Development of a general method for quantifying IgG-based therapeutic monoclonal antibodies in human plasma using protein G purification coupled with a two internal standard calibration strategy using LC-MS/MS

Chiu, Huai-Hsuan; Liao, Hsiao Wei; Shao, Yu‐Yun; Lu, Yen‐Shen; Lin, Ching-Hung; Tsai, I‐Lin; Kuo, Ching‐Hua

doi:10.1016/j.aca.2018.02.040

Cited by 56 publications

(17 citation statements)

References 40 publications

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“…Bioanalytical LC-MS assays for PK studies of these immunotherapies have been published, but these studies focused on antibody quantification in plasma and only rarely in cell fractions. 112,113 However, the quantification of free monoclonal antibodies (mAbs) in plasma for PK assessment can severely be biased by the presence of circulating mAb fragments, post-translationally modified versions of the mAb, mAb immunogenicity, 114 off-target-bound mAbs and mAbs bound to shed soluble molecular targets 115 ( Figure 2). For instance, rituximab PK can largely be modified by the presence of circulating CD20 (cCD20) in the blood.…”

Section: Therapies Applied To Circulating Cells: Ideal Models For Pmentioning

confidence: 99%

Approaching sites of action of drugs in clinical pharmacology: New analytical options and their challenges

Longuespée

Theile

Fresnais

et al. 2020

Brit J Clinical Pharma

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Clinical pharmacology is an important discipline for drug development aiming to define pharmacokinetics (PK), pharmacodynamics (PD) and optimum exposure to drugs, i.e. the concentration-response relationship and its modulators. For this purpose, information on drug concentrations at the anatomical, cellular and molecular sites of action is particularly valuable. In pharmacological assays, the limited accessibility of target cells in readily available samples (i.e. blood) often hampers mass spectrometry-based monitoring of the absolute quantity of a compound and the determination of its molecular action at the cellular level. Recently, new sample collection methods have been developed for the specific capture of rare circulating cells, especially for the diagnosis of circulating tumour cells. In parallel, new advances and developments in mass spectrometric instrumentation now allow analyses to be scaled down to the cellular level. Together, these developments may permit the monitoring of minute drug quantities and show their effect at the cellular level. In turn, such PK/PD associations on a cellular level would not only enrich our pharmacological knowledge of a given compound but also expand the basis for PK/PD simulations. In this review, we describe novel concepts supporting clinical pharmacology at the anatomical, cellular and molecular sites of action, and highlight the new challenges in mass spectrometry-based monitoring. Moreover, we present methods to tackle these challenges and define future needs. K E Y W O R D S clinical pharmacology, mass spectrometry, personalized medicine, review, sites of action 1 | INTRODUCTION Clinical pharmacology studies the relevant parameters of drug fate (pharmacokinetics, PK) and efficacy (pharmacodynamics, PD) in humans in order to establish the dose-response and concentrationresponse relationship. It also defines intraindividual and interindividual modulators of these relationships, leading to different effectiveness, safety and dose requirements. Monitoring drug concentrations and effects is therefore of considerable importance during drug development, starting with the very first clinical trials. 1 Clinical pharmacology clarifies mechanisms of both beneficial and adverse drug effects in order to get closer to drug effectiveness monitoring. A drug's site of action can be defined at different scales: anatomical (the compartment the drug has to reach, e.g. tissues), cellular (the

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Section: Therapies Applied To Circulating Cells: Ideal Models For Pmentioning

confidence: 99%

Approaching sites of action of drugs in clinical pharmacology: New analytical options and their challenges

Longuespée

Theile

Fresnais

et al. 2020

Brit J Clinical Pharma

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“…However, until recently these assays always required specific reagents for affinity purification or specific stable isotope labeled (SIL) ISs. 19,20 Recently, SIL universal mAbs have become commercially available, and an LC-MS/MS assay capable of quantifying multiple co-administered mAbs using nonspecific affinity purification through protein G, and a single commercially available SIL IS, has been described. 21 However, this method has a rather low sensitivity, does not reach the throughput times of LBA, and has not been used for multiplexing the quantification of mAbs that are co-administered in clinical practice.…”

Section: Introductionmentioning

confidence: 99%

Development and validation of an LC-MS/MS method for simultaneous quantification of co-administered trastuzumab and pertuzumab

