Bispecific antigen-binding molecules and methods of use
The present invention provides bispecific antigen-binding molecules having a monovalent arm specific to a first target antigen (e.g., a T cell antigen, such as CD3) and a bivalent arm specific for a second target antigen (e.g., a tumor antigen, such as HER2). Bispecific antigen-binding molecules are useful in the treatment of disorders, such as cancer (e.g., HER2-positive cancer). The invention also features methods of producing bispecific antigen-binding molecules, methods of treating disorders using bispecific antigen-binding molecules, and compositions including bispecific antigen-binding molecules.
1. A bispecific antigen-binding molecule that specifically binds CD3 and HER2 comprising a monovalent arm and a bivalent arm, wherein:
(a) the monovalent arm comprises a Fab A that specifically binds CD3, wherein the C-terminus of the Fab A is fused to an N-terminus of a first Fc subunit, and wherein the Fab A comprises the following HVRs:
(i) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 1,
(ii) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 2,
(iii) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 3,
(iv) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 4,
(v) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 5, and
(vi) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 6;
(b) the bivalent arm comprises a Fab B1 and a Fab B2 that each specifically binds HER2, wherein the C-terminus of the Fab B2 is fused to the N-terminus of the Fab B1 , and the C-terminus of the Fab B1 is fused to an N-terminus of a second Fc subunit, wherein the Fab B1 and the Fab B2 each comprise the following HVRs:
(i) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 11,
(ii) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 36,
(iii) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 37,
(iv) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 38,
(v) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 29, and
(vi) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 26; and
(c) the first Fc subunit is associated with the second Fc subunit to form an Fc domain.
2. The bispecific antigen-binding molecule of claim 1 , wherein:
(a) the Fab A comprises a VH A region and a VL A region, wherein the VH A region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 7, and the VL A region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 8;
(b) the Fab B1 comprises a VH B1 region and a VL B1 region, wherein the VH B1 region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 41, and the VL B1 region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 49; and
(c) the Fab B2 comprises a VH B2 region and a VL B2 region, wherein the VH B2 region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 41, and the VL B2 region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 49.
3. The bispecific antigen-binding molecule of claim 2 , wherein:
(a) the VH A region comprises the amino acid sequence of SEQ ID NO: 7, and the VL A region comprises the amino acid sequence of SEQ ID NO: 8;
(b) the VH B1 region comprises the amino acid sequence of SEQ ID NO: 41, and the VL B1 region comprises the amino acid sequence of SEQ ID NO: 49; and
(c) the VH B2 region comprises the amino acid sequence of SEQ ID NO: 41, and the VL B2 region comprises the amino acid sequence of SEQ ID NO: 49.
4. A bispecific antigen-binding molecule that specifically binds CD3 and HER2 comprising a monovalent arm and a bivalent arm, wherein:
(a) the monovalent arm comprises a Fab A that specifically binds CD3, wherein the C-terminus of the Fab A is fused to an N-terminus of a first Fc subunit, and wherein the Fab A comprises the following HVRs:
(i) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 1,
(ii) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 2,
(iii) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 3,
(iv) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 4,
(v) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 5, and
(vi) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 6;
(b) the bivalent arm comprises a Fab B1 and a Fab B2 that each specifically binds HER2, wherein the C-terminus of the Fab B2 is fused to the N-terminus of the Fab B1 , and the C-terminus of the Fab B1 is fused to an N-terminus of a second Fc subunit, wherein the Fab B1 and the Fab B2 each comprise the following HVRs:
(i) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 11,
(ii) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 19,
(iii) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 20,
(iv) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 21,
(v) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 22, and
(vi) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 26; and
(c) the first Fc subunit is associated with the second Fc subunit to form an Fc domain.
5. The bispecific antigen-binding molecule of claim 4 , wherein:
(a) the Fab A comprises a VH A region and a VL A region, wherein the VH A region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 7, and the VL A region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 8;
(b) the Fab B1 comprises a VH B1 region and a VL B1 region, wherein the VH B1 region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 24, and the VL B1 region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 27; and
(c) the Fab B2 comprises a VH B2 region and a VL B2 region, wherein the VH B2 region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 24, and the VL B2 region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 27.
6. The bispecific antigen-binding molecule of claim 5 , wherein:
(a) the VH A region comprises the amino acid sequence of SEQ ID NO: 7, and the VL A region comprises the amino acid sequence of SEQ ID NO: 8;
(b) the VH B1 region comprises the amino acid sequence of SEQ ID NO: 24, and the VL B1 region comprises the amino acid sequence of SEQ ID NO: 27; and
(c) the VH B2 region comprises the amino acid sequence of SEQ ID NO: 24, and the VL B2 region comprises the amino acid sequence of SEQ ID NO: 27.
7. A bispecific antigen-binding molecule that specifically binds CD3 and HER2 comprising a monovalent arm and a bivalent arm, wherein:
(a) the monovalent arm comprises a Fab A that specifically binds CD3, wherein the C-terminus of the Fab A is fused to an N-terminus of a first Fc subunit, and wherein the Fab A comprises the following HVRs:
(i) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 1,
(ii) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 2,
(iii) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 3,
(iv) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 4,
(v) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 5, and
(vi) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 6;
(b) the bivalent arm comprises a Fab B1 and a Fab B2 that each specifically binds HER2, wherein the C-terminus of the Fab B2 is fused to the N-terminus of the Fab B1 , and the C-terminus of the Fab B1 is fused to an N-terminus of a second Fc subunit, wherein the Fab B1 and the Fab B2 each comprise the following HVRs:
(i) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 11,
(ii) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 19,
(iii) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 32,
(iv) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 21,
(v) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 22, and
(vi) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 23; and
(c) the first Fc subunit is associated with the second Fc subunit to form an Fc domain.
