title: Cytochrome P450 Oxidoreductase Influences CYP2B6 Activity in Cyclophosphamide Bioactivation.
abstract: BACKGROUND
  Cyclophosphamide is commonly used as an important component in conditioning prior to hematopoietic stem cell transplantation, a curative treatment for several hematological diseases. Cyclophosphamide is a prodrug activated mainly by cytochrome P450 2B6 (CYP2B6) in the liver. A high degree of inter- and intra-individual variation in cyclophosphamide kinetics has been reported in several studies.
  
  
  METHODS
  Hydroxylation of cyclophosphamide was investigated in vitro using three microsomal batches of CYP2B6*1 with different ratios of POR/CYP expression levels. Twenty patients undergoing hematopoietic stem cell transplantation were also included in the study. All patients received an i.v. infusion of cyclophosphamide (60 mg/kg/day, for two days) as a part of their conditioning. Blood samples were collected from each patient before cyclophosphamide infusion, 6 h after the first dose and before and 6 h after the second dose. POR gene expression was measured by mRNA analysis and the pharmacokinetics of cyclophosphamide and its active metabolite were determined.
  
  
  RESULTS
  A strong correlation between the in vitro intrinsic clearance of cyclophosphamide and the POR/CYP ratio was found. The apparent Km for CYP2B6.1 was almost constant (3-4 mM), while the CLint values were proportional to the POR/CYP ratio (3-34 μL/min/nmol CYP). In patients, the average expression of the POR gene in blood was significantly (P <0.001) up-regulated after cyclophosphamide infusion, with high inter-individual variations and significant correlation with the concentration ratio of the active metabolite 4-hydroxy-cyclophosphamide/cyclophosphamide. Nine patients were carriers for POR*28; four patients had relatively high POR expression.
  
  
  CONCLUSIONS
  This investigation shows for the first time that POR besides CYP2B6 can influence cyclophosphamide metabolism. Our results indicate that not only CYPs are important, but also POR expression and/or activity may influence cyclophosphamide bioactivation, affecting therapeutic efficacy and treatment related toxicity and hence on clinical outcome. Thus, both POR and CYP genotype and expression levels may have to be taken into account when personalizing treatment schedules to achieve optimal therapeutic drug plasma concentrations of cyclophosphamide.
authors.affiliation: Experimental Cancer Medicine (ECM), Department of Laboratory Medicine, Karolinska Institutet, Huddinge, Stockholm, Sweden.
authors.forename: Ibrahim
authors.initials: I
authors.lastname: El-Serafi
authors.affiliation: Experimental Cancer Medicine (ECM), Department of Laboratory Medicine, Karolinska Institutet, Huddinge, Stockholm, Sweden.
authors.forename: Parvaneh
authors.initials: P
authors.lastname: Afsharian
authors.affiliation: Cancer Center of Karolinska (CCK), Department of Oncology-Pathology, Karolinska Institutet, Solna, Stockholm, Sweden.
authors.forename: Ali
authors.initials: A
authors.lastname: Moshfegh
authors.affiliation: Experimental Cancer Medicine (ECM), Department of Laboratory Medicine, Karolinska Institutet, Huddinge, Stockholm, Sweden.|Department of Clinical Research Centre, Karolinska University Hospital-Huddinge, Stockholm, Sweden.
authors.forename: Moustapha
authors.initials: M
authors.lastname: Hassan
authors.affiliation: Department of Discovery Research, Medivir AB, Huddinge, Sweden.
authors.forename: Ylva
authors.initials: Y
authors.lastname: Terelius
mesh_terms: D000293:Adolescent; D000328:Adult; D002648:Child; D003520:Cyclophosphamide / Q000378:metabolism*; D065702:Cytochrome P-450 CYP2B6 / Q000378:metabolism*; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D006402:Hematologic Diseases / Q000201:enzymology / Q000378:metabolism / Q000628:therapy; D018380:Hematopoietic Stem Cell Transplantation; D006801:Humans; D008861:Microsomes / Q000201:enzymology / Q000378:metabolism; D008875:Middle Aged; D011355:Prodrugs / Q000378:metabolism; D055815:Young Adult
chemical_list: C000602470:POR protein, human; D011355:Prodrugs; D003520:Cyclophosphamide; D003577:Cytochrome P-450 Enzyme System; D065702:Cytochrome P-450 CYP2B6