Multiplex genome editing of mammalian cells for producing recombinant heparin |
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Institution: | 1. TEGA Therapeutics Inc, 3550 General Atomics Court, G02-102, San Diego, CA, 92121, USA;2. Departments of Pediatrics and Bioengineering, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA, 92093, USA;3. Department of Dermatology, University of California, San Diego, School of Medicine, 9500 Gilman Drive, La Jolla, CA, 92093, USA;4. College of Nanoscale Science and Engineering, SUNY Polytechnic Institute, 257 Fuller Road, Albany, NY, 12203, USA;5. Cardiovascular Research Institute, Loyola University Chicago, Health Sciences Division, 2160 S 1st Avenue, Maywood, IL, 60153, USA;6. National Institute for Biological Standards and Control, Blanche Lane, South Mimms, Herts, EN6 3QG, UK;7. Glycobiology Research and Training Center, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA, 92093, USA;8. Department of Cellular and Molecular Medicine, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA, 92093, USA |
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Abstract: | Heparin is an essential anticoagulant used for treating and preventing thrombosis. However, the complexity of heparin has hindered the development of a recombinant source, making its supply dependent on a vulnerable animal population. In nature, heparin is produced exclusively in mast cells, which are not suitable for commercial production, but mastocytoma cells are readily grown in culture and make heparan sulfate, a closely related glycosaminoglycan that lacks anticoagulant activity. Using gene expression profiling of mast cells as a guide, a multiplex genome engineering strategy was devised to produce heparan sulfate with high anticoagulant potency and to eliminate contaminating chondroitin sulfate from mastocytoma cells. The heparan sulfate purified from engineered cells grown in chemically defined medium has anticoagulant potency that exceeds porcine-derived heparin and confers anticoagulant activity to the blood of healthy mice. This work demonstrates the feasibility of producing recombinant heparin from mammalian cell culture as an alternative to animal sources. |
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Keywords: | Heparin Heparan sulfate Anticoagulation Metabolic engineering Mastocytoma Synthetic biology |
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