Siglec neutralizing antibodies

Abstract

This invention relates to agents that bind human Siglecs having inhibitory activity in immune cells, and that neutralize the inhibitory activity of such Siglec. Such agents can be used for the treatment of cancers or infectious disease.

We claim: 
     
         1 . An in vitro method for modulating the activity of monocyte-derived cells and/or lymphocytes, optionally CD56dim NK cells, CD56bright NK cell and/or CD8+ T cells, the method comprising bringing monocyte-derived cells and/or lymphocytes expressing Siglec-9 into contact with an antibody that binds a human Siglec-9 polypeptide and is capable of neutralizing the inhibitory activity of a Siglec-9 polypeptide expressed by a human NK cell, wherein the antibody is selected from the group consisting of:
 (a) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 15 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 16;   (b) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 17 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 18;   (c) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 19 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 20;   (d) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 21 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 22;   (e) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 23 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 24; and   (f) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 25 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 26.   
     
     
         2 . The method of  claim 1 , wherein the cells are present in a biological sample obtained from a subject having a disease, optionally a cancer. 
     
     
         3 . An in vitro method for assessing the activity of NK cells from a subject having disease, the method comprising:
 (i) obtaining a biological sample from a subject comprising NK cells, bringing said cells into contact with an antibody that binds a human Siglec-9 polypeptide and is capable of neutralizing the inhibitory activity of a Siglec-9 polypeptide expressed by a human NK cell, wherein the antibody is selected from the group consisting of:   (a) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 15 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 16;   (b) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 17 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 18;   (c) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 19 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 20;   (d) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 21 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 22;   (e) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 23 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 24; and   (f) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 25 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 26; and   (ii) assessing whether the antibody modulate the activity of the NK cells.   
     
     
         4 . A method for selecting subjects having a cancer that responds to a treatment with an antibody that binds a human Siglec-9 polypeptide and is capable of neutralizing the inhibitory activity of a Siglec-9 polypeptide expressed by a human NK cell, wherein the antibody is selected from the group consisting of:
 (a) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 15 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 16;   (b) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 17 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 18;   (c) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 19 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 20;   (d) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 21 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 22;   (e) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 23 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 24; and   (f) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 25 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 26;   the method comprising determining whether cancer cells in said subject express a ligand of Siglec-9, the expression of sialic acid ligand(s) of Siglec-9 or elevated levels of sialic acid ligand(s) of Siglec-9, being indicative of a responder subject.   
     
     
         5 . The method of  claim 4 , further comprising administering to a responder subject an antibody that binds a human Siglec-9 polypeptide and is capable of neutralizing the inhibitory activity of a Siglec-9 polypeptide expressed by a human NK cell, wherein the antibody is selected from the group consisting of:
 (a) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 15 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 16;   (b) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 17 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 18;   (c) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 19 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 20;   (d) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 21 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 22;   (e) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 23 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 24; and   (f) an antibody comprising (i) a heavy chain variable region comprising CDR 1, 2 and 3 of the heavy chain variable region of SEQ ID NO: 25 and (ii) a light chain variable region comprising CDR 1, 2 and 3 of the light chain variable region of SEQ ID NO: 26.   
     
     
         6 . A method of producing an antibody which neutralizes the inhibitory activity of a Siglec-9 polypeptide, comprising:
 (a) providing a plurality of antibodies that bind a Siglec-9 polypeptide,   (b) selecting antibodies (e.g., those of step of (a)) that neutralize the inhibitory activity of a Siglec-9 polypeptide, and   (c) selecting antibodies (e.g., those of step (b)) that substantially block the interaction between a Siglec-9 polypeptide and a sialic acid ligand thereof, wherein the sialic acid ligand comprises a Neu5Aca2-3Galb1-4GlcNAcb structure.   
     
