Journal Article•
Enzymatic basis of cyclophosphamide activation by hepatic microsomes of the rat
01 Aug 1970-Journal of Pharmacology and Experimental Therapeutics (J Pharmacol Exp Ther)-Vol. 174, Iss: 2, pp 206-210
TL;DR: The data confirm and enhance the thesis that the activation of cyclophosphamide by liver is mediated through the mixed function oxidase system involved in the metabolism of drugs, steroids and carcinogens.
Abstract: Our data confirm and enhance the thesis that the activation of cyclophosphamide by liver is mediated through the mixed function oxidase system involved in the metabolism of drugs, steroids and carcinogens. This is based on the following lines of evidence. First, the enzyme is located primarily if not exclusively in liver microsomes. Second, the enzyme requires reduced triphosphopyridine nucleotide and molecular oxygen for full activity. Third, the enzyme is inhibited in vitro by compounds known to be metabolized by the mixed function oxidase system of liver microsomes such as hexobarbital and steroids and the kinetics of inhibition by hexobarbital is that of a competitive type. Fourth, the enzyme can be inhibited by carbon monoxide as well as other compounds known to interrupt the microsomal electron transport chain such as p -hydroxymercuribenzoate and cytochrome c . Fifth, the enzyme level is markedly increased by phenobarbital administration. Finally, there is a clear sex-related difference in concentration of this enzyme in the rat liver. Evidence is presented that the induction of cyclophosphamide-activating enzyme by phenobarbital can be correlated with an enhancement of the pharmacologic effect of cyclophosphamide in the laboratory animal.
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TL;DR: Variations in the balance between metabolic activation and inactivation of cyclophosphamide owing to autoinduction, dose escalation, drug-drug interactions and individual differences have been reported, suggesting possibilities for optimisation of cyclphosphamide therapy.
Abstract: Cyclophosphamide is an extensively used anticancer and immunosuppressive agent. It is a prodrug undergoing a complicated process of metabolic activation and inactivation. Technical difficulties in the accurate determination of the cyclophosphamide metabolites have long hampered the assessment of the clinical pharmacology of this drug. As these techniques are becoming increasingly available, adequate description of the pharmacokinetics of cyclophosphamide and its metabolites has become possible. There is incomplete understanding on the role of cyclophosphamide metabolites in the efficacy and toxicity of cyclophosphamide therapy. However, relationships between toxicity (cardiotoxicity, veno-occlusive disease) and exposure to cyclophosphamide and its metabolites have been established. Variations in the balance between metabolic activation and inactivation of cyclophosphamide owing to autoinduction, dose escalation, drug-drug interactions and individual differences have been reported, suggesting possibilities for optimisation of cyclophosphamide therapy. Knowledge of the pharmacokinetics of cyclophosphamide, and possibly monitoring the pharmacokinetics of cyclophosphamide in individuals, may be useful for improving its therapeutic index.
489 citations
TL;DR: Clinical correlation between kinetic data and efficacy and/or toxicity awaits studies evaluating the time course of specific cytotoxic metabolites, and conflicting data have been obtained in several animal studies.
Abstract: Cyclosphosphamide is widely used for cancer chemotherapy and for immunosuppression. The parent compound is inactive in vitro and exerts its biological activities through metabolites generated by hepatic microsomal enzymes. The drug may be administered either parenterally or orally. Systemic availability after oral administration is greater than 75% at the low doses which have been studied. Cyclophosphamide is minimally protein bound but some of its metabolites are more than 60% protein bound.
366 citations
287 citations
Journal Article•
TL;DR: Carboxyphosphamide, which has little or no antitumor effect, is much less toxic to clone formation of human epidermoid carcinoma No. 2 cells and to L1210 cells, and administration of pyridoxal in combination with cyclophosphamide increases the life-span of mice implanted with L 1210 cells.
