CD27 single chain molecules and encoding nucleic acids
The present invention refers to single-chain fusion proteins comprising three soluble TNF superfamily (TNFSF) cytokine domains and nucleic acid molecules encoding these fusion proteins. The fusion proteins are substantially non-aggregating and suitable for therapeutic, diagnostic and/or research applications.
1. A single-chain fusion polypeptide comprising:
(a) a first soluble CD27 ligand (CD27L) cytokine domain,
(b) a first peptide linker,
(c) a second soluble CD27L cytokine domain,
(d) a second peptide linker, and
(e) a third soluble CD27L cytokine domain,
wherein each of the soluble CD27L cytokine domains lacks a stalk region and the first and the second peptide linkers independently have a length of 3-8 amino acids.
2. A single-chain fusion polypeptide comprising:
(a) a first soluble CD27 ligand (CD27L) cytokine domain,
(b) a first peptide linker,
(c) a second soluble CD27L cytokine domain,
(d) a second peptide linker, and
(e) a third soluble CD27L cytokine domain,
wherein each of the soluble CD27L cytokine domains lacks a stalk region and the first and the second peptide linkers independently have a length of 3-8 amino acids,
wherein the first, the second, and the third soluble CD27L cytokine domains are independently an N-terminally shortened domain of SEQ ID NO: 7, and optionally consists of an amino acid mutation.
3. The polypeptide of claim 1 , wherein the first, the second, and the third soluble CD27L cytokine domains independently have the amino sequence of 51-193 or 56-193 of SEQ ID NO: 7.
4. The polypeptide of claim 1 , wherein the first, the second, and the third soluble CD27L cytokine domains have the amino sequence of 51-193 of SEQ ID NO: 7.
5. The polypeptide of claim 1 , wherein the first and second peptide linkers are independently linkers comprising glycine and serine.
6. The polypeptide of claim 5 , wherein the linkers further comprise substituted asparagine residues.
7. The polypeptide of claim 1 , wherein the first and second peptide linkers are independently selected from the group consisting of SEQ ID NO: 52-53 and SEQ ID NO: 21-26.
8. The polypeptide of claim 1 , which additionally comprises an N-terminal signal peptide domain.
9. The polypeptide of claim 8 , wherein the N-terminal signal peptide domain comprises a protease cleavage site.
10. The polypeptide of claim 1 , which additionally comprises a further domain at the N-terminal and/or C-terminal end.
11. The polypeptide of claim 10 , wherein the further domain is a Fab or Fc fragment domain.
12. The polypeptide of claim 1 , which additionally comprises a third peptide linker and a domain of Fc fragment.
13. A dimer comprises two polypeptides of claim 12 , wherein the two polypeptides are dimerized via disulfide bridges of the third peptide linkers.
14. A pharmaceutical composition comprising the fusion polypeptide of claim 1 and a pharmaceutically acceptable carrier, diluent and/or adjuvant.
15. An isolated nucleic acid molecule encoding the fusion polypeptide of claim 1 .
16. An isolated host cell or a non-human organism transformed or transfected with the nucleic acid molecule of claim 15 .