Compositions and methods for prevention of escape mutation in the treatment of HER2/NEU over-expressing tumors
This invention provides compositions and methods for treating and vaccinating against an Her2/neu antigen-expressing tumor and inducing an immune response against dominant and several sub-dominant epitopes of the antigen.
1. A recombinant Listeria strain having mutations in the D-alanine racemase (Dal) gene and the D-amino acid transferase (Dat) gene, said recombinant Listeria strain comprising a nucleic acid encoding a first and a second open reading frame, wherein said first open reading frame encodes a recombinant polypeptide comprising SEQ ID NO: 2 fused to a) non-hemolytic LLO protein or N-terminal fragment, b) a PEST sequence, or c) an ActA fragment, and wherein said second open reading frame encodes a metabolic enzyme that complements said mutations, and wherein said recombinant Listeria lacks the ActA virulence gene.
2. An immunogenic composition comprising the recombinant Listeria of claim 1 .
3. A nucleic acid molecule comprising a first open reading frame encoding said fusion polypeptide of claim 1 , wherein said nucleic molecule resides within said recombinant Listeria strain.
4. The recombinant Listeria of claim 1 , wherein said Her2/neu chimeric antigen comprises at least 5, 9, 13, 14, or 17 of the mapped human MHC-class I epitopes.
5. The recombinant Listeria of claim 3 , wherein said nucleic acid molecule is integrated into the Listeria genome.
6. The recombinant Listeria of claim 3 , wherein said nucleic acid molecule is in a plasmid in said recombinant Listeria strain.
7. The recombinant Listeria of claim 6 , wherein said plasmid is stably maintained in said recombinant Listeria strain in the absence of antibiotic selection.
8. The recombinant Listeria of claim 6 , wherein said plasmid does not confer antibiotic resistance upon said recombinant Listeria.
9. The recombinant Listeria of claim 1 , wherein said recombinant Listeria strain is attenuated.
10. The recombinant Listeria strain of claim 1 , wherein said metabolic enzyme is an alanine racemase enzyme or a D-amino acid transferase enzyme.
11. The recombinant Listeria strain of claim 1 , wherein said recombinant Listeria strain has been passaged through an animal host.
12. The immunogenic composition of claim 2 , further comprising an adjuvant.
13. The immunogenic composition of claim 12 , wherein said adjuvant comprises a granulocyte/macrophage colony-stimulating factor (GM-CSF) protein, a nucleotide molecule encoding a GM-CSF protein, saponin QS21, monophosphoryl lipid A, or an unmethylated CpG-containing oligonucleotide.
14. A method of impeding a growth of a Her-2-expressing tumor in a subject, said method comprising the step of administering to said subject the immunogenic composition of claim 2 .
15. A method of eliciting an enhanced immune response to a Her2/neu-expressing tumor in a subject, said method comprising administering to said subject the immunogenic composition of claim 2 .
16. The method of claim 15 , wherein said immune response against said Her2/neu-expressing tumor comprises an immune response to a subdominant epitope of said Her2/neu protein.
17. The method of claim 16 , wherein said immune response against said Her2/neu-expressing tumor comprises an immune response to a subdominant epitope of said Her2/neu protein.
18. The method of claim 15 , wherein said immune response against said Her2/neu-expressing tumor comprises an immune response to a subdominant epitope of said Her2/neu protein.
19. A method of preventing an escape mutation in the treatment of Her2/neu over-expressing tumors, comprising administering to a subject the immunogenic composition of claim 2 .
20. A method of preventing the onset of a Her2/neu antigen-expressing tumor in a subject, said method comprising the step of administering to said subject a composition comprising the immunogenic composition of claim 2 .
21. A method of decreasing the frequency of intra-tumoral T regulatory cells, said method comprising the step of administering to said subject a composition comprising the immunogenic composition of claim 2 .
22. A method of decreasing the frequency of myeloid derived suppressor cells, said method comprising the step of administering to said subject a composition comprising the immunogenic composition of claim 2 .
23. A method of increasing intratumoral ratio of CD8+/T regulatory cells, said method comprising the step of administering to said subject a composition comprising the immunogenic composition of claim 2 .
24. A method of preventing the formation of a Her2/neu-expressing tumor in a subject, said method comprising the step of administering to said subject a composition comprising the immunogenic composition of claim 2 .
25. A method of treating a Her2/neu-expressing tumor in a subject, said method comprising the step of administering to said subject a composition comprising the immunogenic composition of claim 2 .