PRODUCTION OF DIAGNOSTIC AND THERAPEUTIC ANTIBODIES IN PLANTS BY MOLECULAR FARMING

Projektinformationen

ID Finanzhilfevereinbarung: FAIR973110

Projekt abgeschlossen

Startdatum 1 Oktober 1997

Enddatum 30 September 2001

Finanziert unter

• Specific research, technological development and demonstration programme in the field of agriculture and fisheries (including agro-industry, food technologies, forestry, aquaculture and rural development), 1994-1998

Gesamtkosten

Keine Daten

EU-Beitrag

€ 950 000,00

Koordiniert durch

RHEIN.-WESTF. TECHNISCHE HOCHSCHULE AACHEN
Germany

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Verwertbare Ergebnisse

Production of Pharmaceuticals in Plants

The production of a recombinant chimeric mouse-human anti-tumor antibody recognizing the carcino embryonic tumor antigen was successfully achieved in tobacco, on a scale sufficient for preliminary evaluation of performance and efficacy (Vaquero et al, PNAS USA, 1999, in press). This is a first step toward overcoming the bottleneck in the production of therapeutic antibodies, through the large scale molecular farming of recombinant proteins in crop plants. In order to evaluate the expression of different forms of tumor specific antibody in plants, we have developed a recently described Agrobacterium-mediated transient expression system. A recombinant single chain Fv monoclonal antibody T84.66 specific for the human carcinoembryonic antigen (CEA), were engineered into a plant expression vector. Chimeric T84.66 heavy and light chain genes were constructed by exchanging the mouse light and heavy chain constant domain sequences with there human counter parts, and cloned into two independent plant expression vectors. In vivo assembly of full-size cT84.66 was achieved by simultaneous expression of the light and heavy chains after vacuum infiltration of tobacco leaves with two populations of recombinant Agrobacterium. Upscaling the transient system permitted purification of functional recombinant antibodies from the tobacco leaf extracts with in a week. His6-tagged scFv84.66 was purified by immobilized metal affinity chromatography, and cT84.66 by protein A affinity chromatography. Sufficient amounts of recombinant antibodies were recovered for detailed characterization by SDS-PAGE, western blotting and ELISA.