Antibodies to m(h)dm2/4 and their use in diagnosing and treating cancer

Abstract

The present invention relates to certain anti-M(H)DM2/4 antibodies or antigen-binding fragments thereof, pharmaceutical compositions comprising anti-M(H)DM2/4 antibodies or antigen-binding fragments thereof, antibody-drug conjugates comprising anti-M(H)DM2/4 antibodies or antigen-binding fragments thereof bound to a cytotoxic drug, and the use of such antibodies, fragments, compositions and conjugates for treating cancer and/or for preventing metastases. In particular, described herein are certain antibodies or antigen-binding fragments thereof that specifically bind to extracellularly accessible epitopes of M(H)DM2/4 and inhibit tumor growth in vivo, pharmaceutical compositions comprising such antibodies or fragments, antibody-drug conjugates comprising such antibodies or fragments, and the use of such antibodies, fragments, compositions and conjugates for treating cancer or for preventing metastasis.

1 . An antibody or a fragment thereof that specifically binds to an extracellularly accessible epitope of M(H)DM2/4, wherein the antibody or fragment specifically binds to a peptide, wherein the sequence of the peptide consists of MCNTNMSVPTDGAVT (SEQ ID NO:1), TTSQIPASEQE (SEQ ID NO:2), or CPVCRQPIQMIVLTYFP (SEQ ID NO:3). 
     
     
         2 . The antibody or fragment of  claim 1 , wherein the sequence of the peptide is MCNTNMSVPTDGAVT (SEQ ID NO:1). 
     
     
         3 . The antibody or fragment of  claim 1 , wherein the sequence of the peptide is TTSQIPASEQE (SEQ ID NO:2). 
     
     
         4 . The antibody or fragment of  claim 1 , wherein the sequence of the peptide is CPVCRQPIQMIVLTYFP (SEQ ID NO:3). 
     
     
         5 . The antibody or fragment of  claim 1 , which is a humanized antibody or a fragment thereof that specifically binds to HDM2, said antibody or fragment comprising: (i) a heavy chain variable region (VH) comprising VH complementarity determining region (“CDR”) 1, VH CDR2, and VH CDR3; said VH CDR1, VH CDR2 and VH CDR3 being the CDRs of a VH that has an amino acid sequence selected from the group consisting of SEQ ID NO:36, SEQ ID NO:38, and SED ID NO:40; or (ii) a light chain variable region (VL) comprising VL CDR1, VL CDR2, and VL CDR3; said VL CDR1, VL CDR2, and VL CDR3 being the CDRs of a VL that has an amino acid sequence selected from the group consisting of SEQ ID NO:37, SEQ ID NO:39, and SEQ ID NO:41. 
     
     
         6 . The humanized antibody or fragment of  claim 5 , which comprises the VH wherein VH CDR1, VH CDR2 and VH CDR3 are of a VH that has the amino acid sequence of SEQ ID NO:36. 
     
     
         7 . The humanized antibody or fragment of  claim 6 , which comprises the VL wherein VL CDR1, VL CDR2 and VL CDR3 are of a VL that has the amino acid sequence of SEQ ID NO:37. 
     
     
         8 . The humanized antibody or fragment of  claim 5 , which comprises the VH wherein VH CDR1, VH CDR2 and VH CDR3 are of a VH that has the amino acid sequence of SEQ ID NO:38. 
     
     
         9 . The humanized antibody or fragment of  claim 8 , which comprises the VL wherein VL CDR1, VL CDR2 and VL CDR3 are of a VL that has the amino acid sequence of SEQ ID NO:39. 
     
     
         10 . The humanized antibody or fragment of  claim 5 , which comprises the VH wherein VH CDR1, VH CDR2 and VH CDR3 are of a VH that has the amino acid sequence of SEQ ID NO:40. 
     
     
         11 . The humanized antibody or fragment of  claim 10 , which comprises the VL wherein VL CDR1, VL CDR2 and VL CDR3 are of a VL that has the amino acid sequence of SEQ ID NO:41. 
     
