Method for producing antibody using “naked” expression vector expressing type II transmembrane fusion protein
View Patent ↗Methods are disclosed for generating antibodies and an expression vector used to express protein(s) which provoke the antibody response. The expression vector may be useful in generating an antibody directed to an antigen, comprising a gene in operable linkage with a promoter, which gene encodes upon expressing a fusion protein comprising (i) CD134L, a fragment, homologous or orthologues protein thereof as N-terminal moiety of the fusion protein; and (ii) all or part of an antigenic protein as C-terminal moiety of the fusion protein. To generate the antibodies, the vector is injected into a subject animal, which produces a fusion protein, against which antibodies are generated.
1. An expression vector for generating an antibody directed to an antigen in a subject animal, the expression vector comprising a polynucleotide in operable linkage with a promoter, wherein the polynucleotide encodes a fusion protein comprising:
(i) a CD134L polypeptide that is orthologous to human CD134L or a polypeptide having at least 95% amino acid identity to a CD134L polypeptide that is orthologous to human CD134L which anchors in a cell membrane as the N-terminal moiety of the fusion protein; and
(ii) an antigenic protein or an antigenic fragment thereof as the C-terminal moiety of the fusion protein,
wherein the fusion protein further comprises a linker peptide interposed between (i) and (ii).
2. The expression vector of claim 1 , wherein said polypeptide of CD134L that is orthologous to human CD134L or said polypeptide having at least 95% amino acid identity to a CD134L polypeptide that is orthologous to human CD134L which anchors in a cell membrane is from a species different from the species of the subject animal.
3. The expression vector of claim 1 , wherein said N-terminal moiety of the fusion protein comprises the polypeptide of SEQ ID NO: 26 or a polypeptide having at least 95% amino acid sequence identity to the polypeptide of SEQ ID NO. 26 which anchors in a cell membrane.
4. The expression vector of claim 1 , wherein said N-terminal moiety of the fusion protein comprises the polypeptide of SEQ ID NO: 27 or a polypeptide having at least 95% amino acid sequence identity to the polypeptide of SEQ ID NO. 27 which anchors in a cell membrane.
5. The expression vector of claim 1 , wherein said N-terminal moiety of the fusion protein comprises the polypeptide of SEQ ID NO: 28 or a polypeptide having at least 95% amino acid sequence identity to the polypeptide of SEQ ID NO. 28 which anchors in a cell membrane.
6. The expression vector of claim 1 , wherein said N-terminal moiety of the fusion protein comprises the polypeptide of SEQ ID NO: 29 or a polypeptide having at least 95% amino acid sequence identity to the polypeptide of SEQ ID NO. 29 which anchors in a cell membrane.
7. The expression vector of claim 1 , wherein the linker peptide is (G 4 S) 3 (SEQ ID NO: 31).
8. The expression vector of claim 1 , wherein the subject animal is a human.