Mesothelin antibodies and methods for eliciting potent antitumor activity

Patent metadata

Field	Value
Publication number	US-9416190-B2
Application number	US-201314428771-A
Country	US
Kind code	B2
Filing date	Sep 16, 2013
Priority date	Sep 27, 2012
Publication date	Aug 16, 2016
Grant date	Aug 16, 2016

Abstract

Described herein is the use of phage display antibody engineering technology and synthetic peptide screening to identify SD1 and SD2, human single-domain antibodies to mesothelin. SD1 recognizes a conformational epitope at the C-terminal end (residues 539-588) of human mesothelin close to the cell surface. SD2 binds full-length mesothelin. To investigate SD1 as a potential therapeutic agent, a recombinant human Fc (SD1-hFc) fusion protein was generated. The SD1-hFc protein exhibits strong complement-dependent cytotoxicity (CDC), in addition to antibody-dependent cellular cytotoxicity (ADCC), against mesothelin-expressing tumor cells. Furthermore, the SD1-hFc protein causes significant tumor growth inhibition of tumor xenografts in nude mice. SD1 and SD2 are the first human single-domain antibodies targeting mesothelin-expressing tumors.

First claim

The invention claimed is:

1. An isolated human variable heavy (VH) single domain monoclonal antibody that specifically binds human mesothelin, wherein the antibody comprises a complementarity determining region (CDR) 1, a CDR2 and a CDR3 respectively set forth as:
(i) amino acid residues 26-35, 51-58 and 97-103 of SEQ ID NO: 2;
(ii) amino acid residues 31-35, 51-66 and 99-102 of SEQ ID NO: 2;
(iii) amino acid residues 26-33, 51-57 and 96-111 of SEQ ID NO: 15; or
(iv) amino acid residues 31-35, 50-65 and 99-106 of SEQ ID NO: 15.

2. The isolated human VH single domain monoclonal antibody of claim 1 , wherein the amino acid sequence of the antibody is at least 90% or at least 95% identical to SEQ ID NO: 2 or SEQ ID NO: 15.

3. The isolated human VH single domain monoclonal antibody of claim 1 , wherein the amino acid sequence of the antibody comprises SEQ ID NO: 2 or SEQ ID NO: 15.

4. The isolated human VH single domain monoclonal antibody of claim 1 , wherein the antibody is chimeric or synthetic.

5. The isolated human VH single domain monoclonal antibody of claim 1 , wherein the antibody is labeled.

6. The isolated human VH single domain monoclonal antibody of claim 5 , wherein the label is a fluorescent, enzymatic, or radioactive label.

7. An isolated immunoconjugate comprising the human VH single domain monoclonal antibody of claim 1 and an effector molecule.

8. The isolated immunoconjugate of claim 7 , wherein the effector molecule is a toxin.

9. The isolated immunoconjugate of claim 8 , wherein the toxin is Pseudomonas exotoxin or a variant thereof.

10. The isolated immunoconjugate of claim 9 , wherein the Pseudomonas exotoxin or a variant thereof comprises the amino acid sequence of any one of SEQ ID NOs: 3-8.

11. A fusion protein comprising the human VH single domain monoclonal antibody of claim 1 and a heterologous protein.

12. The fusion protein of claim 11 , wherein the heterologous protein is a human Fc protein.

13. A composition comprising a therapeutically effective amount of the human VH single domain monoclonal antibody of claim 1 in a pharmaceutically acceptable carrier.

14. A method of treating a mesothelin-expressing cancer in a subject, comprising selecting a subject having a mesothelin-expressing cancer and administering to the subject a therapeutically effective amount of the human VH single domain monoclonal antibody of claim 1 , wherein the antibody comprises a CDR1, a CDR2 and a CDR3 respectively set forth as:
(i) amino acid residues 26-35, 51-58 and 97-103 of SEQ ID NO: 2; or
(ii) amino acid resides 31-35, 51-66 and 99-102 or SEQ ID NO: 2,
wherein the antibody is fused to a human Fc protein or to a toxin, thereby treating the mesothelin-expressing cancer.

15. A method of inhibiting tumor growth or metastasis of a mesothelin-expressing tumor in a subject, comprising selecting a subject having a mesothelin-expressing tumor and administering to the subject a therapeutically effective amount of the human VH single domain monoclonal antibody of claim 1 , wherein the antibody comprises a CDR1, a CDR2 and a CDR3 respectively set forth as:
(i) amino acid resides 26-35, 51-58 and 97-103 of SEQ ID NO: 2; or
(ii) amino acid residues 31-35, 51-66 and 99-102 of SEQ ID NO: 2,
wherein the antibody is fused to a human Fc protein or to a toxin, thereby inhibiting tumor growth or metastasis.

16. A method of detecting mesothelin in a biological sample, comprising:
contacting the sample with the human VH single domain monoclonal antibody of claim 1 ; and
detecting binding of the antibody to the sample,
wherein an increase in binding of the antibody to the sample as compared to binding of the antibody to a control sample detects mesothelin in the biological sample.

17. The method of claim 14 , wherein the cancer is mesothelioma, prostate cancer, lung cancer, stomach cancer, squamous cell carcinoma, pancreatic cancer, cholangiocarcinoma, triple negative breast cancer or ovarian cancer.

18. An isolated nucleic acid molecule encoding the human VH single domain monoclonal antibody of claim 1 .

19. The isolated nucleic acid molecule of claim 18 , wherein the nucleotide sequence encoding the antibody comprises SEQ ID NO: 1 or SEQ ID NO: 14.

20. The isolated nucleic acid molecule of claim 18 , operably linked to a promoter.

21. An expression vector comprising the isolated nucleic acid molecule of claim 20 .

22. An isolated host cell transformed with the expression vector of claim 21 .

23. The fusion protein of claim 12 , wherein the human Fc protein comprises human IgGγ1 Fc.

24. A chimeric antigen receptor (CAR) comprising the human VH single domain monoclonal antibody of claim 1 .

25. A bispecific antibody comprising the human VH single domain monoclonal antibody of claim 1 .

26. An isolated immunoconjugate comprising the human VH single domain monoclonal antibody of claim 1 and a therapeutic agent.

27. The isolated immunoconjugate of claim 26 , wherein the therapeutic agent comprises a drug.

28. The method of claim 15 , wherein the cancer is mesothelioma, prostate cancer, lung cancer, stomach cancer, squamous cell carcinoma, pancreatic cancer, cholangiocarcinoma, triple negative breast cancer or ovarian cancer.

Assignees

• Us Health

Inventors

• Ho Mitchell
• Pastan Ira H
• Dimitrov Dimiter S
• Tang Zhewei
• Feng Mingqian
• Gao Wei

Classifications

• A61K2039/505

  comprising antibodies · CPC title

• C07K2317/73

  Inducing cell death, e.g. apoptosis, necrosis or inhibition of cell proliferation · CPC title

• C07K2317/734

  Complement-dependent cytotoxicity [CDC] · CPC title

• C07K2317/52

  Constant or Fc region; Isotype · CPC title

• C07K2317/30

  characterized by aspects of specificity or valency · CPC title