     
         12 . The antibody or fragment of  claim 1 , which specifically binds to HDM2, said antibody or fragment comprising a heavy chain variable region (VH) comprising VH complementarity determining region (“CDR”) 1, VH CDR 2, and VH CDR 3, wherein:
 (i) the VH CDR 1 has the amino acid sequence GFTFTHY (SEQ ID NO: 18), the VH CDR 2 has the amino acid sequence RNKAKGYT (SEQ ID NO: 19), and the VH CDR 3 has the amino acid sequence DIGDN (SEQ ID NO:20); 
 (ii) the VH CDR 1 has the amino acid sequence GFTFTHYYMS (SEQ ID NO:42), the VH CDR 2 has the amino acid sequence FIRNKAKGYTAE (SEQ ID NO:45), and the VH CDR 3 has the amino acid sequence DIGDN (SEQ ID NO:20); 
 (iii) the VH CDR 1 has the amino acid sequence HYYMS (SEQ ID NO:43), the VH CDR 2 has the amino acid sequence FIRNKAKGYTAEYSASVKG (SEQ ID NO:46), and the VH CDR 3 has the amino acid sequence DIGDN (SEQ ID NO:20); 
 (iv) the VH CDR 1 has the amino acid sequence THYYMS (SEQ ID NO:44), the VH CDR 2 has the amino acid sequence WLGFIRNKAKGYTAE (SEQ ID NO:47), and the VH CDR 3 has the amino acid sequence ARDIGD (SEQ ID NO:48); or 
 (v) the VH CDR 1 has the amino acid sequence FTFTHYY (SEQ ID NO: 144), the VH CDR 2 has the amino acid sequence IRNKAKGYTA (SEQ ID NO: 145), and the VH CDR 3 has the amino acid sequence ARDIGDN (SEQ ID NO: 146). 
 
     
     
         13 . The antibody or fragment of  claim 1 , which specifically binds to HDM2, said antibody or fragment comprising a heavy chain variable region (VH) comprising VH complementarity determining region (“CDR”) 1, VH CDR 2, and VH CDR 3, wherein:
 (i) the VH CDR 1 has the amino acid sequence GDTLSGS (SEQ ID NO:24), the VH CDR 2 has the amino acid sequence HLNRGT (SEQ ID NO:25), and the VH CDR 3 has the amino acid sequence SPGFAY (SEQ ID NO:26); 
 (ii) the VH CDR 1 has the amino acid sequence GDTLSGSWMH (SEQ ID NO:52), the VH CDR 2 has the amino acid sequence EIHLNRGTTN (SEQ ID NO: 55), and the VH CDR 3 has the amino acid sequence SPGFAY (SEQ ID NO:26); 
 (iii) the VH CDR 1 has the amino acid sequence GSWMH (SEQ ID NO:53), the VH CDR 2 has the amino acid sequence EIHLNRGTTNYNEKFKG (SEQ ID NO:56), and the VH CDR 3 has the amino acid sequence SPGFAY (SEQ ID NO:26); 
 (iv) the VH CDR 1 has the amino acid sequence SGSWMH (SEQ ID NO: 54), the VH CDR 2 has the amino acid sequence WIGEIHLNRGTTN (SEQ ID NO:57), and the VH CDR 3 has the amino acid sequence ARSPGFA (SEQ ID NO: 58); or 
 (v) the VH CDR 1 has the amino acid sequence GDTLSGSW (SEQ ID NO: 148), the VH CDR 2 has the amino acid sequence IHLNRGTT (SEQ ID NO: 143), and the VH CDR 3 has the amino acid sequence ARSPGFA (SEQ ID NO:58). 
 
     
     
         14 . The antibody or fragment of  claim 1 , which specifically binds to HDM2, said antibody or fragment comprising a heavy chain variable region (VH) comprising VH complementarity determining region (“CDR”) 1, VH CDR 2, and VH CDR 3, wherein: (i) the VH CDR 1 has the amino acid sequence GYTFTSY (SEQ ID NO:30), the VH CDR 2 has the amino acid sequence NPRNGG (SEQ ID N0:31), and the VH CDR 3 has the amino acid sequence SGYYAMDY (SEQ ID NO:32);
 (ii) the VH CDR 1 has the amino acid sequence GYTFTSYYMY (SEQ ID NO:62), the VH CDR 2 has the amino acid sequence GINPRNGGTN (SEQ ID NO: 65), and the VH CDR 3 has the amino acid sequence SGYYAMDY (SEQ ID NO:32); 
 (iii) the VH CDR 1 has the amino acid sequence SYYMY (SEQ ID NO:63), the VH CDR 2 has the amino acid sequence GINPRNGGTNFNEKFKN (SEQ ID NO: 66), and the VH CDR 3 has the amino acid sequence SGYY AMDY (SEQ ID NO:32); or 
 (iv) the VH CDR 1 has the amino acid sequence TSYYMY (SEQ ID NO:64), the VH CDR 2 has the amino acid sequence WIGGINPRNGGTN (SEQ ID NO: 67), and the VH CDR 3 has the amino acid sequence TRSGYYAMD (SEQ ID NO:68). 
 
