Human fibronectin type iii protein scaffolds
AssigneeARO Biotherapeutics Company
InventorsRonald V. Swanson, Zhanna Druzina, Yao Xin, Karyn O’Neil
Life of this patent
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Abstract
Protein scaffolds and scaffold libraries based on a fibronectin type III (FN3) domain with an alternative binding surface design, isolated nucleic acids encoding the protein scaffolds, vectors, host cells, methods of making thereof, and uses as therapeutic molecules for treatment and diagnosis of diseases and disorders.
Claims22
A library comprising a plurality of fibronectin type III module (FN3) domains having a diversified C-CD-D-F-FG-G alternative surface comprising a diversified C beta-strand, a CD loop, a D beta-strand, an F beta-strand, an FG loop and a G beta-strand, wherein the polypeptides comprise an amino acid sequence of:
- (SEQ ID NO:
- 44)
- MLSPPSNLRVTDVISTSVTLSWKPPAPITGYXVXYXEXXXXGEW
- KXVXVPGSETSYTVTGLKPGTEYXFXVXAVNGAXXGXPSQXVXV
- TT
- or an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity to the amino acid sequence of SEQ ID NO:
- 44, wherein each X is, independently, any amino acid.
A method of making a library of fibronectin module of type III (FN3) domains having a diversified C-CD-F-FG-G alternative surface comprising a diversified one or more, or each of, C beta-strand, a CD loop, an F beta-strand, an FG loop and G-beta strand, comprising
- a. providing a reference FN3 domain polypeptide having an amino acid sequence at least 80% identical to that of SEQ ID NO:
- 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43 or 44;
- and
- b. introducing diversity into the reference FN3 domain polypeptide by mutating at least one residue in the C beta-strand, CD loop region, F beta-strand region, FG loop region, or G-beta strand region to form the FN3 domain library having the diversified C-CD-F-FG-G alternative surface, wherein:
- the diversified C beta-strand has an amino acid sequence of TGYXVXYXE (SEQ ID NO:
- 45), wherein each X is, independently, any amino acid except a methionine or a cysteine;
- the diversified CD loop has an amino acid sequence of XXXXGE (SEQ ID NO:
- 46), wherein each X is, independently, any amino acid except a methionine or a cysteine;
- the diversified D beta-strand has an amino acid sequence of WKXVXVP (SEQ ID NO:
- 47), wherein each X is, independently, any amino acid except a methionine or a cysteine;
- the diversified F beta-strand has an amino acid sequence of TEYXFXVXAV (SEQ ID NO:
- 48), wherein each X is, independently, is any amino acid except a methionine or a cysteine;
- the diversified FG loop has an amino acid sequence of NGAXXG (SEQ ID NO:
- 49), wherein each X is, independently, any amino acid except a methionine or a cysteine;
- and
- the diversified G beta-strand has an amino acid sequence XPSQXVXVTT (SEQ ID NO:
- 50), wherein each X is, independently, any amino acid except a methionine or a cysteine.
(canceled)
A polypeptide having an amino acid sequence that is at least, or about, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or is identical to a sequence selected from the group consisting of SEQ ID NOS:
- 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, and 43.
A polypeptide comprising an amino acid sequence of:
- (SEQ ID NO:
- 44)
- MLSPPSNLRVTDVTSTSVTLSWKPPAPITGYXVXYXEXXXXGEW
- KXVXVPGSETSYTVTGLKPGTEYXFXVXAVNGAXXGXPSQXVXV
- TT
- wherein each X is, independently, any amino acid.
A polypeptide comprising an amino acid sequence of:
- (SEQ ID NO:
- 74)
- LSPPSNLRVTDVTSTSVTLSWKPPAPITGYXVXYXEXXXXGEWK
- XVXVPGSETSYTVTGLKPGTEYXFXVXAVNGAXXGXPSQXVXVI
- T
- wherein each X is, independently, any amino acid.
-25. (canceled)
Dependencies are inferred from the claim text.