Fibronectin based scaffold proteins

Patent metadata

Field	Value
Publication number	US-2015361159-A1
Application number	US-201414764215-A
Country	US
Kind code	A1
Filing date	Jan 30, 2014
Priority date	Feb 1, 2013
Publication date	Dec 17, 2015
Grant date	—

Abstract

Fibronectin type III ( 10 Fn3) binding domains having novel designs are provided. Also provided are 10 Fn3 domains having combinations of modified regions that can bind to a desired target.

First claim

1 . A polypeptide comprising a 10 Fn3 domain, wherein the 10 Fn3 domain comprises an amino acid sequence wherein
the amino acids corresponding to residues 77-83 of SEQ ID NO: 1 or 6 are represented by x 3-12 , the amino acids corresponding to residues 77-86 of SEQ ID NO: 1 or 6 are represented by x 6-15 , or the amino acids corresponding to residues 75-86 of SEQ ID NO: 1 or 6 are represented by x 4-8 Sx, or the amino acids corresponding to residues 75-83 of SEQ ID NO: 1 or 6 are represented by x 8-14 , or the amino acids corresponding to residues 73-89 of SEQ ID NO: 1 or 6 are represented by xAxxxx 3-12 xIx, or the amino acids corresponding to residues 76-86 of SEQ ID NO: 1 or 6 are represented by xx 3-12 , or the amino acids corresponding to residues 76-86 of SEQ ID NO: 1 or 6 are represented by xx 4-8 Sx, or the amino acids corresponding to residues 76-83 of SEQ ID NO: 1 or 6 are represented by xx 8-14 , or the amino acids corresponding to residues 23-29 of SEQ ID NO: 1 or 6 are represented by x 7 , or the amino acids corresponding to residues 25-29 of SEQ ID NO: 1 or 6 are represented by x 5 , or the amino acids corresponding to residues 26-29 of SEQ ID NO: 1 or 6 are represented by x 4 , or the amino acids corresponding to residues 27-29 of SEQ ID NO: 1 or 6 are represented by x 3 , or the amino acids corresponding to residues 24-28 of SEQ ID NO: 1 or 6 are represented by x 5 , or the amino acids corresponding to residues 25-28 of SEQ ID NO: 1 or 6 are represented by x 4 , or the amino acids corresponding to residues 26-28 of SEQ ID NO: 1 or 6 are represented by x 3 or the amino acids corresponding to residues 17-26 of SEQ ID NO: 1 or 6 are represented by xLxIxWx 4 , or the amino acids corresponding to residues 24-35 of SEQ ID NO: 1 or 6 are represented by x 4 YxIx, or the amino acids corresponding to residues 1-9 of SEQ ID NO: 1 or 6 are represented by x 1-9 Lx, wherein x is any amino acid, and wherein the 10 Fn3 domain binds specifically to a target that is not bound by a 10 Fn3 domain having an amino acid sequence consisting of SEQ ID NO: 1 or 6.

2 - 3 . (canceled)

4 . The polypeptide of claim 1 , wherein the 10 Fn3 domain comprises at least one amino acid variation in the amino acid sequence corresponding to residues 17-30 of SEQ ID NO: 1 or 6.

5 . The polypeptide of claim 1 , wherein the 10 Fn3 domain comprises at least one amino acid variation in the amino acid sequence corresponding to residues 1-9 of SEQ ID NO: 1 or 6.

6 - 7 . (canceled)

8 . The polypeptide of claim 1 , wherein at least 50% of residues x are not the amino acid at the corresponding position in SEQ ID NO: 1 or 6.

9 . The polypeptide of claim 1 , wherein the 10 Fn3 domain binds specifically to a target with a Kd of less than 500 nM.

10 . A library comprising a plurality of polypeptides comprising a 10 Fn3 domain, wherein the 10 Fn3 domain comprises an amino acid sequence wherein
the amino acids corresponding to residues 77-83 of SEQ ID NO: 1 or 6 are represented by x 3-12 , the amino acids corresponding to residues 77-86 of SEQ ID NO: 1 or 6 are represented by x 6-15 , or the amino acids corresponding to residues 75-86 of SEQ ID NO: 1 or 6 are represented by x 4-8 Sx, or the amino acids corresponding to residues 75-83 of SEQ ID NO: 1 or 6 are represented by x 8-14 , or the amino acids corresponding to residues 73-89 of SEQ ID NO: 1 or 6 are represented by xAxxxx 3-12 xIx, or the amino acids corresponding to residues 76-86 of SEQ ID NO: 1 or 6 are represented by xx 3-12 , or the amino acids corresponding to residues 76-86 of SEQ ID NO: 1 or 6 are represented by xx 4-8 Sx, or the amino acids corresponding to residues 76-83 of SEQ ID NO: 1 or 6 are represented by xx 8-14 , or the amino acids corresponding to residues 23-29 of SEQ ID NO: 1 or 6 are represented by x 7 , or the amino acids corresponding to residues 25-29 of SEQ ID NO: 1 or 6 are represented by x 5 , or the amino acids corresponding to residues 26-29 of SEQ ID NO: 1 or 6 are represented by x 4 , or the amino acids corresponding to residues 27-29 of SEQ ID NO: 1 or 6 are represented by x 3 , or the amino acids corresponding to residues 24-28 of SEQ ID NO: 1 or 6 are represented by x 5 , or the amino acids corresponding to residues 25-28 of SEQ ID NO: 1 or 6 are represented by x 4 , or the amino acids corresponding to residues 26-28 of SEQ ID NO: 1 or 6 are represented by x 3 or the amino acids corresponding to residues 17-26 of SEQ ID NO: 1 or 6 are represented by xLxIxWx 4 , or the amino acids corresponding to residues 24-35 of SEQ ID NO: 1 or 6 are represented by x 4 YxIx, or the amino acids corresponding to residues 1-9 of SEQ ID NO: 1 or 6 are represented by x 1-9 Lx, wherein x is any amino acid, and wherein the 10 Fn3 domain binds specifically to a target that is not bound by a 10 Fn3 domain having an amino acid sequence consisting of SEQ ID NO: 1 or 6.

