Anti-5t4 antibodies and uses thereof

Abstract

Anti-5T4 antibodies, anti-5T4 antibody/drug conjugates, and methods for preparing and using the same.

What is claimed: 
     
         1 . An antibody that specifically binds human 5T4 antigen, wherein the antibody:
 (a) comprises an antigen binding domain of murine A1, A2, or A3 antibodies;   (b) competes for 5T4 binding with murine A1, A2, or A3 antibodies;   (c) binds a 5T4 epitope bound by A1, A2, or A3 antibodies; or   (d) comprises a 5T4-binding fragment of an antibody of (a)-(c).   
     
     
         2 . The antibody of  claim 1 , wherein the antibody is a chimeric antibody, a humanized antibody, a single chain antibody, a Fab fragment, a F(ab) 2  fragment, a Fv fragment, a tetrameric antibody, a tetravalent antibody, a multispecific antibody, a domain-specific antibody, a single domain antibody, or a fusion protein. 
     
     
         3 . The antibody of  claim 1 , which is a murine monoclonal antibody. 
     
     
         4 . The antibody of  claim 1 , wherein the antibody has a binding affinity for human 5T4 antigen of at least about 1×10 −7  M to about 1×10 −12  M. 
     
     
         5 . The antibody of  claim 1 , wherein the antibody specifically targets 5T4-expressing cells in vivo. 
     
     
         6 . The antibody of  claim 1 , wherein the heavy chain variable region comprises an amino acid sequence of residues 20-138 of SEQ ID NO:2, residues 19-135 of SEQ ID NO:6, residues 20-141 of SEQ ID NO:10, residues 1-119 of SEQ ID NO:49; or residues of a humanized A1, A2, or A3 heavy chain variable region. 
     
     
         7 . The antibody of  claim 1 , wherein the light chain variable region comprises an amino acid sequence of residues 21-127 of SEQ ID NO:4, residues 23-130 of SEQ ID NO:8, residues 21-127 of SEQ ID NO:12; or residues of a humanized A1, A2, or A3 light chain variable region. 
     
     
         8 . The antibody of  claim 1 , wherein the heavy chain variable region comprises an amino acid sequence of residues 20-138 of SEQ ID NO:2, and wherein the light chain variable region comprises an amino acid sequence of residues 21-127 of SEQ ID NO:4. 
     
     
         9 . The antibody of  claim 1 , wherein the heavy chain variable region comprises an amino acid sequence derived from residues 19-135 of SEQ ID NO:6, and wherein the light chain variable region comprises an amino acid sequence derived from residues 23-130 of SEQ ID NO:8. 
     
     
         10 . The antibody of  claim 1 , wherein the heavy chain variable region comprises an amino acid sequence derived from residues 20-141 of SEQ ID NO:10, and wherein the light chain variable region comprises an amino acid sequence derived from residues 21-127 of SEQ ID NO:12. 
     
     
         11 . The antibody of  claim 2 , which is a chimeric or humanized anti-5T4 antibody. 
     
     
         12 . The chimeric or humanized antibody of  claim 11 , which comprises constant regions derived from human constant regions. 
     
     
         13 . The chimeric or humanized antibody of  claim 12 , wherein the human light chain constant region is derived from human kappa light chain constant region. 
     
     
         14 . The chimeric or humanized antibody or antibody fragment of  claim 12 , wherein the human heavy chain constant region is derived from a human IgG1, IgG2, IgG3, or IgG4 heavy chain constant region. 
     
     
         15 . The chimeric or humanized antibody or antibody fragment of  claim 14 , wherein the human IgG4 heavy chain constant region comprises proline at position 241. 
     
     
         16 . The chimeric or humanized antibody of  claim 11 , wherein the variable region of the at least one light chain or at least one heavy chain comprises:
 (a) framework regions comprising residues of a human antibody framework region; and   (b) one or more CDRs of the light chain variable region of SEQ ID NO:4, 8, or 12, or one or more CDRs of the heavy chain variable region of SEQ ID NO:2, 6, or 10.   
     
     
         17 . The chimeric or humanized antibody of  claim 16 , wherein the human residues are human framework residues selected from the group consisting of:
 (a) a human antibody heavy chain framework region selected from the group consisting of DP-21 (VH7), DP-54 (VH3-07), DP-47 (VH3-23), DP-53 (VH-74), DP-49 (VH3-30), DP-48 (VH3-13), DP-75, DP-8(VH1-2), DP-25, VI-2 b and VI-3 (VH1-03), DP-15 and V1-8 (VH1-08), DP-14 and V1-18 (VH1-18), DP-5 and V1-24P (VH1-24), DP-4 (VH1-45), DP-7 (VH1-46), DP-10, DA-6 and YAC-7 (VH1-69), DP-88 (VH1-e), DP-3 and DA-8 (VH1-f);   (b) a human antibody light chain framework region of a DPK24 subgroup IV germ line clone, a VκIII subgroup (DPK23, DPK22, DPK20, DPK 21), or a VκI subgroup germ line clone (DPK9, DPK1, O2, DPK-7);   (c) a consensus sequence of a heavy chain framework region of (a); or   (d) a framework region that is at least 63% identical to a framework region of (a)-(c).   
     