     
     
         15 . The antibody or fragment of  claim 12 , wherein the antibody or fragment further comprises a light chain variable region (VL) comprising VL complementarity determining region (“CDR”) 1, VL CDR 2, and VL CDR 3, wherein:
 (i) the VL CDR 1 has the amino acid sequence RSSKNLLHSNGITYLY (SEQ ID NO:21), the VL CDR 2 has the amino acid sequence RVSNLAS (SEQ ID NO:22), and the VL CDR 3 has the amino acid sequence AQLLELPYT (SEQ ID NO:23); 
 (ii) the VL CDR 1 has the amino acid sequence LHSNGITYL YWY (SEQ ID NO:49), the VL CDR 2 has the amino acid sequence LLISRVSNLA (SEQ ID NO:50), and the VL CDR 3 has the amino acid sequence AQLLELPY (SEQ ID NO:51); or 
 (iii) the VL CDR 1 has the amino acid sequence KNLLHSNGITY (SEQ ID NO: 147), the VL CDR 2 has the amino acid sequence RVS, and the VL CDR 3 has the amino acid sequence AQLLELPYT (SEQ ID NO:23). 
 
     
     
         16 . The antibody or fragment of  claim 13 , wherein the antibody or fragment further comprises a light chain variable region (VL) comprising VL complementarity determining region (“CDR”) 1, VL CDR 2, and VL CDR 3, wherein:
 (i) the VL CDR 1 has the amino acid sequence RSSKSLLHSNGNSYLY (SEQ ID NO:27), the VL CDR 2 has the amino acid sequence RMSNLAS (SEQ ID NO:28), and the VL CDR 3 has the amino acid sequence MQHLEYPFT (SEQ ID NO:29); 
 (ii) the VL CDR 1 has the amino acid sequence LHSNGNSYLYWF (SEQ ID NO:59), the VL CDR 2 has the amino acid sequence LLIYRMSNLA (SEQ ID NO: 60), and the VL CDR 3 has the amino acid sequence MQHLEYPF (SEQ ID N0:61); or 
 (iii) the VL CDR 1 has the amino acid sequence KSLLHSNGNSY (SEQ ID NO: 141), the VL CDR 2 has the amino acid sequence RMS, and the VL CDR 3 has the amino acid sequence MQHLEYPFT (SEQ ID NO:29). 
 
     
     
         17 . The antibody or fragment of  claim 14 , wherein the antibody or fragment further comprises a light chain variable region (VL) comprising VL complementarity determining region (“CDR”) 1, VL CDR 2, and VL CDR 3, wherein:
 (i) the VL CDR 1 has the amino acid sequence RASQDISNFLN (SEQ ID NO:33), the VL CDR 2 has the amino acid sequence YTSRLHS (SEQ ID NO:34), and the VL CDR 3 has the amino acid sequence QQGNTLPRT (SEQ ID NO: 35); or 
 (ii) the VL CDR 1 has the amino acid sequence SNFLNWY (SEQ ID NO:69), the VL CDR 2 has the amino acid sequence LLIYYTSRLH (SEQ ID NO:70), and the VL CDR 3 has the amino acid sequence QQGNTLPR (SEQ ID NO:71). 
 
     
     
         18 . The antibody or fragment of  claim 12 , which comprises a VH having the amino acid sequence of SEQ ID NO: 36, or a VH having at least 95% sequence identity to the amino acid sequence of SEQ ID NO:36. 
     
     
         19 . The antibody or fragment of  claim 18 , which comprises a light chain variable region (VL) having the amino acid sequence of SEQ ID NO:37, or a VL having at least 95% sequence identity to the amino acid sequence of SEQ ID NO:37. 
     
     
         20 . The antibody or fragment of  claim 13 , which comprises a VH having the amino acid sequence of SEQ ID NO: 38, or a VH having at least 95% sequence identity to the amino acid sequence of SEQ ID NO:38. 
     
