Methods of using steroid-polyanionic polymer-based conjugated targeted to vascular endothelial cells

• US 5,762,918 A
• Filed: 12/05/1994
• Issued: 06/09/1998
• Est. Priority Date: 03/23/1992
• Status: Expired due to Fees

• Current Assignee: Board of Regents of the University of Texas System (University of Texas System)
• Original Assignee: Board of Regents of the University of Texas System (University of Texas System)
• Inventors: Thorpe, Philip E.
• Primary Examiner(s): Page, Thurman K.
• Application Number:
  US08/307,745
• Time in Patent Office: 1,282 Days
• Field of Search: 424/78.08, 424/78.17, 424/78.18, 424/78.19, 424/78.22, 424/422, 424/423, 424/426, 424/451, 424/78.27
• US Class Current: 424/78.17
• CPC Class Codes: A61K 47/58 obtained by reactions only involving carbon-to-carbon unsaturated bonds, e.g. poly[meth]acrylate, polyacrylamide, polystyrene, polyvinylpyrrolidone, polyvinylalcohol or polystyrene sulfonic acid resin
• CPC Class Codes: A61K 47/61 the organic macromolecular compound being a polysaccharide or a derivative thereof

Abstract

This invention discloses new targeted conjugates for the delivery of a compound, and particularly, a steroid, to vascular endothelial cells. The conjugates comprise two components, preferably linked by a selectively-hydrolyzable bond, such as an acid-labile bond or enzyme-sensitive bond. The first component, a polyanionic polymer, and preferably, a polysulphated polymer such as a heparin-derivative, specifically directs the conjugate to vascular endothelial cells. The second component is a selected agent, such as a steroid, which exerts a specific effect on the target cell following its release. In particular, the present invention provides novel conjugated angiogenesis inhibitors, for use in the treatment of pathogenic conditions including cancer, arthritis, and diabetic blindness. An inhibitor comprising a heparin derivative and the anti-angiogenic steroid, cortisol, is herein shown to be markedly acid-labile, to suppress DNA synthesis and cell migration in human umbilical vein endothelial cells, to retard or abolish (depending pending on the route of injection) the vascularization of sponges in vivo and to retard lung tumor growth in mice by 65%. No adverse effects of the conjugate were detected, and equivalent treatments with a mixture of heparin plus cortisol were significantly less effective in all cases.

Claims

1. A method for affecting angiogenesis in an animal, comprising administering to an animal a biologically effective amount of a pharmacological composition comprising a conjugate that comprises a polymer, said polymer being a heparin, conjugated to a selected steroidal agent or agents by means of a biologically releasable bond formed between a reactive group on the steroidal agent or agents and:

a) a free amino group revealed on the polymer through removal of a sulfate, acetyl or acyl group or groups attached to said amino group;

b) a free carboxyl group on the polymer;

c) a reactive group derivatized onto the polymer;

or

d) a combination of a), b) or c);

wherein the conjugate is further characterized as having a molar ratio of agent/polymer of greater than 1 mole agent/mole of polymer.

2. The method of claim 1, wherein the heparin is a non-anticoagulating heparin derivative.

3. The method of claim 2, wherein the conjugate has less than 2.5% of the anticoagulating activity of native heparin.

4. The method of claim 1, wherein said polymer is conjugated to said selected steroidal agent or agents via a disulfide or trisulfide bond.

5. The method of claim 1, wherein said polymer is conjugated to said selected steroidal agent or agents via a selectively hydrolyzable bond.

6. The method of claim 1, wherein said polymer is conjugated to said selected steroidal agent or agents via an acid-labile bond.

7. The method of claim 6, wherein said acid-labile bond is hydrolyzable at a pH of 4.8 or lower.

8. The method of claim 1, wherein said polymer is conjugated to said selected steroidal agent or agents via an enzyme-sensitive bond.

9. The method of claim 8, wherein said enzyme-sensitive bond is a peptide bond.

10. The method of claim 9, wherein said peptide bond includes sequence Leu-Ala-Leu-Ala.

11. The method of claim 8, wherein said enzyme-sensitive bond is an ester, an amide, a phosphodiester or a glycoside.

12. The method of claim 1, wherein said polymer is derivatized to introduce functional groups permitting the attachment of said selected steroidal agent or agents through said biologically releasable bond.

13. The method of claim 12, wherein said polymer is derivatized to introduce side chains terminating in hydrazide, hydrazine, primary amine or secondary amine groups.

14. The method of claim 13, wherein the derivatized polymer is the condensation product of the polymer and adipic dihydrazide, succinic dihydrazide, hydrazine or hydrazine hydrate, and said selected steroidal agent or agents is conjugated to the polymer through a Schiff'"'"'s base linkage.

15. The method of claim 14, wherein said selected steroidal agent or agents is conjugated to said polymer through a hydrazone or acyl hydrazone bond.

16. The method of claim 15, wherein said selected steroidal agent or agents is conjugated to said polymer through a hydrazide linker having the structure:

space="preserve" listing-type="equation">R--CO--NH--NH.sub.2

wherein R=the polymer.

17. The method of claim 14, wherein said selected steroidal agent or agents is conjugated to said polymer by a hydrazide linker having the structure:

space="preserve" listing-type="equation">R--CO--NH--NH--CO--(CH.sub.2).sub.n --CO--NH--NH.sub.2 ;

wherein n=1-50; and

R=the polymer.

18. The method of claim 1, wherein said molar ratio of steroidal agent:

polymer is at least 5 moles steroidal agent/mole polymer.

19. The method of claim 18, wherein said molar ratio of steroidal agent:

polymer ranges from about 5 to about 80 moles steroidal agent/mole polymer.

20. The method of claim 1, further defined as a method for inhibiting angiogenesis, comprising administering to an animal a conjugate comprising said polymer conjugated to an angiostatic steroidal agent in an amount effective to inhibit angiogenesis in said animal.

21. The method of claim 20, further defined as a method for treating cancer, comprising administering to an animal with a solid tumor a conjugate comprising said polymer conjugated to an angiostatic steroidal agent in an amount effective to inhibit angiogenesis in a solid tumor site of said animal.

22. The method of claim 20, wherein said conjugate comprises a polymer conjugated to a steroid from Table II.

23. The method of claim 1, further defined as a method for stimulating angiogenesis, comprising administering to an animal a conjugate comprising said polymer conjugated to an angiogenic steroidal agent in an amount effective to stimulate angiogenesis in said animal.

24. The method of claim 1, wherein said conjugate is formulated into a pharmacological composition suitable for parenteral or topical administration.

25. The method of claim 1, wherein said animal is a human patient.