     
         18 . The chimeric or humanized antibody of  claim 16  comprising at least two CDRs of any one of SEQ ID NOs:2, 4, 6, 8, 10, or 12. 
     
     
         19 . The humanized antibody of  claim 18 , wherein the light chain comprises a variable region comprising at least two of three CDRs of any one of SEQ ID NOs:4, 8, or 12. 
     
     
         20 . The humanized antibody of  claim 19 , wherein the light chain comprises a variable region comprising three CDRs of any one of SEQ ID NOs:4, 8, or 12. 
     
     
         21 . The humanized antibody of  claim 16 , wherein the heavy chain comprises a variable region comprising at least two of three CDRs of any one of SEQ ID NOs:2, 6, or 10. 
     
     
         22 . The humanized antibody of  claim 21 , wherein the heavy chain comprises a variable region comprising three CDRs of any one of SEQ ID NOs:2, 6, or 10. 
     
     
         23 . The humanized antibody of  claim 16 , which comprises the CDRs of SEQ ID NOs:2 and 4, the CDRs of SEQ ID NOs:6 and 8, or the CDRs of SEQ ID NOs:10 and 12. 
     
     
         24 . The chimeric or humanized antibody of  claim 11 , wherein the heavy chain variable region sequence comprises:
 (a) an amino acid sequence of residues 20-138 of SEQ ID NO:2;   (b) an amino acid sequence that is at least 85% identical to residues 20-138 of SEQ ID NO:2;   (c) an amino acid sequence of residues 19-135 of SEQ ID NO:6;   (d) an amino acid sequence that is at least 86% identical to residues 19-135 of SEQ ID NO:6;   (e) an amino acid sequence of residues 20-141 of SEQ ID NO:10;   (f) an amino acid sequence that is at least 91% identical to residues 20-141 of SEQ ID NO:10;   (g) an amino acid sequence of any one of SEQ ID NOs:49, 51, 52, 54, 56, 77, 78, 81, or 82;   (h) an amino acid sequence that is at least 91% identical to SEQ ID NO:51;   (i) an amino acid sequence that is at least 78% identical to SEQ ID NO:54;   (j) an amino acid sequence that is at least 89% identical to SEQ ID NO:77;   (k) an amino acid sequence that is at least 79% identical to SEQ ID NO:78;   (l) an amino acid sequence that is at least 80% identical to SEQ ID NO:81; or   (m) an amino acid sequence that is at least 78% identical to SEQ ID NO:82;   
     
     
         25 . The chimeric or humanized antibody of  claim 11 , wherein the light chain variable region sequence comprises:
 (a) an amino acid sequence of residues 21-127 of SEQ ID NO:4;   (b) an amino acid sequence that is at least 94% identical to residues 21-127 of SEQ ID NO:4;   (c) an amino acid sequence of residues 23-130 of SEQ ID NO:8;   (d) an amino acid sequence that is at least 96% identical to residues 23-130 of SEQ ID NO:8;   (e) an amino acid sequence of residues 21-127 of SEQ ID NO:12;   (f) an amino acid sequence that is at least 98% identical to residues 21-127 of SEQ ID NO:12;   (g) an amino acid sequence of any one of SEQ ID NOs: 58, 60, 62, 64, 66, 68, 70, 72, 74, 76, 79, 80, 83, or 84;   (h) an amino acid sequence that is at least 83% identical to SEQ ID NO:60;   (i) an amino acid sequence that is at least 93% identical to SEQ ID NO:70;   (j) an amino acid sequence that is at least 85% identical to SEQ ID NO:76;   (k) an amino acid sequence that is at least 85% identical to SEQ ID NO:76;   (l) an amino acid sequence that is at least 88% identical to SEQ ID NO:79;   (m) an amino acid sequence that is at least 84% identical to SEQ ID NO:80;   (n) an amino acid sequence that is at least 90% identical to SEQ ID NO:83; or   (o) an amino acid sequence that is at least 91% identical to SEQ ID NO:84.   
     
     
         26 . An antibody/drug conjugate for drug delivery comprising:
 (a) an antibody according to  claim 1 ; and   (b) a drug, which is directly or indirectly bound to the antibody.   
     
     
         27 . The antibody/drug conjugate of  claim 26 , wherein the drug is a therapeutic agent selected from the group consisting of a cytotoxin, a radioisotope, an immunomodulatory agent, an anti-angiogenic agent, an anti-proliferative agent, a pro-apoptotic agent, a chemotherapeutic agent, and a therapeutic nucleic acid. 
     
     
         28 . The antibody/drug conjugate of  claim 27 , wherein the therapeutic agent is a cytotoxin. 
     
