# US 2020/0399359 A1 · Combinatorial antibody libraries and uses thereof

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- Number: US 2020/0399359 A1
- Document ID: US20200399359A1
- Status: Published application
- Kind: A1
- Application number: 17/002,749
- Filed: 2020-08-25
- Published: 2020-12-24
- Earliest claimed: 2008-11-07
- Assignees: Taurus Biosciences, LLC
- Applicants: Taurus Biosciences, LLC
- Inventors: Vaughn SMIDER; James GRAZIANO; Helen Hongyuan MAO; Byeong Doo SONG; Tyson CHASE; Omar BAZIRGAN
- Web page: https://patentgrep.com/patent/US20200399359A1

## CPC

- C07K 16/22: against growth factors {; against growth regulators}
- A61P 9/10: for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
- A61P 15/00: Drugs for genital or sexual disorders (for disorders of sex hormones A61P5/24); Contraceptives
- A61P 19/02: for joint disorders, e.g. arthritis, arthrosis
- A61P 27/02: Ophthalmic agents
- A61P 35/00: Antineoplastic agents
- A61P 43/00: Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
- C07K 16/00: Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies
- C07K 16/18: against material from animals or humans
- C07K 16/28: against receptors, cell surface antigens or cell surface determinants
- C07K 16/2863: {against receptors for growth factors, growth regulators}
- C07K 16/2869: {against hormone receptors (for antibodies against neuromediator receptors C07K16/286)}
- C07K 16/2884: {against CD44}
- C07K 16/2887: {against CD20}
- C07K 16/32: against translation products of oncogenes
- C12N 15/1037: {Screening libraries presented on the surface of microorganisms, e.g. phage display, E. coli display}
- C40B 40/08: Libraries containing RNA or DNA which encodes proteins, e.g. gene libraries
- C40B 50/08: Liquid phase synthesis, i.e. wherein all library building blocks are in liquid phase or in solution during library creation; Particular methods of cleavage from the liquid support
- G01N 33/6845: {Methods of identifying protein-protein interactions in protein mixtures}
- C07K 2317/14: Specific host cells or culture conditions, e.g. components, pH or temperature
- C07K 2317/24: containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
- C07K 2317/51: Complete heavy chain or Fd fragment, i.e. VH + CH1
- C07K 2317/515: Complete light chain, i.e. VL + CL
- C07K 2317/55: Fab or Fab'
- C07K 2317/56: variable (Fv) region, i.e. VH and/or VL
- C07K 2317/73: Inducing cell death, e.g. apoptosis, necrosis or inhibition of cell proliferation
- C07K 2317/74: Inducing cell proliferation
- C07K 2317/76: Antagonist effect on antigen, e.g. neutralization or inhibition of binding
- C07K 2317/92: Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value
- C40B 40/10: Libraries containing peptides or polypeptides, or derivatives thereof
- C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- C40B: COMBINATORIAL CHEMISTRY; LIBRARIES, e.g. CHEMICAL LIBRARIES (in silico combinatorial libraries of nucleic acids, proteins or peptides G16B35/00; in silico combinatorial chemistry G16C20/60)
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)

## Abstract

Methods for making a combinatorial antibody library from human germline segments are provided. Also provided are libraries of nucleic acid molecules compiled from germline segments encoding VL chains and libraries of nucleic acid molecules encoding VH chains, and resulting antibody libraries. The libraries are provided as addressable libraries. Methods for screening antibody libraries against a target protein antigen, and the identified or selected antibodies are provided.

