Site-specific attachment of drugs or other agents to engineered antibodies with C-terminal extensions
View Patent ↗The present invention provides antibodies (e.g., IgG antibodies) having C-terminal cysteine-containing extensions that facilitate antibody conjugation to a partner molecule (e.g. a drug, toxin, marker molecule, protein, radioisotope, or other therapeutic agent). Methods of making, screening and selecting the antibodies of the invention are provided.
1. An antibody-partner molecule conjugate comprising a full-length antibody conjugated to a partner molecule wherein the conjugation occurs via a cysteine-containing extension at the C-terminus of a heavy chain of the antibody, wherein said cysteine-containing extension has an amino acid sequence selected from the group consisting of CAA, CCAA (SEQ ID NO:9), AACAA (SEQ ID NO:7), and GGGGSCAA (SEQ ID NO:8).
2. The antibody-partner molecule conjugate of claim 1 wherein the cysteine-containing extension is introduced by the addition of a cysteine-containing extension to the heavy chain of the antibody.
3. The antibody-partner molecule conjugate of claim 1 wherein the C-terminal cysteine-containing extension is introduced by the replacement of the original C-terminal amino acid residue of the heavy chain of the antibody.
4. The antibody-partner molecule conjugate of claim 1 , wherein the partner molecule is a cytotoxic drug.
5. The antibody-partner molecule conjugate of claim 4 , wherein the drug is selected from the group consisting of an auristatin, a DNA minor groove binding agent, a DNA minor groove alkylating agent, an enediyne, a lexitropsin, a duocarmycin, a taxane, a puromycin, a dolastatin, a maytansinoid, and a vinca alkaloid.
6. The antibody-partner molecule conjugate of claim 1 , further comprising a cleavable linker.
7. The antibody-partner molecule conjugate of claim 6 , wherein the linker is a peptide linker cleavable by an intracellular protease.
8. The antibody-partner molecule conjugate of claim 7 , wherein the peptide linker is a val-cit linker or a phe-lys linker.
9. An antibody-drug conjugate comprising:
a full length antibody that:
(a) binds to PSMA, and
(b) is conjugated to a cytotoxic agent or an immunosuppressive agent,
wherein the antibody-drug conjugate exerts:
(a) a cytotoxic or cytostatic effect on a PSMA-expressing cancer cell line, or
(b) a cytotoxic, cytostatic, or immunosuppressive effect on a PSMA-expressing immune cell,
wherein the conjugation occurs at an introduced cysteine residue at or near the C-terminus of a heavy chain of the antibody, wherein said introduced cysteine residue is in a C-terminal cysteine-containing extension having an amino acid sequence selected from CAA, CCAA (SEQ ID NO:9), AACAA (SEQ ID NO:7), and GGGGSCAA (SEQ ID NO:8).
10. A method of making an antibody-partner molecular conjugate, comprising the steps of:
(a) providing a full-length antibody;
(b) modifying the C-terminus of at least one of the heavy chains of the full-length antibody by adding thereto a cysteine-containing extension having an amino acid sequence selected from the group consisting of CAA, CCAA (SEQ ID NO:9), AACAA (SEQ ID NO:7), and GGGGSCAA (SEQ ID NO:8); and
(c) conjugating the modified full-length antibody to a partner molecule via the cysteine residue of the cysteine-containing extension.
11. A full length antibody, wherein the C-terminus of at least one of its heavy chains has been modified by adding thereto a cysteine-containing extension having an amino acid sequence selected from the group consisting of CAA, CCAA (SEQ ID NO:9), AACAA (SEQ ID NO:7), and GGGGSCAA (SEQ ID NO:8).