Recombinant production of protein having xylosylated n-glycans
AssigneeExpres2ion Biotechnologies ApS
InventorsMagdalena Skrzypczak, Ida Busch Nielsen, Stine Broch Clemmensen, Willem Adriaan De Jongh
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Abstract
Provided are non-plant polypeptides or proteins comprising xylosylated N-linked glycans that comprise β1,2-xylose, wherein at least 25% of the individual protein species comprise xylosylated N-glycans, compositions comprising the non-plant proteins, cells engineered to produce the polypeptide or proteins, and methods of their production and use.
Claims24
A non-plant polypeptide or protein comprising xylosylated N-linked glycans that comprise β1,2-xylose, wherein at least 25% of the individual protein species comprise xylosylated N-glycans, wherein said polypeptide or protein preferably is selected from the group consisting of VAR2CA, HER2 and hEPO;
- a viral protein or polypeptide from HIV, Ebola virus, Zika virus, Chikungunya virus, Dengue virus, Hepatitis A virus, influenza virus, Polio virus, Rabies virus, Measles virus, mumps virus, rubella virus, Rotavirus virus, Smallpox virus, Chickenpox virus, Hepatitis B virus, human papillomavirus, varicella zoster virus, Yellow fever virus, SARS-CoV-1, cytomegalovirus (CMV), and SARS-CoV-2;
- and a bacterial protein or polypeptide from Clostridium tetanii, Corynebacterium diphtheria, Haemophilus influenzae, Bordetella pertussis, Streptococcus pneumoniae, and Neisseria meningitides.
The non-plant polypeptide or protein according to any one of the preceding claims, which of mammalian, crustacean, insect, arachnoid, viral, bacterial, fungal, helminthic, or protozoan origin.
The non-plant polypeptide or protein according to any one of the preceding claims, wherein at least 25% of individual protein/peptide species comprise xylosylated N-glycans, such as at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, 75%, and at least 90%.
The non-plant polypeptide or protein according to any one of the preceding claims, which is obtainable by a method comprising expressing polynucleotide(s) encoding the polypeptide or protein in an S2 cell genetically altered so as to produce an active β1,2-xylosyltransferase and optionally an active UDP-xylose synthase or an increased amount of active UDP-xylose synthase.
The non-plant polypeptide or protein according to any one of the preceding claims, further comprising α1,3-linked fucose.
The non-plant polypeptide or protein according to any one of the preceding claims, which further comprises or is coupled via a non-peptide bond to a heterologous moiety, such as a purification tag, an immunogenic carrier molecule or T-helper lymphocyte epitope, a solubility-modifying group, a protraction group, a targeting moiety, a virus-like particle, and an immune modulating moiety, said heterologous moiety optionally being fused to the polypeptide or protein via a peptide linker.
An immunogenic composition comprising the non-plant polypeptide or protein according to any one of the preceding claims in admixture with at least one immunological adjuvant and optionally a pharmaceutically acceptable carrier and/or diluent and/or excipient.
A genetically modified non-plant eukaryotic cell, such as a mammalian cell or an insect cell or a fungal cell such as a yeast, which comprises at least one heterologous polynucleotide sequence encoding and expressing β1,2-xylosyltransferase, wherein the cell is capable of producing N-glycosylated protein carrying β1,2-xylose groups, and where the heterologous polynucleotide sequence preferably is the XylT gene from Arabidobsis thaliana, or an equivalent polynucleotide encoding a plant β1,2-xylosyltransferase.
Dependencies are inferred from the claim text.