Characterization of the isomeric聽configuration and impurities of (Z)-endoxifen by 2D NMR, high resolution LC猬淢S, and quantitative HPLC analysis

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• 作者：Phyllis Elkins ; Donna Coleman ; Jason Burgess ; Michael Gardner ; John Hines ; Brendan Scott ; Michelle Kroenke ; Jami Larson ; Melissa Lightner ; Gregory Turner ; Jonathan White ; Paul Liu
• 关键词：Endoxifen ; Isomeric characterization ; LC猬淢S ; NMR ; Quantitative HPLC猬淯V analysis
• 刊名：Journal of Pharmaceutical and Biomedical Analysis
• 出版年：25 January, 2014
• 年：2014
• 卷：88
• 期：Complete
• 页码：174-179
• 全文大小：523 K

文摘

(Z)-Endoxifen (4-hydroxy-N-desmethyltamoxifen), an active metabolite generated via actions of CYP3A4/5 and CYP2D6, is a more potent selective estrogen receptor modulator (SERM) than tamoxifen. In the MCF-7 human mammary tumor xenograft model with female athymic mice, (Z)-endoxifen, at an oral dose of 4猬?聽mg/kg, significantly inhibits tumor growth. (Z)-Endoxifen's potential as an alternative therapeutic agent independent of CYP2D6 activities, which can vary widely in ER+ breast cancer patients, is being actively evaluated. This paper describes confirmation of the configuration of the active (Z)-isomer through 2D NMR experiments, including NOE (ROESY) to establish spatial proton猬減roton correlations, and identification of the major impurity as the (E)-isomer in endoxifen drug substance by HPLC/HRMS (HPLC/MS-TOF). Stability of NMR solutions was confirmed by HPLC/UV analysis. For pre-clinical studies, a reverse-phase HPLC猬淯V method, with methanol/water mobile phases containing 10聽mM ammonium formate at pH 4.3, was developed and validated for the accurate quantitation and impurity profiling of drug substance and drug product. Validation included demonstration of linearity, method precision, accuracy, and specificity in the presence of impurities, excipients (for the drug product), and degradation products. Ruggedness and reproducibility of the method were confirmed by collaborative studies between two independent laboratories. The method is being applied for quality control of the API and oral drug product. Kinetic parameters of Z- to E-isomerization were also delineated in drug substance and in aqueous formulation, showing conversion at temperatures above 25聽掳C.