Chimeric protein
The present invention provides a chimeric protein which comprises a multi-spanning transmembrane protein fused to a FAS endodomain, wherein the multi-spanning transmembrane protein binds an extracellular ligand, leading to activation of the FAS endodomain. The chimeric protein is useful as a suicide gene. The invention also provides a cell, such as a T cell comprising such a chimeric protein, which is useful in adoptive T cell immunotherapy approaches.
1. A chimeric protein comprising a multi-spanning transmembrane protein fused to a FAS endodomain,
wherein the multi-spanning transmembrane protein comprises CD20 truncated at the amino-terminus so that it lacks up to 41 amino acids from the CD20 amino-terminus, or at the carboxy-terminus so that it lacks up to 61 amino acids from the CD20 carboxy-terminus, and
wherein the multi-spanning transmembrane protein binds at least one extracellular ligand selected from Rituxumab, Ofatumumab or Veltuzumab, leading to activation of the FAS endodomain.
2. The chimeric protein according to claim 1 , which comprises a FAS endodomain fused to the carboxy-terminus of a truncated version of CD20, lacking up to 41 amino acids from the CD20 amino-terminus.
3. The chimeric protein according to claim 1 , wherein the FAS endodomain comprises the sequence shown in SEQ ID NO: 20.
4. The chimeric protein according to claim 1 , wherein the FAS endodomain comprises the amino acid sequence of SEQ ID NO: 29.
5. A chimeric protein comprising a multi-spanning transmembrane protein that consists of the amino acid sequence of SEQ ID NO: 4, SEQ ID NO: 5, or SEQ ID NO: 6 fused to a FAS endodomain, wherein the multi-spanning transmembrane protein binds at least one extracellular ligand selected from Rituxumab, Ofatumumab or Veltuzumab, leading to activation of the FAS endodomain.
6. The chimeric protein according to claim 5 , wherein the FAS endodomain comprises the sequence shown in SEQ ID NO: 20.
7. The chimeric protein according to claim 5 , wherein the FAS endodomain comprises the amino acid sequence of SEQ ID NO: 29.
8. A chimeric protein comprising:
a multi-spanning transmembrane protein fused to a FAS endodomain,
wherein the multi-spanning transmembrane protein comprises the amino acid sequence of SEQ ID NO: 3 truncated at the amino terminus so that it lacks up to 41 amino acids from the amino-terminus of SEQ ID NO: 3, or truncated at the carboxy-terminus so that it lacks up to 61 amino acids from the carboxy-terminus of SEQ ID NO: 3, and optionally has one or both of the following substitution mutations:
(a) an amino acid substitution at position A170 of SEQ ID NO: 3;
(b) an amino acid substitution at position P172 of SEQ ID NO: 3; and
wherein the multi-spanning transmembrane protein binds at least one extracellular ligand selected from Rituxumab, Ofatumumab or Veltuzumab, leading to activation of the FAS endodomain.
9. The chimeric protein according to claim 8 that comprises the mutation P172W, with position 172 defined with reference to the full length CD20 sequence shown as SEQ ID NO: 3.
10. The chimeric protein according to claim 8 , wherein the FAS endodomain comprises the amino acid sequence of SEQ ID NO: 29.
11. The chimeric protein according to claim 8 , wherein the FAS endodomain comprises the sequence shown in SEQ ID NO: 20.
12. An isolated cell which expresses the chimeric protein according to claim 1 .
13. An isolated cell which expresses the chimeric protein according to claim 5 .
14. An isolated cell which expresses the chimeric protein according to claim 6 .
15. An isolated cell which expresses the chimeric protein according to claim 3 .
16. An isolated cell which expresses the chimeric protein according to claim 4 .
17. An isolated cell which expresses the chimeric protein according to claim 7 .
18. An isolated cell which expresses the chimeric protein according to claim 8 .
19. An isolated cell which expresses the chimeric protein according to claim 9 .
20. An isolated cell which expresses the chimeric protein according to claim 10 .
21. An isolated cell which expresses the chimeric protein according to claim 11 .