Expression and secretion system

Patent metadata

Field	Value
Publication number	US-10633650-B2
Application number	US-201715690544-A
Country	US
Kind code	B2
Filing date	Aug 30, 2017
Priority date	Jul 5, 2012
Publication date	Apr 28, 2020
Grant date	Apr 28, 2020

Abstract

The invention provides an expression and secretion system, and methods of using the same, for the expression and secretion of one fusion protein in prokaryotic cells and a second fusion protein in eukaryotic cells. Also provided herein are nucleic acid molecules, vectors and host cells comprising such vectors and nucleic acid molecules.

First claim

The invention claimed is:

1. A nucleic acid molecule comprising:
(a) a variable light chain (VL) domain-encoding sequence that encodes a VL domain, wherein the VL domain-encoding sequence is 3′ to a first signal sequence that encodes a first signal peptide;
(b) a variable heavy chain (VH) domain-encoding sequence that encodes a VH domain, wherein the VH domain-encoding sequence is 3′ to a second signal sequence that encodes a second signal peptide, wherein each of the first signal peptide and the second signal peptide is functional in both a prokaryotic cell and a eukaryotic cell;
(c) an Fc region-encoding sequence that encodes an Fc region;
(d) a prokaryotic promoter and a eukaryotic promoter, wherein the prokaryotic promoter and the eukaryotic promoter are each operably linked to the VH domain-encoding sequence and/or the VL domain-encoding sequence;
(e) a splice donor 3′ to the VH domain-encoding sequence; and
(f) a splice acceptor 5′ to the Fc region-encoding sequence.

2. The nucleic acid molecule of claim 1 , wherein the prokaryotic promoter is phoA, Tac, Tphac, or Lac promoter.

3. The nucleic acid molecule of claim 1 , wherein the eukaryotic promoter is CMV or SV40.

4. The nucleic acid molecule of claim 1 , wherein the prokaryotic cell is a bacteria cell.

5. The nucleic acid molecule of claim 1 , wherein the eukaryotic cell is a mammalian cell.

6. The nucleic acid molecule of claim 4 , wherein the bacteria cell is an E. coli cell.

7. The nucleic acid molecule of claim 5 , wherein the mammalian cell is a yeast cell, CHO cell, 293 cell, or NSO cell.

8. A vector comprising the nucleic acid molecule of claim 1 .

9. A host cell transformed with the vector of claim 8 .

10. The host cell of claim 9 , wherein the host cell is a bacteria cell.

11. The host cell of claim 10 , wherein the bacteria cell is an E. coli cell.

12. The host cell of claim 9 , wherein the host cell is a eukaryotic cell.

13. The host cell of claim 12 , wherein the eukaryotic cell is a yeast cell, CHO cell, 293 cell, or NSO cell.

14. A process for producing a polypeptide, the process comprising culturing the host cell of claim 9 so that the polypeptide is expressed.

15. The process of claim 14 , further comprising recovering the polypeptide expressed by the host cell.

16. The process of claim 15 , wherein the polypeptide is recovered from the host cell culture medium.

17. The nucleic acid molecule of claim 1 , wherein the nucleic acid molecule encodes an antibody.

18. The nucleic acid molecule of claim 1 , wherein the VL domain is linked to a control protein and the VH domain is linked to the Fc region.

19. The nucleic acid molecule of claim 18 , wherein the control protein is a gD protein or a fragment thereof.

Assignees

• Genentech Inc

Inventors

• Tesar Devin
• Chen Xiaocheng
• Dennis Mark
• Hotzel Isidro

Classifications

• C12N15/85 Primary

  for animal cells · CPC title

• C07K16/00 Primary

  Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies · CPC title

• C07K2319/02

  containing a signal sequence · CPC title

• C07K2319/40

  containing a tag for immunodetection, or an epitope for immunisation · CPC title

• C12N2830/55

  from bacteria · CPC title