Cold insoluble globulin, its purification and use

• US 4,315,906 A
• Filed: 05/21/1979
• Issued: 02/16/1982
• Est. Priority Date: 05/21/1979
• Status: Expired due to Term

• Current Assignee: E. I. du Pont de Nemours and Company (Corteva, Inc.)
• Original Assignee: New England Nuclear Corporation
• Inventors: Gelder, Frank B.
• Primary Examiner(s): Nucker, Christine M.
• Application Number:
  US06/040,995
• Time in Patent Office: 1,002 Days
• Field of Search: 424/1.5, 424/1, 424/12, 424/9, 260/112 R, 260/112 B
• US Class Current: 424/1.49
• CPC Class Codes: A61K 2123/00 Preparations for testing in vivo
• CPC Class Codes: A61K 51/08 Peptides, e.g. proteins , carriers being peptides, polyamino acids, proteins
• CPC Class Codes: C07K 14/78 Connective tissue peptides, e.g. collagen, elastin, laminin, fibronectin, vitronectin or cold insoluble globulin [CIG]
• CPC Class Codes: C07K 16/18 against material from animals or humans

Abstract

A highly purified, biologically active cold insoluble globulin (CIG) preparation is provided having specific opsonic activity of at least 1000 units per mg of CIG. Methods of purifying CIG using affinity antibody columns are described in which the biological activity of the purified CIG is greater than 95% of the activity of the starting material.

The purified CIG can be radiolabelled and radiopharmaceutical preparations containing such CIG are useful as scanning agents for in vivo localizing of areas where CIG-binding determinants are expressed. Such areas include thrombi and damaged tissue such as burns, heart, liver, spleen, muscle, skin, kidney and the like. These areas of localized radiolabelled CIG can be visualized with a gamma camera.

Claims

1. A preparation comprising affinity purified, biologically active cold insoluble globulin having a specific opsonic activity of at least 1000 units per milligram of cold insoluble globulin protein.

2. The preparation of claim 1 wherein said cold insoluble globulin is human cold insoluble globulin.

3. The preparation of claim 1 wherein said specific opsonic activity is at least 2000 units per milligram of protein.

4. The preparation of claim 3 wherein said cold insoluble globulin is human cold insoluble globulin.

5. A method for purifying cold insoluble globulin comprising:

preparing antisera to the cold insoluble globulin to obtain anti-CIG;

attaching said anti-CIG to a substrate to form anti-CIG-substrate;

contacting a fluid comprising cold insoluble globulin to be purified with the anti-CIG-substrate to remove the cold insoluble globulin from the fluid;

separating the anti-CIG-substrate with the cold insoluble globulin attached thereto from the fluid and

eluting purified cold insoluble globulin from the anti-CIG-substrate.

6. The method according to claim 5 further comprising loading said anti-CIG-substrate into a chromatograph column.

7. The method according to claim 5 wherein said cold insoluble globulin is eluted from the anti-CIG-substrate using a solution of glycine and hydrochloric acid.

8. The method according to claim 7 wherein the eluate is neutralized.

9. A radiopharmaceutical preparation comprising a pharmaceutically acceptable carrier containing purified biologically active cold insoluble globulin having a specific opsonic activity of at least 500 units per milligram of cold insoluble globulin protein, said cold insoluble globulin being radiolabelled.

10. The radiopharmaceutical preparation according to claim 9 wherein said cold insoluble globulin is labelled with at least about one atom of radionuclide per ten molecules of cold insoluble globulin.

11. The radiopharmaceutical preparation of claim 9 wherein said cold insoluble globulin is labelled with radioactive iodine.

12. The radiopharmaceutical preparation of claim 9 wherein said cold insoluble globulin has a specific opsonic activity of at least 1000 units per milligram of protein and is labelled with at least about one atom of radioactive iodine per ten molecules of cold insoluble globulin.

13. The radiopharmaceutical preparation of claim 12 wherein said radioactive iodine is ¹²⁵ I or ¹³¹ I.

14. The radiopharmaceutical preparation of claim 12 wherein said cold insoluble globulin is labelled with about 1.5 atoms of radioactive iodine per molecule of protein.

15. The radiopharmaceutical preparation of claim 12 wherein said specific opsonic activity is at least 2000 units per milligram of cold insoluble globulin protein.

