A multiplex liquid chromatography tandem mass spectrometry method for the quantification of seven therapeutic monoclonal antibodies: Application for adalimumab therapeutic drug monitoring in patients with Crohn's disease.

The use of therapeutic monoclonal antibodies (mAbs) is steadily increasing. Previous studies have reported the clinical interest of mAb therapeutic-drug monitoring (TDM), including that of adalimumab, for patients with Crohn's disease (CD). Proof of concept mAb-quantification studies by liquid chromatography mass spectrometry (LC-MS/MS) have been published, but a specific and reliable routine-suited multiplex quantification method is still needed to facilitate mAb TDM. We describe an electrospray ionization LC-MS/MS method for the simultaneous quantification of seven mAbs (adalimumab, cetuximab, infliximab, rituximab, secukinumab, tocilizumab, and trastuzumab) in human plasma. Sample preparation was performed using protein-G purification and trypsin digestion to obtain proteotypic peptides. We retrospectively measured the adalimumab concentration in 65 plasma samples from 56 CD patients and determined the adalimumab therapeutic cut-off concentration associated with biological remission. Calibration curves were linear from 1 to 100 μg mL-1 , except for rituximab (5-100 μg mL-1 ). This method was reproducible, repeatable, and accurate (coefficient of variation and bias < 20%), with no cross contamination. Adalimumab concentrations were significantly higher (p = 0.0198) for patients with biological remission (median: 11.3 μg mL-1 [4.6; 18.3]) than that for patients without a biological response (9.5 μg mL-1 [3.94;17.0]). An adalimumab cut-off concentration of 8.0 μg mL-1 correctly discriminated patients with or without biological remission (sensitivity: 74.1%, specificity: 57.9%). This validated LC-MS/MS routine-suited method is the first allowing simultaneous quantification of up to seven mAbs acting against different pharmacological targets. It opens the field of TDM to numerous mAbs.