Schokker

Bonardi

Molenaar

et al. 2020

mAbs

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Given the increasing use of combination therapy with multiple monoclonal antibodies (mAbs), there is a clinical need for multiplexing assays. For the frequently co-administered anti-human epidermal growth factor receptor 2 (HER2) mAbs trastuzumab and pertuzumab, we developed a high-throughput and robust hybrid ligand-binding liquid chromatography-mass spectrometry (LC-MS)/MS quantitative assay. Nanomolar concentrations of trastuzumab and pertuzumab were determined in 10 µL serum samples after extraction by affinity purification through protein A beads, followed by on-bead reduction, alkylation, and trypsin digestion. After electrospray ionization, quantification was obtained by multiple reaction monitoring LC-MS/MS using SILuMab as an internal standard. The method was validated according to the current guidelines from the US Food and Drug Administration and the European Medicines Agency. Assay linearity was established in the ranges 0.250-250 μg/mL for trastuzumab and 0.500-500 μg/mL for pertuzumab. The method was accurate and selective for the simultaneous determination of trastuzumab and pertuzumab in clinical samples, thereby overcoming the limitation of ligand binding assays that cannot quantify mAbs targeting the same receptor. Furthermore, this method requires a small blood volume, which reduces blood collection time and stress for patients. The assay robustness was verified in a clinical trial where trastuzumab and pertuzumab concentrations were determined in 670 serum samples. As we used commercially available reagents and standards, the described generic bioanalytical strategy can easily be adapted to multiplex quantifications of other mAb combinations in non-clinical and clinical samples.

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“…In addition, when liquid chromatography/mass spectrometry (LC/MS) is used, the inclusion of bracketed standards and/or internal standards, to correct for response shift, is deemed necessary. Due to considerations related to efficiency, costs or availability, in some cases surrogate internal standards are applied 3 . It is, however, generally accepted that stable‐isotope‐labeled internal standard (SIL IS) equivalents of the analytes are the most suitable as they will also correct for any matrix effect 4 .…”

Section: Introductionmentioning

confidence: 99%

Reliable and generic liquid chromatography/mass spectrometry quantification of short peptides using a stable‐isotope‐labeled labeling agent

Kleinnijenhuis¹,

Vergauwen²,

Holthoon³

et al. 2020

Rapid Comm Mass Spectrometry

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Rationale It is important to investigate the behavior of protein hydrolysate components in both in vitro and in vivo studies, to support the elucidation of their biological functions. As protein hydrolysates and biological matrices are highly complex mixtures, it is essential to apply fully reliable and flexible analytical approaches. Methods A novel and generic Liquid Chromatography/Mass Spectrometry methodology was developed to analyze short peptides. A stable‐isotope‐labeled labeling agent 6‐aminoquinolyl‐N‐hydroxysuccinimidyl carbamate (13C3) was synthesized and used to prepare internal standards from non‐labeled analyte peptides. The amino acid and peptides p, pG, Pp, GPp and PpG (where p stands for hydroxyproline) were used for proof of principle. Results The method showed acceptable performance in solvent, in simulated gastrointestinal fluid and in serum. The (linear) dynamic range expanded to over four orders of magnitude, which is very useful when multiple analytes are analyzed in a biological matrix, due to the large differences in concentrations observed for endogenous and protein hydrolysate components. The method provides absolute‐quantitative results and is fully accountable on the single‐sample and single‐component level. Conclusions The methodology can be applied to reliably quantify protein hydrolysate nutraceutical components at various stages during their in vivo processing. Internal standards can also be synthesized for other short peptides whenever they are expected to have biological relevance and require quantification. Overall this provides an excellent analytical tool to support the elucidation of the biological functions of protein hydrolysate components.

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Development of a general method for quantifying IgG-based therapeutic monoclonal antibodies in human plasma using protein G purification coupled with a two internal standard calibration strategy using LC-MS/MS

Cited by 56 publications

References 40 publications

Approaching sites of action of drugs in clinical pharmacology: New analytical options and their challenges

Approaching sites of action of drugs in clinical pharmacology: New analytical options and their challenges

Development and validation of an LC-MS/MS method for simultaneous quantification of co-administered trastuzumab and pertuzumab

Reliable and generic liquid chromatography/mass spectrometry quantification of short peptides using a stable‐isotope‐labeled labeling agent

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