Role of polymorphic human CYP2B6 in cyclophosphamide bioactivation

Abstract

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.

Influence of pharmacogenetics on response and toxicity in breast cancer patients treated with doxorubicin and cyclophosphamide

“…CYP2B6 variants have been associated with both decreased (Lang et al, 2001), (Jinno et al, 2003) and unchanged (Xie et al, 2003) protein expression compared with the wild-type gene. Increased specific activity associated with the variants has also been observed (Lang et al, 2001;Jinno et al, 2003).…”

Association of cyclophosphamide pharmacokinetics to polymorphic cytochrome P450 2C19

“…Some investigators identified CYP2B6 16,17 as the major in vitro catalyst of 4-hydroxy CP, others characterized CYP2C9 as low K m hydroxylase, 12,14 and CYP2C19 as high K m hydroxylase, 12,13 but found also that CYP3A4/5 14 was an important enzyme for CP activation. The polymorphic GSTA1, however, contributes to the detoxification of reactive intermediates, therefore GSTA1 could be possibly associated with side effects of CP treatment.…”

“…11 As activation and elimination of CP is mediated by various polymorphic drugmetabolizing enzymes (Figure 1), the question arises whether hereditary polymorphism could modulate the pharmacokinetics of CP. CP is believed to be predominantly activated to 4-hydroxyphosphamide by the polymorphic cytochrome P450 (CYP) enzymes CYP2C9 [12][13][14] and CYP2B6, [15][16][17] but also by CYP2C19 12,13 and CYP3A. 14,15 CYP3A4, CYP3A5 and CYP3A7 contribute to the heterogeneous expression of the human CYP3A family.…”

Pharmacogenomics and personalized medicine: a review focused on their application in the Chinese population

“…Cyclophosphamide bioactivation was reported to be enhanced in the CYP2B6*6/*6 genotype in vitro and in vivo [85][86][87] . However, contradictory or negative results were presented in other studies of the association between CYP2B6 and pharmacokinetics/clinical outcome [88][89][90][91] .…”