Antigen binding proteins binding to 5T4 and 4-1BB and related compositions and methods
The present disclosure relates to protein molecules that specifically bind to 5T4 and/or 4-1BB. The molecules may have at least one humanized 5T4-binding or 4-1BB-binding domain. Such molecules are useful for the treatment of cancer. The protein molecule binding to 5T4 or 4-1BB may have a second binding domain that binds to another target. The molecules may bind both 5T4-expressing cells and a cell-surface molecule expressed by an effector cell to enhance effector cell activation, proliferation, survival and/or effector-cell mediated cytotoxicity. The disclosure also provides pharmaceutical compositions comprising the 5T4-binding or 4-1BB-binding polypeptide or protein molecules, nucleic acid molecules encoding these polypeptides and methods of making and using these molecules.
1. A multispecific polypeptide comprising a first scFv domain and a second scFv domain, wherein the first and second scFv domains are linked together by a binding domain linker or a binding domain linker and an immunoglobulin Fc domain, wherein the immunoglobulin Fc domain comprises a hinge region and an immunoglobulin constant region; and
wherein the first scFv domain specifically binds to human 5T4 and comprises:
(i) an immunoglobulin heavy chain variable region comprising an HCDR1 amino acid sequence of SEQ ID NO: 30, an HCDR2 amino acid sequence of SEQ ID NO: 32, and an HCDR3 amino acid sequence of SEQ ID NO: 34; and
(ii) an immunoglobulin light chain variable region comprising an LCDR1 amino acid sequence of SEQ ID NO: 42, an LCDR2 amino acid sequence of SEQ ID NO: 10, and an LCDR3 amino acid sequence of SEQ ID NO: 36.
2. The multispecific polypeptide of claim 1 , wherein the 5T4-binding domain comprises a heavy chain variable region comprising an amino acid sequence having at least 95% identity to a sequence selected from the group consisting of SEQ ID NOs: 38 and 46.
3. The multispecific polypeptide of claim 2 , wherein the 5T4-binding domain comprises a heavy chain variable region comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 38 and 46.
4. The multispecific polypeptide of claim 1 , wherein the 5T4-binding domain comprises a light chain variable region comprising an amino acid sequence having at least 95% identity to a sequence selected from the group consisting of SEQ ID NOs: 44, 48 and 50.
5. The multispecific polypeptide of claim 4 , wherein the 5T4-binding domain comprises a light chain variable region comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 44, 48 and 50.
6. The multispecific polypeptide of claim 1 , wherein the 5T4-binding domain comprises:
i) a heavy chain variable region comprising an amino acid sequence having at least 95% identity to SEQ ID NO: 38 and a light chain variable region comprising an amino acid sequence having at least 95% identity to SEQ ID NO: 44; or
ii) a heavy chain variable region comprising an amino acid sequence having at least 95% identity to SEQ ID NO: 46 and a light chain variable region comprising an amino acid sequence having at least 95% identity to SEQ ID NO: 48; or
iii) a heavy chain variable region comprising an amino acid sequence having at least 95% identity to SEQ ID NO: 46 and a light chain variable region comprising an amino acid sequence having at least 95% identity to SEQ ID NO: 50.
7. The multispecific polypeptide of claim 6 , wherein the 5T4-binding domain comprises:
i) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 38 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 44; or
ii) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 46 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 48; or
iii) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 46 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 50.
8. The multispecific polypeptide of claim 1 , wherein the first scFv comprises a sequence having at least 95% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 120, 122 and 124.
9. The multi specific polypeptide of claim 1 , wherein the first scFv domain comprises a mutation in the framework region compared to the framework region of SEQ ID NOs: 130 or 170.
10. The multispecific polypeptide of claim 9 , wherein the mutation introduces a stabilizing disulfide bond.
11. The multispecific polypeptide of claim 1 , wherein the second scFv domain comprises 4-1BB-binding domain and comprises: (i) an immunoglobulin heavy chain variable region (VH) comprising HCDR1, HCDR2, and HCDR3, and (ii) an immunoglobulin light chain variable region (VL) comprising LCDR1, LCDR2, and LCDR3, wherein:
(a) the HCDR1 comprises the amino acid sequence of SEQ ID NO: 2;
(b) the HCDR2 comprises the amino acid sequence of SEQ ID NO: 4;
(c) the HCDR3 comprises the amino acid sequence of SEQ ID NO: 6;
(d) the LCDR1 comprises the amino acid sequence of SEQ ID NO: 8;
(e) the LCDR2 comprises the amino acid sequence of SEQ ID NO: 10; and
(f) the LCDR3 comprises the amino acid sequence of SEQ ID NO: 12.
12. The multispecific polypeptide of claim 11 , wherein the 4-1BB-binding domain comprises a heavy chain variable region comprising an amino acid sequence having at least 95% identity to the sequence of SEQ ID NO: 14.
13. The multispecific polypeptide of claim 12 , wherein the 4-1BB-binding domain comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 14.
14. The multispecific polypeptide of claim 11 , wherein the 4-1BB-binding domain comprises a light chain variable region comprising an amino acid sequence having at least 95% identity to the amino acid sequence of SEQ ID NO: 16.
15. The multispecific polypeptide of claim 12 , wherein the 4-1BB-binding domain comprises a light chain variable region comprising the amino acid sequence of SEQ ID NO: 16.
16. The multispecific polypeptide of claim 11 , wherein the 4-1BB-binding domain comprises:
a heavy chain variable region comprising an amino acid sequence having at least 95% identity to SEQ ID NO: 14 and a light chain variable region comprising an amino acid sequence having at least 95% identity to SEQ ID NO: 16.
17. The multispecific polypeptide of claim 16 , wherein the 4-1BB-binding domain comprises:
a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 14 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 16.
18. The multispecific polypeptide of claim 11 , wherein the second scFv comprises a sequence having at least 95% identity to the amino acid sequence of SEQ ID NO: 110.
19. The multispecific polypeptide of claim 1 , wherein the Fc domain comprises the sequence set forth in SEQ ID NO: 158 or SEQ ID NO: 160.
20. A method for treating cancer in a subject in need thereof, wherein the cancer is characterized by expression of 5T4, the method comprising administering to the subject a therapeutically effective amount of the polypeptide of claim 1 .
21. The method of claim 20 , wherein the cancer is breast cancer, pancreatic cancer, ovarian cancer, non-small cell lung cancer, mesothelioma, chronic lymphocytic leukemia (CLL), mantle cell leukemia (MCL), acute lymphoblastic leukemia (ALL), squamous cell carcinoma, melanoma, adrenal cancer, bladder cancer, cervical cancer, renal cancer, gastric cancer, prostate cancer, thyroid cancer, liver cancer, uterine cancer, neurofibroma, sarcoma, carcinoma, or head and neck cancer.
22. An isolated nucleic acid molecule encoding the polypeptide of claim 1 .
23. An expression vector comprising an isolated nucleic acid molecule of claim 22 .
24. A recombinant host cell comprising the expression vector of claim 23 .