Recently, immune checkpoint inhibitors, including anti-programmed cell death protein 1 (PD-1) antibodies, have dramatically changed treatment strategies for several cancers. In pharmacokinetic/pharmacodynamic studies, experiments using a variety of animal species are assumed.

We have identified optimal multiple reaction monitoring transitions for signature candidate peptides of nivolumab in human, mouse, and rat plasma and developed a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to quantify nivolumab (an anti-PD-1 antibody) using trastuzumab as the internal standard.

Calibration curves were linear in the range of 1–200 µg/mL. The intra- and inter-day precision and accuracy in human plasma fulfilled Food and Drug Administration guideline criteria for bioanalytical validation. There was no need to change the measurement method in mouse plasma. On the other hand, in rat plasma, an interference peak was observed at a retention time similar to that of the surrogate peptide ASGITFSNSGMHWVR (550.75 > 661.50) employed in human and mouse plasma. Therefore, we confirmed that ASQSVSSYLAWYQQKPGQAPR (785.0 > 940.2) can be used as an alternate nivolumab surrogate peptide in rat plasma at the same concentration range as used in human and mouse plasma. Using our method, the concentration range and a gradual increase in trough value were confirmed in clinical samples from two antibody-treated patients, including one with gastric cancer and one with non-small-cell lung cancer. The time course and blood concentration transition also were evaluated in nivolumab administration experiments in mouse and rat.

The present study showed that the selection of the optimal peptide is essential for accurate LC-MS/MS measurement of nivolumab concentration in human, mouse, and rat plasma. The method developed here is expected to be of use in non-clinical and clinical pharmacokinetic studies.

最近，免疫检查点抑制剂，包括抗程序性细胞死亡蛋白1（PD-1）抗体，已经大大改变了几种癌症的治疗策略。在药代动力学/药效学研究中，假定使用多种动物进行实验。

我们已经确定了人类，小鼠和大鼠血浆中的nivolumab标志候选肽的最佳多反应监测过渡，并开发了液相色谱-串联质谱（LC-MS / MS）方法来定量nivolumab（抗PD-1抗体） ）使用曲妥珠单抗作为内标。

校准曲线在1–200 µg / mL范围内呈线性。人体血浆中日间和日间的精确度和准确度均达到了美国食品药品管理局（FDA）的生物分析验证准则。无需更改小鼠血浆中的测量方法。另一方面，在大鼠血浆中，在保留时间观察到干扰峰，类似于在人和小鼠血浆中使用的替代肽ASGITFSNSGMHWVR（550.75> 661.50）。因此，我们确认了ASQSVSSYLAWYQQKPGQAPR（785.0> 940.2）可以在大鼠血浆中以与人类和小鼠血浆相同的浓度范围用作替代的nivolumab替代肽。使用我们的方法，从两名接受抗体治疗的患者的临床样品中确认了浓度范围和谷值的逐渐增加，包括一名患有胃癌和一名非小细胞肺癌。在小鼠和大鼠的nivolumab给药实验中还评估了时程和血药浓度变化。

本研究表明，最佳肽段的选择对于准确LC-MS / MS测量人，小鼠和大鼠血浆中的蚕lum浓度至关重要。预期本文开发的方法可用于非临床和临床药代动力学研究。