A liquid chromatography-tandem mass spectrometry method for the determination of 5-fluorouracil degradation rate by intact peripheral blood mononuclear cells

Alfonso M. Lostia, Luana Lionetto, Cristiano Ialongo, Giovanna Gentile, Antonella Viterbo, Paola Malaguti, Ida Paris, Luca Marchetti, Paolo Marchetti, Antonio De Blasi, Maurizio Simmaco

Research output: Contribution to journalArticle

Abstract

5-Fluorouracil (5-FU) is a major chemotherapy drug used for the treatment of tumors. It is catabolized mainly by dihydropyrimidine dehydrogenase, and patients with a complete or partial deficiency of dihydropyrimidine dehydrogenase activity are at risk of developing severe 5-FU-associated toxicity. The aim of this study was to demonstrate that intact peripheral blood mononuclear cells (PBMCs) can be an effective model to evaluate the degradation rate of 5-FU. We developed a sensitive and specific liquid chromatography-tandem mass spectrometry method to measure in vitro the rate of 5-FU degradation by intact PBMC. 5-FU degradation rate was determined by measuring the decrease of a fixed amount of 5-FU (10 μg/mL) added to a solution of PBMC, after 2 hours incubation, expressed as nanogram per milliliter of 5-FU degraded per minute × 10 cells. Freshly prepared intact PBMC can degrade efficiently in vitro-added 5-FU. The assay consists of 3 steps: (1) PBMC isolation from peripheral blood, (2) PBMC incubation with 5-FU in vitro for different times, and (3) determination of 5-FU amount to calculate the degradation rate. 5-FU was analyzed by a Q Trap 2000 triple quadrupole/ion trap mass spectrometer in the multiple-reaction-monitoring modes. The chromatographic separation was accomplished using a C18 column with a run time of 16 minutes. By analyzing samples from 39 patients with no 5-FU toxicity, the mean 5-FU degradation rate was 1.85 ± 0.50 ng•mL•min × 10 cells. The assessment of a test to measure 5-FU degradation rate in PBMC of patients before 5-FU administration could represent a prescreening method for evaluating the possible toxicity of this drug as an aid to set up a personalized medicine approach for each patient.

Original languageEnglish
Pages (from-to)482-488
Number of pages7
JournalTherapeutic Drug Monitoring
Volume31
Issue number4
DOIs
Publication statusPublished - Aug 2009

Keywords

  • 5-fluorouracil
  • Dihydropyrimidine dehydrogenase
  • LC-MS/MS
  • Peripheral blood mononuclear cells, therapeutic drug monitoring
  • Personalized medicine

ASJC Scopus subject areas

  • Pharmacology (medical)
  • Pharmacology

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