Anti-siglec-8 antibodies and methods of use thereof

Abstract

The invention provides humanized anti-Siglec-8 antibodies and their use in treating and preventing eosinophil-mediated disorders and/or mast cell-mediated disorders, as well as compositions and kits comprising the humanized anti-Siglec-8 antibodies.

1 . A humanized antibody that binds to a human Siglec-8, wherein the antibody comprises a human IgG1 Fc region; and wherein at least one or two of the heavy chains of the antibody is non-fucosylated. 
     
     
         2 . (canceled) 
     
     
         3 . (canceled) 
     
     
         4 . The antibody of  claim 1 , wherein the antibody comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises (i) HVR-H1 comprising the amino acid sequence of SEQ ID NO:61, (ii) HVR-H2 comprising the amino acid sequence of SEQ ID NO:62, and (iii) HVR-H3 comprising the amino acid sequence of SEQ ID NO:63; and wherein the light chain variable region comprises (i) HVR-L1 comprising the amino acid sequence of SEQ ID NO:64, (ii) HVR-L2 comprising the amino acid sequence of SEQ ID NO:65, and (iii) HVR-L3 comprising the amino acid sequence of SEQ ID NO:66. 
     
     
         5 . The antibody of  claim 1 , wherein the antibody comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO:6; and a light chain variable region comprising the amino acid sequence of SEQ ID NO:16 or 21. 
     
     
         6 - 8 . (canceled) 
     
     
         9 . The antibody of  claim 1 , wherein the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:75; and a light chain comprising the amino acid sequence of SEQ ID NO:76 or 77. 
     
     
         10 . (canceled) 
     
     
         11 . The antibody of  claim 1 , wherein the antibody has been engineered to improve antibody-dependent cell-mediated cytotoxicity (ADCC) activity. 
     
     
         12 - 21 . (canceled) 
     
     
         22 . A nucleic acid encoding the antibody of  claim 1 . 
     
     
         23 . A vector comprising the nucleic acid of  claim 22 . 
     
     
         24 . The vector of  claim 23 , which is an expression vector. 
     
     
         25 . A host cell comprising the nucleic acid of  claim 22 . 
     
     
         26 . A method of producing an antibody comprising culturing the host cell of  claim 25  under a condition that produces the antibody. 
     
     
         27 . The method of  claim 26 , further comprising recovering the antibody produced by the host cell. 
     
     
         28 . An anti-Siglec-8 antibody produced by the method of  claim 26 . 
     
     
         29 . A pharmaceutical composition comprising the antibody of  claim 1  and a pharmaceutically acceptable carrier. 
     
     
         30 . A composition comprising an antibody that specifically binds to human Siglec-8, wherein the antibody comprises a human IgG1 Fc region and N-glycoside-linked carbohydrate chains linked to the Fc region, wherein less than 50% of the N-glycoside-linked carbohydrate chains contain a fucose residue. 
     
     
         31 . The composition of  claim 30 , wherein substantially none of the N-glycoside-linked carbohydrate chains contain a fucose residue. 
     
     
         32 . The composition of  claim 30 , wherein the antibody is a humanized antibody or a human antibody. 
     
     
         33 . The composition of  claim 30 , wherein the antibody comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises (i) HVR-H1 comprising the amino acid sequence of SEQ ID NO:61, (ii) HVR-H2 comprising the amino acid sequence of SEQ ID NO:62, and (iii) HVR-H3 comprising the amino acid sequence of SEQ ID NO:63; and wherein the light chain variable region comprises (i) HVR-L1 comprising the amino acid sequence of SEQ ID NO:64, (ii) HVR-L2 comprising the amino acid sequence of SEQ ID NO:65, and (iii) HVR-L3 comprising the amino acid sequence of SEQ ID NO:66. 
     
     
         34 . The composition of  claim 33 , wherein the antibody comprises a heavy chain variable region comprising the amino acid sequence selected from SEQ ID NOs:2-10; and a light chain variable region comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:16-22. 
     
