Monoclonal antibodies against claudin-18 for treatment of cancer
The present invention provides antibodies useful as therapeutics for treating and/or preventing diseases associated with cells expressing CLD18, including tumor-related diseases such as gastric cancer, esophageal cancer, pancreatic cancer, lung cancer, ovarian cancer, colon cancer, hepatic cancer, head-neck cancer, and cancer of the gallbladder.
1. A method of producing an antibody, or an antigen binding fragment thereof, that binds to CLD18A2 but not to CLD18A1, the method comprising the steps of:
(a) culturing a human host cell transformed with one or more expression vectors under conditions in which the host cell expresses the antibody or antigen binding fragment thereof; and
(b) harvesting a preparation of the antibody or antigen binding fragment thereof expressed by the human host cell;
wherein the one or more expression vectors comprise:
(i) a nucleic acid sequence encoding a polypeptide comprising the antibody heavy chain CDR1, CDR2, and CDR3 regions having the amino acid sequences of positions 45-52, positions 70-77, and positions 116-124 of SEQ ID NO: 117, respectively; and a nucleic acid sequence encoding a polypeptide comprising the antibody light chain CDR1, CDR2, and CDR3 regions having the amino acid sequences of positions 47-52, positions 70-72, and positions 109-117 of SEQ ID NO: 123, respectively; or
(ii) a nucleic acid sequence encoding a polypeptide comprising the antibody heavy chain CDR1, CDR2, and CDR3 regions having the amino acid sequences of positions 45-52, positions 70-77, and positions 116-126 of SEQ ID NO: 116, respectively; and a nucleic acid sequence encoding a polypeptide comprising the antibody light chain CDR1, CDR2, and CDR3 regions having the amino acid sequences of positions 47-58, positions 76-78, and positions 115-123 of SEQ ID NO: 121, respectively; or
(iii) a nucleic acid sequence encoding a polypeptide comprising the antibody heavy chain CDR1, CDR2, and CDR3 regions having the amino acid sequences of positions 45-52, positions 70-77, and positions 116-126 of SEQ ID NO: 118, respectively; and a nucleic acid sequence encoding a polypeptide comprising the antibody light chain CDR1, CDR2, and CDR3 regions having the amino acid sequences of positions 47-58, positions 76-78, and positions 115-123 of SEQ ID NO: 125, respectively.
2. The method of claim 1 , wherein the human host cell is a lymphocytic cell.
3. The method of claim 1 , wherein the human host cell is a B cell.
4. The method of claim 1 , wherein the expression vector comprises a promoter sequence, a leader sequence, a translation initiation sequence, a light chain constant region, a heavy chain constant region, 3′ untranslated sequence, a polyadenylation sequence, or a transcription termination sequence.
5. The method of claim 1 , wherein the antigen binding fragment is a Fab, F(ab) 2 , Fv, or single chain Fv.
6. A recombinant nucleic acid comprising a combination of a first and a second nucleic acid sequence, wherein the combination is selected from the group consisting of:
(i) a first nucleic acid sequence encoding a polypeptide comprising heavy chain CDR1, CDR2, and CDR3 regions having the amino acid sequences of positions 45-52, positions 70-77, and positions 116-124 of SEQ ID NO: 117, respectively, and a second nucleic acid sequence encoding a polypeptide comprising light chain CDR1, CDR2, and CDR3 regions having the amino acid sequences of positions 47-52, positions 70-72, and positions 109-117 of SEQ ID NO: 123, respectively;
(ii) a first nucleic acid sequence encoding a polypeptide comprising heavy chain CDR1, CDR2, and CDR3 regions having the amino acid sequences of positions 45-52, positions 70-77, and positions 116-126 of SEQ ID NO: 116, respectively, and a second nucleic acid sequence encoding a polypeptide comprising light chain CDR1, CDR2, and CDR3 regions having the amino acid sequences of positions 47-58, positions 76-78, and positions 115-123 of SEQ ID NO: 121, respectively;
(iii) a first nucleic acid sequence encoding a polypeptide comprising heavy chain CDR1, CDR2, and CDR3 regions having the amino acid sequences of positions 45-52, positions 70-77, and positions 116-126 of SEQ ID NO: 118, respectively, and a second nucleic acid sequence encoding a polypeptide comprising light chain CDR1, CDR2, and CDR3 regions having the amino acid sequences of positions 47-58, positions 76-78, and positions 115-123 of SEQ ID NO: 125, respectively.
7. The recombinant nucleic acid of claim 6 , wherein the first nucleic acid sequence encodes a heavy chain variable region comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 133, 134, and 135.
8. The recombinant nucleic acid of claim 6 , wherein the second nucleic acid sequence encodes a light chain variable region comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 138, 140, and 142.
9. The recombinant nucleic acid of claim 6 , wherein the first or second nucleic acid sequence is operatively linked to expression control sequences.
10. A transformed human cell comprising the recombinant nucleic acid of claim 6 .
11. The transformed cell of claim 10 , wherein the cell is a lymphocytic cell.
12. A polypeptide encoded by the recombinant nucleic acid of claim 6 .
13. The polypeptide of claim 12 , further comprising an immunoglobulin hinge region.
14. A recombinant nucleic acid comprising a nucleic acid sequence selected from the group consisting of:
(i) a nucleic acid sequence encoding a polypeptide comprising a heavy chain variable region having the amino acid sequence of SEQ ID NO: 134 and a light chain variable region having the amino acid sequence of SEQ ID NO: 140;
(ii) a nucleic acid sequence encoding a polypeptide comprising a heavy chain variable region having the amino acid sequence of SEQ ID NO: 133 and a light chain variable region having the amino acid sequence of SEQ ID NO: 138; and
(iii) a nucleic acid sequence encoding a polypeptide comprising a heavy chain variable region having the amino acid sequence of SEQ ID NO: 135 and a light chain variable region having the amino acid sequence of SEQ ID NO: 142.
15. The recombinant nucleic acid of claim 14 , wherein the nucleic acid sequence is operatively linked to expression control sequences.
16. A transformed human cell comprising the recombinant nucleic acid of claim 14 .
17. The transformed cell of claim 16 , wherein the cell is a lymphocytic cell.
18. A polypeptide encoded by the recombinant nucleic acid of claim 14 .
19. The polypeptide of claim 18 , further comprising an immunoglobulin hinge region.
20. An anti-CLD18A2 antibody expressed by the transformed human cell of claim 16 , wherein the antibody binds to CLD18A2 but not to CLD18A1.
21. An antigen binding fragment expressed by the transformed human cell of claim 16 , wherein the antigen binding fragment binds to CLD18A2 but not to CLD18A1.
22. The antigen binding fragment of claim 21 , wherein the antigen binding fragment is a Fab, F(ab) 2 , Fv, or single chain Fv.
23. The antigen binding fragment of claim 21 , wherein the antigen binding fragment is fused to an immunoglobulin hinge region.
24. A method of producing a recombinant eukaryotic host cell, comprising the steps of:
a. transforming a eukaryotic cell with an expression vector comprising the recombinant nucleic acid of claim 6 or 14 ; and
b. obtaining the transformed cell, wherein the transformed cell comprises the recombinant nucleic acid.
25. The method of claim 24 , wherein the recombinant eukaryotic host cell is a human cell.
26. The method of claim 24 , wherein the recombinant eukaryotic host cell is a lymphocytic cell.
27. A recombinant human cell comprising the polypeptide of claim 12 .
28. A recombinant human cell comprising the polypeptide of claim 18 .