Claims
- 1. An isolated nucleic acid sequence comprising a sequence that encodes a vascular endothelial cell growth factor having a molecular weight of about 45,000 daltons under non-reducing conditions and about 23,000 under reducing conditions as measured by SDS-PAGE.
- 2. sequence of claim 1 that is a DNA sequence.
- 3. An isolated DNA sequence comprising a sequence that hybridizes to the DNA sequence:
5′-CCTATGGCTGAAGGCGGCCAGAAGCCTCACGAAGTGGTGAAGTTCATGGACGTGTATCA-3′ when incubated therewith at 42° C. in 20% formamide, 5×SSC, 50 mM sodium phosphate pH 6.8, 0.1% sodium pyrophosphate, 5×Denhardt's solution, and 50 μg/ml salmon sperm DNA, and washed with 2×SSC, 0.1% SDS at 42° C., wherein said isolated sequence contains at least about ten nucleotides.
- 4. The sequence of claim 3 that contains at least about twenty nucleotides.
- 5. The sequence of claim 3 that hybridizes to the DNA sequence of FIG. 2 when incubated therewith at 42° C. in 20% formamide, 5×SSC, 50 mM sodium phosphate pH 6.8, 0.1% sodium pyrophosphate, 5×Denhardt's solution, and 50 μg/ml salmon sperm DNA, and washed with 2×SSC, 0.1% SDS at 42° C., wherein said isolated sequence contains at least about ten nucleotides.
- 6. The sequence of claim 5 that contains at least about twenty nucleotides.
- 7. The sequence of claim 6 that contains at least about thirty nucleotides.
- 8. The nucleic acid sequence of claim 3 further comprising a promoter operably linked to said nucleic acid sequence.
- 9. An expression vector comprising the nucleic acid sequence of claim 3 operably linked to control sequences recognized by a host transformed by the vector.
- 10. A host cell transformed with the expression vector of claim 9.
- 11. The host cell of claim 10 wherein the cell is eukaryotic.
- 12. The lost cell of claim 10 wherein the cell is prokaryotic.
- 13. An isolated DNA sequence comprising a DNA sequence encoding a vascular endothelial cell growth factor having an amino acid sequence sufficiently duplicative of that of vascular endothelial cell growth factor to allow it to possess one or both of the biological properties of (a) promoting growth selectively of vascular endothelial cells but not bovine corneal endothelial cells, lens epithelial cells, adrenal cortex cells, BHK-21 fibroblasts, or keratinocytes, or (b) cross-reacting immunologically with an antibody raised against at least one epitope of the corresponding native protein.
- 14. The sequence of claim 13 that is a cDNA sequence.
- 15. The sequence of claim 13 further comprising a signal sequence-encoding region N-terminal to the cell growth factor-encoding sequence.
- 16. The sequence of claim 15 wherein the signal sequence is recognized by mammalian cells.
- 17. The sequence of claim 13 that is a genomic sequence.
- 18. The sequence of claim 13 that is covalently bound to a detectable moiety.
- 19. An expression vector comprising the DNA sequence of claim 13 operably linked to control sequences recongized by a host transformed with the vector.
- 20. The vector of claim 19 that is a plasmid.
- 21. A host cell transformed with the expression vector of claim 19.
- 22. The host cell of claim 21 that is eukaryotic.
- 23. The host cell of claim 22 that is mammalian.
- 24. A method of producing a vascular endothelial cell growth factor, which method comprises culturing the cells of claim 21 to express the growth factor in the host cell culture.
- 25. The method of claim 24 further comprising the step of recovering the growth factor from the host cell culture.
- 26. Vascular endothelial cell growth factor that is unaccompanied by associated native glycosylation, that has at least about 80% homology with the amino acid sequence of the mature protein shown in FIG. 2, and that possesses one or both of the biological properties of (a) promoting growth selectively of vascular endothelial cells but not bovine corneal endothelial cells, lens epithelial cells, adrenal cortex cells, BHK-21 fibroblasts, or keratinocytes, or (b) cross-reacting immunologically with an antibody raised against at least one epitope of the corresponding native protein.
- 27. The cell growth factor of claim 26 that is of bovine origin.
- 28. The cell growth factor of claim 26 that is of human origin.
- 29. Vascular endothelial cell growth factor that is completely free of source proteins.
- 30. The cell growth factor of claim 29 that is of bovine origin.
- 31. The cell growth factor of claim 29 that is of human origin.
- 32. (Amended) A pharmaceutical composition useful for promotion of vascular endothelial cell growth comprising:
(a) a therapeutically effective amount of [the protein of claim 26] vascular endothelial cell growth factor that is unaccompanied by native glycosylation, that has at least about 80% homology with the amino acid sequence of the mature protein shown in FIG. 2. and that possesses the biological property of promoting growth selectively of vascular endothelial cells and (b) [in] a pharmaceutically acceptable carrier. A method for treating trauma affecting the vascular endothelium comprising administering to an animal or human suffering from said trauma an effective amount of the composition of claim 32.
- 37. A method for treating trauma affecting the vascular endothelium comprising administering to an animal or human suffering from said trauma an effective amount of the composition of claim 33.
- 38. The method of claim 36 further comprising administering to said animal or human an effective amount of another cell growth factor.
- 39. The method of claim 36 wherein the trauma is diabetic ulcers or a wound of the blood vessels or heart.
- 40. The method of claim 36 wherein a human is treated.
Parent Case Info
[0001] This application is a continuation of co-pending application Ser. No. 08/904,334 filed on Jul. 31, 1997, which is a continuation application of Ser. No. 08/561,560 filed on Nov. 22, 1995, now abandoned, which is a continuation application of Ser. No. 08/306,213 filed on Sep. 14, 1994, now abandoned, which is a continuation application of Ser. No. 08/047,756 filed on Apr. 15, 1993, now abandoned, which is a continuation application of Ser. No. 07/351,117 filed on May 12, 1989, now abandoned.
Continuations (5)
|
Number |
Date |
Country |
Parent |
08904334 |
Jul 1997 |
US |
Child |
10166941 |
Jun 2002 |
US |
Parent |
08561560 |
Nov 1995 |
US |
Child |
08904334 |
Jul 1997 |
US |
Parent |
08306213 |
Sep 1994 |
US |
Child |
08561560 |
Nov 1995 |
US |
Parent |
08047756 |
Apr 1993 |
US |
Child |
08306213 |
Sep 1994 |
US |
Parent |
07351117 |
May 1989 |
US |
Child |
08047756 |
Apr 1993 |
US |