Chemically defined non-polymeric valency platvorm molecules and conjugates thereof

• US 20050026856A1
• Filed: 07/30/2003
• Published: 02/03/2005
• Est. Priority Date: 07/15/1992
• Status: Active Grant

• Current Assignee: La Jolla Pharmaceutical Company (Innoviva, Inc.)
• Original Assignee: La Jolla Pharmaceutical Company (Innoviva, Inc.)
• Inventors: Yu, Lin, Jones, David S., Livingston, Douglas Alan, Coutts, Stephen M.
• Granted Patent: US 7,351,855 B2
• Time in Patent Office: Days
• US Class Current: 514/44.R
• CPC Class Codes: A61K 2039/6093 Synthetic polymers, e.g. polyethyleneglycol [PEG], Polymers or copolymers of (D) glutamate and (D) lysine
• CPC Class Codes: A61K 39/0008 Antigens related to auto-immune diseases; Preparations to induce self-tolerance
• CPC Class Codes: A61K 39/35 Allergens
• CPC Class Codes: A61K 47/54 the modifying agent being an organic compound

Abstract

Chemically-defined, non-polymeric valency platform molecules and conjugates comprising chemically-defined valency platform molecules and biological or chemical molecules including polynucleotide duplexes of at least 20 base pairs that have significant binding activity for human lupus anti-dsDNA autoantibodies.

Claims

1. A conjugate of at least one biological molecule and a valency platform, said valency platform having the formula:

Q₁-(CH₂CH₂O)_nCH₂CH₂-Q₁

wherein each Q₁ is a chemical moiety independently selected from the group consisting of;

wherein each Q₂ is a chemical moiety independently selected from the group consisting of;

wherein each Q₃ is a chemical moiety independently selected from the group consisting of;

wherein each T is a chemical moiety independently selected from the group consisting of;

wherein;

n is an integer from 0 to 300;

m is in integer from 1 to 10;

p is an integer from 1 to 10;

each X is independently F, Cl, Br, I, or other good leaving group;

each R^ALK is independently a linear, branched, or cyclic alkyl group of 1 to 20 carbon atoms;

each R^SUB is independently H, a linear, branched, or cyclic alkyl group of 1 to 20 carbon atoms, an aryl group of 6 to 20 carbon atoms, or an alkaryl group of 7 to 30 carbon atoms;

each R^ESTER is independently N-succinimidyl, p-nitrophenyl, pentafluorophenyl, tetrafluorophenyl, 2,4,5-trichlorophenyl, 2,4-dinitrophenyl, cyanomethyl, or other activating group; and

each R^B is independently a chemical moiety comprising 1 to 50 atoms selected from the group consisting of C, H, N, O, Si, P, and S.

2. A conjugate according to claim 1, wherein each Q₂ is:

3. A conjugate according to claim 2, wherein each Q₁ is:

4. A conjugate according to claim 2, wherein each Q₁ is:

5. A conjugate according to claim 2, wherein each Q₁ is:

6. A conjugate according to any one of claims 3 to 5, wherein each Q₃ is a chemical moiety independently selected from the group consisting of:

7. A conjugate according to any one of claims 3 to 5, wherein each Q₃ is a chemical moiety independently selected from the group consisting of:

8. A conjugate according to claim 6, wherein each T is a chemical moiety independently selected from the group consisting of:

9. A conjugate according to claim 7, wherein each T is a chemical moiety independently selected from the group consisting of:

10. A conjugate according to any one of claims 1 to 5, wherein each said biological molecule is selected from the group consisting of a carbohydrate, a lipid, a lipopolysaccharide, a peptide, a protein, a glycoprotein, a single stranded oligonucleotide, a double-stranded oligonucleotide, a hapten, and an analog thereof or a combination thereof.

11. A conjugate according to claim 8, wherein each said biological molecule is selected from the group consisting of a carbohydrate, a lipid, a lipopolysaccharide, a peptide, a protein, a glycoprotein, a single stranded oligonucleotide, a double-stranded oligonucleotide, a hapten, and an analog thereof or a combination thereof.

12. A conjugate according to claim 9, wherein each said biological molecule is selected from the group consisting of a carbohydrate, a lipid, a lipopolysaccharide, a peptide, a protein, a glycoprotein, a single stranded oligonucleotide, a double-stranded oligonucleotide, a hapten, and an analog thereof or a combination thereof.

13. A conjugate according to any one of claims 1 to 5, wherein each said biological molecule is an analog of an immunogen wherein (a) said analog binds specifically to B cells to which said immunogen binds specifically and (b) said conjugate lacks a T cell epitope.

14. A conjugate according to claim 8, wherein each said biological molecule is an analog of an immunogen wherein (a) said analog binds specifically to B cells to which said immunogen binds specifically and (b) said conjugate lacks a T cell epitope.

15. A conjugate according to claim 7, wherein each said biological molecule is an analog of an immunogen wherein (a) said analog binds specifically to B cells to which said immunogen binds specifically and (b) said conjugate lacks a T cell epitope.

