Beumer Jan Hendrik, Rosing Hilde, Hillebrand Michel J X, Nan-Offeringa Lianda G A H, Foley Karen, Yule S Murray, Heck Albert J R, Schellens Jan H M, Beijnen Jos H
Department of Pharmacy & Pharmacology, Slotervaart Hospital/The Netherlands Cancer Institute, Louwesweg 6, 1066 EC Amsterdam, The Netherlands.
Rapid Commun Mass Spectrom. 2004;18(23):2839-48. doi: 10.1002/rcm.1699.
E7070 (indisulam) is a novel anticancer drug currently undergoing clinical investigation. We present a sensitive and specific method for the quantitative determination of E7070 and its metabolite M1 (1,4-benzenedisulphonamide) in human plasma, urine and faeces. The analytes and their tetra-deuterated analogues, which were used as internal standards, were isolated from the biological matrix by solid-phase extraction with OASIS cartridges (0.5 mL plasma or 1 mL urine) and by liquid-liquid extraction with ethyl acetate at pH 5 (1 mL faecal homogenate). The analytes were separated on a C8 reversed-phase chromatographic column and analyzed using electrospray ionization and tandem mass spectrometric detection in the negative ion mode. The validated concentration ranges in plasma were 0.1-20 microg/mL for E7070 and 0.01-2 microg/mL for M1. In urine and faecal homogenate, a concentration range from 0.05-10 microg/mL or microg/g, respectively, was validated for both analytes. Validation of the plasma assay was performed according to the most recent FDA guidelines. The assay fulfilled all generally accepted requirements for linearity (r > 0.99, residuals between -8 and 10%), accuracy (-13.5 to 1.4%) and precision (all less than 11%) in the tested matrices. We investigated recovery, stability (working solutions at -20 degrees C and at room temperature, biological matrices at -20 degrees C, room temperature and after 3 freeze/thaw cycles; final extracts at room temperature) and robustness. All these parameters were found acceptable. This method is suited for mass balance studies or therapeutic drug monitoring, as demonstrated by a case example showing plasma concentrations and cumulative excretion of E7070 and M1 in urine and faeces. Furthermore, we show the presence of E7070 metabolites in patient urine.
E7070（茚地那韦）是一种目前正在进行临床研究的新型抗癌药物。我们提出了一种灵敏且特异的方法，用于定量测定人血浆、尿液和粪便中的E7070及其代谢物M1（1,4 - 苯二磺酰胺）。将用作内标的分析物及其四氘代类似物通过使用OASIS小柱（0.5 mL血浆或1 mL尿液）进行固相萃取，以及在pH 5时用乙酸乙酯（1 mL粪便匀浆）进行液 - 液萃取，从生物基质中分离出来。分析物在C8反相色谱柱上分离，并使用电喷雾电离和串联质谱检测在负离子模式下进行分析。血浆中经验证的浓度范围为E7070为0.1 - 20μg/mL，M1为0.01 - 2μg/mL。在尿液和粪便匀浆中，两种分析物经验证的浓度范围分别为0.05 - 10μg/mL或μg/g。血浆分析方法的验证是根据美国食品药品监督管理局（FDA）的最新指南进行的。该分析方法在测试基质中满足了线性（r > 0.99，残差在 - 8%至10%之间）、准确度（ - 13.5%至1.4%）和精密度（均小于11%）的所有普遍接受的要求。我们研究了回收率、稳定性（ - 20℃和室温下的工作溶液、 - 20℃、室温下以及3次冻融循环后的生物基质；室温下的最终提取物）和耐用性。所有这些参数均被认为是可接受的。如一个病例示例所示，该方法适用于质量平衡研究或治疗药物监测，该病例示例显示了E7070和M1在尿液和粪便中的血浆浓度以及累积排泄情况。此外，我们还展示了患者尿液中E7070代谢物的存在。
