generally considered as an accurate assessment of an individual's metabolic capacity of CYP2D6. Based on the metabolic ratios, individuals are phenotypically classified as being poor metabolizer (PM), extensive metabolizer (EM), or ultra-rapid metabolizer (UM) phenotypes, each displaying characteristic pharmacokinetic patterns and clinical outcomes (Eichelbaum and Gross, 1990;Dahl et al., 1995;Meyer and Zanger, 1997). The debrisoquine pharmacokinetic profile of an individual with EM phenotype can be changed to a pattern similar to that of an individual with PM phenotyp...

show abstract

Enzymes in addition to CYP3A4 and 3A5 mediateN-demethylation of dextromethorphan in human liver microsomes

Wang

Unadkat

1999

Biopharm. Drug Dispos.

View full text Add to dashboard Cite

Both indinavir and troleandomycin (CYP3A inhibitors) are incapable of completely inhibiting dextromethorphan metabolism to 3‐methoxymorphinan in human liver microsomes. It is hypothesized that CYPs in addition to CYP3A4 and 3A5 contribute to this biotransformation. The effect of CYP‐selective inhibitors on the residual 3‐methoxymorphinan activity in human liver microsomes (i.e. in the presence of 30 μm indinavir, a selective CYP3A4 and 3A5 inhibitor) was measured to identify these enzymes. At this concentration, indinavir completely inhibited the formation of 3‐methoxymorphinan by rCYP3A4 and rCYP3A5. In addition, the formation kinetics of 3‐methoxymorphinan in rCYPs was measured. Only CYP2B6, 2C8 and 2C18 were considered likely candidates as contributors to residual 3‐methoxymorphinan activity. The residual 3‐methoxymorphinan activity was highly correlated with CYP2B6 activity as measured by CYP2B6 antibody (r2=0.90, p<0.001) and by orphenadrine (r2=0.97, p<0.001), but was not correlated (r2=0.12, p>0.05) with CYP2C8 activity. Collectively, these findings suggest that CYP2B6 is a major contributor towards residual 3‐methoxymorphinan activity, while CYP2C8 and 2C18 are either minor contributors or do not contribute to this metabolic process. Copyright © 1999 John Wiley & Sons, Ltd.

show abstract

Inhibitory Monoclonal Antibody to Human Cytochrome P450 2B6

Cited by 40 publications

References 21 publications

Role of polymorphic human CYP2B6 in cyclophosphamide bioactivation

Role of polymorphic human CYP2B6 in cyclophosphamide bioactivation

Cytochrome P450 In Vitro Reaction Phenotyping: A Re-evaluation of Approaches Used for P450 Isoform Identification

Enzymes in addition to CYP3A4 and 3A5 mediateN-demethylation of dextromethorphan in human liver microsomes

Contact Info

Product

Resources

About