التفاصيل البيبلوغرافية
| العنوان: |
Pharmaceutical preparation and method of treatment of human malignancies with arginine deprivation |
| Document Number: |
20050244398 |
| تاريخ النشر: |
November 3, 2005 |
| Appl. No: |
10/518223 |
| Application Filed: |
June 20, 2003 |
| مستخلص: |
The present invention provides an isolated and substantially purified recombinant human arginase having sufficiently high enzymatic activity and stability to maintain Adequate Arginine Depletion in a patient. The present invention also provides a pharmaceutical composition comprising the modified invention enzyme and method for treatment of diseases using the pharmaceutical composition. |
| Inventors: |
Cheng, Ning Man (Hong Kong, CN); Leung, Yun Chung (Hong Kong, CN); Lo, Wai Hung (Hong Kong, CN) |
| Claim: |
1. An isolated recombinant human arginase I, comprising substantially the same amino acid sequence as set forth in SEQ ID NO: 9 and having a purity of 80-100%. |
| Claim: |
2. The recombinant human arginase I according to claim 1 further comprising six histidines attached to the amino terminal end thereof. |
| Claim: |
3. The recombinant human arginase I according to claim 1 having a specific activity of at least 250 I.U./mg. |
| Claim: |
4. The recombinant human arginase I according to claim 3 having a specific activity of 500 to 600 I.U./mg. |
| Claim: |
5. The recombinant human arginase I according to claim 4, comprising a modification that results in an in vitro plasma half-life of at least approximately 3 days. |
| Claim: |
6. An isolated recombinant human arginase I according to claim 1, having a purity of at least 90%. |
| Claim: |
7. The recombinant human arginase I according to claim 5, wherein said modification is pegylation. |
| Claim: |
8. The recombinant human arginase I according to claim 7, wherein said pegylation results from covalently attaching at least one polyethylene glycol (PEG) moiety to said arginase using a coupling agent. |
| Claim: |
9. The recombinant human arginase I according to claim 8, wherein said coupling agent is selected from the group consisting of 2,4,6-trichloro-s-triazine (cyanuric chloride, CC) and succinimide propionic acid (SPA). |
| Claim: |
10. A method of producing recombinant protein comprising: (a) cloning a gene encoding said protein; (b) constructing a recombinant Bacillus subtilis strain for expression of said protein; (c) fermenting said recombinant Bacillus subtilis cells using fed-batch fermentation; (d) heat-shocking said recombinant Bacillus subtilis cells to stimulate expression of said recombinant protein; and (e) purifying said recombinant protein from the product of said fermentation. |
| Claim: |
11. The method according to claim 10 wherein said Bacillus subtilis is a prophage. |
| Claim: |
12. The method according to claim 10 wherein said protein is human arginase I. |
| Claim: |
13. The method according to claim 12 wherein said human arginase I comprises six histidines linked to the amino-terminus thereof, and said purifying step comprises affinity chromatography in a chelating column. |
| Claim: |
14. The method according to claim 12 wherein said fermenting step is performed using a feeding medium consisting essentially of 180-320 g/L glucose, 2-4 g/L MgSO4.7H2O, 45-80 g/L tryptone, 7-12 g/L K2HPO4 and 3-6 g/L KH2PO4. |
| Claim: |
15. A pharmaceutical composition comprising an isolated and substantially purified arginase. |
| Claim: |
16. The pharmaceutical composition according to claim 15 wherein said recombinant human arginase is human arginase I. |
| Claim: |
17. The pharmaceutical composition according to claim 15 wherein said recombinant human arginase is human arginase I, further comprising eentaifing six additional histidines attached to the amino terminal end thereof. |
| Claim: |
18. The pharmaceutical composition according to claim 15, wherein said composition is further formulated in a pharmaceutically acceptable carrier. |
| Claim: |
19. The pharmaceutical composition according to claim 15, wherein the formulation of said pharmaceutical composition is in a form suitable for oral use, for a sterile injectable solution or a sterile injectable suspension. |
| Claim: |
20. The pharmaceutical composition according to claim 16, wherein said recombinant human arginase I has a specific enzyme activity of at least 250 I.U./mg. |
| Claim: |
21. The pharmaceutical composition according to claim 20, wherein said recombinant human arginase I has a specific enzyme activity of 500 to 600 I.U./mg. |
| Claim: |
22. The pharmaceutical composition according to claim 16, wherein said recombinant human arginase I has a half-life in patient plasma of at least 3 days. |
| Claim: |
23. The pharmaceutical composition according to claim×22, wherein said recombinant human arginase I has a half-life in patient plasma of approximately at least 1 day. |
| Claim: |
24. A method of treatment of human malignancies, comprising administering human arginase I. |
| Claim: |
25. A method of treatment of human malignancies comprising administering the pharmaceutical composition of claim 15. |
| Claim: |
26. The use according to method of claim 25, wherein said human malignancies are selected from the group consisting of: liver tumor, breast cancer, colon cancer and rectal cancer. |
| Claim: |
27. A method of treatment of human malignancies comprising administering recombinant human arginase to a patient. |
| Claim: |
28. A method of treatment of human malignancies in a patient comprising administering a pharmaceutical composition that reduces the physiological arginine level in said patient to below 10 μM for at least 3 days. |
| Current U.S. Class: |
424094/400 |
| رقم الانضمام: |
edspap.20050244398 |
| قاعدة البيانات: |
USPTO Patent Applications |