Anti-TNF alpha fibronectin type III domain based scaffold compositions, methods and uses
A protein scaffold based on a consensus sequence of the tenth fibronectin type III (FN3) repeat from human fibronectin, preferably human Tenascin, that binds to human TNFα including isolated nucleic acids that encode a protein scaffold, vectors, host cells, and methods of making and using thereof have applications in diagnostic and/or therapeutic compositions, methods and devices.
1. An isolated protein scaffold based on a fibronectin type III (FN3) domain, wherein the isolated protein scaffold comprises:
a backbone portion comprising the amino acid sequence of residues 1-21, 29-74, and 82-89 of SEQ ID NO: 16;
a B:C loop portion between the backbone portion residues corresponding to residues 21 and 29 of SEQ ID NO: 16, wherein the B:C loop portion has an amino acid sequence that has at least 75% identity to the amino acid sequence of SEQ ID NO: 22, 23, 36, 41, 42, 44, 45, 52, 58, 63, or 64; and
an F:G loop portion between the backbone portion residues corresponding to residues 74 and 82 of SEQ ID NO: 16, wherein the F:G loop portion has an amino acid sequence that has at least 75% identity to the amino acid sequence of SEQ ID NO: 78, 80, 96, 97, 112, 114, 122, 127, 145, 157, or 167, wherein the protein scaffold is capable of binding to human TNFα.
2. The protein scaffold of claim 1 , comprising:
a backbone portion comprising the amino acid sequence of residues 1-21, 29-74, and 82-89 of SEQ ID NO: 16;
a B:C loop portion between the backbone portion residues corresponding to residues 21 and 29 of SEQ ID NO: 16, wherein the B:C loop portion comprises the amino acid sequence of SEQ ID NO: 22, 23, 36, 41, 42, 44, 45, 52, 58, 63, or 64; and
an F:G loop portion between the backbone portion residues corresponding to residues 74 and 82 of SEQ ID NO: 16, wherein the F:G loop portion comprises the amino acid sequence of SEQ ID NO: 78, 80, 96, 97, 112, 114, 122, 127, 145, 157, or 167, wherein the protein scaffold is capable of binding to human TNFα.
3. The protein scaffold of claim 2 , wherein said scaffold binds human TNFα with at least one affinity selected from a K D of less than or equal to 10 −9 M, less than or equal to 10 −10 M, less than or equal to 10 −11 M, less than or equal to 10 −12 M, less than or equal to 10 −13 M, less than or equal to 10 −14 M, and less than or equal to 10 −15 M, as determined by surface plasmon resonance or the Kinetic Exclusion Assay method.
4. A composition comprising the protein scaffold according to claim 1 or 2 and at least one pharmaceutically acceptable carrier or diluent.
5. The composition according to claim 4 , further comprising at least one compound or polypeptide selected from a detectable label or reporter, an anti-infective drug, a cardiovascular (CV) system drug, a central nervous system (CNS) drug, an autonomic nervous system (ANS) drug, a respiratory tract drug, a gastrointestinal (GI) tract drug, a hormonal drug, a drug for fluid or electrolyte balance, a hematologic drug, an antineoplastic, an immunomodulation drug, an opthalmic, otic or nasal drug, a topical drug, a nutritional drug, a cytokine, and a cytokine antagonist.