Optimization of bioprocess conditions improves production of a CHO cell-derived, bioengineered heparin
Jong Youn Baik,Hussain Dahodwala,Eziafa I. Oduah,Lee Talman,Trent R. Gemmill,Trent R. Gemmill,Leyla Gasimli,Payel Datta,Bo Yang,Guoyun Li,Fuming Zhang,Lingyun Li,Robert J. Linhardt,Andrew M. Campbell,Stephen F. Gorfien,Susan T. Sharfstein +15 more
TLDR
Bioprocess optimization, in parallel with metabolic engineering refinements, will play a substantial role in developing a bioengineered heparin to replace the current animal-derived drug.Abstract:
Heparin is the most widely used anticoagulant drug in the world today. Heparin is currently produced from animal tissues, primarily porcine intestines. A recent contamination crisis motivated development of a non-animal-derived source of this critical drug. We hypothesized that Chinese hamster ovary (CHO) cells could be metabolically engineered to produce a bioengineered heparin, equivalent to current pharmaceutical heparin. We previously engineered CHO-S cells to overexpress two exogenous enzymes from the heparin/heparan sulfate biosynthetic pathway, increasing the anticoagulant activity ∼100-fold and the heparin/heparan sulfate yield ∼10-fold. Here, we explored the effects of bioprocess parameters on the yield and anticoagulant activity of the bioengineered GAGs. Fed-batch shaker-flask studies using a proprietary, chemically-defined feed, resulted in ∼two-fold increase in integrated viable cell density and a 70% increase in specific productivity, resulting in nearly three-fold increase in product titer. Transferring the process to a stirred-tank bioreactor increased the productivity further, yielding a final product concentration of ∼90 μg/mL. Unfortunately, the product composition still differs from pharmaceutical heparin, suggesting that additional metabolic engineering will be required. However, these studies clearly demonstrate bioprocess optimization, in parallel with metabolic engineering refinements, will play a substantial role in developing a bioengineered heparin to replace the current animal-derived drug.read more
Citations
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Heparin: Past, Present, and Future
TL;DR: A variety of strategies have been proposed to produce a bioengineered heparin including microbial production, mammalian cell production, and chemoenzymatic modification, and strategies for creating “designer” heparins and heparan-sulfates with various biochemical and physiological properties are proposed.
Journal ArticleDOI
Chemoenzymatic Synthesis of Glycosaminoglycans.
TL;DR: Improvements in chemoenzymatic synthesis of GAGs have successfully resulted in multigram-scale synthesis of low-molecular-weight heparins (LMWHs), with some showing excellent anticoagulant activity and even resulting in more effective protamine reversal than commercial, animal-sourced LMWH drugs.
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The road to animal-free glycosaminoglycan production: current efforts and bottlenecks.
TL;DR: It is envisioned that new engineering approaches together with advances in the basic biology and chemistry of GAGs will move GAG production beyond its currently limited supply chain.
Journal ArticleDOI
Bio-Based Strategies for Producing Glycosaminoglycans and Their Oligosaccharides
TL;DR: This review summarizes recently developed bio-based strategies for producing GAGs, highlighting in particular enzymatic, metabolic engineering, and synthetic biology strategies.
Journal ArticleDOI
Heparan Sulfate Domains Required for Fibroblast Growth Factor 1 and 2 Signaling through Fibroblast Growth Factor Receptor 1c
Victor Schultz,Mathew Suflita,Xinyue Liu,Xing Zhang,Yanlei Yu,Lingyun Li,Dixy E. Green,Yongmei Xu,Fuming Zhang,Paul L. DeAngelis,Jian Liu,Robert J. Linhardt +11 more
TL;DR: It is suggested that it is now possible to chemoenzymatically synthesize precise HS polysaccharides that can selectively mediate growth factor signaling and might be useful in both understanding and controlling the growth, proliferation, and differentiation of cells in stem cell therapies, wound healing, and the treatment of cancer.
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