Persistence and Fab-specific interaction of acyl-protein thioesterase-1 with monoclonal antibodies during downstream processing

Šprager, Ernest (avtor), Krajnc, Aleksander (avtor), Lunder, Mojca (avtor), Vašl, Jožica (avtor), Bratkovič, Tomaž (avtor)

Difficult-to-remove host cell proteins (HCPs) can compromise monoclonal antibody (mAb) product quality and stability. This study investigated the persistence of acyl-protein thioesterase-1, a polysorbate-degrading HCP, during downstream processing and its interaction with a set of mAbs. Acyl-protein thioesterase-1 remained in the eluates after reprocessing a specific mAb with protein A affinity liquid chromatography, even when various wash buffer additives were used, indicating a strong product:HCP interaction that resists conventional purification strategies. Upon cleaving the mAb and purifying its Fab and Fc fragments, we used biolayer interferometry to show that acyl-protein thioesterase-1 selectively and tightly binds to the Fab region. Domain-specific sensor-immobilization approach suggested that the binding site was located on the antibody's CH1 domain, and the interaction was influenced by preincubation with the mAb's cognate antigen. These findings highlight the importance of characterizing HCP:mAb interactions as a basis for improving HCP reduction strategies.

• Jezik: Angleški jezik
• Ključne besede: monoclonal antibodies, chromatography, host cell proteins, acyl-protein thioesterase-1, protein-protein interactions, biolayer interferometry
• Vrsta gradiva: Članek v reviji
• Tipologija: 1.01 - Izvirni znanstveni članek
• Organizacija: FFA - Fakulteta za farmacijo
• Status publikacije: Objavljeno
• Različica publikacije: Objavljena publikacija
• Leto izida: 2026
• Št. strani: Str. 1-7
• Številčenje: Vol. 241, art. 106920
• PID: 20.500.12556/RUL-181020
• UDK: 577
• ISSN pri članku: 1046-5928
• DOI: 10.1016/j.pep.2026.106920
• COBISS.SI-ID: 272547075
• Datum objave v RUL: 23.03.2026

Gradivo je del revije

• Naslov: Protein expression and purification
• Skrajšan naslov: Protein expr. purif.
• Založnik: Academic Press
• ISSN: 1046-5928
• COBISS.SI-ID: 2627623

Licence

• Licenca: CC BY 4.0, Creative Commons Priznanje avtorstva 4.0 Mednarodna
• Povezava: http://creativecommons.org/licenses/by/4.0/deed.sl
• Opis: To je standardna licenca Creative Commons, ki daje uporabnikom največ možnosti za nadaljnjo uporabo dela, pri čemer morajo navesti avtorja.

Sekundarni jezik

• Jezik: Slovenski jezik
• Ključne besede: monoklonska protitelesa, kromatografija, proteini gostiteljskih celic, acil-protein tioesteraza-1, interakcije protein-protein, interferometrija biološke plasti

Projekti

• Financer: ARIS - Javna agencija za znanstvenoraziskovalno in inovacijsko dejavnost Republike Slovenije
• Številka projekta: P1-0420
• Naslov: Napredna imunološka zdravila in celični pristopi v farmaciji