## Claims

### Claim 1

A combinatorial human antibody library, wherein the library is an addressable library and is selected from the group consisting of:
A. a combinatorial human antibody addressable library comprising a plurality of antibody or antigen-binding antibody fragments, wherein each member in the library is a functional antibody or functional antigen-binding antibody fragment, and:
a) each antibody or antigen-binding antibody fragment contains a variable light (VL) chain and a variable heavy (VH) chain or a sufficient portion thereof to form an antigen binding site; wherein:
i) each VL chain is encoded by a nucleic acid molecule that comprises a Vκ and a Jκ human germline segment or degenerate codons thereof, or a Vλ and a Jλ human germline segment or degenerate codons thereof, wherein the segments are linked in-frame without a stop codon; and
ii) each VH chain is encoded by a nucleic acid molecule that comprises a human VH and a human JH germline segment and any sequence of nucleotides between the VH and JH germline segments, wherein the segments are linked in-frame without a stop codon; and
b) the library comprises at least about or 50 or 100 or more different antibody or antigen-binding antibody fragments;
B. a combinatorial human antibody addressable library comprising a plurality of antibody or antigen-binding antibody fragments, wherein each member in the library is a functional antibody or functional antigen-binding antibody fragment, and:
a) each antibody or antigen-binding antibody fragment contains a modified variable light (VL) chain and/or a modified variable heavy chain (VH) chain or a sufficient portion thereof to form an antigen binding site; wherein:
i) each VL chain is encoded by a nucleic acid molecule that comprises a Vκ and a Jκ human germline segment or degenerate codons thereof or a Vλ and a Jλ human germline segment or degenerate codons thereof, wherein the segments are linked in-frame without a stop codon;
ii) each VH chain is encoded by a nucleic acid molecule that comprises a VH, DH and a JH human germline segment or degenerate codons thereof, wherein the segments are linked in-frame without a stop codon; and
b) the VL chain and/or VH chain is/are modified by replacement or insertion of at least one amino acid into at least one complementarity determining region (CDR); and
c) the library comprises at least about or 50 or 100 or more different antibody or antigen-binding antibody fragments.

### Claim 2 (depends on 1)

The combinatorial human antibody library according to claim 1, wherein:
the sequence of nucleotides between the VH and JH germline segments encodes a peptide mimetic; or
the amino acids that are inserted or replaced correspond to a peptide mimetic.

### Claim 3 (depends on 2)

The combinatorial human antibody library according to claim 2, wherein:
the peptide mimetic is selected from the group consisting of a peptide mimetic of TPO, EPO, G-CSF, IL-5, human brain natriuretic peptide (hBNP-32), exendin 4, GLP-1, GLP-2, glucagon, PACAP-38, CD209L, TNF, VEGF, MMP inhibitor, and CTLA-4; or
the peptide mimetic has the sequence of amino acids selected from among any of SEQ ID NOS: 891 and 987-1014.

### Claim 4 (depends on 1)

The combinatorial human antibody library according to claim 1, wherein the antibody or antigen-binding antibody fragment at each address is the same antibody or antigen-binding antibody fragment and is different from the antibody or antigen-binding antibody fragment at all other addresses.

### Claim 5 (depends on 1)

The combinatorial human antibody library according to claim 1, wherein:
all or a subset of germline VH segments are linked with all or a subset of DH segments which are linked with all or a subset of germline JH segments to generate the plurality of nucleic acid molecules encoding a VH chain; and
all or a subset of germline Vκ segments are linked with all or a subset of germline Jκ segments, or all or a subset of germline Vλ segments are linked to all or a subset of germline Jλ segments to generate a plurality of nucleic acid molecules encoding a VL chain.

### Claim 6 (depends on 1)

The combinatorial human antibody library according to claim 1, wherein
each VH germline segment is selected from the group consisting of IGHV1, IGHV2, IGHV3, IGHV4, IGHV5, IGHV6, IGHV7, and an allelic variant of any of the foregoing; and/or
each DH germline segment is selected from the group consisting of IGHD1, IGHD2, IGHD3, IGHD4, IGHD5, IGHD6, IGHD7, and an allelic variant of any of the foregoing; and/or
each JH germline segment is selected from the group consisting of IGHJ1, IGHJ2, IGHJ3, IGHJ4, IGHJ5, IGHJ6, and an allelic variant of any of the foregoing; and/or
each Vκ germline segment is selected from the group consisting of IGKV1, IGKV2, IGKV3, IGKV4, IGKV5, IGKV6, and an allelic variant of any of the foregoing; and/or
each Jκ germline segment is selected from the group consisting of IGKJ1, IGKJ2, IGKJ3, IGKJ4, IGKJ5, and an allelic variant of any of the foregoing and/or
each Vλ germline segment is selected from the group consisting of IGLV1, IGLV2, IGLV3, IGLV4, IGLV5, IGLV6, IGLV7, IGLV8, IGLV9, IGLV10, IGLV11, and an allelic variant of any of the foregoing; and/or
each Jλ germline segment selected from the group consisting of IGLJ1, IGLJ2, IGLJ3, IGLJ4, IGLJ5, IGLJ6, IGLJ7, and an allelic variant of any of the foregoing.