11 - 12 . (canceled)

13 . A method for identifying a polypeptide that binds to a target comprising screening the library of claim 10 to identify a polypeptide that binds to the target.

14 - 274 . (canceled)

275 . A polypeptide comprising a 10 Fn3 domain, wherein the 10 Fn3 domain comprises at least one variable amino acid sequence, wherein
the amino acids corresponding to residues 1-9 of SEQ ID NO: 1 or 6 are represented by x 1-9 Lx; or the amino acids corresponding to residues 1-9 of SEQ ID NO: 1 or 6 consist of the amino acid sequence GVSDVPGGSG, or a portion thereof, linked to an amino acid sequence represented by x 1-9 Lx; and/or the amino acids corresponding to residues 23-29 of SEQ ID NO: 1 or 6 are represented by x 7 ; or the amino acids corresponding to residues 25-29 of SEQ ID NO: 1 or 6 are represented by x 5 ; or the amino acids corresponding to residues 26-29 of SEQ ID NO: 1 or 6 are represented by x 4 ; or the amino acids corresponding to residues 27-29 of SEQ ID NO: 1 or 6 are represented by x 3 ; or the amino acids corresponding to residues 24-28 of SEQ ID NO: 1 or 6 are represented by x 5 ; or the amino acids corresponding to residues 25-28 of SEQ ID NO: 1 or 6 are represented by x 4 ; or the amino acids corresponding to residues 26-28 of SEQ ID NO: 1 or 6 are represented by x 3 ; or the amino acids corresponding to residues 17-26 of SEQ ID NO: 1 or 6 are represented by xLxIxWx 4 ; or the amino acids corresponding to residues 24-35 of SEQ ID NO: 1 or 6 are represented by x 6 YxIx; and/or the amino acids corresponding to residues 52-55 of SEQ ID NO: 1 or 6 are represented by x 4 ; or the amino acids corresponding to residues 47-55 of SEQ ID NO: 1 or 6 are represented by xFxVx 4 ; or the amino acids corresponding to residues 51-60 of SEQ ID NO: 1 or 6 are represented by x 6 AxIx; and/or the amino acids corresponding to residues 77-83 of SEQ ID NO: 1 or 6 are represented by x 3-12 ; or the amino acids corresponding to residues 77-86 of SEQ ID NO: 1 or 6 are represented by x 6-15 ; or the amino acids corresponding to residues 75-86 of SEQ ID NO: 1 or 6 are represented by x 4-8 Sx; or the amino acids corresponding to residues 75-83 of SEQ ID NO: 1 or 6 are represented by x 8-14 ; or the amino acids corresponding to residues 73-89 of SEQ ID NO: 1 or 6 are represented by xAxxx 3-12 xIx; or the amino acids corresponding to residues 76-86 of SEQ ID NO: 1 or 6 are represented by xx 3-12 ; or the amino acids corresponding to residues 76-86 of SEQ ID NO: 1 or 6 are represented by xx 4-8 Sx; or the amino acids corresponding to residues 76-83 of SEQ ID NO: 1 or 6 are represented by xx 8-14 ; and
x is any amino acid; and the 10 Fn3 domain binds specifically to a t

Assignees

• Bristol Myers Squibb Co

Inventors

• Lipovsek Dasa
• Davis Jonathan H

Classifications

• C07K2317/92

  Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value · CPC title

• C07K16/2857

  against nuclear receptors, e.g. retinoic acid receptor [RAR], RXR, orphan receptor · CPC title

• C07K14/78 Primary

  Connective tissue peptides, e.g. collagen, elastin, laminin, fibronectin, vitronectin or cold insoluble globulin [CIG] · CPC title

• C07K16/244

  Interleukins [IL] · CPC title

• C07K16/2833

  against MHC-molecules, e.g. HLA-molecules · CPC title