Coupling CRISPR interference with FACS enrichment : new approach in glycoengineering of CHO cell lines for therapeutic glycoprotein production

Glinšek, Katja (avtor), Kramer, Lovro (avtor), Krajnc, Aleksander (avtor), Krajnc, Eva (avtor), Pirher, Nina (avtor), Marušič, Jaka (avtor), Hellmann, Leon (avtor), Podobnik, Barbara (avtor), Štrukelj, Borut (avtor), Ausländer, David (avtor), Gaber, Rok (avtor)

Difficulties in obtaining and maintaining the desired level of the critical quality attributes (CQAs) of therapeutic proteins as well as the pace of the development are major challenges of current biopharmaceutical development. Therapeutic proteins, both innovative and biosimilars, are mostly glycosylated. Glycans directly influence the stability, potency, plasma half-life, immunogenicity, and effector functions of the therapeutic. Hence, glycosylation is widely recognized as a process-dependent CQA of therapeutic glycoproteins. Due to the typically high heterogeneity of glycoforms attached to the proteins, control of glycosylation represents one of the most challenging aspects of biopharmaceutical development. Here, we explored a new glycoengineering approach in therapeutic glycoproteins development, which enabled us to achieve the targeted glycoprofile of the Fc-fusion protein in a fast manner. Coupling CRISPRi technology with lectin-FACS sorting enabled downregulation of the endogenous gene involved in fucosylation and further enrichment of CHO cells producing Fc-fusion proteins with reduced fucosylation levels. Enrichment of cells with targeted glycoprofile can lead to time-optimized clone screening and speed up cell line development. Moreover, the presented approach allows isolation of clones with varying levels of fucosylation, which makes it applicable to a broad range of glycoproteins differing in target fucosylation level.

• Jezik: Angleški jezik
• Ključne besede: CHO cells, cell line development, fucosylation, glycoengineering, therapeutic glycoproteins
• 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: 2022
• Št. strani: 13 str.
• Številčenje: Vol. 17, iss. 7, art. e2100499
• PID: 20.500.12556/RUL-138877
• UDK: 542:547.96
• ISSN pri članku: 1860-7314
• DOI: 10.1002/biot.202100499
• COBISS.SI-ID: 112903939
• Datum objave v RUL: 24.08.2022

GLINŠEK, Katja, KRAMER, Lovro, KRAJNC, Aleksander, KRAJNC, Eva, PIRHER, Nina, MARUŠIČ, Jaka, HELLMANN, Leon, PODOBNIK, Barbara, ŠTRUKELJ, Borut, AUSLÄNDER, David in GABER, Rok, 2022, Coupling CRISPR interference with FACS enrichment : new approach in glycoengineering of CHO cell lines for therapeutic glycoprotein production. Biotechnology journal [na spletu]. 2022. Vol. 17, no. 7,  e2100499. [Dostopano 19 september 2025]. DOI 10.1002/biot.202100499. Pridobljeno s: https://repozitorij.uni-lj.si/IzpisGradiva.php?lang=slv&id=138877

Gradivo je del revije

• Naslov: Biotechnology journal
• Skrajšan naslov: Biotechnol. j.
• Založnik: Wiley-VCH
• ISSN: 1860-7314
• COBISS.SI-ID: 522332953

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.
• Začetek licenciranja: 24.08.2022

Sekundarni jezik

• Jezik: Slovenski jezik
• Ključne besede: celice CHO, razvoj celične linije, fukozilacija, glikoinženiring, terapevtski glikoproteini

Projekti

• Financer: Drugi - Drug financer ali več financerjev
• Številka projekta: OP20.04327(C3330-19-952017)

• Financer: Drugi - Drug financer ali več financerjev
• Številka projekta: OP20.04327(C3330-19-952017)