TCR libraries
The present invention relates to a library of particles, the library displaying a plurality of different T cell receptors (TCRs), wherein the plurality of TCRs consists essentially of TCRs comprising an alpha chain comprising an alpha chain variable domain and a beta chain comprising a beta chain variable domain and the library comprises more than one TRAV gene product and/or more than one TRBV gene product, wherein the beta chain variable domain does not comprise one or more of a TRBV5-1, 5-3, 5-4, 5-5, 5-6, 5-7 or 5-8 gene product and wherein the plurality of TCRs do not consist essentially of TCRs comprising a TRAV12-2 gene product from a natural repertoire and a TRBV6 gene product from a natural repertoire and TCRs comprising a TRAV21 gene product from a natural repertoire and a TRBV6 gene product from a natural repertoire.
1. A library of particles displaying a plurality of T cell receptors (TCRs), wherein the plurality of TCRs:
i) consists essentially of TCRs comprising
(a) an alpha chain, the alpha chain comprising an alpha chain variable domain from a natural repertoire of TCRs that have undergone thymic selection in a human donor, and
(b) a beta chain, the beta chain comprising a beta chain variable domain from a natural repertoire of TCRs that have undergone thymic selection in a human donor and that does not comprise one or more of a TRBV5-1, 5-3, 5-4, 5-5, 5-6, 5-7 or 5-8 gene product; and
ii) comprises more than one TRAV gene product and/or more than one TRBV gene product,
wherein the TCRs are covalently attached to the particle, and
wherein the particles are phage particles, ribosomes, yeast cells, or mammalian cells.
2. The library of claim 1 , wherein the beta chain variable domain does not comprise either of a TRBV10-2 gene product and/or a TRBV10-3 gene product.
3. The library of claim 1 , wherein the beta chain variable domain does not comprise a TRBV2 gene product, a TRBV9 gene product, and/or a TRBV14 gene product.
4. The library of claim 1 , wherein the beta chain variable domain does not comprise a TRBV5 gene product.
5. The library of claim 1 , wherein the beta chain variable domain comprises a TRBV5-1 gene product.
6. The library of claim 1 , wherein the framework region, constant region, and CDR1, CDR2 and/or CDR3 sequence of the alpha and/or beta variable domain comprises at least one non-natural mutation.
7. The library of claim 1 , wherein the alpha chain variable domain and the beta chain variable domain are displayed as a single polypeptide chain.
8. The library of claim 1 , wherein the TCRs comprise a non-native disulfide bond between a constant region of the alpha chain and a constant region of the beta chain.
9. The library of claim 1 , wherein the TCRs comprise a native disulfide bond between a constant region of the alpha chain and a constant region of the beta chain.
10. The library of claim 1 , wherein each alpha chain and each beta chain comprises a dimerization domain.
11. The library according to claim 10 , wherein the dimerization domain is heterologous.
12. The library of claim 1 , wherein the particles are phage particles.
13. The library of claim 1 , wherein the particles are ribosomes.
14. The library of claim 1 , wherein the particles are yeast cells.
15. The library of claim 1 , wherein the particles are mammalian cells.
16. A method of obtaining a T cell receptor (TCR) that specifically binds a peptide antigen, comprising screening the library of phage particles of claim 12 with the peptide antigen, the method comprising:
a) panning the library using as a target the peptide antigen;
b) repeating step a) one or more times;
c) screening the phage particles identified in step a) or b); and
d) identifying a TCR that specifically binds the peptide antigen.
17. A method of making the library of particles of claim 1 , the library displaying a plurality of different TCRs, the method comprising:
i) obtaining a plurality of nucleic acids from a natural repertoire that encode different TRAV alpha chain variable domains;
ii) obtaining a plurality of nucleic acids from a natural repertoire that encode different TRBV beta chain variable domains, excluding nucleic acids that encode one or more of TRBV5-1, 5-3, 5-4, 5-5, 5-6, 5-7 or 5-8 beta chain variable domains;
iii) cloning the TRAV alpha chain variable domain encoding nucleic acids into expression vectors;
iv) cloning the TRBV beta chain variable domain encoding nucleic acids into the same or different vectors; and
v) expressing the vectors in particles, thereby generating a library consisting essentially of TCRs comprising an alpha chain variable domain and a beta chain variable domain encoded by the nucleic acids.
18. A method of making the library of particles of claim 1 , the library displaying a plurality of different TCRs, the method comprising:
i) obtaining a plurality of nucleic acids from a natural repertoire that encode different TRAV alpha chain variable domains using primers that hybridize to nucleic acids encoding TRAV alpha chain variable domains;
ii) obtaining a plurality of nucleic acids from a natural repertoire that encode different TRBV beta chain variable domains using primers that hybridize to nucleic acids encoding TRBV beta chain variable domains, excluding primers that hybridize to one or more nucleic acids encoding a TRBV5-1, 5-3, 5-4, 5-5, 5-6, 5-7 or 5-8 beta chain variable domains;
iii) cloning the TRAV alpha chain variable domain encoding nucleic acids into expression vectors;
iv) cloning the TRBV beta chain variable domain encoding nucleic acids into the same or different vectors; and
v) expressing the vectors in particles, thereby generating a library consisting essentially of TCRs comprising an alpha chain variable domain and a beta chain variable domain encoded by the nucleic acids to which said primers hybridize.
19. The library of claim 1 , wherein at least a plurality of the particles display a TRBV3-1 gene product.