Development of capillary size exclusion chromatography for the analysis of monoclonal antibody fragments extracted from human vitreous humor

Antibody Drug Conjugates (ADCs) are innovative biopharmaceuticals gaining increasing attention over the last two decades. The concept of ADCs lead to new therapy approaches in numerous oncological indications as well in infectious diseases. Currently, around 60 CECs are in clinical trials indicating the expanding importance of this class of protein therapeutics. ADCs show unprecedented intrinsic heterogeneity and address new quality attributes which have to be assessed. Liquid chromatography is one of the most frequently used analytical method for the characterization of ADCs. This review summarizes recent results in the chromatographic characterization of ADCs and supposed to provide a general overview on the possibilities and limitations of current approaches for the evaluation of drug load distribution, determination of average drug to antibody ratio (DAR), and for the analysis of process/storage related impurities. Hydrophobic interaction chromatography (HIC), reversed phase liquid chromatography (RPLC), size exclusion chromatography (SEC) and multidimensional separations are discussed focusing on the analysis of marketed ADCs. Fundamentals and aspects of method development are illustrated with applications for each technique. Future perspectives in hydrophilic interaction chromatography (HILIC), HIC, SEC and ion exchange chromatography (IEX) are also discussed.

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“…Despite this limitation, 300 mm x 300 m i.d. SEC capillary columns were however applied for the separation of mAbs fragments [93,94].…”

Section: Secmentioning

confidence: 99%

Current possibilities of liquid chromatography for the characterization of antibody-drug conjugates

Bobály

Fleury-Souverain²,

Beck

et al. 2018

Journal of Pharmaceutical and Biomedical Analysis

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show abstract

“…So far, the use of a capillary SEC column (300 μm I.D.) has been demonstrated for the characterization of intact monoclonal antibodies (mAbs) [23]. However, capillary SEC columns are not yet commercially available, and they present significant manufacturing challenges [24].…”

Section: Introductionmentioning

confidence: 99%

Micro-flow size-exclusion chromatography for enhanced native mass spectrometry of proteins and protein complexes

Ventouri

Veelders

Passamonti

et al. 2023

Analytica Chimica Acta

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“…SEC capillary columns were successfully applied for the separation of mAbs fragments. 58,59 Finally, SEC has also been employed for ADC characterization, but regular SEC conditions often provide poor peak shapes of ADCs and unacceptable resolution between aggregates and monomeric ADC products. This can probably be explained by the nonspecific interactions between the hydrophobic cytotoxic drugs and the surface of the stationary phase.…”

Section: ■ Chromatographic Approachesmentioning

confidence: 99%

“…Until now, the number of publications in this field is rather limited, but 300 mm × 300 μm i.d. SEC capillary columns were successfully applied for the separation of mAbs fragments. , …”

Section: Chromatographic Approachesmentioning

confidence: 99%

Chromatographic, Electrophoretic, and Mass Spectrometric Methods for the Analytical Characterization of Protein Biopharmaceuticals

et al. 2015

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Development of capillary size exclusion chromatography for the analysis of monoclonal antibody fragments extracted from human vitreous humor

Cited by 11 publications

References 17 publications

Current possibilities of liquid chromatography for the characterization of antibody-drug conjugates

Current possibilities of liquid chromatography for the characterization of antibody-drug conjugates

Micro-flow size-exclusion chromatography for enhanced native mass spectrometry of proteins and protein complexes

Chromatographic, Electrophoretic, and Mass Spectrometric Methods for the Analytical Characterization of Protein Biopharmaceuticals

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