UPLC-MS/MS-Based Analysis of Trastuzumab in Plasma Samples: Application in Breast Cancer Patients Sample Monitoring

Trastuzumab is a target-based recombinant humanized IgG1 monoclonal antibody (mAbs), extensively employed for treatment of metastatic breast cancer with human epidermal growth receptor 2 (HER2) overexpression. Studies around the world have reported that mAbs have substantial inter-patient unpredictable absorption, distribution, metabolism, and excretion (ADME-pharmacokinetics) because of multiple elements manipulating the concentration of mAbs in plasma. Herein, we have established a bioanalytical technique using UPLC-MS/MS with an easy sample workup method and in-solution digestion protocol to assay the trastuzumab plasma samples from breast cancer patients in clinical studies. Surrogated proteolytic peptides were used for accurate quantification of trastuzumab (CanMab) with a trastuzumab signature peptide with [13C6,15N4 ]-arginine and [13C6,15N2 ]-lysine stable isotope-labeled (SIL) peptide. Experiments to validate the method were accurately carried out according to the guidelines mentioned in the bioanalytical method validation protocol. The evaluation established excellent linearity over a wide range of 5–500 µg/mL. The experimental procedure was efficaciously performed in a pilot study of five breast cancer patients and residual concentrations of drugs from responding and non-responding subjects were compared. The receiver operating characteristic (ROC) examination displayed that 52.25 µg/mL was the Cmin threshold predictive response with a satisfactory sensitivity of 88.58% and specificity of 79.25%.