### Claim 7 (depends on 1)

The combinatorial human antibody library according to claim 1, wherein the plurality of nucleic acid molecules encoding a VH chain and/or a VL chain is/are generated from a subset of germline segments selected based on sequence similarities or differences, gene family, length, composition, CDR length or composition, species, functionality, specificity, group, or subgroup.

### Claim 8 (depends on 1)

The combinatorial human antibody library according to claim 1 that comprises at or about or more than at or about 50, 102, 103, 104, 2×104, 3×104, 4×104, 5×104, 6×104, 7×104, 8×104, 9×104, 105, 106, 107, 108, 109 different antibody or antigen-binding antibody fragments.

### Claim 9 (depends on 1)

The combinatorial human antibody library of claim 1, wherein the CDR is CDRH3.

### Claim 10 (depends on 1)

The combinatorial human antibody library of claim 1, wherein the sequence of nucleotides between the VH and JH germline segments is a DH germline segment or degenerate codons thereof or is an inverted D—H germline segment.

### Claim 11 (depends on 1)

The combinatorial human antibody library of claim 1, wherein the antibodies or antigen-binding fragments are arranged in a spatial array.

### Claim 12 (depends on 1)

The combinatorial human antibody library of claim 1, wherein the antibodies or antigen-binding fragments are immobilized on a solid support or are in solution.

### Claim 13 (depends on 1)

The combinatorial human antibody library of claim 1, wherein the antibodies or antigen-binding fragments are identifiably labeled.

### Claim 14 (depends on 6)

The combinatorial human antibody library of claim 6, wherein:
each VH germline segment is selected from among any of SEQ ID NOS: 10-238; and/or
each DH germline segment is selected from among any of SEQ ID NOS: 239-272; and/or
each JH germline segment is selected from among any of SEQ ID NOS: 273-285; and/or
each Vκ germline segment is selected from among any of SEQ ID NOS: 286-355 and 868; and/or
each Jκ germline segment is selected from among any of SEQ ID NOS: 356-364; and/or
each Vλ germline segment is selected from among any of SEQ ID NOS: 365-441; and/or
each Jλ germline segment is selected from among any of SEQ ID NOS: 442-451.

### Claim 15 (depends on 1)

The combinatorial human antibody library of claim 1, wherein the antibody or antigen-binding antibody fragments are full length antibodies.

### Claim 16 (depends on 1)

The combinatorial human antibody library of claim 1, wherein the antibody or antigen-binding antibody fragments are antigen-binding antibody fragments.

### Claim 17 (depends on 16)

The combinatorial human antibody library of claim 16, wherein the antigen-binding antibody fragment is a Fab.

### Claim 18 (depends on 1)

A library of nucleic acid molecules that encodes the antibody and/or antigen-binding antibody fragments of a combinatorial human antibody library according to claim 1.

### Claim 19 (depends on 18)

The library of nucleic acid molecules of claim 18, wherein the library of nucleic acid molecules are carried in vectors.

### Claim 20 (depends on 19)

The library of nucleic acid molecules of claim 19, wherein the vector is an expression vector.

### Claim 21

A library of nucleic acid molecules encoding a combinatorial human antibody library, wherein the encoded combinatorial human antibody library is an addressable library and comprises a plurality of antibody or antigen-binding antibody fragments, wherein each member in the library is a functional antibody or functional antigen-binding antibody fragment, and:
a) each antibody or antigen-binding antibody fragment contains a variable light (VL) chain and a variable heavy (VH) chain or a sufficient portion thereof to form an antigen binding site; wherein:
i) each VL chain is encoded by a nucleic acid molecule that comprises a Vκ and a Jκ human germline segment or degenerate codons thereof or a Vλ and a Jλ human germline segment or degenerate codons thereof, wherein the segments are linked in-frame without a stop codon;
ii) each VH chain is encoded by a nucleic acid molecule that comprises a human VH, DH, and a JH germline segment or degenerate codons thereof, wherein the segments are linked in-frame without a stop codon; and
b) the library comprises at least about or 50 or 100 or more different antibody or antigen-binding antibody fragments.

