cAMP-dependent phosphorylation of CYP2B1 as a functional switch for cyclophosphamide activation and its hormonal control in vitro and in vivo.
An important feature of cytochrome P450 (CYP) 2B1 is its high ability to convert the prodrug cyclophosphamide (CPA) to therapeutically cytotoxic metabolites, resulting in interstrand DNA-cross-linking and cell death. We have examined whether and how the phosphorylation of CYP2B1 influences CPA metabolic activation in vitro and in vivo. We found first that only part of the total CYP2B1 pool undergoes phosphorylation. This part is fully inactivated. Second, phosphorylation of CYP2B1 in intact hepatocytes reduced by up to 75% toxification of CPA to mutagenic metabolites (totally dependent on the same preferentially CYP2B-catalyzed 4-hydroxylation of CPA as is the generation of highly cytotoxic species). Third, the phosphoacceptor-serine 128 of CYP2B1 in the consensus sequence for interaction with the protein kinase A represents an on/off switch for the activation of CPA depending on the phosphorylation conditions in the cell. Fourth, evidence is presented that the above-described events also occur in vivo. In conclusion, a successful therapy with CPA, helped by forced expression of CYP2B1 in tumor cells (as recently proposed) will, in addition, be profoundly modified by its phosphorylation status.
Oesch-Bartlomowicz
B
B
Institute of Toxicology, University of Mainz, Mainz, Germany. oeschb@mail.uni-mainz.de
Richter
B
B
Becker
R
R
Vogel
S
S
Padma
P R
PR
Hengstler
J G
JG
Oesch
F
F
D000818:Animals; D018906:Antineoplastic Agents, Alkylating / Q000378:metabolism*; D001711:Biotransformation; D000242:Cyclic AMP / Q000502:physiology*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000502:physiology; D003520:Cyclophosphamide / Q000378:metabolism*; D019362:Cytochrome P-450 CYP2B1 / Q000737:chemistry / Q000378:metabolism*; D005260:Female; D005934:Glucagon / Q000494:pharmacology; D022781:Hepatocytes / Q000187:drug effects / Q000378:metabolism; D008297:Male; D009153:Mutagens / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000502:physiology; D010766:Phosphorylation; D051381:Rats; D017207:Rats, Sprague-Dawley; D012694:Serine; D012737:Sex Factors; D013329:Structure-Activity Relationship; D013997:Time Factors
D018906:Antineoplastic Agents, Alkylating; D009153:Mutagens; D012694:Serine; D003520:Cyclophosphamide; D005934:Glucagon; D000242:Cyclic AMP; D019362:Cytochrome P-450 CYP2B1; D017868:Cyclic AMP-Dependent Protein Kinases; D010749:Phosphoprotein Phosphatases