Single chain fusion polypeptides comprising soluble light cytokine domains
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:
(i) a first soluble LIGHT cytokine domain,
(ii) a first peptide linker,
(iii) a second soluble LIGHT cytokine domain,
(iv) a second peptide linker, and
(v) a third soluble LIGHT cytokine domain,
wherein each of the soluble LIGHT cytokine domains lacks a stalk region and the first and the second peptide linkers independently have a length of 3-8 amino acids.
2. The polypeptide of claim 1 , wherein the first soluble LIGHT cytokine domain starts at an amino acid residue of Glu91 or Ala95, or between Glu91 and Ala95 of human LIGHT (SEQ ID NO:16).
3. The polypeptide of claim 1 , wherein the first soluble LIGHT cytokine domain starts at amino acid Glu91 of human LIGHT (SEQ ID NO:16).
4. The polypeptide of claim 1 , wherein the second and/or third soluble LIGHT cytokine domain is an N-terminally shortened domain and optionally comprises amino acid sequence mutations.
5. The polypeptide of claim 1 , wherein the second and/or third soluble LIGHT cytokine domain starts at amino acid Pro94 or Ala95 of human LIGHT (SEQ ID NO:16).
6. The polypeptide of claim 1 , wherein the first, second and/or third soluble LIGHT cytokine domain is the human LIGHT of SEQ ID NO: 16, and said domain ends at amino acid Val420.
7. The polypeptide of claim 1 , which additionally comprises an N-terminal signal peptide domain.
8. The polypeptide of claim 7 , wherein the N-terminal signal peptide domain comprises a protease cleavage site.
9. The polypeptide of claim 1 , which additionally comprises a further domain at the N-terminal and/or C-terminal end.
10. The polypeptide of claim 9 , wherein the further domain is a Fab or Fc fragment domain.
11. The polypeptide of claim 1 , wherein the first and second peptide linkers are independently glycine/serine linkers.
12. The polypeptide of claim 11 , wherein the glycine/serine linkers comprise substituted asparagine residues.
13. An isolated nucleic acid molecule encoding the fusion polypeptide of claim 1 .
14. An isolated host cell or a non-human organism transformed or transfected with the nucleic acid molecule of claim 13 .
15. A pharmaceutical composition comprising the fusion polypeptide of claim 1 and a pharmaceutically acceptable carrier, diluent and/or adjuvant.