Nijenhuis C M, Haverkate H, Rosing H, Schellens J H M, Beijnen J H
Department of Pharmacy & Pharmacology, Antoni van Leeuwenhoek/The Netherlands Cancer Institute and MC Slotervaart, Amsterdam, The Netherlands.
Department of Pharmacy & Pharmacology, Antoni van Leeuwenhoek/The Netherlands Cancer Institute and MC Slotervaart, Amsterdam, The Netherlands.
J Pharm Biomed Anal. 2016 Jun 5;125:270-9. doi: 10.1016/j.jpba.2016.03.049. Epub 2016 Mar 25.
Dabrafenib (Tafinlar(®)) and trametinib (Mekinist(®)) are registered for the treatment of patients with BRAF V600 mutation positive unresectable or metastatic melanoma. To support therapeutic drug monitoring (TDM) and clinical pharmacological trials, an assay to simultaneously quantify dabrafenib and trametinib in human plasma using liquid chromatography tandem mass spectrometry was developed and validated. Human plasma samples were collected on an outpatient base and stored at nominally -20°C. Analytes and internal standards (stable isotope labeled compounds) were extracted with TBME. After snap freezing the samples in a dry ice-ethanol bath, the organic layer was transferred to a clean tube and evaporated under a gentle stream of nitrogen gas. The dry extract was then reconstituted with 100μL acetonitrile and 5μL of the final extract was injected and separated on a C18 column with gradient elution, and analyzed with triple quadrupole mass spectrometry in positive-ion mode. The validated assay ranges from 50 to 5000ng/mL for dabrafenib and 0.5-50ng/mL for trametinib were linear, and correlation coefficient (r(2)) of 0.996 or better. At all concentrations of both analytes the biases were within ±15% of the nominal concentrations and precisions were ≤15%. All results were within the acceptance criteria of the latest US FDA guidance and EMA guidelines on method validation. Dabrafenib was found to degrade under the influence of light in different organic solvents and at least seven degradation products were detected. In conclusion, the described method to simultaneously quantify dabrafenib and trametinib in human plasma was successfully validated and applied for therapeutic drug monitoring in cancer patients treated with dabrafenib and trametinib.
达拉非尼（泰菲乐（®））和曲美替尼（迈吉宁（®））已获批用于治疗BRAF V600突变阳性的不可切除或转移性黑色素瘤患者。为支持治疗药物监测（TDM）和临床药理学试验，开发并验证了一种使用液相色谱串联质谱法同时定量人血浆中达拉非尼和曲美替尼的检测方法。在门诊采集人血浆样本，并储存于标称温度为-20°C的环境中。分析物和内标（稳定同位素标记化合物）用叔丁基甲醚萃取。在干冰-乙醇浴中对样本进行速冻后，将有机层转移至干净试管中，在温和的氮气流下蒸发。然后将干燥提取物用100μL乙腈复溶，取5μL最终提取物进样，在C18柱上进行梯度洗脱分离，并采用正离子模式的三重四极杆质谱进行分析。验证后的检测方法中，达拉非尼的线性范围为50至5000ng/mL，曲美替尼的线性范围为0.5至50ng/mL，相关系数（r²）为0.996或更高。在两种分析物的所有浓度下，偏差均在标称浓度的±15%以内，精密度≤15%。所有结果均符合美国FDA最新指南和EMA关于方法验证的指导原则中的验收标准。发现达拉非尼在不同有机溶剂中受光照影响会降解，且至少检测到七种降解产物。总之，所描述的同时定量人血浆中达拉非尼和曲美替尼的方法已成功验证，并应用于接受达拉非尼和曲美替尼治疗的癌症患者的治疗药物监测。
