Repotrectinib, a next-generation ROS1/TRK/ALK tyrosine kinase inhibitor, overcomes resistance due to acquired solvent-front mutations involving ROS1, NTRK1-3, and ALK. A bioanalytical assay for quantification of repotrectinib in mouse plasma and seven tissue-related matrices (brain, liver, spleen, kidney, small intestinal tissue, small intestinal content, and testis homogenates) was developed and validated using liquid chromatography with tandem mass spectrometric detection in a high-throughput 96-well format. Protein precipitation was performed by adding acetonitrile, also containing the internal standard axitinib, to 10-µl samples for all matrices. Chromatographic separation of analytes was done on an ACQUITY UPLC® BEH C18 column by gradient elution using ammonium hydroxide in water and methanol. Compounds were monitored with positive electrospray ionization using a triple quadruple mass spectrometer in selected reaction monitoring mode. The method was successfully validated in the 1-1000 ng/ml calibration range. Precisions (intra- and interday) were in the range of 1.3-8.7% and accuracies were in between 90.5% and 107.3% for all levels in all matrices. The developed method was successfully applied to investigate the plasma pharmacokinetics and tissue accumulation of repotrectinib in wild-type mice.

中文翻译：

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使用液相色谱-串联质谱法对小鼠血浆和组织匀浆中的新一代ROS1 / TRK / ALK抑制剂repotrectinib进行生物分析测定。

下一代ROS1 / TRK / ALK酪氨酸激酶抑制剂Repotrectinib克服了由于涉及ROS1，NTRK1-3和ALK的溶剂前突变引起的耐药性。建立了一种定量分析小鼠血浆和七个组织相关基质（脑，肝，脾，肾，小肠组织，小肠内容物和睾丸匀浆）中repotrectinib的生物分析方法，并通过液相色谱和串联质谱检测进行了验证高通量96孔格式。通过向所有基质的10 µl样品中添加乙腈（也包含内标阿西替尼）来进行蛋白质沉淀。使用水和甲醇中的氢氧化铵通过梯度洗脱在ACQUITYUPLC®BEH C18色谱柱上进行分析物的色谱分离。在选定的反应监测模式下，使用三重四极杆质谱仪通过正电喷雾电离监测化合物。该方法已在1-1000 ng / ml校准范围内成功验证。在所有矩阵中，所有级别的精度（日内和日间）均在1.3-8.7％的范围内，准确度在90.5％至107.3％之间。所开发的方法已成功应用于研究repotrectinib在野生型小鼠中的血浆药代动力学和组织蓄积。