RU-2026110003-A - GLYCOENGINEERING CELLS AND METHODS OF APPLICATION

Inventors

• ЛИ, Чжэн, Шин
• ВОН, Чи-Хуэй
• ВАН, Сцзу-Вэнь

Assignees

• РОК БАЙОМЕДИКАЛ, ИНК.

Dates

Publication Date

20260505

Application Date

20240619

Priority Date

20230908

Claims (20)

1. 1. A cell for expressing a sialylated glycoprotein, wherein the cell constitutively and/or controllably expresses an exogenous catalytic peptide of a sialyltransferase and an exogenous catalytic peptide of a galactosyltransferase, wherein the exogenous catalytic peptide of a sialyltransferase and the exogenous catalytic peptide of a galactosyltransferase are translated in close proximity to each other.
2. 2. The cell according to claim 1, characterized in that the exogenous catalytic peptide of sialyltransferase and the exogenous catalytic peptide of galactosyltransferase are expressed in one transcript.
3. 3. The cell of claim 1, comprising a first nucleic acid configured to express an exogenous catalytic peptide of a sialyltransferase and a second nucleic acid configured to express an exogenous catalytic peptide of a galactosyltransferase, wherein the first nucleic acid and the second nucleic acid are transcriptionally controlled by the same promoter.
4. 4. The cell of claim 3, wherein the first nucleic acid and the second nucleic acid are linked to each other by a connecting nucleic acid that is configured to encode a ribosome-shifting peptide.
5. 5. The cell of claim 3, wherein the exogenous catalytic peptide of sialyltransferase and the exogenous catalytic peptide of galactosyltransferase are co-assembled for expression as a fusion protein.
6. 6. The cell of claim 5, wherein the fusion protein comprises a first portion containing a catalytic peptide domain of a sialyltransferase and a second portion containing a catalytic peptide domain of a galactosyltransferase, wherein the first portion and the second portion are connected to each other via a cleavable linker, except wherein the cleavable linker is configured to allow post-translational cleavage of the fusion protein, resulting in the release of the catalytic peptide of a sialyltransferase and the catalytic peptide of a galactosyltransferase as separate proteins upon cleavage.
7. 7. The cell according to claim 3, characterized in that the first nucleic acid is obtained from the ST6Ga11 gene or the PspST gene.
8. 8. The cell according to claim 3, characterized in that the second nucleic acid is obtained from the B4GALT1 gene.
9. 9. The cell according to claim 3, characterized in that the promoter is a constitutive promoter or an activatable promoter.
10. 10. The cell of claim 1, wherein the sialyltransferase is beta-galactoside-alpha-2,6-sialyltransferase 1.
11. 11. The cell according to claim 1, characterized in that the catalytic peptide of galactosyltransferase is beta-1,4-galactosyltransferase 1.
12. 12. The cell according to claim 1, characterized in that the cell is deficient in fucosyltransferase activity.
13. 13. A method for glycoengineering a recombinant glycoprotein, comprising
14. delivering an expression vector to a cell according to any one of claims 1 to 12, wherein the expression vector comprises a payload in the form of a nucleic acid configured to encode a recombinant glycoprotein; and
15. expressing a nucleic acid payload in a cell to thereby produce a plurality of recombinant glycoproteins, wherein at least one of the plurality of recombinant glycoproteins is conjugated to a sialylated glycan.
16. 14. The method according to claim 13, characterized in that the sialylated glycan is an α2-6-sialylated complex type (SCT) glycan.
17. 15. The method according to claim 13, characterized in that the sialylated glycan in the cortex does not contain fucose.
18. 16. The method according to claim 13, wherein at least 50% of the totality of recombinant glycoproteins are conjugated with a sialylated glycan.
19. 17. The method of claim 13, further comprising obtaining the aggregate of recombinant glycoproteins within 200 hours after expression of the nucleic acid payload in the cell.
20. 18. The method according to claim 13, characterized in that the recombinant glycoprotein is an antibody or an antigen-binding fragment thereof.