Anti-VEGF protein compositions and methods for producing the same
The present disclosure pertains to compositions comprising anti-VEGF proteins.
1. An oxo-aflibercept composition produced in a cell culture media comprising a chemically defined media (CDM), wherein one or more amino acid residues of aflibercept is oxidized and
wherein said one or more amino acid residues is histidine and/or tryptophan, and
wherein said oxo-aflibercept has a glycosylation profile based on molar percentage selected from the group consisting of about 40% to about 50% total fucosylated glycans, about 30% to about 55% total sialylated glycans, about 2% to about 15% mannose-5, and about 60% to about 79% galactosylated glycans.
2. The oxo-aflibercept composition of claim 1 , wherein said oxo-aflibercept has about 40% to about 50% total fucosylated glycans.
3. The oxo-aflibercept composition of claim 1 , wherein said oxo-aflibercept has about 30% to about 55% total sialylated glycans.
4. The oxo-aflibercept composition of claim 1 , wherein said oxo-aflibercept has about 2% to about 15% mannose-5.
5. The oxo-aflibercept composition of claim 1 , wherein said oxo-aflibercept has about 60% to about 79% galactosylated glycans.
6. The oxo-aflibercept composition of claim 1 , wherein said oxo-aflibercept is glycosylated at one or more Asparagines as follows: G0-GlcNAc glycosylation; G1-GlcNAc glycosylation; G1S-GlcNAc glycosylation; G0 glycosylation; G1 glycosylation; G1S glycosylation; G2 glycosylation; G2S glycosylation; G2S2 glycosylation; G0F glycosylation; G2F2S glycosylation; G2F2S2 glycosylation; G1F glycosylation; G1FS glycosylation; G2F glycosylation; G2FS glycosylation; G2FS2 glycosylation; G3FS glycosylation; G3FS3 glycosylation; G0-2GlcNAc glycosylation; Man4 glycosylation; Man4_A1G1 glycosylation; Man4_A1G1S1 glycosylation; Man5 glycosylation; Man5_A1G1 glycosylation; Man5_A1G1S1 glycosylation; Man6 glycosylation; Man6_G0+Phosphate glycosylation; Man6+Phosphate glycosylation; and/or Man7 glycosylation.
7. A culture media composition comprising chemically defined media (CDM) and oxo-aflibercept, wherein one or more amino acid residues of aflibercept is oxidized and
wherein said one or more amino acid residues is histidine and/or tryptophan, and
wherein said oxo-aflibercept has a glycosylation profile based on molar percentage selected from the group consisting of about 40% to about 50% total fucosylated glycans, about 30% to about 55% total sialylated glycans, about 2% to about 15% mannose-5, and about 60% to about 79% galactosylated glycans.
8. The oxo-aflibercept composition of claim 7 , wherein said oxo-aflibercept has about 40% to about 50% total fucosylated glycans.
9. The oxo-aflibercept composition of claim 7 , wherein said oxo-aflibercept has about 30% to about 55% total sialylated glycans.
10. The oxo-aflibercept composition of claim 7 , wherein said oxo-aflibercept has about 2% to about 15% mannose-5.
11. The oxo-aflibercept composition of claim 7 , wherein said oxo-aflibercept has about 60% to about 79% galactosylated glycans.
12. The oxo-aflibercept composition of claim 7 , wherein said oxo-aflibercept is glycosylated at one or more Asparagines as follows: G0-GlcNAc glycosylation; G1-GlcNAc glycosylation; G1S-GlcNAc glycosylation; G0 glycosylation; G1 glycosylation; G1S glycosylation; G2 glycosylation; G2S glycosylation; G2S2 glycosylation; G0F glycosylation; G2F2S glycosylation; G2F2S2 glycosylation; G1F glycosylation; G1FS glycosylation; G2F glycosylation; G2FS glycosylation; G2FS2 glycosylation; G3FS glycosylation; G3FS3 glycosylation; G0-2GlcNAc glycosylation; Man4 glycosylation; Man4_A1G1 glycosylation; Man4_A1G1S1 glycosylation; Man5 glycosylation; Man5_A1G1 glycosylation; Man5_A1G1S1 glycosylation; Man6 glycosylation; Man6_G0+Phosphate glycosylation; Man6+Phosphate glycosylation; and/or Man7 glycosylation.
