Galactosylation of human erythropoietin produced by chimeric chickens expressing galactosyltransferase

Kojima, Yoshitane; Wakita, Jun; Inayoshi, Yujin; Suzuki, Ryosuke; Yamada, Yuki; Kaneoka, Hidenori; Nishijima, Ken‐ichi; Iijima, Shinji

doi:10.1016/j.jbiosc.2013.11.010

Cited by 8 publications

(10 citation statements)

References 20 publications

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“…Although a detailed analysis of glycosylation of hIFN-β in KI egg white would be required to address this issue, the lack of addition of these two terminal sugars may underlie our observed aggregation of hIFN-β in egg white as well as other possible alterations such as a reduction in protein half-life 13 . In this respect, transgenic chickens that ectopically express galactosyltransferase may constitute a platform to generate a bioreactor system based on KI hens 49 , 50 . Hence, proper galactosylation followed by sialyation in the oviduct magnum might resolve the issue of protein aggregation in egg white.…”

Section: Discussionmentioning

confidence: 99%

Efficient production of human interferon beta in the white of eggs from ovalbumin gene–targeted hens

Oishi

Yoshii

Miyahara

et al. 2018

Sci Rep

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Transgenic chickens could potentially serve as bioreactors for commercial production of recombinant proteins in egg white. Many transgenic chickens have been generated by randomly integrating viral vectors into their genomes, but transgene expression has proved insufficient and/or limited to the initial cohort. Herein, we demonstrate the feasibility of integrating human interferon beta (hIFN-β) into the chicken ovalbumin locus and producing hIFN-β in egg white. We knocked in hIFN-β into primordial germ cells using a CRISPR/Cas9 protocol and then generated germline chimeric roosters by cell transplantation into recipient embryos. Two generation-zero founder roosters produced hIFN-β knock-in offspring, and all knock-in female offspring produced abundant egg-white hIFN-β (~3.5 mg/ml). Although female offspring of the first generation were sterile, their male counterparts were fertile and produced a second generation of knock-in hens, for which egg-white hIFN-β production was comparable with that of the first generation. The hIFN-β bioactivity represented only ~5% of total egg-white hIFN-β, but unfolding and refolding of hIFN-β in the egg white fully recovered the bioactivity. These results suggest that transgene insertion at the chicken ovalbumin locus can result in abundant and stable expression of an exogenous protein deposited into egg white and should be amenable to industrial applications.

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Section: Discussionmentioning

confidence: 99%

Efficient production of human interferon beta in the white of eggs from ovalbumin gene–targeted hens

Oishi

Yoshii

Miyahara

et al. 2018

Sci Rep

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“…Human monoclonal antibodies and therapeutic proteins produced in egg whites contain nearly correct post‐translational modifications (PTM) that enable them to be effective therapeutic agents . Although prediction of the protein structure via bioinformatics tools has enabled changes to the hydrophobic areas in the final products of recombinant proteins , some important molecular events such as appropriate glycosylation occur in a species‐specific manner . Glycosylation is one of the most common PTM which transpires in the endoplasmic reticulum (ER) and continues in the Golgi apparatus in eukaryotic cells.…”

Section: Avian Eggs Are Natural Bioreactors For the Production Of Thementioning

confidence: 99%

“…This form of glycosylation does not exist in human proteins and may cause an allergic reaction . However, in chicken humanized glycan N ‐acetylneuraminic acid (NANA) is added to the terminal residue of N ‐glycans . The necessary absence of certain glycol units on human proteins, in particular the epitopes of alpha‐galactose (α‐Gal), were observed on egg white‐derived recombinant proteins .…”

Section: Avian Eggs Are Natural Bioreactors For the Production Of Thementioning

confidence: 99%

“…Transgenic avians, because of their short breeding time, simple management, and high productivity are of interest for recombinant therapeutic production [6,30]. Human monoclonal antibodies and therapeutic proteins produced in egg whites contain nearly correct posttranslational modifications (PTM) that enable them to be effective therapeutic agents [31][32][33]. Although prediction of the protein structure via bioinformatics tools has ena- bled changes to the hydrophobic areas in the final products of recombinant proteins [34], some important molecular events such as appropriate glycosylation occur in a species-specific manner [31][32][33].…”

Section: Avian Eggs Are Natural Bioreactors For the Production Of Thementioning

confidence: 99%

“…This form of glycosylation does not exist in human proteins and may cause an allergic reaction [33,37]. However, in chicken humanized glycan N-acetylneuraminic acid (NANA) is added to the terminal residue of N-glycans [31][32][33]36]. The necessary absence of certain Chicken pluripotent stem cell lines as a convenient platform for production of recombinant proteins and vaccines.…”

Section: Avian Eggs Are Natural Bioreactors For the Production Of Thementioning

confidence: 99%

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Avian embryos and related cell lines: A convenient platform for recombinant proteins and vaccine production

Farzaneh

Hassani

Mozdziak³

et al. 2017

Biotechnology Journal

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Chick embryos are a significant historical research model in basic and applied sciences. The embryonated eggs have been used for virus inoculation in order to vaccine production for nearly a century. Recently, avian eggs and cell lines derived from embryonated eggs have found wide application in biotechnology. This review will discuss about the unique characteristics of avian eggs in terms of safety, large scale and economical production of recombinant proteins. This system also provides the human-like glycosylation on target proteins and therefore can be considered as a suitable host for biomanufacturing of humanized monoclonal antibodies and therapeutic proteins. Avian derived cell lines are an alternative for rapid vaccine manufacturing during a pandemic. Based on the latest knowledge in cell and animal transgenesis, the currently available germ cell-mediated gene transfer system provides a more efficient strategy in gene targeting and creation of transgenic birds that lead to advancements in industrial, biotechnology, and biological research applications. This review covers the recent development of avian fertilized eggs and related cell lines in a variety of human biopharmaceuticals and viral vaccine manufacturing.

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Evaluation of expression systems for recombinant protein production in chicken egg bioreactors

Kajihara,

Ezaki,

Watanabe

et al. 2023

Biotechnology Journal

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Chicken eggs have gained attention as excellent bioreactors because of their genetic modifications. However, the development of chicken egg bioreactors requires a long time from the construction of the production system to the evaluation of the products. Therefore, in this study, we established a chicken cell line producing ovalbumin (OVA) and constructed a system for the rapid evaluation of the production system. First, we knocked in the EF1α promoter upstream of the OVA locus in chicken DF‐1 cells for continuous OVA expression. Furthermore, we analyzed and identified an ideal position at the OVA locus for the insertion of useful protein genes to maximize recombinant protein yield. We found that knocking in the EF1α promoter upstream of exon1 yielded the maximum production of OVA protein. In addition, we found that production efficiency was higher with linking a recombinant hFGF2 cDNA to the 5′ side of the OVA.Therefore, we established an OVA expressing cell line and an evaluation system for proteins in chicken egg bioreactors. Our findings may improve the efficiency of chicken expression systems and expand their applications in protein production. 　This article is protected by copyright. All rights reserved

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Galactosylation of human erythropoietin produced by chimeric chickens expressing galactosyltransferase

Cited by 8 publications

References 20 publications

Efficient production of human interferon beta in the white of eggs from ovalbumin gene–targeted hens

Efficient production of human interferon beta in the white of eggs from ovalbumin gene–targeted hens

Avian embryos and related cell lines: A convenient platform for recombinant proteins and vaccine production

Evaluation of expression systems for recombinant protein production in chicken egg bioreactors

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