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Simultaneous analysis of mitotane and its main metabolites in human blood and urine samples by SPE-HPLC technique
Ana Mornar1, Miranda Sertić, Nikša Turk
1University of Zagreb, Faculty of Pharmacy and Biochemistry, A. Kovačića 1, 10 000, Zagreb, Croatia. amornar@pharma.hr
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.
Insights
This study developed a method to measure mitotane and its metabolites in biological fluids for treating adrenocortical carcinoma (ACC). The method found high mitotane levels but no metabolites in one patient, suggesting further research into metabolism is needed.
Area of Science:
- Pharmacology
- Analytical Chemistry
- Oncology
Background:
- Adrenocortical carcinoma (ACC) is a rare cancer with poor outcomes.
- Mitotane is a key treatment for ACC, but its mechanism and metabolism are unclear.
- Understanding mitotane metabolism is crucial for optimizing therapy.
Observation:
- A novel SPE-HPLC method was developed and validated for simultaneous mitotane and metabolite determination.
- The method was applied to plasma, red blood cells, and urine samples.
- Analysis of a patient treated with mitotane revealed high drug levels but no detectable metabolites.
Findings:
- The developed SPE-HPLC method effectively quantifies mitotane and its metabolites in biological samples.
- A patient with high plasma mitotane concentrations showed no detectable levels of the investigated metabolites.
- Polymorphisms in CYP2C9 and CYP2C19 enzymes were investigated for potential roles in mitotane metabolism.
Implications:
- This validated method can aid in therapeutic drug monitoring for mitotane in ACC patients.
- The absence of detected metabolites warrants further investigation into mitotane's metabolic pathways.
- Understanding genetic variations in drug-metabolizing enzymes may personalize mitotane treatment strategies.
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