Multivalent fibronectin based scaffold domain proteins
AssigneeBristol-Myers Squibb Company
InventorsRay CAMPHAUSEN, Eric FURFINE, Irvith M. CARVAJAL, H. Nicholas MARSH, Marco GOTTARDIS, Joan CARBONI, Ricardo ATTAR
Life of this patent
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Abstract
The present invention relates to multivalent polypeptides comprising at least two fibronectin scaffold domains connected via a polypeptide linker. The invention also relates to multivalent polypeptides for use in diagnostic, research and therapeutic applications. The invention further relates to cells comprising such proteins, polynucleotide encoding such proteins or fragments thereof, and to vectors comprising the polynucleotides encoding the innovative proteins.
Claims16
1-24. (canceled)
A polypeptide comprising:
- (a) an N-terminal domain comprising a first fibronectin type III tenth domain (10Fn3), wherein the 10Fn3 domain (i) comprises a loop, AB;
- a loop, BC;
- a loop, CD;
- a loop, DE;
- a loop EF;
- and a loop FG;
- (ii) has at least one loop selected from loop BC, DE, and FG with an altered amino acid sequence relative to the sequence of the corresponding loop of the human 10Fn3 domain, (iii) comprises an amino acid sequence that is at least 60% identical to SEQ ID NO:
- 1, and (iv) binds to a first target molecule with a KD of less than 500 nM;
- (b) a C-terminal domain comprising a second fibronectin type III tenth domain (10Fn3), wherein the 10Fn3 domain (i) comprises a loop, AB;
- a loop, BC;
- a loop, CD;
- a loop, DE;
- a loop EF;
- and a loop FG;
- (ii) has at least one loop selected from loop BC, DE, and FG with an altered amino acid sequence relative to the sequence of the corresponding loop of the human 10Fn3 domain, (iii) comprises an amino acid sequence that is at least 60% identical to the amino acid sequence of SEQ ID NO:
- 1, and (iv) binds to a second target molecule with a KD of less than 500 nM;
- wherein the first and second 10Fn3 domains are linked via a polypeptide selected from a glycine-serine based linker, a glycine-proline based linker, a proline-alanine based linker, or the amino acid sequence of SEQ ID NO:
- 20.
The polypeptide of claim 1, wherein the first and second 10Fn3 domains comprise an amino acid sequence that is at least 70% identical to the amino acid sequence of SEQ ID NO:
- 1.
The polypeptide of claim 1, wherein the first 10Fn3 domain binds to a first target molecule with a KD of less than 100 nM and the second 10Fn3 domain binds to a second target molecule with a KD of less than 100 nM.
The polypeptide of claim 1, wherein the loop BC and loop FG of the first and second 10Fn3 domains have an altered amino acid sequence relative to the sequence of the corresponding loops of the human 10Fn3 domain.
The polypeptide of claim 1, wherein the first and second target molecules are the same.
The polypeptide of claim 1, wherein the first and second target molecules are different.
The polypeptide of claim 1, wherein the first and/or second 10Fn3 domain comprises an N-terminal extension sequence, such as M, MG, G, or SEQ ID NO:
- 45, 46, or 48.
The polypeptide of claim 1, wherein the first and/or second 10Fn3 domain is linked at its C-terminus to the amino acid sequence of E, EI, ES, EC, EGS, EGC, EID, SEQ ID NO:
- 17, 18, 19, 50, 51, 52, 71, or 72.
The polypeptide of claim 1, wherein the first and second 10Fn3 domains are linked via a glycine-serine based linker, such as a glycine-serine based linker selected from the amino acid sequence of SEQ ID NOs:
- 21, 22, 23, 24, and 25.
The polypeptide of claim 1, wherein the first and second 10Fn3 domains are linked via a glycine-proline based linker, such as a glycine-proline based linker is selected from the amino acid sequence of SEQ ID NOs:
- 32, 33, and 34.
The polypeptide of claim 1, wherein the first and second 10Fn3 domains are linked via a proline-alanine based linker, such as a proline-alanine based linker is selected from the amino acid sequence of SEQ ID NOs:
- 60, 61, and 62.
The polypeptide of claim 1, further comprising one or more pharmacokinetic (PK) moieties selected from:
- a polyoxyalkylene moiety, such as polyethylene glycol, a human serum albumin binding protein, sialic acid, human serum albumin, IgG, an IgG binding protein, transferrin, and an Fc fragment.
The polypeptide of claim 1, wherein said polypeptide has been deimmunized to remove one or more T-cell epitopes.
A pharmaceutically acceptable composition comprising the polypeptide of claim 1.
Dependencies are inferred from the claim text.