In vivo evaluation and in vitro metabolism of leuprolide in mice--mass spectrometry-based biomarker measurement for efficacy and toxicity.
The study of pharmacologically active peptides is central for the understanding of cancer and the development of novel therapeutic approaches. In this context, both qualitative and quantitative determination of bioactive peptides in biological fluids/tissues and their effect on endogenous factors (e.g. hormones) are of great importance. A mass spectrometry-based approach was developed and applied towards the measurement of leuprolide, a peptide drug for the treatment of prostate cancer, in mouse plasma. High-pressure liquid chromatography coupled to a hybrid quadrupole linear ion trap (QqLIT) mass spectrometer, a platform that combines the benefits of triple QqLIT instruments, was employed for the study. Using the described methodology, we established that picomolar concentrations of leuprolide could be measured in mouse plasma (limit of quantification of 0.1 ng/ml). In order to optimize pharmacokinetic properties of analogs of leuprolide, a facile in vivo mouse model was developed and leuprolide concentrations were determined in mouse plasma following intraperitoneal administration. In the same animal model, we demonstrated the versatility of the described MS-based approach by the determination of plasma concentrations of testosterone, an established biomarker for the treatment of prostate cancer. Following dosing with leuprolide, circulating testosterone was increased significantly in comparison to vehicle-treated mice. Finally, in vitro metabolism of leuprolide was evaluated by incubation of leuprolide with mouse kidney membranes, followed by identification of major metabolites by MS. Such studies provide the framework for future evaluation of novel leuprolide analogs with potential therapeutic advantages.
Sofianos
Zacharias D
ZD
Division of Pharmacology-Pharmacotechnology, Biomedical Research Foundation, Academy of Athens, Athens 11527, Greece.
Katsila
Theodora
T
Kostomitsopoulos
Nikolaos
N
Balafas
Vaggelis
V
Matsoukas
John
J
Tselios
Theodoros
T
Tamvakopoulos
Constantin
C
D000818:Animals; D018931:Antineoplastic Agents, Hormonal / Q000378:metabolism* / Q000493:pharmacokinetics / Q000633:toxicity*; D015415:Biomarkers / Q000378:metabolism; D002138:Calibration; D005615:Freezing; D007202:Indicators and Reagents; D007668:Kidney / Q000378:metabolism; D016729:Leuprolide / Q000378:metabolism* / Q000493:pharmacokinetics / Q000633:toxicity*; D008297:Male; D008566:Membranes / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D012015:Reference Standards; D015203:Reproducibility of Results; D013739:Testosterone / Q000097:blood
D018931:Antineoplastic Agents, Hormonal; D015415:Biomarkers; D007202:Indicators and Reagents; D013739:Testosterone; D016729:Leuprolide