METABOLISM OF CYCLOPHOSPHAMIDE BY LIPOXYGENASES

Cyclophosphamide (CP) can be cooxidized by several peroxidases. A numb er of xenobiotics also undergo cooxidation during metabolism of unsatu rated fatty acids by lipoxygenases. The current study examined linolei c acid-dependent cooxidation of CP by soybean lipoxygenase and rabbit 15-lipoxygenase. Incubations of CP (0-5 mM) with soybean lipoxygenase (0-1 x 10(4) units) and linoleic acid (0-5 mM) resulted in the generat ion of acrolein, an easily detected breakdown product of the initial u nstable hydroxylated CP metabolite. Acrolein formation was dependent o n time and concentrations of the enzyme, linoleic acid, and CP. The fo rmation of acrolein was complete by 3 min and was favored at pHs above 7. The lipoxygenase inhibitors BW755C (0.1 mM), BWA4C (0.1 mM), and p heridione (0.01 mM) significantly inhibited the metabolism of CP to ac rolein by 92, 66, and 57%, respectively. Vitamin E (0.1 mM), superoxid e dismutase (10 units), and nordihydroguaiaretic acid (0.1 mM) signifi cantly decreased acrolein generation by 74, 76, and 77%, respectively. Glutathione (1 mM) significantly decreased measurable acrolein by 80% , perhaps by binding to this reactive aldehyde. One mM cysteine, merca ptoethane sulfonic acid, glutathione disulfide, or albumin also decrea sed measurable acrolein (16-36%), but significantly less efficiently t han glutathione. Incubation of rabbit leukocyte 15-lipoxygenase with l inoleic acid and CP also resulted in the generation of acrolein. These results indicate that lipoxygenases can cooxidize CP. The inhibitory effects of antioxidants suggest that this catalysis may involve free r adicals.