Efficient mAb production in CHO cells incorporating PEI‐mediated transfection, mild hypothermia and the co‐expression of XBP‐1

Codamo, Joe; Hou, Jeff Jia Cheng; Hughes, Benjamin S.; Gray, Peter P.; Munro, Trent P.

doi:10.1002/jctb.2572

Cited by 23 publications

(21 citation statements)

References 64 publications

(97 reference statements)

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“…Improved transient expression was demonstrated in modified HEK293 cells expressing Epstein-Barr virus nuclear antigen 1 [27]. A similar approach in CHO cells using the Epi-CHO transient gene expression (TGE) system showed up to a 64% increase in monoclonal antibody titer when compared to previously reported systems [28,29]. These tools are summarized in Table 1 and collectively provide significant flexibility and precision in genome editing and represent significant improvements over standard practices such as homologous recombination or illegitimate integration.…”

Section: Site-specific Genome Editingmentioning

confidence: 85%

Emerging synthetic biology tools for engineering mammalian cell systems and expediting cell line development

Lanza

Cheng

Alper

2012

Current Opinion in Chemical Engineering

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Section: Site-specific Genome Editingmentioning

confidence: 85%

Emerging synthetic biology tools for engineering mammalian cell systems and expediting cell line development

Lanza

Cheng

Alper

2012

Current Opinion in Chemical Engineering

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“…81 Mammalian expression and purification Transient expression of all BsAbs was performed using PEImediated transfection of suspension adapted CHO cells as previously described. 82 KIH chains were co-transfected at a 1:1 DNA ratio (1H10-Vh : 1H10-Vk). Culture supernatants were harvested 7 -10 d post-transfection by centrifugation, then filtered and stored frozen at -80 C until purification could be completed.…”

Section: Sequence Designmentioning

confidence: 99%

Nanocell targeting using engineered bispecific antibodies

Taylor

Howard

Jones

et al. 2015

mAbs

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There are many design formats for bispecific antibodies (BsAbs), and the best design choice is highly dependent on the final application. Our aim was to engineer BsAbs to target a novel nanocell (EnGeneIC Delivery Vehicle or EDV TM nanocell) to the epidermal growth factor receptor (EGFR). EDV TM nanocells are coated with lipopolysaccharide (LPS), and BsAb designs incorporated single chain Fv (scFv) fragments derived from an anti-LPS antibody (1H10) and an anti-EGFR antibody, ABX-EGF. We engineered various BsAb formats with monovalent or bivalent binding arms and linked scFv fragments via either glycine-serine (G4S) or Fc-linkers. Binding analyses utilizing ELISA, surface plasmon resonance, bio-layer interferometry, flow cytometry and fluorescence microscopy showed that binding to LPS and to either soluble recombinant EGFR or MDA-MB-468 cells expressing EGFR, was conserved for all construct designs. However, the Fc-linked BsAbs led to nanocell clumping upon binding to EDV TM nanocells. Clumping was eliminated when additional disulfide bonds were incorporated into the scFv components of the BsAbs, but this resulted in lower BsAb expression. The G4S-linked tandem scFv BsAb format was the optimal design with respect to EDV binding and expression yield. Doxorubicin-loaded EDV TM nanocells actively targeted with tandem scFv BsAb in vivo to MDA-MB-468-derived tumors in mouse xenograft models enhanced tumor regression by 40% compared to passively targeted EDV TM nanocells. BsAbs therefore provide a functional means to deliver EDV TM nanocells to target cells.

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“…Instead, transient clones are primarily used to rapidly investigate the effect of a particular gene on the target product’s productivity. For example, transient overexpression of the X-box binding protein 1 in CHO cells expressing a monoclonal antibody increased productivity (Codamo et al, 2011). There are also examples of stable expressing clones, for example the stable overexpression of Mcl-1 in CHO cells expressing a monoclonal antibody increased cell viability (Majors et al, 2009), and the stable overexpression of Akt in CHO cells expressing a monoclonal antibody was observed to decrease apoptosis (Hwang and Lee, 2009).…”

Section: Applied Genomics In Bioprocessingmentioning

confidence: 99%

Genomics in mammalian cell culture bioprocessing

Wuest

Harcum

Lee

2012

Biotechnology Advances

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Explicitly identifying the genome of a host organism including sequencing, mapping, and annotating its genetic code has become a priority in the field of biotechnology with aims at improving the efficiency and understanding of cell culture bioprocessing. Recombinant protein therapeutics, primarily produced in mammalian cells, constitute a $108 billion global market. The most common mammalian cell line used in biologic production processes is the Chinese hamster ovary (CHO) cell line, and although great improvements have been made in titer production over the past 25 years, the underlying molecular and physiological factors are not well understood. Confident understanding of CHO bioprocessing elements (e.g. cell line selection, protein production, and reproducibility of process performance and product specifications) would significantly improve with a well understood genome. This review describes mammalian cell culture use in bioprocessing, the importance of obtaining CHO cell line genetic sequences, and the current status of sequencing efforts. Furthermore, transcriptomic techniques and gene expression tools are presented, and case studies exploring genomic techniques and applications aimed to improve mammalian bioprocess performance are reviewed. Finally, future implications of genomic advances are surmised.

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Efficient mAb production in CHO cells incorporating PEI‐mediated transfection, mild hypothermia and the co‐expression of XBP‐1

Cited by 23 publications

References 64 publications

Emerging synthetic biology tools for engineering mammalian cell systems and expediting cell line development

Emerging synthetic biology tools for engineering mammalian cell systems and expediting cell line development

Nanocell targeting using engineered bispecific antibodies

Genomics in mammalian cell culture bioprocessing

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