Department Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia, PA, USA.
Ther Drug Monit. 2011 Aug;33(4):460-3. doi: 10.1097/IAE.0b013e3182219af8.
Everolimus was approved for the prevention of organ rejection of kidney transplants in adult patients in April 2010 by the US Food and Drug Administration. Therapeutic drug monitoring of everolimus is recommended for all patients receiving this immunosuppressant. The goal of this study was to improve and revalidate our previously published high-performance liquid chromatography tandem mass spectrometry method for the measurement of everolimus and to assess the performance of the recently introduced isotopically labeled internal standard (IS).
For this method, the following innovative changes were incorporated: (1) sample preparation includes the addition of water to hemolyze red blood cells; (2) a separate extraction column is added to the system; (3) different settings of the switching valve are used to minimize dead volume; (4) a more sensitive mass spectrometer is used (API 5000) for drug detection; (5) isotopically labeled everolimus is used as the IS.
The addition of water to blood before adding ZnSO4 with methanol improves absolute recovery of everolimus from 77.3% ± 6.18% to 82.3% ± 6.3%. The use of the more sensitive mass spectrometer permitted the use of a smaller sample volume and lowered the lower limit of quantification from 1.0 to 0.5 ng/mL. Between-day precision for quality control samples is below 9%, and the accuracy ranged from 94.8% to 106.4%. Neither carryover nor matrix effects were observed. A comparison study showed very good agreement between the results obtained by our laboratory and those obtained by a reference laboratory (r = 0.93). The performance of the new IS, [13c2d4] RAD001, was compared with that of SDZ RAD 223-756. Everolimus concentration results obtained using the isotopically labeled IS agreed more closely with the results from the reference laboratory (using 13c2d4) as compared with everolimus concentration results obtained using the analog (SDZ RAD 223-756).
This revised methodology together with the comparison study data proved that our novel method for the measurement of everolimus is sensitive, precise, reliable, and robust.
依维莫司于 2010 年 4 月获得美国食品和药物管理局批准，用于预防成人肾移植患者的器官排斥反应。所有接受这种免疫抑制剂治疗的患者均建议进行依维莫司治疗药物监测。本研究的目的是改进和重新验证我们之前发表的高效液相色谱串联质谱法，以测量依维莫司，并评估最近引入的同位素标记内标（IS）的性能。
该方法进行了以下创新改进：（1）样品制备包括加入水以溶解红细胞；（2）在系统中添加单独的提取柱；（3）使用不同的切换阀设置以最小化死体积；（4）使用更灵敏的质谱仪（API 5000）进行药物检测；（5）使用同位素标记的依维莫司作为 IS。
在加入甲醇中的 ZnSO4 之前向血液中加入水，可将依维莫司的绝对回收率从 77.3%±6.18%提高至 82.3%±6.3%。使用更灵敏的质谱仪允许使用更小的样品量，并将定量下限从 1.0 降低至 0.5ng/mL。质控样品的日间精密度低于 9%，准确度范围为 94.8%至 106.4%。未观察到交叉污染或基质效应。比较研究表明，我们实验室与参考实验室获得的结果非常吻合（r=0.93）。新的 IS[13c2d4]RAD001 的性能与 SDZ RAD 223-756 的性能进行了比较。与使用模拟物（SDZ RAD 223-756）相比，使用同位素标记 IS 获得的依维莫司浓度结果与参考实验室的结果更为一致（使用 13c2d4）。
该修订方法和比较研究数据证明，我们用于测量依维莫司的新方法灵敏、准确、可靠且稳健。
