title: Simultaneous analysis of mitotane and its main metabolites in human blood and urine samples by SPE-HPLC technique.
abstract: Adrenocortical carcinoma (ACC) is a rare malignancy with an incompletely understood pathogenesis and a poor prognosis. The adrenalytic activity of mitotane has made it the most important single drug in the treatment of ACC. Unfortunately, the exact mechanism of mitotane action is still unknown. It is believed that mitotane belongs to the class of drugs that require metabolic transformation by cytochrome P450 for therapeutic action; therefore determination of plasma levels of not only mitotane but also its metabolites would help in carrying out the treatment. The objective of this work was to develop and validate an SPE-HPLC method for simultaneous determination of mitotane and its metabolites in different biological fluids. The sample preparation consisted of a solid-phase extraction on a Discovery DSC(18) cartridge, while analysis of extracts was performed on a Symmetry C(18) column. The usefulness of the proposed method was confirmed by analysis of plasma, red cell and urine samples from patient chronically treated with 1.5 g of mitotane. The patient involved in this study had a high plasma concentration of mitotane and none of the investigated metabolites were found. In order to investigate whether the polymorphism of CYP2C9 and CYP2C19 enzymes could be related to the metabolism of mitotane, RT-PCR analysis was performed.
authors.affiliation: University of Zagreb, Faculty of Pharmacy and Biochemistry, A. Kovačića 1, 10 000, Zagreb, Croatia. amornar@pharma.hr
authors.forename: Ana
authors.initials: A
authors.lastname: Mornar
authors.forename: Miranda
authors.initials: M
authors.lastname: Sertić
authors.forename: Nikša
authors.initials: N
authors.lastname: Turk
authors.forename: Biljana
authors.initials: B
authors.lastname: Nigović
authors.forename: Mirko
authors.initials: M
authors.lastname: Koršić
mesh_terms: D000284:Administration, Oral; D000306:Adrenal Cortex Neoplasms / Q000097:blood / Q000188:drug therapy / Q000235:genetics / Q000652:urine; D018268:Adrenocortical Carcinoma / Q000097:blood / Q000188:drug therapy / Q000235:genetics / Q000652:urine; D018931:Antineoplastic Agents, Hormonal / Q000097:blood* / Q000652:urine*; D001189:Aryl Hydrocarbon Hydroxylases / Q000235:genetics; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D065731:Cytochrome P-450 CYP2C19; D065729:Cytochrome P-450 CYP2C9; D004355:Drug Stability; D005260:Female; D006801:Humans; D016014:Linear Models; D008875:Middle Aged; D008939:Mitotane / Q000097:blood* / Q000652:urine*; D060888:Real-Time Polymerase Chain Reaction; D015203:Reproducibility of Results; D052616:Solid Phase Extraction / Q000379:methods*
chemical_list: D018931:Antineoplastic Agents, Hormonal; D008939:Mitotane; C450260:CYP2C9 protein, human; D065729:Cytochrome P-450 CYP2C9; D001189:Aryl Hydrocarbon Hydroxylases; C045793:CYP2C19 protein, human; D065731:Cytochrome P-450 CYP2C19