Recombniant method for making leukemia inhibitor factor

• US 5,427,925 A
• Filed: 05/06/1993
• Issued: 06/27/1995
• Est. Priority Date: 04/02/1987
• Status: Expired due to Term

• Current Assignee: EMD Millipore Corporation (Merck & Co., Inc.)
• Original Assignee: Zenyth Therapeutics Limited (CSL Limited)
• Inventors: Gearing, David P., Nicola, Nicos A., Gough, Nicholas M., King, Julie A., Nice, Edouard C., Metcalf, Donald, Willson, Tracy A., Hilton, Douglas J., Simpson, Richard J.
• Primary Examiner(s): Walsh, Stephen G.
• Application Number:
  US08/058,979
• Time in Patent Office: 782 Days
• Field of Search: 435/69.5, 435/69.1, 435/69.8, 435/240.2, 435/252.33, 435/255, 530/351, 530/395
• US Class Current: 435/69.5
• 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 35/02 specific for leukemia
• CPC Class Codes: C07K 14/5415 Leukaemia inhibitory factor [LIF]
• CPC Class Codes: C07K 2319/00 Fusion polypeptide
• CPC Class Codes: C07K 2319/02 containing a signal sequence
• CPC Class Codes: C07K 2319/23 containing a GST-tag
• CPC Class Codes: C07K 2319/50 containing protease site
• CPC Class Codes: C07K 2319/74 containing a fusion for binding to a cell surface receptor
• CPC Class Codes: C07K 2319/95 containing a motif/fusion for degradation (ubiquitin fusions, PEST sequence)
• CPC Class Codes: C12N 15/81 for yeasts
• CPC Class Codes: C12N 15/86 Viral vectors
• CPC Class Codes: C12N 2740/13043 viral genome or elements thereof as genetic vector
• CPC Class Codes: C12P 21/02 having a known sequence of two or more amino acids, e.g. glutathione
• CPC Class Codes: C12P 21/06 produced by the hydrolysis of a peptide bond, e.g. hydrolysate products preparing foodstuffs by protein hydrolysis A23J3/00

Abstract

The present invention provides a method for producing recombinant leukemia inhibitory factor (LIF) by the expression, in suitable host cells, of recombinant DNA molecules encoding LIF.

Claims

1. A method of producing human leukemia inhibitory factor (LIF) comprising the steps of

(a) transforming or transfecting suitable host cells with a recombinant DNA molecule comprising a nucleotide sequence which codes for human LIF,

(b) culturing the host cells of step (a) under conditions in which said cells express the recombinant DNA and produce human LIF, and

c) recovering said human LIF.

2. The method of claim 1 in which said recombinant DNA molecule codes for human LIF having the amino acid sequence set forth in FIG. 26.

3. The method of claim 1 in which said recombinant DNA molecule comprises a nucleotide sequence the complement of a DNA sequence coding for human LIF.

4. The method of claims 1, 2, or 3, wherein the recombinant DNA molecule further comprises a promoter sequence operably linked to said nucleotide sequence to allow expression of the recombinant DNA and production of human LIF by said host cells.

5. The method of claim 4, wherein said nucleotide sequence further codes for a signal sequence contiguous with the amino acid sequence of human LIF, wherein said signal sequence directs secretion of human LIF from said host cells.

6. The method according to claim 5 wherein said signal sequence is cleavable from the human LIF polypeptide such that human LIF is recoverable as a mature biologically active polypeptide.

7. The method of claim 1, wherein the host cell is selected from the group consisting of bacteria, yeast and mammalian cells.

8. The method of claim 7 wherein the host cell is E. coli.

9. The method of claim 7 wherein the host cell is Saccharomyces cerevisiae.

10. The method of claim 7 wherein the host cell is a hematopoietic cell.

11. The method of claims 9 or 10 wherein said human LIF is glycosylated.

12. A method of producing murine leukemia inhibitory factor (LIF) comprising the steps of

(a) transforming or transfecting suitable host cells with a recombinant DNA molecule comprising a nucleotide sequence which codes for murine LIF,

(b) culturing the host cells of step (a) under conditions in which said cells express the recombinant DNA and produce murine LIF, and

(c) recovering said murine LIF.

13. The method of claim 12 in which said recombinant DNA molecule codes for murine LIF having the amino acid sequence set forth in FIG. 26.

14. The method of claim 12 in which said recombinant DNA molecule comprises a nucleotide sequence the complement of a DNA sequence coding for murine LIF.

15. The method of claims 12, 13 or 14, wherein the recombinant DNA molecule further comprises a promoter sequence operably linked to said nucleotide sequence to allow expression of the recombinant DNA and production of murine LIF by said host cells.

16. The method of claim 15, wherein said nucleotide sequence further codes for a signal sequence contiguous with the amino acid sequence of murine LIF, wherein said signal sequence directs secretion of murine LIF from said host cells.

17. The method according to claim 16 wherein said signal sequence is cleavable from the murine LIF polypeptide such that murine LIF is recoverable as a mature biologically active polypeptide.

18. The method of claim 12 wherein the host cell is selected from the group consisting of bacteria, yeast and mammalian cells.

19. The method of claim 18 wherein the host cell is E. coli.

20. The method of claim 18 wherein the host cell is Saccharomyces cerevisiae.

21. The method of claim 18 wherein the host cell is a hematopoietic cell.

22. The method of claims 20 or 21 wherein said murine LIF is glycosylated.