### Claim 22 (depends on 21)

The nucleic acid library of claim 21, wherein in the encoded combinatorial human antibody library the antibody or antigen-binding antibody fragment at each address is the same antibody or antigen-binding antibody fragment and is different from the antibody or antigen-binding antibody fragment at all other addresses.

### Claim 23 (depends on 21)

The nucleic acid library according to claim 21, wherein in the encoded combinatorial human antibody library:
all or a subset of germline VH segments are linked with all or a subset of DH segments which are linked with all or a subset of germline JH segments to generate the plurality of nucleic acid molecules encoding a VH chain; and
all or a subset of germline Vκ segments are linked with all or a subset of germline Jκ segments, or all or a subset of germline Vλ segments are linked to all or a subset of germline Jλ segments to generate a plurality of nucleic acid molecules encoding a VL chain.

### Claim 24 (depends on 21)

The nucleic acid library according to claim 21, wherein in the encoded combinatorial human antibody library:
each VH germline segment is selected from the group consisting of IGHV1, IGHV2, IGHV3, IGHV4, IGHV5, IGHV6, IGHV7, and an allelic variant of any of the foregoing; and/or
each DH germline segment is selected from the group consisting of IGHD1, IGHD2, IGHD3, IGHD4, IGHD5, IGHD6, IGHD7, and an allelic variant of any of the foregoing; and/or
each JH germline segment is selected from the group consisting of IGHJ1, IGHJ2, IGHJ3, IGHJ4, IGHJ5, IGHJ6, and an allelic variant of any of the foregoing; and/or
each Vκ germline segment is selected from the group consisting of IGKV1, IGKV2, IGKV3, IGKV4, IGKV5, IGKV6, and an allelic variant of any of the foregoing; and/or
each Jκ germline segment is selected from the group consisting of IGKJ1, IGKJ2, IGKJ3, IGKJ4, IGKJ5, and an allelic variant of any of the foregoing and/or
each Vλ germline segment is selected from the group consisting of IGLV1, IGLV2, IGLV3, IGLV4, IGLV5, IGLV6, IGLV7, IGLV8, IGLV9, IGLV10, IGLV11, and an allelic variant of any of the foregoing; and/or
each Jλ germline segment selected from the group consisting of IGLJ1, IGLJ2, IGLJ3, IGLJ4, IGLJ5, IGLJ6, IGLJ7, and an allelic variant of any of the foregoing.

### Claim 25 (depends on 21)

The nucleic acid library of claim 21, wherein the encoded combinatorial human antibody library comprises at or about or more than at or about 50, 102, 103, 104, 2×104, 3×104, 4×104, 5×104, 6×104, 7×104, 8×104, 9×104, 105, 106, 107, 108, 109 different antibodies or antigen-binding antibody fragments.

### Claim 26 (depends on 21)

The nucleic acid library of claim 21, wherein in the encoded combinatorial human antibody library:
each VH germline segment is selected from among any of SEQ ID NOS: 10-238; and/or
each DH germline segment is selected from among any of SEQ ID NOS: 239-272; and/or
each JH germline segment is selected from among any of SEQ ID NOS: 273-285; and/or
each Vκ germline segment is selected from among any of SEQ ID NOS: 286-355 and 868; and/or
each Jκ germline segment is selected from among any of SEQ ID NOS: 356-364; and/or
each Vλ germline segment is selected from among any of SEQ ID NOS: 365-441; and/or
each Jλ germline segment is selected from among any of SEQ ID NOS: 442-451.

### Claim 27 (depends on 21)

The nucleic acid library of claim 21, wherein in the encoded combinatorial human antibody library each of the antibodies or antigen-binding antibody fragments are full length antibodies.

