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AssigneeGlycoFi, Inc.
InventorsStefan Wildt, Robert Miele, Juergen Nett, Robert Davidson
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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 GlcNAcMan3GlcNAc2 core 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.
A method for producing a human-like glycoprotein in a non-human eukaryotic host cell comprising the step of diminishing or depleting the activity of one or more enzymes in the host cell that transfers a sugar residue to the 1,6 arm of a lipid-linked oligosaccharide structure.
A nucleic acid molecule comprising or consisting of at least forty-five consecutive nucleotide residues of FIG. 6 (P. pastoris ALG 3 gene).
A P.pastoris cell in which the sequences of FIG. 6 (P. pastoris ALG 3 gene), are mutated whereby the glycosylation pattern of the cell is altered.
A method to enhance the degree of glucosylation of lipid-linked oligosaccharides comprising the step of increasing alpha-1,3 glucosyltransferase activity in a host cell.
A method to enhance the degree of glucosylation of lipid-linked oligosaccharides comprising decreasing the substrate specificity of oligosaccharyl transferase activity in a host cell.
A method for producing in a non-mammalian host cell an immunoglobulin polypeptide having an N-glycan comprising a bisecting GlcNAc, the method comprising the step of expressing in the host cell a GnTIII activity.
A non-mammalian host cell that produces an immunoglobulin having an N-glycan comprising a bisecting GlcNAc.
A method for producing in a non-human host cell a polypeptide having an N-glycan comprising a bisecting GlcNAc, the method comprising the step of expressing in the host cell a GnTIII activity.
A non-human host cell that produces a polypeptide having an N-glycan comprising a bisecting GlcNAc.
A method for producing a human-like glycoprotein in a non-human eukaryotic host cell comprising the step of diminishing or depleting from the host cell an alg gene activity and introducing into the host cell at least one glycosidase activity.
A method for producing a human-like glycoprotein having an N-glycan comprising at least two GlcNAcs attached to a trimannose core.
Dependencies are inferred from the claim text.