16. The radiopharmaceutical preparation of claim 9 wherein said cold insoluble globulin is human cold insoluble globulin.

17. The radiopharmaceutical preparation of claim 16 wherein said specific opsonic activity is at least 1000 units per milligram of cold insoluble globulin protein.

18. The radiopharmaceutical preparation of claim 16 wherein said cold insoluble globulin is labelled with radioactive iodine.

19. A radiopharmaceutical preparation comprising a radiolabelled biologically active cold insoluble globulin in a pharmaceutically acceptable carrier, said radiolabelled cold insoluble globulin having been made by radiolabelling cold insoluble globulin having a specific opsonic activity of at least 1000 units per milligram of cold insoluble globulin.

20. The radiopharmaceutical preparation of claim 19 wherein said cold insoluble globulin is human cold insoluble globulin.

21. The radiopharmaceutical preparation of claim 20 wherein said specific opsonic activity is at least 2000 units per milligram of cold insoluble globulin protein.

22. The radiopharmaceutical preparation of claim 20 wherein said cold insoluble globulin is labelled with radioactive iodine.

23. A method for in vivo visualizing a site of protein or cellular components expressing CIG-binding determinants in a higher animal, said method comprising injecting said animal with a radiopharmaceutical preparation containing radiolabelled affinity purified cold insoluble globulin having a specific opsonic activity of at least 1000 units per milligram of protein, localizing the cold insoluble globulin at said site, and visualizing said site with radioscintigraphic imaging apparatus.

24. The method according to claim 23 wherein said higher animal is a human being and said cold insoluble globulin is human cold insoluble globulin.

25. A method for in vivo visualizing a site of thrombi or tissue damage in a higher animal, said method comprising injecting said animal with a radiopharmaceutical preparation containing radiolabelled cold insoluble globulin, localizing the cold insoluble globulin at said site of thrombi or tissue damage, and visualizing said site of thrombi or tissue damage with radioscintigraphic imaging apparatus.

26. A method for in vivo visualizing a burn site in a higher animal, said method comprising intravenously injecting said animal with a radiopharmaceutical preparation containing radiolabelled cold insoluble globulin, localizing the cold insoluble globulin at said burn site, and visualizing said site with radioscintigraphic imaging apparatus.

27. A method for in vivo visualizing a site of thrombi in a higher animal, said method comprising intravenously injecting said animal with a radiopharmaceutical preparation containing radiolabelled cold insoluble globulin, localizing the cold insoluble globulin at the site of said thrombi, and visualizing said site with radioscintigraphic imaging apparatus.

28. A method for in vivo visualizing a site of an ulcer in a higher animal, said method comprising intravenously injecting said animal with a radiopharmaceutical preparation containing radiolabelled cold insoluble globulin, localizing the cold insoluble globulin at said site of said ulcer, and visualizing said site with radioscintigraphic imaging apparatus.

29. A method for in vivo visualizing an infarct in a higher animal, said method comprising intravenously injecting said animal with a radiopharmaceutical preparation containing radiolabelled cold insoluble globulin, localizing the cold insoluble globulin at said infarct, and visualizing said infarct with radioscintigraphic imaging apparatus.

30. A method for in vivo visualizing a site of lymph node metastases in a higher animal, said method comprising lymphatically injecting said animal with a radiopharmaceutical preparation containing radiolabelled cold insoluble globulin, localizing the cold insoluble globulin at said site of said lymph node metastases, and visualizing said site with radioscintigraphic imaging apparatus.

31. A method for in vivo visualizing of tumors in a higher animal, said method comprising when intravenously injecting said animal with a radiopharmaceutical preparation containing radiolabelled cold insoluble globulin, localizing the cold insoluble globulin at the site of said tumor, and visualizing said tumor with radioscintigraphic imaging apparatus.

32. A preparation comprising biologically active cold insoluble globulin having a specific opsonic activity of at least 1000 units per milligram of protein.

33. A preparation according to claim 32, wherein said cold insoluble globulin is human cold insoluble globulin.

34. A preparation according to claim 32, wherein said specific opsonic activity is at least 2000 units per milligram of protein.

35. A preparation according to claim 34, wherein said cold insoluble globulin is human cold insoluble globulin.

36. A preparation in accord with any of claims 32, 33, 34 or 35, wherein said cold insoluble globulin is purified by an affinity purification method.