Antibody fragment-polymer conjugates and uses of same

A conjugate consisting essentially of an antibody fragment covalently modified by one or two nonproteinaceous polymer molecules at a free sulfhydryl group of a cysteine residue within the hinge region of the antibody fragment,

• wherein said antibody fragment is engineered to both (1) provide an unpaired cysteine within the hinge region, so as to provide said free sulfhydryl group, and (2) to avoid disulfide bridge formation between said cysteine and an amino acid in the opposite chain of said antibody fragment,
• wherein the conjugate binds the same antigen as the parental molecule that is not covalently modified by one or two nonproteinaceous polymer molecules, and wherein the antibody fragment comprises an antigen binding site that binds to a human tumor necrosis factor-α (TNF-α) polypeptide.

A conjugate consisting essentially of an antibody fragment covalently modified by one or two nonproteinaceous polymer molecules at a free sulfhydryl group of a cysteine residue within the hinge region of the antibody fragment,

• wherein said antibody fragment is engineered to both (1) provide an unpaired cysteine within the hinge region, so as to provide said free sulfhydryl group, and (2) to avoid disulfide bridge formation between said cysteine and an amino acid in the opposite chain of said antibody fragment,
• wherein the apparent molecular weight of the conjugate, as determined by size exclusion chromatography is at least 500 kD, and the conjugate binds the same antigen as the parental molecule that is not covalently modified by one or two nonproteinaceous polymer molecules, and wherein the antibody fragment comprises an antigen binding site that binds to a human tumor necrosis factor-α (TNF-α) polypeptide.

A conjugate consisting essentially of an antibody fragment covalently modified by one or two nonproteinaceous polymer molecules at a free sulfhydryl group of a cysteine residue within the hinge region of the antibody fragment,

• wherein said antibody fragment is engineered to both (1) provide an unpaired cysteine within the hinge region, so as to provide said free sulfhydryl group, and (2) to avoid disulfide bridge formation between said cysteine and an amino acid in the opposite chain of said antibody fragment,
• wherein (a) the apparent molecular weight of the conjugate, as determined by size exclusion chromatography is at least 500 kD, (b) the average actual molecular weight of each nonproteinaceous polymer molecule is at least 20 kD, and (c) the conjugate binds the same antigen as, the parental molecule that is not covalently modified by one or two nonproteinaceous polymer molecules, and wherein the antibody fragment comprises an antigen binding site that binds to a human tumor necrosis factor-α (TNF-α) polypeptide.

A conjugate consisting essentially of an antibody fragment covalently modified by one or two nonproteinaceous polymer molecules at a free sulfhydryl group of a cysteine residue within the hinge region of the antibody fragment,

• wherein said antibody fragment is engineered to both (1) provide an unpaired cysteine within the hinge region, so as to provide said free sulfhydryl group, and (2) to avoid disulfide bridge formation between said cysteine and an amino acid in the opposite chain of said antibody fragment,
• (a) the apparent molecular weight of the conjugate, as determined by size exclusion chromatography is at least 500 kD, (b) the average actual molecular weight of each nonproteinaceous polymer molecule is at least 20 kD, (c) wherein the conjugate has an apparent size that is about 8-fold greater than the apparent size of the parental antibody fragment, wherein the conjugate binds the same antigen as the parental molecule that is not covalently modified by one or two nonproteinaceous polymer molecules, and wherein the antibody fragment comprises an antigen binding site that binds to a human tumor necrosis factor-α (TNF-α) polypeptide.