     
         29 . The antibody/drug conjugate of  claim 26 , wherein the cytotoxin is an antibiotic, an inhibitor of tubulin polymerization, an alkylating agent, a protein synthesis inhibitor, a protein kinase inhibitor, a phosphatase inhibitor, a topoisomerase inhibitor, or an enzyme. 
     
     
         30 . The antibody/drug conjugate of  claim 27 , wherein the cytotoxin is an antibiotic. 
     
     
         31 . The antibody/drug conjugate of  claim 30 , wherein the antibiotic is calicheamicin. 
     
     
         32 . The antibody/drug conjugate of  claim 31 , wherein the calicheamicin is an N-acetyl derivative or disulfide analog of calicheamicin. 
     
     
         33 . The antibody/drug conjugate of  claim 32 , wherein the calicheamicin is N-acetyl-γ-calicheamicin. 
     
     
         34 . The antibody/drug conjugate of  claim 26 , wherein the drug is bound to the antibody via a linker. 
     
     
         35 . The antibody/drug conjugate of  claim 34 , wherein the linker is selected from the group consisting of 4-(4′ acetylphenoxy)butanoic acid (AcBut), 3-acetylphenyl acidic acid (AcPac), 4-mercapto-4-methyl-pentanoic acid (Amide), and derivatives thereof. 
     
     
         36 . A method for delivering a drug to 5T4-expressing cells comprising contacting the cells with an antibody/drug conjugate comprising (i) an anti-5T4 antibody of  claim 1 , and (ii) a drug which is bound to the anti-5T4 antibody directly or indirectly. 
     
     
         37 . The method of  claim 36 , wherein the drug is internalized in a target cell. 
     
     
         38 . A method for treating a subject having a 5T4-positive cancer, said method comprising administering to the subject a therapeutically effective amount of an anti-5T4 antibody/drug conjugate comprising (i) an anti-5T4 antibody of  claim 1 , and (ii) a therapeutic agent which is bound to the anti-5T4 antibody directly or indirectly. 
     
     
         39 . The method of  claim 38 , wherein the anti-5T4 antibody/drug conjugate is an anti-5T4 antibody/calicheamicin conjugate, and further comprising administering a second therapeutic agent, wherein the anti-5T4/calicheamicin conjugate and the second therapeutic agent are administered concurrently or consecutively in either order. 
     
     
         40 . An antibody that specifically binds to a human 5T4 epitope comprising residues 173-252 and 276-355, excepting residues 213-214, of SEQ ID NO:87. 
     
     
         41 . An antibody that specifically binds to a human 5T4 epitope comprising amino acids 276-355 of SEQ ID NO:87. 
     
     
         42 . An antibody that specifically binds to a human 5T4 epitope comprising amino acids 83-163 of SEQ ID NO:87. 
     
     
         43 . An antibody that specifically binds to cynomologous monkey 5T4 having an amino acid sequence of SEQ ID NO:86. 
     
     
         44 . An isolated 5T4 protein comprising:
 (a) an amino acid sequence of SEQ ID NO:85 or 86; or   (b) an amino acid sequence that is at least 95% identical to SEQ ID NO:85.   
     
     
         45 . An antibody that specifically binds the isolated 5T4 protein of  claim 44 . 
     
     
         46 . An isolated nucleic acid encoding an anti-5T4 heavy chain variable region comprising:
 (a) a nucleotide sequence of nucleotides 58-414 of SEQ ID NO:1;   (b) a nucleotide sequence of nucleotides 55-405 of SEQ ID NO:5;   (c) a nucleotide sequence of nucleotides 58-423 of SEQ ID NO:9;   (d) a nucleotide sequence encoding any one of SEQ ID NOs:48, 50, 53, or 55;   (e) a nucleotide sequence that is 89% identical to SEQ ID NO:50 when the query coverage is 100%;   (f) a nucleotide sequence that is 82% identical to SEQ ID NO:53 when the query coverage is 100%; or   (g) a nucleic acid that specifically hybridizes to the complement of any one of (a)-(d) under stringent hybridization conditions.   
     
     
         47 . An isolated nucleic acid encoding an anti-5T4 light chain variable region comprising:
 (a) a nucleotide sequence of nucleotides 61-381 of SEQ ID NO:3;   (b) a nucleotide sequence of nucleotides 67-390 of SEQ ID NO:7;   (c) a nucleotide sequence of nucleotides 61-381 of SEQ ID NO:11;   (d) a nucleotide sequence encoding a humanized A1, A2, or A3 light chain variable region of any one of SEQ ID NOs: 57, 59, 61, 63, 65, 67, 69, 71, 73, or 75;   (e) a nucleotide sequence that is 84% identical to SEQ ID NO:59 when the query coverage is 100%;   (f) a nucleotide sequence that is 86% identical to SEQ ID NO:69 when the query coverage is 100%;   (g) a nucleotide sequence that is 85% identical to SEQ ID NO:75 when the query coverage is 100%; or   (h) a nucleic acid that specifically hybridizes to the complement of any one of (a)-(d) under stringent hybridization conditions.