HIGH RESOLUTION MASS SPECTROMETRY OF IMATINIB BIOTRANSFORMATION PRODUCTS

ABSSUB-4439

Background: Therapeutic drug monitoring is widely applied useful tool for treatment individualization in order to achieve optimal clinical response and avoid toxicity. Due to drug metabolizing enzymes several bioactive or toxic metabolites may be produced. Therefore determination of not only parent drug but also of metabolites may have clinical relevance and could be helpful for treatment adjustment.

Aims: To assess a detailed profile of imatinib (IMA) metabolites in plasma of patients with chronic myeloid leukemia (CML).

Methods: Before analysis plasma proteins were precipitated by methanol and supernatant was injected. Separation was performed on Phenomenex Kinetex C18 column (100 x 2.1 mm; 1.7 μm) using UltiMate 3000 RS (Thermo Scientific) liquid chromatography. For detection Orbitrap Elite mass spectrometer (Thermo Scientific) based on exact mass measurement was used. Scan range of m/z 350 – 1200 was chosen and the resolution was set at 60,000 FWHM. MS/MS data were acquired in data-dependent strategy based on fragmentation of 25 selected exact m/z values of potential metabolites. Collision energy for CID was set at 35 eV. Dynamic exclusion duration of 3 s was enabled; mass accuracy was 5 ppm. Data were evaluated using Excalibur 2.2 SP1, MetWorks 1.3 SP3 and Mass Frontier 7.0 software. Eight samples of 7 CML patients with optimal or suboptimal response to treatment were analyzed (age 41-77 years, 400 mg IMA/day; sampling time 20-27 h after IMA ingestion).

Results: Contrary to previously described 14 IMA metabolites in plasma (Rochat B. et al. Mass Spectrom. 2008;43:736) we have found 100 potential metabolites in concentration range of 4 orders of magnitude (0.001 – 6.630 %) related to IMA plasma levels (635 – 1360 ng/mL). All metabolites found in plasma were identified on the base of exact m/z values and confirmed by MS2 and MS3 fragmentation experiments. Importantly, differences in the profiles of oxidized and dioxidized metabolites in optimally and sub-optimally responding patients were observed.

Summary/Conclusion: The new generation of high resolution mass spectrometry offers sensitive tool for detail study of drug metabolisation in biofluids. IMA is metabolised to many metabolites which have the potential to be diagnostically useful. Further studies should be focused on elucidation of clinical significance of newly discovered metabolites. Acknowledgements: The work was supported by grant LF UP 2013-004 and 2013-010, grant of IGA Ministry of Health, Czech Republic NT12218-4/2011. Infrastructural part of this project (Institute of Molecular and Translational Medicine) was supported from the Operational Program Research and Development for Innovations; Project CZ.1.05/2.1.00/01.0030.

Keywords: None

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