Recombinant production of polyanionic polymers, and uses thereof

• US 20050118136A1
• Filed: 09/14/2004
• Published: 06/02/2005
• Est. Priority Date: 03/21/2001
• Status: Abandoned Application

• Current Assignee: Cell Therapeutics, Inc.
• Original Assignee: Cell Therapeutics, Inc.
• Inventors: Waggoner, David W. Jr., Leung, David W., Pietz, Gregory E., Bergman, Philip A., Tompkins, Christopher K., Lofquist, Alan
• Application Number:
  US10/939,988
• Publication Number:
  US 20050118136A1
• Time in Patent Office: Days
• US Class Current: 424/85.100
• CPC Class Codes: A61K 47/645 Polycationic or polyanionic oligopeptides, polypeptides or polyamino acids, e.g. polylysine, polyarginine, polyglutamic acid or peptide TAT
• CPC Class Codes: C07K 14/001 by chemical synthesis
• CPC Class Codes: C07K 2319/00 Fusion polypeptide
• CPC Class Codes: C07K 7/06 having 5 to 11 amino acids
• CPC Class Codes: C07K 7/08 having 12 to 20 amino acids gastrins C07K14/595; somatostatins C07K14/655; melanotropins C07K14/68

Abstract

A polyanionic polymer can improve the bioactivity and water-solubility properties of a drug to which it is joined. The inventive method provides a monodispersed preparation of a recombinantly-produced polyanionic polymer that can be easily manipulated, such as lengthened. An active moiety may be chemically or recombinantly joined to a polyanionic polymer to increase its biological half-life and/or solubility. The instant invention also provides a method for targeting the delivery of a polyanionic polymer conjugate or fusion protein to a specific cell type or tissue.

Claims

1. A recombinant fusion protein comprising a polyanionic polypeptide and another polypeptide at either one end or at both ends thereof.

2. The recombinant fusion protein according to claim 1, comprising a first polypeptide at the amino-terminal end of said polyanionic polypeptide and a second polypeptide at the carboxyl-terminal end, wherein said first polypeptide and said second polypeptide are the same or are different.

3. The recombinant fusion protein according to claim 2, wherein each of said first polypeptide and said second polypeptide is selected from the group consisting of a targeting polypeptide and a therapeutic polypeptide.

4. The recombinant fusion protein according to claim 3, wherein said first polypeptide and said second polypeptide are different.

5. The recombinant fusion protein according to claim 1, wherein the other polypeptide is selected from the group consisting of an interferon, interferon-α

, and granulocyte colony stimulating factor.

6. The recombinant fusion protein according to claim 1, wherein the other polypeptide is an anti-angiogenic protein selected from the group consisting of a pigment epithelium-derived factor, vascular endothelial growth inhibitor, the domain 5 region of high molecular weight kininogen, endostatin, restin, plasminogen kringle 1 domain, plasminogen kringle 5 domain, and angiostatin.

7. The recombinant fusion protein according to claim 1, wherein the other polypeptide is a recognition motif, selected from the group consisting of an antibody, an antibody fragment, folate, AGCKNFFWKTFTSC, ALNGREESP, CNGRC, ATWLPPR and CTTHWGFTLC.

8. The recombinant fusion protein according to claim 1, wherein the polyanionic polymer is polyglutamic acid or polyaspartic acid and the other protein is interferon-α

.

11. The recombinant fusion protein according to claim 1, wherein the polyanionic polymer is polyglutamic acid or polyaspartic acid and the other protein is granulocyte colony stimulating factor.

15. The recombinant fusion protein according to claim 1, further comprising a spacer amino acid, selected from the group consisting of glycine, an alanine, a β

-alanine, a glutamate and leucine.

62. The recombinant fusion protein according to claim 1, wherein said polyanionic polypeptide is larger than 10 kD.

9. (canceled)

10. (canceled)

12-14. -14. (canceled)

16-48. -48. (canceled)

49. A method for treating a disease or ailment in an individual comprising administering to said individual an effective amount of a recombinantly-produced polyanionic polymer conjugate or fusion protein, wherein the polyanionic polymer is either polyglutamic acid or polyaspartic acid.

50. The method according to claim 49, wherein the recombinantly-produced polyanionic polymer is conjugated to interferon-α

or granulocyte colony stimulating factor.

51. The method according to claim 49, wherein the recombinantly-produced polyanionic polymer is recombinantly linked to any one of interferon-α

or, granulocyte colony stimulating factor.

52. The method according to claim 49, wherein the polyanionic polymer further comprises at least one spacer selected from the group consisting of a glycine, an alanine, a P-alanine, a glutamate, leucine, or an isoleucine, diols, aminothiols, hydroxythiols, aminoalcohols, and an spacer comprising the formula, —

[NH—

(CHR′

)p-CO]n-, wherein R′

is a side chain of a naturally occurring amino acid, n is an integer between 1 and 10, most preferably between 1 and 3; and

p is an integer between 1 and 10, most preferably between 1 and 3;

hydroxyacids of the general formula —

[O—

(CHR′

)p-CO]n-, wherein R′

is a side chain of a naturally occurring amino acid, n is an integer between 1 and 10, most preferably between 1 and 3; and

p is an integer between 1 and 10, most preferably between 1 and 3.

53. The method according to claim 49, wherein the polyanionic polymer is joined to interferon-a.

56. The method according to claim 49, wherein the polyanionic polymer is joined to granulocyte colony stimulating factor.

54. (canceled)

55. (canceled)

57-61. -61. (canceled)