Purification process development of a recombinant monoclonal antibody expressed in glycoengineered Pichia pastoris

Jiang, Yi; Li, Fang; Zha, Dongxing; Potgieter, Thomas I.; Mitchell, Teresa; Moore, Renée; Cukan, Michael; Houston-Cummings, Nga Rewa; Nylen, Adam; Drummond, James E.; McKelvey, Troy W.; d’Anjou, Marc; Stadheim, Terrance A.; Sethuraman, Natarajan; Li, Huijuan

doi:10.1016/j.pep.2010.11.004

Cited by 27 publications

(16 citation statements)

References 22 publications

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“…Anti-HER2 mAb produced in YGLY8316 was purified through affinity capture using protein A beads and further purified by ion exchange chromatography. 30 Antibody purity by SDS-PAGE and its spectrum from size exclusion chromatography HPLC are shown in Figure 1A and B in comparison to trastuzumab. Purified anti-HER2 mAb was composed of more than 99% fully assembled antibody including double heavy and light chain, and the quality of the antibody profile was comparable to that of trastuzumab.…”

Section: Resultsmentioning

confidence: 99%

Glycoengineered Pichia produced anti-HER2 is comparable to trastuzumab in preclinical study

Zhang¹,

Li²,

Dumitru³

et al. 2011

mAbs

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Section: Resultsmentioning

confidence: 99%

Glycoengineered Pichia produced anti-HER2 is comparable to trastuzumab in preclinical study

Zhang¹,

Li²,

Dumitru³

et al. 2011

mAbs

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“…Fed-batch fermentations, secreted recombinant protein purifications, N-glycan characterizations, and all other analytical assays were performed as described previously (6,11,24).…”

Section: Methodsmentioning

confidence: 99%

Inactivation of a GAL4 -Like Transcription Factor Improves Cell Fitness and Product Yield in Glycoengineered Pichia pastoris Strains

Jiang

Argyros

Bukowski

et al. 2015

Appl Environ Microbiol

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bWith a completely reengineered and humanized glycosylation pathway, glycoengineered Pichia pastoris has emerged as a promising production host for the manufacture of therapeutic glycoproteins. However, the extensive genetic modifications have also negatively affected the overall fitness levels of the glycoengineered host cells. To make glycoengineered Pichia strains more compatible with a scalable industrial fermentation process, we sought to identify genetic solutions to broadly improve cell robustness during fermentation. In this study, we report that mutations within the Pichia pastoris ATT1 (PpATT1) gene (a homolog of the Saccharomyces cerevisiae GAL4 [ScGAL4] transcriptional activator) dramatically increased the cellular fitness levels of glycoengineered Pichia strains. We demonstrate that deletion of the PpATT1 gene enabled glycoengineered Pichia strains to improve their thermal tolerance levels, reduce their cell lysis defects, and greatly improve fermentation robustness. The extension of the duration of fermentation enabled the PpATT1-modified glycoengineered Pichia strains to increase their product yields significantly without any sacrifice in product quality. Because the ATT1 gene could be deleted from any Pichia strains, including empty hosts and protein-expressing production strains alike, we suggest that the findings described in this study are broadly applicable to any Pichia strains used for the production of therapeutic proteins, including monoclonal antibodies, Fc fusions, peptides, hormones, and growth factors. Since the approval of the first biopharmaceutical product (recombinant insulin) in 1982, biopharmaceuticals, as a prescription drug class, have enjoyed the highest growth rate within the pharmaceutical industry (1). With more than 230 approved products currently on the market, biopharmaceuticals are playing vital roles in the prevention and treatment of a wide variety of diseases, ranging across infectious diseases, inflammatory disorders, metabolic diseases, and cancer. Most biopharmaceuticals are manufactured from one of three different expression host systems: mammalian cells, yeasts, and bacteria (2). Bacterial systems (e.g., Escherichia coli) are the industry standard expression hosts for proteins that do not require posttranslational modifications (e.g., insulin, growth hormone, granulocyte colony-stimulating factor [G-CSF]). To date, mammalian cells (e.g., CHO) have been the preferred expression system for glycoproteins and monoclonal antibodies (MAbs), primarily due to their ability to produce glycosylation patterns adequate for drug safety and efficacy. Despite the advantages of being unicellular eukaryotes with efficient protein-folding and posttranslational-modification capabilities, yeasts are used for the production of only a small number of therapeutic proteins, because their high-mannose glycosylation patterns are not optimal for drug safety and efficacy.Glycoengineered Pichia pastoris has recently emerged as a promising production host for the manufacture of therapeutic ...

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“…In this context, similar considerations apply to the HCP contamination levels of mAbs eluted from Protein A columns, where in this case biospecific affinity interaction are exploited for the purification. Previous studies [32][33][34][35][36] have shown that mAbs purified with Protein A adsorbents can preferentially bind to subsets of HCPs, and the extent of interaction and identity of these associated HCPs varying significantly according to the structure and isotype of the specific IgG mAb and the composition of the different cell culture feedstocks. Whether the same subsets of HCPs are associated with mAbs purified with these new pyridinyl based adsorbents as found with Protein A affinity resins is the subject of on-going investigations.…”

Section: Efficient Removal Of Cho Host Cell Proteins By 4 0 -Terpsea mentioning

confidence: 99%

Purification of monoclonal antibodies by chemical affinity mixed mode chromatography

Zhang

Fredericks

Campi

et al. 2015

Separation and Purification Technology

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Purification process development of a recombinant monoclonal antibody expressed in glycoengineered Pichia pastoris

Cited by 27 publications

References 22 publications

Glycoengineered Pichia produced anti-HER2 is comparable to trastuzumab in preclinical study

Glycoengineered Pichia produced anti-HER2 is comparable to trastuzumab in preclinical study

Inactivation of a GAL4 -Like Transcription Factor Improves Cell Fitness and Product Yield in Glycoengineered Pichia pastoris Strains

Purification of monoclonal antibodies by chemical affinity mixed mode chromatography

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