Materials and methods for improved immunoglycoproteins
View Patent ↗Immunoglycoproteins, including antibodies, with improved ADCC and altered glycosylation patterns are provided. Also provided are cell culturing methods and media for producing such immunoglycoproteins, and therapeutic uses of such immunoglycoproteins.
1. A composition comprising recombinant immunoglycoprotein molecules comprising a N-linked glycosylation site for linkage of oligosaccharide, wherein said immunoglycoprotein molecules have a glucose content characterized by a ratio of glucose molecules per said N-linked glycosylation site of at least 1.5.
2. The composition of claim 1 , wherein said ratio is at least 2.
3. The composition of claim 1 , wherein said ratio is at least 2.5.
4. The composition of any of claims 1 , 2 , and 3 wherein said recombinant immunoglycoprotein molecules in a reduced state each comprise a single N-linked glycosylation site.
5. A composition of recombinant immunoglycoprotein molecules comprising a N-linked glycosylation site for linkage of oligosaccharide, wherein at least 60% of said N-linked oligosaccharides of said N-linked glycosylation site have a glucose content from two to three glucose molecules.
6. A composition of recombinant immunoglycoprotein molecules comprising a N-linked glycosylation site for linkage of oligosaccharide, wherein at least 60% of said N-linked oligosaccharides of said N-linked glycosylation site have a glucose content of three glucose molecules.
7. A composition of any of claims 1 , 2 , 3 , 5 , and 6 wherein said immunoglycoprotein molecules exhibit at least 5-fold higher ADCC compared to control immunoglycoprotein molecules of the same encoded amino acid sequence produced in CHO-K1 cells in the absence of carbohydrate modifier.
8. A composition of any of claims 1 , 2 , 3 , 5 , and 6 wherein at least 60% of the N-linked oligosaccharides of said N-linked glycosylation site contain no fucose.
9. A composition comprising recombinant immunoglycoprotein molecules comprising a N-linked glycosylation site for linkage of oligosaccharide wherein said molecules exhibit the following characteristics:
a) said immunoglycoprotein molecules have a glucose content characterized by a ratio of glucose molecules per said N-linked glycosylation site of at least 1.5;
b) at least 60% of said N-linked oligosaccharides of said N-linked glycosylation site have a glucose content from two to three glucose molecules;
c) at least 5-fold higher ADCC compared to control immunoglycoprotein molecules of the same encoded amino acid sequence produced in CHO-K1 cells in the absence of castanospermine; and
d) at least 60% of the N-linked oligosaccharides of said N-linked glycosylation site contain no fucose.
10. A composition of any of claims 1 , 2 , 5 , 6 , and 9 and a sterile pharmaceutically acceptable carrier or diluent.
11. The composition of claim 10 which is sterile.
12. A composition of any of claims 1 , 2 , 5 , 6 , and 9 comprising at least 100 liters of culture medium.
13. A composition of any of claims 1 , 2 , 5 , 6 , and 9 comprising at least 100 g of immunoglycoprotein molecules and a pharmaceutically acceptable carrier or diluent.
14. The composition of claim 10 wherein the recombinant immunoglycoprotein molecules are bispecific in antigen-binding.
15. The composition of claim 10 wherein the recombinant immunoglycoprotein molecules are trispecific in antigen-binding.
16. The composition of any of claims 1 , 2 , 5 , 6 , and 9 wherein the recombinant immunoglycoprotein molecule is a small modular immunopharmaceutical.
17. The composition of claim 16 wherein the small modular immunopharmaceutical comprises amino acids 24-496 of SEQ ID NO: 2.