### Claim 28 (depends on 21)

The nucleic acid library of claim 21, wherein in the encoded combinatorial human antibody library each of the antibody or antigen-binding antibody fragments are antigen-binding antibody fragments.

### Claim 29 (depends on 28)

The nucleic acid library of claim 28, wherein the antigen-binding antibody fragment is a Fab.

### Claim 30

A method of generating a combinatorial human antibody library, comprising:
a) combining a VH, a DH, and a JH human germline segment or portion thereof in frame to generate a sequence of a nucleic acid molecule encoding a VH chain or a portion thereof;
b) combining a Vκ and a Jκ human germline segment or portion thereof, or a Vλ and a Jλ human germline segment or portion thereof in frame to generate a sequence of a nucleic acid molecule encoding a VL chain or a portion thereof, wherein:
in step a) and b) each of the portions of the VH, DH, JH, Vκ, Jκ, Vλ or Jλ are
sufficient to produce an antibody or portion thereof containing a VH or VL or portion thereof that forms a sufficient antigen binding site;
c) repeating step a) and b) a plurality of times to generate sequences of a plurality of different nucleic acid molecules;
d) synthesizing the nucleic acid molecules to produce two libraries, wherein:
the first library comprises nucleic acid molecules encoding a VH chain or a portion thereof; and
the second library comprises nucleic acid molecules encoding a VL chain or a portion thereof;
e) expressing the nucleic acid molecules of the first library and the second library of part (d) and generating an antibody or antigen-binding antibody fragment formed by the VH chains and VL chains or portions thereof, thereby generating the combinatorial human antibody library.

### Claim 31 (depends on 30)

The method of generating a combinatorial human antibody library of claim 30, wherein expressing the nucleic acid molecules comprises introducing a nucleic acid molecule from the first library and from the second library into a cell and growing the cells under conditions to express the antibodies or portions thereof in each cell, and repeating this a plurality of times to produce a library of cells, wherein each cell contains nucleic acid molecules encoding a different combination of VH and VL from every other cell in the library of cells.

### Claim 32 (depends on 30)

The method of generating a combinatorial human antibody library of claim 30, wherein the library is an addressable library, wherein in step d) the synthesized nucleic acid sequences are individually addressed, thereby generating a first addressed nucleic acid library and a second addressed nucleic acid library.

### Claim 33 (depends on 30)

The method of generating a combinatorial human antibody library of claim 30, wherein the expressed antibody is a full length antibody, or a fragment or portion thereof sufficient to form an antigen binding site.

### Claim 34 (depends on 30)

The method of generating a combinatorial human antibody library of claim 30, wherein the expressed antibody is a Fab.

### Claim 35 (depends on 30)

The method of generating a combinatorial human antibody library of claim 30, further purifying the antibodies or portions thereof.

### Claim 36 (depends on 30)

A human combinatorial antibody library produced by the method of claim 30.

### Claim 37 (depends on 1)

A method of screening a combinatorial human antibody library for binding or activity against a target protein, comprising:
a) contacting one or more members of a combinatorial human antibody library according to claim 1 with a target protein, wherein the target protein is a membrane-bound protein, cell surface receptor (CSR), or a CSR ligand, a cytokine receptor, a receptor kinase, a receptor phosphatase, a receptor involved in cell-cell interactions, and a cellular adhesion molecule, wherein the target protein is selected from the group consisting of VEGFR-1, VEGFR-2, VEGFR-3, a epidermal growth factor receptor (EGFR), ErbB-2, ErbB-3, IGF-R1, C-Met, TNF-R1, TNF-R2, BTLA, HVEM, LT-13R, CD20, CD3, CD25, NOTCH, DLL4, G-CSF-R, GM-CSF-R, EPO-R, a cadherin, an integrin, CD52 and CD44, a VEGF-A, VEGF-B, VEGF-C, VEGF-D, PIGF, EGF, HGF, TNF-a, LIGHT, lymphotoxin (LT), IgE, G-CSF, GM-CSF, and EPO; and
b) determining whether any member of the combinatorial human antibody library binds to or modulates a functional activity of the target protein, wherein the functional activity is selected from the group consisting of cellular proliferation, lymphoma apoptosis, chemotaxis, cancer cell invasion, matrigel, endothelial proliferation, tube formation, and signal transduction;
wherein the method further comprises identifying the antibody or antigen-binding antibody fragment that binds the target protein.

