TNF superfamily member light fusion proteins
The present invention refers to fusion proteins comprising a TNF superfamily (TNFSF) cytokine or a receptor binding domain thereof fused to a trimerization domain and a nucleic acid molecule encoding the fusion protein. The fusion protein is present as a trimeric complex or as an oligomer thereof and is suitable for therapeutic, diagnostic and/or research applications.
1. A fusion protein comprising:
(i) a receptor binding domain of a TNF-superfamily cytokine of LIGHT having the amino residues 91-240 of SEQ ID NO: 40;
(iii) a bacteriophage RB69 fibritin trimerization domain having 20-30 amino acids residues comprising the amino acid residues 455-480 of SEQ ID NO:24, or the amino acid residues 456-480 of SEQ ID NO:24; and
(ii) a flexible linker between (i) and (iii), wherein the linker has 5-20 amino acid residues having the amino acid sequence (GSS) a (SSG) b (GS) c (S) d , and a, b, c, d are independently 0, 1, 2, 3, 4, or 5; wherein the receptor binding domain of the TNF-superfamily cytokine is fused to the N-terminus of the linker and the linker is fused to the N-terminus of the fibritin trimerization domain.
2. The fusion protein according to claim 1 , wherein the linker has the amino acid sequence of GSS GSS GSS GS (SEQ ID NO: 46).
3. The fusion protein according to claim 1 , further comprising:
(iv) a N-terminal signal peptide.
4. The fusion protein according to claim 3 , further comprising:
(v) a protease cleavage site C-terminal to the signal peptide.
5. The fusion protein according to claim 1 , further comprising:
(iv) a C-terminal flexible element.
6. The fusion protein according to claim 5 , further comprising:
(v) a recognition or purification domain fused in frame to the C-terminal flexible element.
7. The fusion protein according to claim 1 , wherein the fusion protein forms a trimeric complex or an oligomer of a trimeric complex.
8. The fusion protein according to claim 7 , wherein the complex consists of three identical fusion proteins.
9. An isolated nucleic acid encoding the fusion protein according to claim 1 .
10. An expression vector comprising the nucleic acid according to claim 9 .
11. A cell transformed or transfected with the expression vector according to claim 10 .
12. A pharmaceutical composition comprising:
the fusion protein according to claim 1 , and a pharmaceutically acceptable carrier, diluent or adjuvant.
13. The fusion protein according to claim 1 , wherein the fibritin trimerization domain comprises SEQ ID NO: 9.
14. A fusion protein comprising (i) the amino acid residues 91-240 of SEQ ID NO: 40, (ii) SEQ ID NO: 46 and (iii) SEQ ID NO: 9, in this particular order, starting from the N-terminus of the fusion protein.