Investigation of Antibody-Drug Conjugates by Mass Spectrometry

2019 ◽  
pp. 251-263
Author(s):  
Madhuri Jayathirtha ◽  
Costel C. Darie
Keyword(s):  
Mass Spectrometry ◽  
Antibody Drug Conjugates ◽  
Drug Conjugates ◽  
Antibody Drug

scholarly journals Application of triple quadrupole mass spectrometry for the characterization of antibody–drug conjugates

2019 ◽  
Vol 411 (12) ◽  
pp. 2569-2576 ◽  
Author(s):  
Malin Källsten ◽  
Matthijs Pijnappel ◽  
Rafael Hartmann ◽  
Fredrik Lehmann ◽  
Lucia Kovac ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Triple Quadrupole ◽  
Quadrupole Mass ◽  
Triple Quadrupole Mass Spectrometry ◽  
Antibody Drug Conjugates ◽  
Quadrupole Mass Spectrometry ◽  
Drug Conjugates ◽  
Antibody Drug

scholarly journals Multilevel characterization of antibody-ligand conjugates by CESI-MS

2020 ◽  
Vol 20 ◽  
Author(s):  
Bryan Fonslow ◽  
Gabor Jarvas ◽  
Marton Szigeti ◽  
Andras Guttman
Keyword(s):  
Mass Spectrometry ◽  
Capillary Electrophoresis ◽  
Peptide Mapping ◽  
Site Occupancy ◽  
Intact Protein ◽  
Ionization Mass ◽  
Antibody Drug Conjugates ◽  
Drug Conjugates ◽  
Antibody Drug

Aims: Demonstrating the capabilities of our new capillary electrophoresis – mass spectrometry method, which facilitates highly accurate relative quantitation of modification site occupancy of antibody-ligand (e.g., antibody-drug) conjugates. Background: Antibody-drug conjugates play important roles in medical discovery for imaging and therapeutic intervention. The localization and stoichiometry of the conjugation can affect the orientation, selectivity, specificity, and strength of molecular interactions, influencing biochemical function. Objective: To demonstrate the option to analyze the localization and stoichiometry of antibody-ligand conjugates by using essentially the same method at all levels including ligand infusion, peptide mapping, as well as reduced and intact protein analysis. Materials and Methods: Capillary electrophoresis coupled to electrospray ionization mass spectrometry was used to analyze the antibodyligand conjugates. Results: We identified three prevalent ligand conjugation sites with estimated stoichiometries of 73, 14, and 6% and an average ligand-antibody ratio of 1.37, illustrating the capabilities of CE-ESI-MS for rapid and efficient characterization of antibody-drug conjugates. Conclusion: The developed multilevel analytical method offers a comprehensive way to determine the localization and stoichiometry of antibody-drug conjugates for molecular medicinal applications. In addition, a significant advantage of the reported approach is that small, hydrophilic, unmodified peptides well separated from the neutrals, which is not common with other liquid phase separation methods such as LC.

scholarly journals Investigating the Impact of Sample Preparation on Mass Spectrometry-Based Drug-To-Antibody Ratio Determination for Cysteine- and Lysine-Linked Antibody–Drug Conjugates

Antibodies ◽  
2020 ◽  
Vol 9 (3) ◽  
pp. 46
Author(s):  
Malin Källsten ◽  
Rafael Hartmann ◽  
Lucia Kovac ◽  
Fredrik Lehmann ◽  
Sara Bergström Lind ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Sample Preparation ◽  
Reversed Phase ◽  
Sample Treatment ◽  
Antibody Molecule ◽  
Antibody Drug Conjugates ◽  
Drug Conjugates ◽  
The Impact ◽  
Antibody Ratio ◽  
Antibody Drug

Antibody–drug conjugates (ADCs) are heterogeneous biotherapeutics and differ vastly in their physicochemical properties depending on their design. The number of small drug molecules covalently attached to each antibody molecule is commonly referred to as the drug-to-antibody ratio (DAR). Established analytical protocols for mass spectrometry (MS)-investigation of antibodies and ADCs often require sample treatment such as desalting or interchain disulfide bond reduction prior to analysis. Herein, the impact of the desalting and reduction steps—as well as the sample concentration and elapsed time between synthesis and analysis of DAR-values (as acquired by reversed phase liquid chromatography MS (RPLC–MS))—was investigated. It was found that the apparent DAR-values could fluctuate by up to 0.6 DAR units due to changes in the sample preparation workflow. For methods involving disulfide reduction by means of dithiothreitol (DTT), an acidic quench is recommended in order to increase DAR reliability. Furthermore, the addition of a desalting step was shown to benefit the ionization efficiencies in RPLC–MS. Finally, in the case of delayed analyses, samples can be stored at four degrees Celsius for up to one week but are better stored at −20 °C for longer periods of time. In conclusion, the results demonstrate that commonly used sample preparation procedures and storage conditions themselves may impact MS-derived DAR-values, which should be taken into account when evaluating analytical procedures.

Mass Spectrometry of Antibody–Drug Conjugates in Plasma and Tissue in Drug Development

2013 ◽  
pp. 279-304 ◽  
Author(s):  
Surinder Kaur ◽  
Keyang Xu ◽  
Ola Saad ◽  
Luna Liu ◽  
Tim Slattery ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Drug Development ◽  
Antibody Drug Conjugates ◽  
Drug Conjugates ◽  
Antibody Drug

scholarly journals Cutting-edge mass spectrometry methods for the multi-level structural characterization of antibody-drug conjugates

2016 ◽  
Vol 13 (2) ◽  
pp. 157-183 ◽  
Author(s):  
Alain Beck ◽  
Guillaume Terral ◽  
François Debaene ◽  
Elsa Wagner-Rousset ◽  
Julien Marcoux ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Structural Characterization ◽  
Cutting Edge ◽  
Antibody Drug Conjugates ◽  
Drug Conjugates ◽  
Multi Level ◽  
Antibody Drug
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