title: Transient expression in tobacco leaves of an aglycosylated recombinant antibody against the epidermal growth factor receptor.
abstract: When generating stably transformed transgenic plants, transient gene expression experiments are especially useful to rapidly confirm that the foreign molecule of interest is correctly assembled and retains its biological activity. TheraCIM(R) (CIMAB S.A., Havana) is a recombinant humanized antibody against the Epidermal Growth Factor receptor (EGF-R), now in clinical trials for cancer therapy in Cuba and other countries. An aglycosylated version (Asn 297 was mutated for Gln 297) of this antibody was transiently expressed in tobacco leaves after vacuum-mediated infiltration of recombinant Agrobacterium tumefaciens that carried a binary plasmid bearing the antibody heavy and light chain genes and plant regulation signals. Protein extracts from "agroinfiltrated" leaves were tested by ELISA and Western blot, showing that the fully assembled antibody was accumulated in plant tissues. The absence of plant specific glycans did not interfere in the assembling or in the activity of the plantibody, as demonstrated in this work. Indirect immunofluorescence demonstrated that the aglycosylated antibody expressed in plants recognizes the EGF-R expressed on the surface of A431 human tumor culture cells.
authors.affiliation: Center for Genetic Engineering and Biotechnology, P.O. Box 6162, Havana 10600, Cuba.
authors.forename: Meilyn
authors.initials: M
authors.lastname: Rodríguez
authors.forename: Nadia I
authors.initials: NI
authors.lastname: Ramírez
authors.forename: Marta
authors.initials: M
authors.lastname: Ayala
authors.forename: Freya
authors.initials: F
authors.lastname: Freyre
authors.forename: Lincidio
authors.initials: L
authors.lastname: Pérez
authors.forename: Ada
authors.initials: A
authors.lastname: Triguero
authors.forename: Cristina
authors.initials: C
authors.lastname: Mateo
authors.forename: Guillermo
authors.initials: G
authors.lastname: Selman-Housein
authors.forename: Jorge V
authors.initials: JV
authors.lastname: Gavilondo
authors.forename: Merardo
authors.initials: M
authors.lastname: Pujol
mesh_terms: D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000235:genetics* / Q000276:immunology; D061067:Antibodies, Monoclonal, Humanized; D002478:Cells, Cultured; D066246:ErbB Receptors / Q000276:immunology*; D018506:Gene Expression Regulation, Plant / Q000502:physiology; D018515:Plant Leaves / Q000235:genetics / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000378:metabolism*; D015202:Protein Engineering / Q000379:methods*; D013997:Time Factors; D014026:Nicotiana / Q000235:genetics* / Q000378:metabolism*
chemical_list: D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; C501637:TheraCIM; D066246:ErbB Receptors