16. A conjugate according to any one of claims 1 to 5, comprising a plurality of biological molecules, wherein each of said biological molecules comprises a polynucleotide duplex, said duplex having a significant binding activity for human systemic lupus erythematosus anti-double stranded DNA autoantibodies.

17. A conjugate according to claim 16, wherein each of said duplexes are coupled to said valency platform via a linker group.

18. A conjugate according to claim 16, wherein each of said duplexes are coupled to said valency platform via a linker group derived from a thio-6 carbon chain phosphate or a thio-6 carbon chain phosphorothioate.

19. A conjugate according to claim 16, wherein said duplexes are substantially homogeneous in length.

20. A conjugate according to claim 16, wherein said duplexes are substantially homogeneous in nucleotide composition.

21. A conjugate according to claim 16, wherein said duplexes are 20 to 50 base pairs in length.

22. A conjugate according to claim 16, wherein said duplexes are covalently bonded to said valency platform at or proximate to one of their ends.

23. A conjugate according to claim 16, wherein said conjugate suppresses antibody production associated with human systemic lupus erythematosus.

24. A conjugate according to claim 16, wherein said duplexes have a B-DNA helical structure and a significant binding activity for human systemic lupus erythematosus anti-double stranded DNA autoantibodies.

25. A composition comprising a conjugate according to claim 16 formulated with a pharmaceutically acceptable injectable vehicle.

26. A composition comprising a conjugate according to claim 13 formulated with a pharmaceutically acceptable injectable vehicle.

27. A method of suppressing antibody production in an individual in need thereof comprising administering the composition according to claim 43 to the individual in an effective amount such that antibody production is suppressed.

28. A method of suppressing antibody production in an individual in need thereof, said method comprising administering the composition according to claim 43 to the individual in an effective amount such that antibody production contributing to thyroiditis, stroke, or myasthenia gravis is suppressed.

29. A method of suppressing antibody production in an individual in need thereof comprising administering the composition according to claim 44 to the individual in an effective amount such that antibody production contributing to human systemic lupus erythematosus is suppressed.

30. A method of making a conjugate according to claim 16, said method comprising the steps of:

(a) reacting a plurality of first single-stranded polynucleotides to said valency platform for a time and under conditions effective to form a conjugate between the valency platform and the plurality of first single-stranded polynucleotides;

(b) annealing a complementary single-stranded polynucleotide to said first single-stranded polynucleotides for a time and under conditions effective to form the conjugate according to claim 66; and

(c) isolating the conjugate of step (b).

31. A conjugate according to claim 13, wherein the valency platform molecule is derivatized by a reagent selected from the group consisting of 3,5-bis-(iodoacetamido) benzoyl chloride, 3-carboxypropionamide-N,N-bis[(6‘

-N’

-carboxybenzyloxyaminohexyl)acetamide] 4″

-nitrophenyl ester, 3-carboxypropionamide-N,N-bis-[(8‘

-N’

-carbobenzyloxyamino-3′

,6′

-dioxaoctyl)acetamide]4″

-nitrophenyl ester, and N,N-di(2-[6‘

-N’

-carbobenzyloxy-aminohexanoamido]ethyl)amine.

32. A conjugate according to claim 16, wherein the polynucleotide duplex is (CA)₁₀:

(TG)₁₀ and the valency platform molecule is derivatized triethylene glycol.

33. A conjugate according to claim 32, wherein the molar ratio of duplex to valency platform molecule is in the range of 2:

1 to 8;

1.

34. A conjugate according to claim 32, wherein the molar ratio of duplex to valency platform molecule is 4:

1.

35. A conjugate according to claim 16, wherein the polynucleotide duplex is (CA)₁₀:

(TG)₁₀ and the valency platform molecule is derivatized 2,2′

-ethylenedioxydiethylamine.

36. A conjugate according to claim 35, wherein the molar ratio of duplex to valency platform molecule is in the range of 2:

1 to 8;

1.

37. A conjugate according to claim 35, wherein the molar ratio of duplex to valency platform molecule is 4:

1.

38. A conjugate according to claim 13, wherein the immunogen is an external immunogen.

39. A conjugate according to claim 38, wherein the external immunogen is a biological drug, an allergen or a Rhesus (Rh/D) immunogen associated with Rh hemolytic disease, α

-sperm associated with male infertility or the carbohydrate complex associated with rheumatic fever.

40. A conjugate according to claim 13, wherein the immunogen is a self-immunogen.

41. A conjugate according to claim 38, wherein the self-immunogen contributes to thyroiditis, diabetes, stroke or myasthenia gravis.

42. A conjugate according to claim 13, wherein the immunogen and the analog are polypeptides.

43. A composition comprising a conjugate according to claim 13 formulated with a pharmaceutically acceptable carrier.

44. A composition comprising a conjugate according to claim 16 formulated with a pharmaceutically acceptable carrier.

45. The conjugate according to claim 16 wherein said polynucleotide duplex contains at least 20 base pairs.