UNIGE document Scientific Article
previous document  unige:152867  next document
add to browser collection
Title

State-of-the-Art Native Mass Spectrometry and Ion Mobility Methods to Monitor Homogeneous Site-Specific Antibody-Drug Conjugates Synthesis

Authors
Deslignière, Evolène
Ehkirch, Anthony
Toftevall, Hanna
Sjögren, Jonathan
Beck, Alain
show hidden authors show all authors [1 - 10]
Published in Pharmaceuticals. 2021, vol. 14, no. 6
Abstract Antibody-drug conjugates (ADCs) are biotherapeutics consisting of a tumor-targeting monoclonal antibody (mAb) linked covalently to a cytotoxic drug. Early generation ADCs were predominantly obtained through non-selective conjugation methods based on lysine and cysteine residues, resulting in heterogeneous populations with varying drug-to-antibody ratios (DAR). Site-specific conjugation is one of the current challenges in ADC development, allowing for controlled conjugation and production of homogeneous ADCs. We report here the characterization of a site-specific DAR2 ADC generated with the GlyCLICK three-step process, which involves glycan-based enzymatic remodeling and click chemistry, using state-of-the-art native mass spectrometry (nMS) methods. The conjugation process was monitored with size exclusion chromatography coupled to nMS (SEC-nMS), which offered a straightforward identification and quantification of all reaction products, providing a direct snapshot of the ADC homogeneity. Benefits of SEC-nMS were further demonstrated for forced degradation studies, for which fragments generated upon thermal stress were clearly identified, with no deconjugation of the drug linker observed for the T-GlyGLICK-DM1 ADC. Lastly, innovative ion mobility-based collision-induced unfolding (CIU) approaches were used to assess the gas-phase behavior of compounds along the conjugation process, highlighting an increased resistance of the mAb against gas-phase unfolding upon drug conjugation. Altogether, these state-of-the-art nMS methods represent innovative approaches to investigate drug loading and distribution of last generation ADCs, their evolution during the bioconjugation process and their impact on gas-phase stabilities. We envision nMS and CIU methods to improve the conformational characterization of next generation-empowered mAb-derived products such as engineered nanobodies, bispecific ADCs or immunocytokines.
Keywords native mass spectrometrysize-exclusion chromatography (SEC)ion mobility-mass spectrometry (IM-MS)collision-induced unfolding (CIU)antibody-drug conjugate (ADC)site-specific conjugation
Identifiers
PMID: 34073805
Full text
Article (Published version) (2.3 MB) - public document Free access
Structures
Research group Sciences analytiques
Citation
(ISO format)
DESLIGNIÈRE, Evolène et al. State-of-the-Art Native Mass Spectrometry and Ion Mobility Methods to Monitor Homogeneous Site-Specific Antibody-Drug Conjugates Synthesis. In: Pharmaceuticals, 2021, vol. 14, n° 6. doi: 10.3390/ph14060498 https://archive-ouverte.unige.ch/unige:152867

39 hits

14 downloads

Update

Deposited on : 2021-07-09

Export document
Format :
Citation style :