     
         35 . The composition of  claim 30 , wherein the antibody comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises (i) HVR-H1 comprising the amino acid sequence of SEQ ID NO:61, (ii) HVR-H2 comprising the amino acid sequence of SEQ ID NO:62, and (iii) HVR-H3 comprising the amino acid sequence selected from SEQ ID NOs:67-70; and wherein the light chain variable region comprises (i) HVR-L1 comprising the amino acid sequence of SEQ ID NO:64, (ii) HVR-L2 comprising the amino acid sequence of SEQ ID NO:65, and (iii) HVR-L3 comprising the amino acid sequence of SEQ ID NO:71. 
     
     
         36 . The composition of  claim 35 , wherein the antibody comprises a heavy chain variable region comprising the amino acid sequence selected from SEQ ID NOs:11-14; and a light chain variable region comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:23-24. 
     
     
         37 . The composition of  claim 30 , wherein the antibody comprises a heavy chain variable region comprising the amino acid sequence selected from SEQ ID NOs:2-14; and a light chain variable region comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:16-24. 
     
     
         38 . The composition of  claim 30 , further comprising a pharmaceutically acceptable carrier. 
     
     
         39 . The composition of  claim 30 , wherein the binding affinity or avidity of the antibody to a human Siglec-8 is higher than the binding affinity or avidity of antibody 2E2 or 2C4 to the human Siglec-8. 
     
     
         40 . The composition of  claim 30 , wherein the antibody has a Tm of at least about 70° C. in a thermal shift assay. 
     
     
         41 . The composition of  claim 40 , wherein the antibody has a Tm of at least about 70° C. to at least about 72° C. in a thermal shift assay. 
     
     
         42 - 76 . (canceled) 
     
     
         77 . A method of treating or preventing a disease mediated by cells expressing Siglec-8 in a subject, the method comprising administering to the subject an effective amount of the composition of  claim 30 . 
     
     
         78 . The method of  claim 77 , wherein the disease is an eosinophil mediated-disease. 
     
     
         79 . The method of  claim 77 , wherein the disease is a mast cell mediated-disease. 
     
     
         80 . The method of  claim 77 , wherein the antibody inhibits one or more symptoms of an allergic reaction. 
     
     
         81 . The method of  claim 80 , wherein the allergic reaction is a Type I hypersensitivity reaction. 
     
     
         82 . The method of  claim 77 , wherein the disease is selected from the group consisting of: asthma, allergic rhinitis, nasal polyposis, atopic dermatitis, chronic urticaria, mastocytosis, eosinophilic leukemia, and hypereosinophilic syndrome. 
     
     
         83 . The method of  claim 77 , wherein the disease is selected from the group consisting of: pauci granulocytic asthma, acute or chronic airway hypersensitivity, eosinophilic esophagitis, Churg-Strauss syndrome, inflammation associated with a cytokine, inflammation associated with cells expressing Siglec-8, malignancy associated with cells expressing Siglec-8, physical urticaria, cold urticaria, pressure-urticaria, bullous pemphigoid, food allergy, and allergic bronchopulmonary aspergillosis (ABPA). 
     
     
         84 . The method of  claim 77 , wherein the subject is suffering from asthma that is not adequately controlled by an inhaled corticosteroid, a short acting β2 agonist, a long acting β2 agonist, or a combination thereof. 
     
     
         85 - 109 . (canceled) 
     
     
         110 . The antibody of  claim 1 , wherein the antibody is produced in a cell line having a alpha1,6-fucosyltransferase (Fut8) knockout. 
     
     
         111 . The antibody of  claim 1 , wherein the antibody is produced in a cell line overexpressing β1,4-N-acetylglycosminyltransferase III (GnT-III). 
     
     
         112 . The antibody of  claim 111 , wherein the cell line additionally overexpresses Golgi μ-mannosidase II (ManII). 
     
     
         113 . The composition of  claim 30 , wherein the antibody is produced in a cell line having a alpha1,6-fucosyltransferase (Fut8) knockout. 
     
     
         114 . The composition of  claim 30 , wherein the antibody is produced in a cell line overexpressing β1,4-N-acetylglycosminyltransferase III (GnT-III). 
     
     
         115 . The composition of  claim 114 , wherein the cell line additionally overexpresses Golgi μ-mannosidase II (ManII).