     
         21 . The antibody or fragment of  claim 20 , which comprises a VL having the amino acid sequence of SEQ ID NO: 39, or a VL having at least 95% sequence identity to the amino acid sequence of SEQ ID NO:39. 
     
     
         22 . The antibody or fragment of  claim 14 , which comprises a VH having the amino acid sequence of SEQ ID NO:40, or a VH having at least 95% sequence identity to the amino acid sequence of SEQ ID NO:40. 
     
     
         23 . The antibody or fragment of  claim 22 , which comprises a VL having the amino acid sequence of SEQ ID NO:41, or a VL having at least 95% sequence identity to the amino acid sequence of SEQ ID NO:41. 
     
     
         24 . The antibody or fragment of  claim 1 , wherein the antibody is a humanized antibody. 
     
     
         25 . The antibody or fragment of  claim 1 , wherein the antibody is a human antibody. 
     
     
         26 . The antibody or fragment of  claim 1 , wherein the antibody is a chimeric antibody. 
     
     
         27 . The antibody or fragment of  claim 1 , wherein the antibody is a monoclonal antibody. 
     
     
         28 . The antibody or fragment of  claim 1 , which is an immunoglobulin. 
     
     
         29 . The antibody or fragment of  claim 28 , wherein the immunoglobulin is an IgG. 
     
     
         30 . The antibody or fragment of  claim 28 , wherein the immunoglobulin is of IgG1 isotype. 
     
     
         31 . The antibody or fragment of  claim 28 , wherein the immunoglobulin is of IgG3 isotype. 
     
     
         32 . The antibody or fragment of  claim 1 , which comprises an Fe region, and wherein the Fe region is a human IgG1, a human IgG2, a human IgG3, a human IgG4, or a human IgM Fc region. 
     
     
         33 . The antibody or fragment of  claim 32 , wherein the antibody or fragment mediates complement-dependent cytotoxicity (CDC) or antibody-dependent cell-mediated cytotoxicity (ADCC). 
     
     
         34 . The antibody or fragment of  claim 33 , wherein the antibody or fragment mediates complement-dependent cytotoxicity (CDC). 
     
     
         35 . The antibody or fragment of  claim 1 , which is an Fv fragment, a Fab fragment, a Fab′ fragment, a F(ab′) 2 fragment, a single chain antibody molecule, or a single chain Fv (scFv). 
     
     
         36 . The antibody or fragment of  claim 1 , wherein the HDM2 is an HDM2 variant that lacks a nuclear localization signal domain, an HDM2 variant that lacks the sequence of amino acids 179 to 185 of SEQ ID NO:4, and/or an HDM2 variant that lacks the sequence of amino acids 464 to 471 of SEQ ID NO:4. 
     
     
         37 . The antibody or fragment of  claim 1 , which is purified. 
     
     
         38 . The antibody or fragment of  claim 1 , which inhibits tumor cell proliferation in vivo. 
     
     
         39 . (canceled) 
     
     
         40 . (canceled) 
     
     
         41 . An antibody-drug conjugate comprising the antibody or fragment of  claim 1 , bound to a cytotoxic drug. 
     
     
         42 . A pharmaceutical composition comprising a therapeutically effective amount of the antibody or fragment of  claim 1 . 
     
     
         43 .- 110 . (canceled) 
     
     
         111 . A method of diagnosing cancer in a subject, said method comprising:
 (a) detecting whether the antibody or fragment of  claim 1  binds to the surface of intact cells of the subject; and   (b) diagnosing the subject with cancer if binding is detected in step (a).   
     
     
         112 . The method of  claim 111 , which is an ex vivo method. 
     
     
         113 . The method of  claim 111 , further comprising obtaining intact cells from the subject before step (a). 
     
     
         114 . The method of  claim 111 , which comprises administering the antibody or fragment to the subject before the detecting in step (a), and wherein the detecting is performed by in vivo imaging of the subject. 
     
     
         115 . The method of  claim 111 , wherein, in step (a), the antibody or fragment is labeled. 
     
     
         116 . The method of  claim 111 , wherein the subject is a human. 
     
     
         117 . The antibody or a fragment thereof of  claim 1 , wherein the antibody or a fragment thereof specifically binds to an extracellularly accessible epitope of HDM2. 
     
     
         118 . (canceled)