13. The oxo-aflibercept composition of claim 7 , wherein the N-glycosylation of the F(ab′) 2 portion of said oxo-aflibercept at Asn36 is between 98.3% and 99.4%.
14. The oxo-aflibercept composition of claim 7 , wherein the N-glycosylation of the F(ab′) 2 portion of said oxo-aflibercept at Asn68 is between 51.9% and 64.9%.
15. The oxo-aflibercept composition of claim 7 , wherein the N-glycosylation of the F(ab′) 2 portion of said oxo-aflibercept at Asn123 is between 98.4% and 99.9%.
16. The oxo-aflibercept composition of claim 7 , wherein the N-glycosylation of the F(ab′) 2 portion of said oxo-aflibercept at Asn196 is between 55.1% and 98.6%.
17. A composition comprising oxo-aflibercept, wherein one or more amino acid residues of aflibercept is oxidized and
wherein said one or more amino acid residues is histidine and/or tryptophan, and
wherein said oxo-aflibercept has a glycosylation profile based on molar percentage selected from the group consisting of about 40% to about 52.4% total fucosylated glycans, about 30% to about 59.2% total sialylated glycans, about 5.2% to about 15% mannose-5, and about 60% to about 80.6% galactosylated glycans.
18. The oxo-aflibercept composition of claim 17 , wherein said oxo-aflibercept has about 40% to about 50% total fucosylated glycans.
19. The oxo-aflibercept composition of claim 17 , wherein said oxo-aflibercept has about 30% to about 55% total sialylated glycans.
20. The oxo-aflibercept composition of claim 17 , wherein said oxo-aflibercept has about 2% to about 15% mannose-5.
21. The oxo-aflibercept composition of claim 17 , wherein said oxo-aflibercept has about 60% to about 79% galactosylated glycans.
22. The oxo-aflibercept composition of claim 17 , wherein said oxo-aflibercept has about 51.4% to about 52.4% total fucosylated glycans.
23. The oxo-aflibercept composition of claim 17 , wherein said oxo-aflibercept has about 58.2% to about 59.2% total sialylated glycans.
24. The oxo-aflibercept composition of claim 17 , wherein said oxo-aflibercept has about 5.19% to about 5.24% mannose-5.
25. The oxo-aflibercept composition of claim 17 , wherein said oxo-aflibercept has about 79.6% to about 80.6% galactosylated glycans.
26. The oxo-aflibercept composition of claim 17 , wherein said oxo-aflibercept is glycosylated at one or more Asparagines as follows: G0-GlcNAc glycosylation; G1-GlcNAc glycosylation; G1S-GlcNAc glycosylation; G0 glycosylation; G1 glycosylation; G1S glycosylation; G2 glycosylation; G2S glycosylation; G2S2 glycosylation; G0F glycosylation; G2F2S glycosylation; G2F2S2 glycosylation; G1F glycosylation; G1FS glycosylation; G2F glycosylation; G2FS glycosylation; G2FS2 glycosylation; G3FS glycosylation; G3FS3 glycosylation; G0-2GlcNAc glycosylation; Man4 glycosylation; Man4_A1G1 glycosylation; Man4_A1G1S1 glycosylation; Man5 glycosylation; Man5_A1G1 glycosylation; Man5_A1G1S1 glycosylation; Man6 glycosylation; Man6_G0+Phosphate glycosylation; Man6+Phosphate glycosylation; and/or Man7 glycosylation.
27. The oxo-aflibercept composition of claim 13 , wherein the N-glycosylation of F(ab′) 2 portion of said oxo-aflibercept at SEQ ID NO. 100 is between 98.3% and 99.4%.
28. The oxo-aflibercept composition of claim 14 , wherein the N-glycosylation of F(ab′) 2 portion of said oxo-aflibercept at SEQ ID NO. 101 is between 51.9% and 64.9%.
29. The oxo-aflibercept composition of claim 15 , wherein the N-glycosylation of F(ab′) 2 portion of said oxo-aflibercept at SEQ ID NO. 102 is between 98.4% and 99.9%.
30. The oxo-aflibercept composition of claim 16 , wherein the N-glycosylation of F(ab′) 2 portion of said oxo-aflibercept at SEQ ID NO. 103 is between 55.1% and 98.6%.