This invention discloses the development of a novel platform for recombinant production of bioactive glycoproteins and
cancer specific vaccines in plants. Plants and
plant cell cultures have been humanized with respect to human
mucin-type
protein O-
glycosylation. A panel of
plant cell factories for production of recombinant glycoproteins with designed human O-
glycosylation, including an improved
cancer vaccine candidate, has been developed. The platform provides basis for i) production of an essentially unlimited array of O-glycosylated human
glycoprotein therapeutics, such as human
interferon α2B and podoplanin, and ii) for further
engineering of additional
cancer specific O-glycans on glycoproteins of therapeutical value. Currently, mammalian cells are required for human O-
glycosylation, but plants offer a unique
cell platform for
engineering O-glycosylation since they do not perform
human type O-glycosylation. Introduction of O-glycosylation into
plant cells requires i) that wild-type plant cells do not modify the
target peptide substrates and ii) that the appropriate enzymes and substrates are introduced into of plant cells such that O-glycosylation in the
secretory pathway proceed and the glycosylated
peptide substrates are preferentially exported to the exterior of the cell or accumulated in the cell. In this invention i) the integrity of transiently and stably expressed ‘
mucin’ type target peptides in plants cells has been determined and ii)
mucin-type O-glycosylation has been established in plants by transient and stable introduction of a
Pseudomonas aeruginosa C4-epimerase, the human polypeptide GalNAc-transferases T2 and T4 (GalNAc-T2 and T4) and various human target peptides or proteins. In the present invention GalNAc-T2 and -T4 have been used to produce a Tn cancer glycoform of MUC1.