Cyclophosphamide (CPA)

1. P450-Catalyzed Deactivation Cyclophosphamide (CPA) Dechloroethyl-CPA Chloroacetylaldehyde CYP3A4 CYP2B6 CYP2B6 CYP2C9 CYP3A4 CYP2A6 CYP2C8 CYP2C19 P450-Catalyzed Activation Acrolein Aldophosphamide 4-Hydroxy-CPA DNA Alkylation Inactivation ALDH1A1 ALDH3A1 ALDH5A1 Phosphoramidemustard GSTA1 GSTP1 Conjugation with glutathione Carboxyphosphamide

2. Figure 1: Main metabolism pathway of Cyclophosphamide (CPA) • Activation of CPA to 4-hydroxycyclophosphamide is catalyzed by the hepatic cytochrome P450 (CYP) isozymes. Competing with C-4 hydroxylation of CPA is a minor (~10%) oxidative pathway that leads to N-dechloroethylation and the formation of the neurotoxic chloroacetaldehyde. 4-Hydroxycyclophosphamide interconverts rapidly with its tautomer, aldophosphamide. Aldophosphamide undergoes a spontaneous (non-enzymatic) elimination reaction to yield acrolein (associated with bladder toxicity) and the ultimate active alkylating cytotoxic compound phosphoramide mustard. A major detoxification route is the oxidation of aldophosphamide to the inactive carboxyphosphamide. Multiple CPA metabolites can react with glutathione (GSH), a tripeptidewithanunusual peptide linkagebetween the amine group of cysteine (which is attachedbynormal peptide linkageto a glycine) and the carboxylgroup of the glutamate side-chain, resulting in the formation of various conjugates at different sites along the pathway. Some of these reactions with GSH may be reversible while others are irreversible; the latter are associated with detoxification pathways. • Genes involved in this metabolism process are mentioned.

3. CATABOLIC Pathway ANABOLIC Pathway Incorporationinto DNA FdUDP FdUTP 5 FU FdUMP DHFU DPD Thymidylate Synthase dUMP dTMP DNA Synthesis Dihydrofolate 5’,10’-methylene THF MTHFR Folic acid 5’-methylTHF

4. Figure 2: Main metabolism pathway of 5-Fluorouracil (5-FU) There are several routes for metabolism of 5-FU, some of which lead to activation and pharmacodynamic actions of the drug. The rate-limiting step of 5-FU catabolism is dihydropyrimidine dehydrogenase (DPYD) conversion of 5-FU to dihydrofluorouracil (DHFU). The main mechanism of 5-FU activation is conversion to fluorodeoxyuridine monophosphate (FdUMP) which inhibits the enzyme thymidylate synthase (TYMS), an important part of the folate-homocysteine cycle and purine and pyrimidine synthesis. Methylenetetrahydrofolatereductase (MTHFR) irreversibly reduces 5,10-methylenetetrahydrofolate (substrate) to 5-methyltetrahydrofolate (product). 5,10-methylenetetrahydrofolate is used to convert dUMP to dTMP for de novo thymidine synthesis. Fluorodexoyuridinediphosphate (FdUDP) can be converted to Fluorodeoxyuridinetriphosphate (FdUTP) and incorporated into DNA. Genes involved in this metabolism process are mentioned.

5. ABCC/MRP 1 Intercalationinto DNA and RNA EPIRUBICINE Epirubicin CYP 450 reductase UGT Intercalationinto DNA and RNA Aglycones Epirubicin glucuronide Epirubicinol UGT Epirubicinol glucuronide

6. Figure 3: Main metabolism pathway of Epirubicin The metabolism of epirubicin is characterised by complex biotransformation to relatively or totally inactive metabolites. Quantitatively, the glucuronides of epirubicin and epirubicinol are very important. The four main metabolic routes are: (1) reduction of the C-13 keto-group with the formation of the 13(S)-dihydro derivative, epirubicinol; (2) conjugation of both the unchanged drug and epirubicinol with glucuronic acid; (3) loss of the amino sugar moiety through a hydrolytic process or a redox process with the formation of aglycones. Genes involved in this metabolism process are mentioned