Multichain polypeptides or proteins and processes for their production

• US 4,816,397 A
• Filed: 11/14/1984
• Issued: 03/28/1989
• Est. Priority Date: 03/25/1983
• Status: Expired due to Term

• Current Assignee: Celltech Group Limited (UCB SA)
• Original Assignee: Celltech Group Limited (UCB SA)
• Inventors: Wood, Clive R., Kenten, John H., Boss, Michael A., Emtage, John S.
• Primary Examiner(s): Hazel, Blondel
• Application Number:
  US06/672,265
• Time in Patent Office: 1,595 Days
• Field of Search: 435/68, 435/172.3, 435/243, 435/320, 435/253, 435/255
• US Class Current: 435/69.6
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: A61P 7/00 Drugs for disorders of the blood or the extracellular fluid
• CPC Class Codes: B01D 15/3809 of the antigen-antibody type, e.g. protein A, G, L chromatography
• CPC Class Codes: C07K 16/462 Igs containing a variable region (Fv) from one specie and a constant region (Fc) from another
• CPC Class Codes: C07K 2317/24 containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
• CPC Class Codes: C12N 15/63 Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
• CPC Class Codes: G01N 33/531 Production of immunochemical test materials

Abstract

Multichain polypeptides or proteins and processes for their production in cells of host organisms which have been transformed by recombinant DNA techniques. According to a first aspect of the present invention, there is provided a process for producing a heterologous multichain polypeptide or protein in a single host cell, which comprises transforming the host cell with DNA sequences coding for each of the polypeptide chains and expressing said polypeptide chains in said transformed host cell. According to another aspect of the present invention there is provided as a product of recombinant DNA technology an Ig heavy or light chain or fragment thereof having an intact variable domain. The invention also provides a process for increasing the level of protein expression in a transformed host cell and vectors and transformed host cells for use in the processes.

Claims

1. A process for producing an Ig molecule or an immunologially functional Ig fragment comprising at least the variable domains of the Ig heavy and light chains, in a single host cell, comprising the steps of:

(i) transforming said single host cell with a first DNA sequence encoding at least the variable domain of the Ig heavy chain and a second DNA sequence encoding at least the variable domain of the Ig light chain, and

(ii) independently expessing said first DNA sequence and said second DNA sequence so that said Ig heavy and light chains are produced as separate molecules in said transformed single host cell.

2. The process according to claim 1 wherein said first and second DNA sequences are present in different vectors.

3. The process according to claim 1 wherein said first and second DNA sequences are present in a single vector.

4. A process according to claim 2 wherein the or each vector is a plasmid.

5. A process according to claim 4, wherein the plasmid is col E1, pcR1, pBR322, RP4 or phage DNA.

6. A process according to claim 1 wherein the host cell is a baceterium or a yeast.

7. A process according to claim 6, wherein the host cell is E. Coli, B. subtilis or S. cerevisiae.

8. A process according to claim 7, wherein the host cell is E. Coli strain HB101, X1776, X2887, PS410, MRC 1 RV308, B or E1035.

9. A process according to claim 1 wherein the Ig heavy and light chains are expressed in the host cell and secreted therefrom as an immunologically functional Ig molecule or Ig fragment.

10. A process according to claim 1 wherein the Ig heavy and light chains are produced in insoluble or membrane bound form and are solubilised and allowed to refold in solution to form an immunologically functional Ig molecule or Ig fragment.

11. A process according to claim 1, wherein the DNA sequences code for the complete Ig heavy and light chains.

12. A process according to claim 1, wherein said first or said second DNA sequence further encodes at least one constant domain, wherein the constant domain is derived from the same source as the variable domain to which it is attached.

13. A process according to claim 1, wherein said first or said second DNA sequence further encodes at least one constant domain, wherein the constant domain is derived from a source different from that from which the variable domain to which it is attached is derived.

14. A process according to claim 1, wherein said first and second DNA sequences are derived from one or more monoclonal antibody producing hybridomas.

15. A vector comprising a first DNA sequence encoding at last a variable domain of an Ig heavy chain and a second DNA sequence encoding at least a variable domain of an Ig light chain wherein said first DNA sequence and said second DNA sequence are located in said vector at different insertion sites.

16. A vector according to claim 15, which is a plasmid.

17. A host cell transformed with a vector according to claims 15.

18. A transformed host cell comprising at least two vectors, at least one of said vectors comprising a DNA sequence encoding at least a variable domain of an Ig heavy chain and at least another one of said vectors comprising a DNA sequence encoding at least the variable domain of an Ig light chain.