Single domain ligands, receptors comprising said ligands methods for their production, and use of said ligands and receptors

• US 6,248,516 B1
• Filed: 06/06/1995
• Issued: 06/19/2001
• Est. Priority Date: 11/11/1988
• Status: Expired due to Term

• Current Assignee: Stratagene California, The Scripps Research Institute, UK Research and Innovation
• Original Assignee: Medical Research Council (Government of United Kingdom)
• Inventors: Güssow, Detlef, Ward, Elizabeth Sally, Winter, Gregory Paul
• Primary Examiner(s): KETTER, JAMES S
• Application Number:
  US08/470,031
• Time in Patent Office: 2,205 Days
• Field of Search: 435/240.2, 435/252.3, 435/252.33, 435/6, 435/69.6, 435/441, 435/446, 536/23.7, 536/23.4, 536/23.5, 536/23.6
• US Class Current: 435/6.16
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: C07K 16/461 Igs containing Ig-regions, -domains or -residues form different species
• CPC Class Codes: C07K 2317/24 containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
• CPC Class Codes: C12Q 1/6876 Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes

Abstract

The present invention relates to single domain ligands derived from molecules in the immunoglobulin (Ig) superfamily, receptors comprising at least one such ligand, methods for cloning, amplifying and expressing DNA sequences encoding such ligands, preferably using the polymerase chain reaction, methods for the use of said DNA sequences in the production of Ig-type molecules and said ligands or receptors, and the use of said ligands or receptors in therapy, diagnosis and catalysis.

Claims

1. A library for expression of immunoglobulin heavy chain variable domains (VH domains), said library comprising a repertoire of nucleic acid sequences encoding a third CDR of an immunoglobulin heavy chain variable domain, each member of said repertoire being flanked by VH sequences so as to provide nucleic acid encoding a repertoire of immunoglobulin heavy chain variable domains which are identical except for said third CDR.

2. A library according to claim 1 wherein said third CDRs are derived from preexisting repertoires of CDRs.

3. A library according to claim 1 wherein said third CDRs comprise random sequences.

4. A library according to claim 1 wherein said nucleic acid encoding a repertoire of immunoglobulin heavy chain variable domains further comprises a sequence encoding one or more constant domains for expression of Ig-type chains.

5. A method for generating an antibody variable domain expression library having a diversity of CDR3 sequences, said method comprising:

providing expression vectors, said vectors comprising a variable domain encoding sequence of an antibody;

introducing by mutagenesis a diversity of CDR3 sequences into said variable domain encoding sequence; and

recovering an expression library having a diversity of binding activities.

6. The method of claim 5 wherein said antibody variable domain is a VH domain.

7. The method of claim 5 wherein said expression vector encodes an Fab antibody fragment.

8. The method of claim 5 wherein said expression vector encodes a scFv fragment.

18. An expression library produced by the method of claim 5.

19. An expression library produced by the method of claim 6.

20. An expression library produced by the method of claim 7.

21. An expression library produced by the method of claim 8.

9. An expression library which expresses antibody variable domains, said library comprising a universal set of framework regions carrying a diversity of CDR3 sequences, said library having a diversity of binding activities.

10. The expression library of claim 9 wherein said antibody variable domains are VH domains.

11. The expression library of claim 9 wherein said antibody variable domains are VL domains.

12. The expression library of claim 9 wherein said variable domains are expressed in the form of Fab antibody fragments.

13. An expression library which expresses antibody variable domains having CDR diversity in only the CDR3 sequences, said library having a diversity of binding activities.

14. The expression library of claim 13 wherein said antibody variable domains are VH domains.

15. The expression library of claim 13 wherein said antibody variable domains are VL domains.

16. The expression library of claim 13 wherein said variable domains are expressed in the form of Fab antibody fragments.

17. The expression library of claim 13 wherein said variable domains are expressed in the form of scFv antibody fragments.