### Claim 38 (depends on 36)

A method of screening a combinatorial human antibody library for binding or activity against a target protein, comprising:
a) contacting one or more members of a combinatorial human antibody library of claim 36 with a target protein, wherein the target protein is a membrane-bound protein, cell surface receptor (CSR), or a CSR ligand, a cytokine receptor, a receptor kinase, a receptor phosphatase, a receptor involved in cell-cell interactions, and a cellular adhesion molecule, wherein the target protein is selected from the group consisting of VEGFR-1, VEGFR-2, VEGFR-3, a epidermal growth factor receptor (EGFR), ErbB-2, ErbB-3, IGF-R1, C-Met, TNF-R1, TNF-R2, BTLA, HVEM, LT-13R, CD20, CD3, CD25, NOTCH, DLL4, G-CSF-R, GM-CSF-R, EPO-R, a cadherin, an integrin, CD52 and CD44, a VEGF-A, VEGF-B, VEGF-C, VEGF-D, PIGF, EGF, HGF, TNF-a, LIGHT, lymphotoxin (LT), IgE, G-CSF, GM-CSF, and EPO; and
b) determining whether any member of the combinatorial human antibody library binds to or modulates a functional activity of the target protein, wherein the functional activity is selected from the group consisting of cellular proliferation, lymphoma apoptosis, chemotaxis, cancer cell invasion, matrigel, endothelial proliferation, tube formation, and signal transduction;
wherein the method further comprises identifying the antibody or antigen-binding antibody fragment that binds the target protein.

### Claim 39

A method of screening a combinatorial human antibody library for binding or activity against a target protein, comprising:
1) contacting one or more members of a combinatorial human antibody library with a target protein, wherein:
the combinatorial human antibody library is an addressable library and comprises a plurality of antibody or antigen-binding antibody fragments, wherein each member in the library is a functional antibody or functional antigen-binding antibody fragment, and:
a) each antibody or antigen-binding antibody fragment contains a variable light (VL) chain and a variable heavy (VH) chain or a sufficient portion thereof to form an antigen binding site; wherein:
i) each VL chain is encoded by a nucleic acid molecule that comprises a Vκ and a Jκ human germline segment or degenerate codons thereof or a Vλ and a Jλ human germline segment or degenerate codons thereof, wherein the segments are linked in-frame without a stop codon;
ii) each VH chain is encoded by a nucleic acid molecule that comprises a human VH, DH, and a JH germline segment or degenerate codons thereof, wherein the segments are linked in-frame without a stop codon; and
b) the library comprises at least about or 50 or 100 or more different antibody or antigen-binding antibody fragments; and
the target protein is a membrane-bound protein, cell surface receptor (CSR), or a CSR ligand, a cytokine receptor, a receptor kinase, a receptor phosphatase, a receptor involved in cell-cell interactions, and a cellular adhesion molecule, wherein the target protein is selected from the group consisting of VEGFR-1, VEGFR-2, VEGFR-3, a epidermal growth factor receptor (EGFR), ErbB-2, ErbB-3, IGF-R1, C-Met, TNF-R1, TNF-R2, BTLA, HVEM, LT-13R, CD20, CD3, CD25, NOTCH, DLL4, G-CSF-R, GM-CSF-R, EPO-R, a cadherin, an integrin, CD52 and CD44, a VEGF-A, VEGF-B, VEGF-C, VEGF-D, PIGF, EGF, HGF, TNF-a, LIGHT, lymphotoxin (LT), IgE, G-CSF, GM-CSF, and EPO; and
2) determining whether any member of the combinatorial human antibody library binds to or modulates a functional activity of the target protein, wherein the functional activity is selected from the group consisting of cellular proliferation, lymphoma apoptosis, chemotaxis, cancer cell invasion, matrigel, endothelial proliferation, tube formation, and signal transduction;
wherein the method further comprises identifying the antibody or antigen-binding antibody fragment that binds the target protein.