ABSTRACT
Title
Therapeutic drug monitoring of imatinib in leukemic patients: comparison between an HPLC–UV method and an LC-MS/MS method.
Authors
E. Pirro1, S. De Francia1, F.M. Piccione1, R. Arvonio2,3, G. Muccioli2,3, M. Ruoppolo2,3, F. Di Carlo1
1Laboratory of Clinical Pharmacology, Department of Clinical & Biological Sciences, University of Turin, AOU San Luigi Gonzaga, Orbassano, Italy.
2Department of Biochemistry and Medical Biotechnology, University of Naples “Federico II”, Naples, Italy.
3CEINGE Advanced Biotechnologys.c.a.r.l., Naples, Italy.
1Laboratory of Clinical Pharmacology, Department of Clinical & Biological Sciences, University of Turin, AOU San Luigi Gonzaga, Orbassano, Italy.
2Department of Biochemistry and Medical Biotechnology, University of Naples “Federico II”, Naples, Italy.
3CEINGE Advanced Biotechnologys.c.a.r.l., Naples, Italy.
Abstract
Chronic myeloid leukemia (CML) is a myeloproliferative disorder, characterized by the presence of the Philadelphia chromosome, consequence of a reciprocal translocation between the long arms of chromosomes 9 and 22, producing a fusion oncogene referred to as BCR-ABL. Current frontline therapy for CML is imatinib, a 2-phenylaminopyrimidine-type inhibitorof the Bcr-Abl kinase, that competitively inhibits the binding of ATP to the ATP binding pocket of Bcr-Abl.
Therapeutic drug monitoring of imatinib can be useful to evaluate patient adherence to daily oral therapy, potential drug–drug interactions, treatment efficacy, and severe drug-related adverse events.
This study was undertaken to compare a new HPLC coupled with UV detection method and an LC –MS/MS method for the quantification of imatinib in human plasma.The HPLC-UV method is based on a liquid extraction from 500 μl plasma by organic protein precipitation. Chromatographic separation was achieved on a C18 3μ reverse phase analytical column, the mobile phase consisted of solvent A (Water:Methanol:Triethylamine/72.5:25:2.5) 40%,methanol 20%, acetonitrile 40% at constant flow rate of 0.5 ml/min at 35°C and the eluate was monitored at 267 nm. Internal standard was nilotinib. All analysis were performed in duplicate.
The LC –MS/MS method is based on a liquid extraction from 200 μl plasma by organic protein precipitation. Chromatographic separation was achieved on a C18 5μ reverse phase analytical column, the mobile phase was a gradient of solvent A (0.05% formic acid in ammonium formate 4 mM) and solvent B(0.05% formic acid in acetonitrile) at constant flow rate of 0.3 ml/min. Internal standard was deuterated imatinib mesylate. All analyses were performed in triplicate.
166 blood samples were obtained from CML patients, followed by several Italian hospitals, and analysed by HPLC-UV in Orbassano and by LC-MS/MS in Naples. Median results of each samples from the two laboratories were correlated and a significative correlation was found (r:0.918, P<0.001).
So, in order to monitor imatinib, two methods allowing simple and reproducible assays can be used. Several reports [1-4] describe methods using LC-MS but that is a technology not yet widely available and so the HPLC-UV method can be easily applied in laboratories without MS technology for routine clinical use.
[1] R. Bakhtiar, L. Khemani, M. Hayes, T. Bedman, F. Tse, J Pharm Biomed Anal., 28, 1183 (2002).
[2] K. Titier, S. Picard, D. Ducint, E. Teilhet, N. Moore, P. Berthaud, F.X. Mahon, M. Molimard, Ther Drug Monit., 27, 634 (2005).
[3] R. Bakhtiar, J. Lohne, L. Ramos, L. Khemani, M. Hayes, F. Tse, J Chromatogr B Analyt Technol Biomed Life Sci., 768, 325 (2002).
[4] S. De Francia, A. D'Avolio, F. De Martino, E. Pirro, L. Baietto, M. Siccardi, M. Simiele, S. Racca, G. Saglio, F. Di Carlo, G. Di Perri, J Chromatogr B Analyt Technol Biomed Life Sci., 877, 1721 (2009).
Therapeutic drug monitoring of imatinib can be useful to evaluate patient adherence to daily oral therapy, potential drug–drug interactions, treatment efficacy, and severe drug-related adverse events.
This study was undertaken to compare a new HPLC coupled with UV detection method and an LC –MS/MS method for the quantification of imatinib in human plasma.The HPLC-UV method is based on a liquid extraction from 500 μl plasma by organic protein precipitation. Chromatographic separation was achieved on a C18 3μ reverse phase analytical column, the mobile phase consisted of solvent A (Water:Methanol:Triethylamine/72.5:25:2.5) 40%,methanol 20%, acetonitrile 40% at constant flow rate of 0.5 ml/min at 35°C and the eluate was monitored at 267 nm. Internal standard was nilotinib. All analysis were performed in duplicate.
The LC –MS/MS method is based on a liquid extraction from 200 μl plasma by organic protein precipitation. Chromatographic separation was achieved on a C18 5μ reverse phase analytical column, the mobile phase was a gradient of solvent A (0.05% formic acid in ammonium formate 4 mM) and solvent B(0.05% formic acid in acetonitrile) at constant flow rate of 0.3 ml/min. Internal standard was deuterated imatinib mesylate. All analyses were performed in triplicate.
166 blood samples were obtained from CML patients, followed by several Italian hospitals, and analysed by HPLC-UV in Orbassano and by LC-MS/MS in Naples. Median results of each samples from the two laboratories were correlated and a significative correlation was found (r:0.918, P<0.001).
So, in order to monitor imatinib, two methods allowing simple and reproducible assays can be used. Several reports [1-4] describe methods using LC-MS but that is a technology not yet widely available and so the HPLC-UV method can be easily applied in laboratories without MS technology for routine clinical use.
[1] R. Bakhtiar, L. Khemani, M. Hayes, T. Bedman, F. Tse, J Pharm Biomed Anal., 28, 1183 (2002).
[2] K. Titier, S. Picard, D. Ducint, E. Teilhet, N. Moore, P. Berthaud, F.X. Mahon, M. Molimard, Ther Drug Monit., 27, 634 (2005).
[3] R. Bakhtiar, J. Lohne, L. Ramos, L. Khemani, M. Hayes, F. Tse, J Chromatogr B Analyt Technol Biomed Life Sci., 768, 325 (2002).
[4] S. De Francia, A. D'Avolio, F. De Martino, E. Pirro, L. Baietto, M. Siccardi, M. Simiele, S. Racca, G. Saglio, F. Di Carlo, G. Di Perri, J Chromatogr B Analyt Technol Biomed Life Sci., 877, 1721 (2009).