title: Purification of the therapeutic antibody trastuzumab from genetically modified plants using safflower Protein A-oleosin oilbody technology.
abstract: Production of therapeutic monoclonal antibodies using genetically modified plants may provide low cost, high scalability and product safety; however, antibody purification from plants presents a challenge due to the large quantities of biomass that need to be processed. Protein A column chromatography is widely used in the pharmaceutical industry for antibody purification, but its application is limited by cost, scalability and column fouling problems when purifying plant-derived antibodies. Protein A-oleosin oilbodies (Protein A-OB), expressed in transgenic safflower seeds, are relatively inexpensive to produce and provide a new approach for the capture of monoclonal antibodies from plants. When Protein A-OB is mixed with crude extracts from plants engineered to express therapeutic antibodies, the Protein A-OB captures the antibody in the oilbody phase while impurities remain in the aqueous phase. This is followed by repeated partitioning of oilbody phase against an aqueous phase via centrifugation to remove impurities before purified antibody is eluted from the oilbodies. We have developed this purification process to recover trastuzumab, an anti-HER2 monoclonal antibody used for therapy against specific breast-cancers that over express HER2 (human epidermal growth factor receptor 2), from transiently infected Nicotiana benthamiana. Protein A-OB overcomes the fouling problem associated with traditional Protein A chromatography, allowing for the development of an inexpensive, scalable and novel high-resolution method for the capture of antibodies based on simple mixing and phase separation.
authors.affiliation: School of Environmental Sciences, University of Guelph, 50 Stone Road East, Guelph, ON N1G 2W1, Canada. mdmclean@uoguelph.ca
authors.forename: Michael D
authors.initials: MD
authors.lastname: McLean
authors.forename: Rongji
authors.initials: R
authors.lastname: Chen
authors.forename: Deqiang
authors.initials: D
authors.lastname: Yu
authors.forename: Kor-Zheng
authors.initials: KZ
authors.lastname: Mah
authors.forename: John
authors.initials: J
authors.lastname: Teat
authors.forename: Haifeng
authors.initials: H
authors.lastname: Wang
authors.forename: Steve
authors.initials: S
authors.lastname: Zaplachinski
authors.forename: Joseph
authors.initials: J
authors.lastname: Boothe
authors.forename: J Christopher
authors.initials: JC
authors.lastname: Hall
mesh_terms: D000595:Amino Acid Sequence; D061067:Antibodies, Monoclonal, Humanized / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000276:immunology*; D031187:Carthamus tinctorius / Q000737:chemistry*; D002846:Chromatography, Affinity; D006801:Humans; D008969:Molecular Sequence Data; D015388:Organelles / Q000378:metabolism*; D046549:Plantibodies / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D013205:Staphylococcal Protein A / Q000276:immunology*; D014026:Nicotiana / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D000068878:Trastuzumab
chemical_list: D061067:Antibodies, Monoclonal, Humanized; D029681:Arabidopsis Proteins; D046549:Plantibodies; D013205:Staphylococcal Protein A; C079353:oleosin protein, Arabidopsis; D000068878:Trastuzumab