Vectors for expression of hml-2 polypeptides

• US 20100086565A1
• Filed: 06/13/2003
• Published: 04/08/2010
• Est. Priority Date: 06/13/2002
• Status: Active Grant

• Current Assignee: Novartis Vaccines And Diagnostics Incorporated (Novartis Ag)
• Original Assignee: Novartis Vaccines And Diagnostics Incorporated (Novartis Ag)
• Inventors: zur Megede, Jan, Donnelly, John, Hardy, Stephen F.
• Granted Patent: US 8,518,694 B2
• Time in Patent Office: Days
• US Class Current: 424/204.100
• CPC Class Codes: A61K 2039/5258 Virus-like particles
• CPC Class Codes: A61K 2039/53 DNA (RNA) vaccination
• CPC Class Codes: A61K 2039/54 characterised by the route of administration
• CPC Class Codes: A61K 2039/545 characterised by the dose, timing or administration schedule
• CPC Class Codes: A61K 39/12 Viral antigens
• CPC Class Codes: A61K 39/21 Retroviridae, e.g. equine infectious anemia virus
• CPC Class Codes: A61P 13/08 of the prostate
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 25/28 for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
• CPC Class Codes: A61P 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 37/04 Immunostimulants
• CPC Class Codes: C07K 14/005 from viruses
• CPC Class Codes: C12N 2740/10022 New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
• CPC Class Codes: C12N 2740/10034 Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein

Abstract

A nucleic acid vector comprising: (i) a promoter; (ii) a sequence encoding a HML-2 polypeptide operably linked to said promoter; and (iii) a selectable marker. Preferred vectors comprise: (I) a eukaryotic promoter; (ii) a sequence encoding a HML-2 polypeptide downstream of and operably linked to said promoter, (iii) a prokaryotic selectable marker; (iv) a prokaryotic origin of replication; and (v) a eukaryotic transcription terminator downstream of and operably linked to said sequence encoding a HML-2 polypeptide. Vectors of the invention are particularly useful for expression of HML-2 polypeptides either in vitro (e.g. for later purification). Or in vivo (e.g. for nucleic acid immunization). They are well suited to nucleic acid immunization against prostrate tumors. A preferred HML-2 is PCAV, which is located in chromosome 22 at 20.428 megabases (22q11.2).

Claims

1. A nucleic acid vector comprising:

(i) a promoter;

(ii) a sequence encoding a polypeptide from a member of the HML-2 subgroup of the HERV-K family of endogenous retroviruses, said sequence being operably linked to said promoter; and

(iii) a selectable marker.

2. The vector of claim 1, further comprising:

(iv) an origin of replication; and

(v) a transcription terminator downstream of and operably linked to (ii).

3. The vector of claim 2, wherein:

(i) &

(v) are eukaryotic; and

(iii) &

(iv) are prokaryotic.

4. The vector of claim 1 wherein the HML-2 is PCAV from human chromosome 22.

5. The vector of claim 1 wherein the promoter is functional in vivo in a human.

6. The vector of claim 1 wherein the promoter is a viral promoter.

7. The vector of claim 6, wherein the viral promoter is from cytomegalovirus (CMV).

8. The vector of claim 1 comprising transcriptional regulatory sequences in addition to the promoter.

9. The vector of claim 1 wherein the HML-2 polypeptide is a gag, prt, pol, env, cORF or PCAP polypeptide.

10. The vector of claim 9, wherein the HML-2 polypeptide:

(a) has at least 65% identity to one or more of SEQ ID NOS ;

1-50, 69-74, 78 and 79; and

/or (b) comprises a fragment of at least 7 amino acids from one or more of SEQ ID NOS ;

1-50, 69-74, 78 and 79.

11. The vector of claim 1 wherein the selectable marker functions in a bacterium.

12. The vector of claim 1 wherein the selectable marker is an antibiotic resistance genes.

13. The vector of claim 1 wherein the vector is a plasmid.

14. The vector of claim 1 wherein the vector comprises an origin of replication.

15. The vector of claim 14, wherein the origin of replication is active in prokaryotes but not in eukaryotes.

16. The vector of claim 1 further comprising a eukaryotic transcriptional terminator sequence downstream of the HML2-coding sequence.

17. The vector of claim 1 further comprising a multiple cloning site.

18. The vector of claim 1 further comprising an IRES upstream of a second sequence encoding a eukaryotic polypeptide.

19. A pharmaceutical composition comprising the vector of claim 1.

22. A method for raising an immune response, comprising administering an immunogenic dose of the vector of claim 1 to an animal.

23. A method for treating a patient with a prostate tumor, comprising administering to them the pharmaceutical composition of claim 19.

20-21. -21. (canceled)

24. A virus-like particle (VLP) comprising HML-2 gag polypeptides.

27. A method of raising an immune response in an animal, comprising administering to the animal the VLP of claim 24.

28. A method for treating a patient with a prostate tumor, comprising administering to them the VLP of claim 24.

29. A method for diagnosing cancer in a patient, comprising the step of (a) contacting antibodies from the patient with the VLP of claim 24, and/or (b) contacting antibodies against the VLP of claim 24 with a patient sample.

25-26. -26. (canceled)