8. The bispecific antigen-binding molecule of claim 7 , wherein:
(a) the Fab A comprises a VH A region and a VL A region, wherein the VH A region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 7, and the VL A region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 8;
(b) the Fab B1 comprises a VH B1 region and a VL B1 region, wherein the VH B1 region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 33, and the VL B1 region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 25; and
(c) the Fab B2 comprises a VH B2 region and a VL B2 region, wherein the VH B2 region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 33, and the VL B2 region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 25.
9. The bispecific antigen-binding molecule of claim 8 , wherein:
(a) the VH A region comprises the amino acid sequence of SEQ ID NO: 7, and the VL A region comprises the amino acid sequence of SEQ ID NO: 8;
(b) the VH B1 region comprises the amino acid sequence of SEQ ID NO: 33, and the VL B1 region comprises the amino acid sequence of SEQ ID NO: 25; and
(c) the VH B2 region comprises the amino acid sequence of SEQ ID NO: 33, and the VL B2 region comprises the amino acid sequence of SEQ ID NO: 25.
10. A bispecific antigen-binding molecule that specifically binds CD3 and HER2 comprising a monovalent arm and a bivalent arm, wherein:
(a) the monovalent arm comprises a Fab A that specifically binds CD3, wherein the C-terminus of the Fab A is fused to an N-terminus of a first Fc subunit, and wherein the Fab A comprises the following HVRs:
(i) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 1,
(ii) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 2,
(iii) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 3,
(iv) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 4,
(v) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 5, and
(vi) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 6;
(b) the bivalent arm comprises a Fab B1 and a Fab B2 that each specifically binds HER2, wherein the C-terminus of the Fab B2 is fused to the N-terminus of the Fab B1 , and the C-terminus of the Fab B1 is fused to an N-terminus of a second Fc subunit, wherein the Fab B1 and the Fab B2 each comprise the following HVRs:
(i) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 11,
(ii) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 36,
(iii) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 43,
(iv) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 21,
(v) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 29, and
(vi) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 26; and
(c) the first Fc subunit is associated with the second Fc subunit to form an Fc domain.
11. The bispecific antigen-binding molecule of claim 10 , wherein:
(a) the Fab A comprises a VH A region and a VL A region, wherein the VH A region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 7, and the VL A region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 8;
(b) the Fab B1 comprises a VH B1 region and a VL B1 region, wherein the VH B1 region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 44, and the VL B1 region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 48; and
(c) the Fab B2 comprises a VH B2 region and a VL B2 region, wherein the VH B2 region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 44, and the VL B2 region comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 48.
12. The bispecific antigen-binding molecule of claim 11 , wherein:
(a) the VH A region comprises the amino acid sequence of SEQ ID NO: 7, and the VL A region comprises the amino acid sequence of SEQ ID NO: 8;
(b) the VH B1 region comprises the amino acid sequence of SEQ ID NO: 44, and the VL B1 region comprises the amino acid sequence of SEQ ID NO: 48; and
(c) the VH B2 region comprises the amino acid sequence of SEQ ID NO: 44, and the VL B2 region comprises the amino acid sequence of SEQ ID NO: 48.
13. An immunoconjugate comprising the bispecific antigen-binding molecule of claim 1 and a cytotoxic agent.
14. An immunoconjugate comprising the bispecific antigen-binding molecule of claim 4 and a cytotoxic agent.
15. An immunoconjugate comprising the bispecific antigen-binding molecule of claim 7 and a cytotoxic agent.
16. An immunoconjugate comprising the bispecific antigen-binding molecule of claim 10 and a cytotoxic agent.
17. A composition comprising the bispecific antigen-binding molecule of claim 1 and a pharmaceutically acceptable carrier, excipient, or diluent.
18. A composition comprising the bispecific antigen-binding molecule of claim 4 and a pharmaceutically acceptable carrier, excipient, or diluent.
19. A composition comprising the bispecific antigen-binding molecule of claim 7 and a pharmaceutically acceptable carrier, excipient, or diluent.
20. A composition comprising the bispecific antigen-binding molecule of claim 10 and a pharmaceutically acceptable carrier, excipient, or diluent.
21. A kit comprising:
(a) the composition of claim 17 ; and
(b) a package insert comprising instructions for administering the composition to a subject to treat or delay progression of a HER2-positive cancer.
22. A kit comprising:
(a) the composition of claim 18 ; and
(b) a package insert comprising instructions for administering the composition to a subject to treat or delay progression of a HER2-positive cancer.
23. A kit comprising:
(a) the composition of claim 19 ; and
(b) a package insert comprising instructions for administering the composition to a subject to treat or delay progression of a HER2-positive cancer.
24. A kit comprising:
(a) the composition of claim 20 ; and
(b) a package insert comprising instructions for administering the composition to a subject to treat or delay progression of a HER2-positive cancer.