T cell receptor-like antibodies specific for a WTI peptide presented by HLA-A2
The present invention provides antigen binding proteins that specifically bind to Wilms' tumor protein (WT1), including humanized, chimeric and fully human antibodies against WT1, antibody fragments, chimeric antigen receptors (CARs), fusion proteins, and conjugates thereof. The antigen binding proteins and antibodies bind to HLA-A0201-restricted WT1 peptide. Such antibodies, fragments, fusion proteins and conjugates thereof are useful for the treatment of WT1 associated cancers, including for example, breast cancer, ovarian cancer, prostate cancer, chronic myelocytic leukemia, multiple myeloma, acute lymphoblastic leukemia (ALL), acute myeloid/myelogenous leukemia (AML) and myelodysplastic syndrome (MDS). In more particular embodiments, the anti-WT1/A antibodies may comprise one or more framework region amino acid substitutions designed to improve protein stability, antibody binding and/or expression levels.
1. A chimeric antigen receptor comprising an antigen binding fragment, comprising one of:
(a) a heavy chain (HC) variable region comprising HC-CDR1, HC-CDR2 and HC-CDR3 and HC-CDR3 respectively, comprising amino acid sequences SEQ ID NOS: 2, 3, and 4; and a light chain (LC) variable region comprising LC-CDR1, LC-CDR2 and LC-CDR3 respectively, comprising amino acid sequences SEQ ID NOS: 8, 9 and 10;
(b) a heavy chain (HC) variable region comprising HC-CDR1, HC-CDR2 and HC-CDR3 respectively, comprising amino acid sequences SEQ ID NOS: 20, 21 and 22; and a light chain (LC) variable region comprising LC-CDR1, LC-CDR2 and LC-CDR3 respectively, comprising amino acid sequences SEQ ID NOS: 26, 27 and 28;
(c) a heavy chain (HC) variable region comprising HC-CDR1, HC-CDR2 and HC-CDR3 respectively, comprising amino acid sequences SEQ ID NOS: 38, 39 and 40; and a light chain (LC) variable region comprising LC-CDR1, LC-CDR2 and LC-CDR3 respectively, comprising amino acid sequences selected from SEQ ID NOS: 44, 45 and 46;
(d) a heavy chain (HC) variable region comprising HC-CDR1, HC-CDR2 and HC-CDR3 respectively, comprising amino acid sequences SEQ ID NOS: 56, 57 and 58; and a light chain (LC) variable region comprising LC-CDR1, LC-CDR2 and LC-CDR3 respectively, comprising amino acid sequences SEQ ID NOS: 62, 63 and 64;
(e) a heavy chain (HC) variable region comprising HC-CDR1, HC-CDR2 and HC-CDR3 respectively, comprising amino acid sequences SEQ ID NOS: 74, 75 and 76; and a light chain (LC) variable region comprising LC-CDR1, LC-CDR2 and LC-CDR3 respectively, comprising amino acid sequences SEQ ID NOS: 80, 81 and 82; or
(f) a heavy chain (HC) variable region comprising HC-CDR1, HC-CDR2 and HC-CDR3 respectively, comprising amino acid sequences SEQ ID NOS: 92, 93 and 94; and a light chain (LC) variable region comprising LC-CDR1, LC-CDR2 and LC-CDR3 respectively, comprising amino acid sequences SEQ ID NOS: 98, 99 and 100.
2. The chimeric antigen receptor of claim 1 , comprising an antigen binding fragment, comprising a V H and V L comprising first and second amino acid sequences, respectively, selected from SEQ ID NOS: 14 and 16; 32 and 34; 50 and 52; 68 and 70; 86 and 88; and 104 and 106.
3. The chimeric antigen receptor of claim 1 , comprising an antigen binding fragment, comprising an amino acid sequence selected from SEQ ID NOS: 18, 36, 54, 72, 90, and 108.
4. The chimeric antigen receptor of claim 1 , wherein the chimeric antigen receptor specifically binds to an WT1 peptide bound to HLA-A2.
5. The chimeric antigen receptor of claim 4 , wherein said WT1 peptide has the amino acid sequence RMFPNAPYL (SEQ ID NO: 1).
6. The chimeric antigen receptor of claim 4 , wherein said HLA-A2 is HLA-A0201.
7. A recombinant cell comprising the chimeric antigen receptor of claim 1 .
8. The recombinant cell of claim 7 that is a recombinant immune cell.
9. The recombinant immune cell of claim 8 that is a recombinant T cell.
10. The recombinant immune cell of claim 9 , wherein the cell is cytotoxic to a cell expressing WT1 bound to HLA-A2.
11. The recombinant immune cell of claim 10 , wherein said HLA-A2 is HLA-A0201.
12. The recombinant cell of claim 7 , further comprising nucleic acid encoding the chimeric antigen receptor.
13. The recombinant cell of claim 12 , wherein the nucleic acid comprises:
(i) first, second and third nucleotide sequences selected from the group consisting of SEQ ID NOS: 5, 6 and 7; 23, 24 and 25; 41, 42 and 43; 58, 59 and 60; 77, 78 and 79; and 95, 96 and 97; and
(ii) fourth, fifth and sixth nucleotide sequences selected from the group consisting of SEQ ID NOS: 11, 12 and 13; 29, 30 and 31; 47, 48 and 49; 65, 66 and 67; 83, 84 and 85; and 101, 102 and 103.
14. The recombinant cell of claim 12 , wherein the nucleic acid comprises first and second nucleotide sequences selected from the group consisting of SEQ ID NOS: 15 and 17; 33 and 35; 51 and 53; 69 and 71; 87 and 89 and 105 and 107; or a nucleotide sequence selected from the group consisting of SEQ ID NOS: 19, 37, 55, 73, 91 and 109.
15. A method for treatment of a subject having a WT1-positive disease, comprising administering to the subject a therapeutically effective amount of the recombinant cell of claim 7 .
16. The method of claim 15 , wherein the WT1-positive disease is a chronic leukemia or acute leukemia or WT1 + cancer.
17. The method of claim 15 , wherein the WT1-positive disease is selected from the group consisting of chronic myelocytic leukemia, multiple myeloma (MM), acute lymphoblastic leukemia (ALL), acute myeloid/myelogenous leukemia (AML), myelodysplastic syndrome (MDS), mesothelioma, ovarian cancer, gastrointestinal cancers, breast cancer, prostate cancer and glioblastoma.
18. A pharmaceutical composition comprising the recombinant cell of claim 7 .
19. A nucleic acid encoding the chimeric antigen receptor of claim 1 .
20. The nucleic acid of claim 19 , comprising:
(i) first, second and third nucleotide sequences selected from the group consisting of SEQ ID NOS: 5, 6 and 7; 23, 24 and 25; 41, 42 and 43; 58, 59 and 60; 77, 78 and 79; and 95, 96 and 97; and fourth, fifth and sixth nucleotide sequences selected from the group consisting of SEQ ID NOS: 11, 12 and 13; 29, 30 and 31; 47, 48 and 49; 65, 66 and 67; 83, 84 and 85; and 101, 102 and 103;
(ii) first and second nucleotide sequences selected from the group consisting of SEQ ID NOS: 15 and 17; 33 and 35; 51 and 53; 69 and 71; 87 and 89 and 105 and 107; or
(iii) a nucleotide sequence selected from the group consisting of SEQ ID NOS: 19, 37, 55, 73, 91 and 109.