Glycofi, Incorporated

Merck Sharp + Dohme Corp.

Juergen Nett

Robert Davidson

Stefan Wildt

Piotr Bobrowicz

Tillman U Gerngross

Stephen Robin Hamilton

Byung-Kwon Choi

Natarajan Sethuraman

Byung-kwon Choi

William James Cook

Terrance A Stadheim

Robert Gordon Miele

Piotri Bobrowicz

Method and system for expression systems, based on ade1 and ade2 auxotrophic strains of yeast and fungi, includingare disclosed. The expression systems are useful for increased cellular productivity of transformed cell lines and for production of recombinant glycoproteins at industrial scale.

Yeast strains for protein production

Lower eukaryotic host cells have been recombinantly engineered to produce glycoprotein having human-like O-glycosylation. The glycoproteins are useful for the production of glycoprotein compositions with advantages for the production of human therapeutics.

Yeast strain for the production of proteins with modified O-glycosylation

The present invention relates to host cells having modified lipid-linked oligosaccharides which may be modified further by heterologous expression of a set of glycosyltransferases, sugar transporters and mannosidases to become host-strains for the production of mammalian, e.g., human therapeutic glycoproteins. The process provides an engineered host cell which can be used to express and target any desirable gene(s) involved in glycosylation. Host cells with modified lipid-linked oligosaccharides are created or selected. N-glycans made in the engineered host cells have a GlcNAcManGlcNAccore structure which may then be modified further by heterologous expression of one or more enzymes, e.g., glycosyl-transferases, sugar transporters and mannosidases, to yield human-like glycoproteins. For the production of therapeutic proteins, this method may be adapted to engineer cell lines in which any desired glycosylation structure may be obtained.

Method to engineer mammalian-type carbohydrate structures

The present invention relates to eukaryotic host cells having modified oligosaccharides which may be modified further by heterologous expression of a set of glycosyltransferases, sugar transporters and mannosidases to become host-strains for the production of mammalian, e.g., human therapeutic glycoproteins. The invention provides nucleic acid molecules and combinatorial libraries which can be used to successfully target and express mammalian enzymatic activities such as those involved in glycosylation to intracellular compartments in a eukaryotic host cell. The process provides an engineered host cell which can be used to express and target any desirable gene(s) involved in glycosylation. Host cells with modified oligosaccharides are created or selected. N-glycans made in the engineered host cells have a ManGlcNAccore structure which may then be modified further by heterologous expression of one or more enzymes, e.g., glycosyltransferases, sugar transporters and mannosidases, to yield human-like glycoproteins. For the production of therapeutic proteins, this method may be adapted to engineer cell lines in which any desired glycosylation structure may be obtained.

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The present invention relates to eukaryotic host cells having modified oligosaccharides which may be modified further by heterologous expression of a set of glycosyltransferases, sugar transporters and mannosidases to become host-strains for the production of mammalian, e.g., human therapeutic glycoproteins. The invention provides nucleic acid molecules and combinatorial libraries which can be used to successfully target and express mammalian enzymatic activities such as those involved in glycosylation to intracellular compartments in a eukaryotic host cell. The process provides an engineered host cell which can be used to express and target any desirable gene(s) involved in glycosylation. Host cells with modified oligosaccharides are created or selected. N-glycans made in the engineered host cells have a ManGlcNAccore structure which may then be modified further by heterologous expression of one or more enzymes, e.g., glycosyltransferases, sugar transporters and mannosidases, to yield human-like glycoproteins.

Combinatorial DNA library for producing modified N-glycans in lower eukaryotes

The present invention provides a novel lower eukaryotic host cell producing human-like glycoproteins characterized as having a terminal &#x3b2;-galactose residue and essentially lacking fucose and sialic acid residues. The present invention also provides a method for catalyzing the transfer of a galactose residue from UDP-galactose onto an acceptor substrate in a recombinant lower eukaryotic host cell, which can be used as a therapeutic glycoprotein.

Production of galactosylated glycoproteins in lower eukaryotes

Methods and materials are provided for integrating heterologous nucleic acids into the genome of a cell or virus without disrupting expression of genes adjacent to the insertion site.

Recombinant vectors

The present invention relates to eukaryotic host cells, especially lower eukaryotic host cells, having modified oligosaccharides which may be modified further by heterologous expression of a set of glycosyltransferases, sugar and sugar nucleotide transporters to become host-strains for the production of mammalian, e.g., human therapeutic glycoproteins. The process provides an engineered host cell which can be used to express and target any desirable gene(s) involved in glycosylation. Host cells with modified lipid-linked oligosaccharides are created or selected. N-glycans made in the engineered host cells exhibit GnTIII, GnTIV, GnTV, GnT VI or GnTIX activity, which produce bisected and/or multiantennary N-glycan structures and may be modified further by heterologous expression of one or more enzymes, e.g., glycosyltransferases, sugar, sugar nucleotide transporters, to yield human-like glycoproteins. For the production of therapeutic proteins, this method may be adapted to engineer cell lines in which any desired glycosylation structure may be obtained.

Production of modified glycoproteins having multiple antennary structures

A novel gene encodingorotate-phosphoribosyl transferase (URA5) is disclosed. Methods for producing and selecting yeast strains capable of stable genetic integration of heterologous sequences into the host genome are also provided.

The present invention relates to eukaryotic host cells having modified oligosaccharides which may be modified further by heterologous expression of a set of glycosyltransferases, sugar transporters and mannosidases to become host-strains for the production of mammalian, e.g., human therapeutic glycoproteins. The process provides an engineered host cell which can be used to express and target any desirable gene(s) involved in glycosylation. Host cells with modified lipid-linked oligosaccharides are created or selected. N-glycans made in the engineered host cells exhibit GnTIII activity, which produce bisected N-glycan structures and may be modified further by heterologous expression of one or more enzymes, e.g., glycosyltransferases, sugar transporters and mannosidases, to yield human-like glycoproteins. For the production of therapeutic proteins, this method may be adapted to engineer cell lines in which any desired glycosylation structure may be obtained.

N-acetylglucosaminyltransferase III expression in lower eukaryotes

Methods and materials are provided for the production of compositions of erythropoietin protein, wherein said compositions comprise a pre-selected N-linked glycosylation pattern as the predominant N-glycoform.

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Grantham, NH, United States of America

Juergen Nett