Abstract: Summary Cyclophosphamide is converted by enzymes of mouse liver into two metabolites. Production of the first (aldophosphamide), which is uncharged, requires TPNH, is inhibited by CO, and is accomplished predominantly by the microsomal fraction. With the microsomal enzyme, the Km for cyclophosphamide is 0.5 mm; nicotine, atropine, ephedrine, apomorphine, and cocaine are potent inhibitors. Phenobarbital, cytochrome c, 2-diethylaminoethyl-2,2-diphenylvalerate, and some steroid hormones also inhibit the reaction. Aldophosphamide is very toxic, as judged by inhibition of clone formation of human epidermoid carcinoma No. 2 cells and by toxicity to L1210 leukemia cells. The initial metabolite is further converted to 2-carboxyethyl N,N-bis-(2-chloroethyl)phosphorodiamidate (carboxyphosphamide) by an enzyme in the soluble portion of the cell. This enzyme can be replaced by purified aldehyde oxidase (aldehyde:oxygen oxidoreductase, EC 1.2.3.1). Carboxyphosphamide, which has little or no antitumor effect, is much less toxic to clone formation of human epidermoid carcinoma No. 2 cells and to L1210 cells. Administration of pyridoxal, which could saturate the endogenous aldehyde oxidase and thus delay the production of carboxyphosphamide, in combination with cyclophosphamide increases the life-span of mice implanted with L1210 cells. The metabolic conversion of nicotine to cotinine by liver proceeds in the same manner as cyclophosphamide oxidation. Nicotine is also oxidized by an amine oxidase to nicotine 1′-oxide. Lung homogenates accomplish the initial oxidation of both cyclophosphamide and nicotine but do not metabolize the products further. Kidney homogenates contain the amine oxidase producing nicotine 1′-oxide. Several other tissues are not active in the metabolism of either cyclophosphamide or nicotine.
189 citations
Cites background from "Enzymatic basis of cyclophosphamide..."
...antitumor agent (10, 20, 36, 37, 40), has little cytotoxic or alkylating activity until it is acted upon by an enzyme of liver microsomes (4, 6)....
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...Inhibition of Cyclophosphamide metabolism by CO has been previously noted (6)....
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...Cohen and Jao (6) find a ratio of 5:1....
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...Inhibition of the activation of cyclophosphamide by testosterone, prednisolone, and cytochrome c has been noted previously (6, 12)....
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Journal Article•
TL;DR: Findings and 2-substrate kinetic studies support the conclusion that the same enzyme system or systems with a common rate-limiting step catalyze the biotransformation of cyclophosphamide, ethylmorphine, and many other drugs in rat liver.
Abstract: Summary The conversion of cyclophosphamide to its alkylating metabolite(s) by microsomes obtained from male rat liver was characterized. Under the conditions used, the reaction was found (a) to be temperature dependent; (b) to be enzyme concentration dependent; (c) to proceed in a linear fashion for 20 min; (d) to be localized to the hepatic microsomal fraction (no activity was found in Walker 256 carcinosarcoma cell fractions or in thymus, adrenal, and kidney cell fractions); (e) to require NADPH (NADH could not serve as an electron donor); (f) to be inhibited by carbon monoxide; (g) to be inhibited noncompetitively by nicotinamide; and (h) to be greatly depressed if previously frozen rather than fresh microsomes were used. Cyclophosphamide bound to microsomal hemoprotein elicited a Type I spectrum. As determined by a Lineweaver-Burk plot, the Vmax was 4.20 µmoles/g/hr and the Km was 1.39 mm. Microsomes from livers of female rats and mice metabolized cyclophosphamide with a Vmax of 0.66 and 9.69 µmoles/g/hr, respectively, and a Km of 0.67 and 0.68 mm, respectively. Cyclophosphamide inhibited metabolism of ethylmorphine competively and vice versa. The Ki9s when each was used as an inhibitor for the other9s metabolism equaled their respective Km9s. These findings and 2-substrate kinetic studies support the conclusion that the same enzyme system or systems with a common rate-limiting step catalyze the biotransformation of cyclophosphamide, ethylmorphine, and many other drugs in rat liver. While large differences in rates of metabolism were observed in vitro between male and female rats, proportional differences were not found in vivo as estimated by blood levels of alkylating activity following a single injection of cyclophosphamide i.p.
165 citations