Evaluation of “Difficult‐to‐Express” Monoclonal Antibodies in a CHO‐Based Hybrid Site‐Specific Integration System Under Industrially Relevant Conditions
Year: 2025
Type: article
Abstract: Variation in the primary sequence of monoclonal antibodies (mAbs) can negatively affect their behavior in biopharmaceutical manufacturing platforms, and efforts to identify mAbs with poor "developability" characteristics lack robust methods for assessing mAb expression from an industrially relevant platform. Recent advancements in site-specific integration-based (SSI) platforms in Chinese hamster ovary (CHO) cells can mitigate the high transcriptional variation observed with random integration and the low industrial relevance of transient expression by providing a flexible platform for mAb expression from a consistent clonal background. This work applies a novel SSI-based expression system capable of generating isogenic cell pools in less than 1 month to systematically compare the expression of ten sequence variants of two therapeutically relevant mAbs from two genomic loci under industrially relevant culture conditions. Eight single amino acid mutations in trastuzumab resulted in reduced productivity compared to the wild-type mAb in batch cultures, and three mutations maintained a low-expressing phenotype in fed-batch cultures. The mutations resulted in variant-specific patterns of decreased domain stability and increased ER stress. The application of industrially relevant SSI systems in developability workflows could strengthen the understanding of the sequence determinants of mAb expression to improve mAb design, candidate selection, and process development decisions.
Source: Biotechnology Journal
DOI: 10.1002/biot.70102
Authors: Alana C. Szkodny, Kelvin H. Lee
Institution: University of Delaware
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: hybrid
Estimated APC paid: $4,780
Awards: IUCRC Phase II+: University of Delaware: Advanced Mammalian Biomanufacturing Innovation Center (AMBIC), G3567290435, THE NATIONAL INSTITUTE FOR INNOVATION IN MANUFACTURING BIOPHARMACEUTICALS (NIIMBL), Phase I I/UCRC University of Delaware Site: Advanced Mammalian Biomanufacturing Innovation Center (AMBIC)
OpenAlex ID: W4413298628