     
         7 . The method of  claim 6 , wherein assessing ability to neutralize the inhibitory activity of a Siglec polypeptide comprises selecting an antibody capable of causing an increase in cytotoxicity, optionally a marker of cytotoxicity, in NK cells purified from human donors that express Siglec-9, when the NK cells are brought into contact with a target human cell bearing a ligand of the Siglec on the target cell surface. 
     
     
         8 . An antibody or antibody fragment that binds a human Siglec-9 polypeptide, comprising: a heavy chain CDR1 comprising the amino acid sequence SYWMH (SEQ ID NO: 75); a heavy chain CDR2 comprising the amino acid sequence EINPSNGHTNYNEKFES (SEQ ID NO: 78); a heavy chain CDR3 comprising the amino acid sequence GVESYDFDDALDY (SEQ ID NO: 80); a light chain CDR1 comprising the amino acid sequence RASQDINNYLN (SEQ ID NO: 83); a light chain CDR2 comprising the amino acid sequence YTSRLHS (SEQ ID NO: 58); a light chain CDR3 comprising the amino acid sequence QQGNTLPFT (SEQ ID NO: 86); and human heavy and light chain framework sequences. 
     
     
         9 . The antibody of  claim 8 , wherein the antibody is an antibody having an Fc domain that is modified to reduce binding between the Fc domain and an Fcγ receptor. 
     
     
         10 . The antibody of  claim 8 , wherein said antibody is an antibody fragment. 
     
     
         11 . An isolated Siglec binding protein comprising an antibody fragment of  claim 10 . 
     
     
         12 . A pharmaceutical composition comprising an antibody or antibody fragment according to  claim 8 , and a pharmaceutically acceptable carrier. 
     
     
         13 . A kit comprising the antibody or antibody fragment of  claim 8  and a labelled secondary anti-IgG antibody that specifically recognizes said antibody. 
     
     
         14 . A nucleic acid encoding a heavy and/or light chain of an antibody or antibody fragment of  claim 8 . 
     
     
         15 . A hybridoma or recombinant host cell producing the antibody or antibody fragment of  claim 8 . 
     
     
         16 . A method for the treatment of a cancer in a patient in need thereof, the method comprising administering to said patient an effective amount of an antibody or antibody fragment of  claim 8 . 
     
     
         17 . An antibody or antibody fragment that binds a human Siglec-9 polypeptide, comprising: a heavy chain CDR1 comprising the amino acid sequence DYSMH (SEQ ID NO: 112); a heavy chain CDR2 comprising the amino acid sequence WIITETGEPTYADDFRG (SEQ ID NO: 115); a heavy chain CDR3 comprising the amino acid sequence DFDGY (SEQ ID NO: 117); a light chain CDR1 comprising the amino acid sequence RASENIYSYLA (SEQ ID NO: 119); a light chain CDR2 comprising the amino acid sequence NAKTLTE (SEQ ID NO: 122); a light chain CDR3 comprising the amino acid sequence QHHYGFPWT (SEQ ID NO: 123); and human heavy and light chain framework sequences. 
     
     
         18 . The antibody of  claim 17 , wherein the antibody is an antibody having an Fc domain that is modified to reduce binding between the Fc domain and an Fcγ receptor. 
     
     
         19 . The antibody of  claim 17 , wherein said antibody is an antibody fragment. 
     
     
         20 . An isolated Siglec binding protein comprising an antibody fragment of  claim 19 . 
     
     
         21 . A pharmaceutical composition comprising an antibody or antibody fragment according to  claim 17 , and a pharmaceutically acceptable carrier. 
     
     
         22 . A kit comprising the antibody or antibody fragment of  claim 17  and a labelled secondary anti-IgG antibody that specifically recognizes said antibody. 
     
     
         23 . A nucleic acid encoding a heavy and/or light chain of an antibody or antibody fragment of  claim 17 . 
     
     
         24 . A hybridoma or recombinant host cell producing the antibody or antibody fragment of  claim 17 . 
     
     
         25 . A method for the treatment of a cancer in a patient in need thereof, the method comprising administering to said patient an effective amount of an antibody or antibody fragment of  claim 17 .