Production of recombinant erythropoietin

• US 5,441,868 A
• Filed: 10/23/1987
• Issued: 08/15/1995
• Est. Priority Date: 12/13/1983
• Status: Expired due to Term

• Current Assignee: Amgen Inc.
• Original Assignee: Kirin-Amgen Inc. (Amgen Inc.)
• Inventors: Lin, Fu-Kuen
• Primary Examiner(s): Martinell, James
• Application Number:
  US07/113,179
• Time in Patent Office: 2,853 Days
• Field of Search: 435/70, 435/69.5, 435/172.3, 435/69.1, 435/69.4, 435/69.6, 435/240.2, 435/320.1, 935/50, 935/13, 536/27, 536/23.5, 536/23.51
• US Class Current: 435/69.4
• 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 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 7/00 Drugs for disorders of the blood or the extracellular fluid
• CPC Class Codes: A61P 7/06 Antianaemics
• CPC Class Codes: C07K 14/505 Erythropoietin [EPO]
• CPC Class Codes: C07K 16/22 against growth factors ; against growth regulators
• CPC Class Codes: C12N 15/70 Vectors or expression systems specially adapted for E. coli
• CPC Class Codes: C12N 15/81 for yeasts
• CPC Class Codes: C12N 15/85 for animal cells
• CPC Class Codes: C12N 2800/108 episomal vectors

Abstract

Disclosed are novel polypeptides possessing part or all of the primary structural conformation and one or more of the biological properties of mammalian erythropoietin ("EPO") which are characterized in preferred forms by being the product of procaryotic or eucaryotic host expression of an exogenous DNA sequence. Illustratively, genomic DNA, cDNA and manufactured DNA sequences coding for part or all of the sequence of amino acid residues of EPO or for analogs thereof are incorporated into autonomously replicating plasmid or viral vectors employed to transform or transfect suitable procaryotic or eucaryotic host cells such as bacteria, yeast or vertebrate cells in culture. Upon isolation from culture media or cellular lysates or fragments, products of expression of the DNA sequences display, e.g., the immunological properties and in vitro and in vivo biological activities of EPO of human or monkey species origins. Disclosed also are chemically synthesized polypeptides sharing the biochemical and immunological properties of EPO. Also disclosed are improved methods for the detection of specific single stranded polynucleotides in a heterologous cellular or viral sample prepared from, e.g., DNA present in a plasmid or viral-borne cDNA or genomic DNA "library".

Claims

1. A process for the production of a glycosylated erythropoietin polypeptide having the in vivo biological property of causing bone marrow cells to increase production of reticulocytes and red blood cells comprising the steps of:

(a) growing, under suitable nutrient conditions, mammalian host cells transformed or transfected with an isolated DNA sequence encoding human erythropoietin; and

(b) isolating said glycosylated erythropoietin polypeptide therefrom.

2. The process according to claim 1 wherein said host cells are CHO cells.

3. The process according to claim 1 wherein said host cells are COS cells.

4. The process according to claim 1 wherein said DNA is cDNA.

5. The process according to claim 1 wherein said DNA is genomic DNA.