# US 8,771,693 B2 · Methods and compositions for the generation and use of conformation-specific antibodies

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- Number: US 8,771,693 B2
- Document ID: US8771693B2
- Status: Granted
- Kind: B2
- Application number: 13/504,700
- Filed: 2010-10-26
- Published: 2013-01-31
- Granted: 2014-07-08
- Earliest claimed: 2009-10-27
- Assignees: Beth Israel Deaconess Medical Center, Inc.
- Applicants: Kun Ping Lu; Xiao Zhen Zhou
- Inventors: Kun Ping Lu; Xiao Zhen Zhou
- Web page: https://patentgrep.com/patent/US8771693B2

## CPC

- C07K 16/40: against enzymes
- A61P 11/06: Antiasthmatics
- A61P 25/28: for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
- A61P 35/00: Antineoplastic agents
- A61P 37/04: Immunostimulants
- C07K 14/4711: {Alzheimer's disease; Amyloid plaque core protein}
- C07K 16/44: against material not provided for elsewhere {, e.g. haptens, metals, DNA, RNA, amino acids}
- G01N 33/573: for enzymes or isoenzymes
- C07K 2317/14: Specific host cells or culture conditions, e.g. components, pH or temperature
- G01N 2333/99: Isomerases (5.)
- 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
- 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

The present invention features methods and compositions for the generation and use of conformation-specific anti-bodies or fragments thereof.

## Claims

### Claim 1

An isolated conformation-specific antibody or fragment thereof that specifically binds to a Xaa-Pro motif of a polypeptide, wherein: said polypeptide is a PPlase substrate, the peptidyl-prolyl bond of said Xaa-Pro motif is in a cis conformation, said Xaa is serine or threonine and is phosphorylated, and said antibody or fragment thereof binds to the cis conformation of said Xaa-Pro motif of said polypeptide with at least 10- to 100-fold greater affinity than to the trans conformation of said Xaa-Pro motif of said polypeptide.

### Claim 2 (depends on 1)

The isolated antibody of claim 1, wherein said PPlase substrate is a Pin1 substrate.

### Claim 3 (depends on 2)

The isolated antibody of claim 2, wherein said Pin1 substrate is NIMA, RAB4, CDC25, WEE1, PLK1, MYT1, CDC27, CENP-F, Incenp, RBP1, NHERF-1, KRMP1, CK2, TopoIIα, DAB2, p54nrb, Sil, EMI1, cyclin D1, Ki67, c-Myc, cyclin E, c-Jun, β-catenin, Cf-2, NF-κB, RAF1, c-Fos, RARα, AIB1/SRC-3, HBx, STAT3, p53, Bcl-2, p73, BimEL, p66Shc, CHE1, tau, amyloid precursor protein (APP), APP fragment, synphilin-1, gephyrin, MCL1, NFAT, AUF1, IRF3, BTK, SIN3-RPD3, or hSpt5.

### Claim 4 (depends on 1)

The isolated antibody of claim 1, produced by a process which comprises the steps of:
(i) providing an antibody library;
(ii) contacting said antibody library with a polypeptide comprising a Xaa-Pro motif;
(iii) determining binding of an antibody from said antibody library to said Xaa-Pro motif of said polypeptide, wherein said antibody specifically binds to the cis conformation of said Xaa-Pro motif of said polypeptide; and
(iv) isolating said antibody, wherein said antibody is a conformation-specific antibody.

### Claim 5 (depends on 1)

An isolated antibody composition of claim 1 to induce passive immunity against a disorder associated with a deregulation of PPlase activity.

### Claim 6 (depends on 3)

The isolated antibody of claim 3, wherein said Pin1 substrate is tau.

### Claim 7 (depends on 3)

The isolated antibody of claim 3, wherein said Pin1 substrate is APP.

### Claim 8 (depends on 3)

The isolated antibody of claim 3, wherein said Pin1 substrate is c-Jun.

### Claim 9 (depends on 3)

The isolated antibody of claim 3, wherein said Pin1 substrate is NF-κB.

### Claim 10 (depends on 3)

The isolated antibody of claim 3, wherein said Pin1 substrate is p53.

### Claim 11

An isolated conformation-specific antibody or fragment thereof that specifically binds to a Xaa-Pro motif of a polypeptide, wherein: said polypeptide is a PPlase substrate, the peptidyl-prolyl bond of said Xaa-Pro motif is in a trans conformation, said Xaa is serine or threonine and is phosphorylated, and said antibody or fragment thereof binds to the trans conformation of said Xaa-Pro motif of said polypeptide with at least 10- to 100-fold greater affinity than to the cis conformation of said Xaa-Pro motif of said polypeptide.

### Claim 12 (depends on 11)

The isolated antibody of claim 11, wherein said PPlase substrate is a Pin1 substrate.

### Claim 13 (depends on 12)

The isolated antibody of claim 12, wherein said Pin1 substrate is NIMA, RAB4, CDC25, WEE1, PLK1, MYT1, CDC27, CENP-F, Incenp, RBP1, NHERF-1, KRMP1, CK2, TopoIIα, DAB2, p54nrb, Sil, EMI1, cyclin D1, Ki67, c-Myc, cyclin E, c-Jun, β-catenin, Cf-2, NF-κB, RAF1, c-Fos, RARα, AIB1/SRC-3, HBx, STAT3, p53, Bcl-2, p73, BimEL, p66Shc, CHE1, tau, amyloid precursor protein (APP), APP fragment, synphilin-1, gephyrin, MCL1, NFAT, AUF1, IRF3, BTK, SIN3-RPD3, or hSpt5.

### Claim 14 (depends on 13)

The isolated antibody of claim 13, wherein said Pin1 substrate is tau.

### Claim 15 (depends on 13)

The isolated antibody of claim 13, wherein said Pin1 substrate is APP.

### Claim 16 (depends on 13)

The isolated antibody of claim 13, wherein said Pin1 substrate is c-Jun.

### Claim 17 (depends on 13)

The isolated antibody of claim 13, wherein said Pin1 substrate is NF-κB.

### Claim 18 (depends on 13)

The isolated antibody of claim 13, wherein said Pin1 substrate is p53.

### Claim 19 (depends on 11)

The isolated antibody of claim 11, produced by a process which comprises the steps of:
(i) providing an antibody library;
(ii) contacting said antibody library with a polypeptide comprising a Xaa-Pro motif;
(iii) determining binding of an antibody from said antibody library to said Xaa-Pro motif of said polypeptide, wherein said antibody specifically binds to the trans conformation of said Xaa-Pro motif of said polypeptide; and
(iv) isolating said antibody, wherein said antibody is a conformation-specific antibody.

### Claim 20 (depends on 11)

An isolated antibody composition of claim 11 to induce passive immunity against a disorder associated with a deregulation of PPlase activity.