Succinylated interleukin-2 for pharmaceutical compositions

• US 4,931,544 A
• Filed: 01/27/1988
• Issued: 06/05/1990
• Est. Priority Date: 09/04/1986
• Status: Expired due to Term

• Current Assignee: Chiron Corporation (Bayer AG)
• Original Assignee: Cetus Corporation (Bayer AG)
• Inventors: Katre, Nandini, Knauf, Michael J.
• Primary Examiner(s): Draper, Garnette D.
• Application Number:
  US07/148,106
• Time in Patent Office: 860 Days
• Field of Search: 530/350, 530/351, 530/404, 530/405, 530/406, 530/402, 530/408, 530/409, 530/410, 604/890, 604/891, 514/2, 514/8, 514/12, 514/21, 424/422, 424/443, 424/464, 484/85.1, 484/85.2
• US Class Current: 530/351
• 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: C07K 14/55 IL-2

Abstract

A pharmaceutical composition is prepared wherein biologically active conjugated interleukin-2 is dissolved in an aqueous carrier medium without the presence of a solubilizing agent. The unconjugated IL-2, which is not water soluble or not readily soluble in water at pH 6-8 without such solubilizing agent, is selectively conjugated to one or more succinyl groups by reaction with succinic anhydride.

Claims

1. A biologically active, conjugated, recombinant interleukin2 that is covalently and selectively conjugated to one or more succinyl moieties, wherein said interleukin-2 in its unconjugated form is hydrophobic and insoluble or not readily soluble in water or an aqueous carrier medium under ambient conditions of room temperature and atmospheric pressure at a pH of between about 6 and 8 in the absence of a solubilizing agent.

2. A pharmaceutical composition comprising a non-toxic, inert, pharmaceutically acceptable aqueous carrier medium in which is dissolved the interleukin-2 of claim 1.

3. The composition of claim 2 wherein the interleukin-2 is conjugated to one or two succinyl moieties.

4. The composition of claim 2 wherein the interleukin 2 is conjugated to more than two succinyl moieties.

5. The composition of claim 2 wherein the medium is at a pH of about 5-8 and contains a buffer.

6. The composition of claim 2 wherein the medium is at a pH of about 6.8-7.8 and contains a buffer.

7. The composition of claim 2 wherein the interleukin-2 is a recombinant interleukin-2 from a human source.

8. The composition of claim 7 wherein the interleukin-2 is an interleukin-2 mutein.

9. The composition of claim 8 wherein said mutein has the amino acid(s) that are at position 104 or 125 or positions 104 and 125 in the native protein substituted with a neutral amino acid.

10. The composition of claim 9 wherein said mutein is ser₁₂₅ IL-2, des-ala-₁ ser₁₂₅ IL-2, des-ala₁ ala-₁₀₄ IL-2, or des-ala₁ ala₁₀₄ ser₁₂₅ IL-2.

17. The composition of claim 1 that is formulated as an injectable microcapsule that with a single injection delivers the IL-2 in a total dose of about 2×

10⁵ to about 4×

10⁷ units for 15 to 30 days.

18. The composition of claim 1 that is formulated as an injectable microcapsule that with a single injection delivers the IL-2 in an amount of about 10³ to 10² units/kg/day.

19. The composition of claim 1 that is formulated as an injectable microcapsule comprising a powder of particles 20 to 100 μ

m in diameter that comprise 0.5 to 5% of said IL-2 encapsulated in a biodegrabable, biocompatible polyester.

20. The composition of claim 19 wherein the polyester is poly(DL-lactide-co-glycolide).

11. A process for preparing a biologically active, succinylated interleukin-2 comprising:

(a) reacting succinic anhydride with biologically active, normally hydrophobic, recombinant interleukin-2 that before succinylation is insoluble or not readily soluble in water or an aqueous medium under ambient conditions of room temperature and atmospheric pressure at a pH of between about 6 and 8; and

(b) isolating the succinylated interleukin-2.

13. The process of claim 11 wherein in step (a) a total of at least 2 moles of succinic anhydride is employed per mole of interleukin-2.

12. A process for preparing a pharmaceutical composition comprising:

(a) reacting succinic anhydride with biologically active, normally hydrophobic, recombinant interleukin-2 that before succinylated is insoluble or not readily soluble in water or an aqueous medium under ambient conditions of room temperature and atmospheric pressure at a pH of between about 6 and 8;

(b) isolating the succinylated interleukin-2; and

(c) formulating said succinylated interleukin-2 in a non-toxic, inert, pharmaceutically acceptable aqueous carrier medium.

15. The process of claim 12 wherein the succinylated interleukin-2 is formulated at a pH of 6-8 and the interleukin-2 is from a human source.

16. The process of claim 15 wherein said interleukin-2 is recombinant interleukin-2.

14. The process of claim 112 wherein in step (a) a total of at least 2 moles of succinic anhydride is employed per mole of interleukin-2.