EPH RECEPTOR Fc VARIANTS WITH ENHANCED ANTIBODY DEPENDENT CELL-MEDIATED CYTOTOXICITY ACTIVITY

Abstract

The present invention relates to novel Fc variants that immuno-specifically bind to an Eph receptor. The Fc variants comprise a binding region that immunospecifically binds to an Eph receptor and an Fc region that further comprises at least one novel amino acid residue which may provide for enhanced effector function. More specifically, this invention provides Fc variants that have modified binding affinity to one or more Fc ligand (e.g., FcγR, C1q). Additionally, the Fc variants have altered antibody-dependent cell-mediated cytotoxicity (ADCC) and/or complement dependent cytotoxicity (CDC) activity. The invention further provides methods and protocols for the application of said Fc variants that immunospecifically bind to an Eph receptor, particularly for therapeutic purposes.

1 . An antibody comprising an IgG 1  Fc region, wherein the Fc region comprises at least the high effector function amino acid residue 332E, as numbered by the EU index as set forth in Kabat, wherein the antibody comprising at least the high effector function amino acid residue 332E has an altered binding affinity for one or more FcγRs as compared to the same antibody not comprising at least the high effector function amino acid residue 332E. 
     
     
         2 . The antibody of  claim 1 , wherein the Fc region further comprises at least the high effector function amino acid residues 239D and 330L, as numbered by the EU index as set forth in Kabat. 
     
     
         3 . The antibody of  claim 1 , wherein the high effector function amino acid residue is selected from the group consisting of: 234E, 235R, 235A, 235W, 235P, 235V, 235Y, 236E, 239D, 265L, 269S, 269G, 298I, 298T, 298F, 327N, 327G, 327W, 328S, 328V, 329H, 329Q, 330K, 330V, 330G, 330Y, 330T, 330L, 330I, 330R, 330C, 332E, 332H, 332S, 332W, 332F, 332D, and 332Y, wherein the numbering system is that of the EU index as set forth in Kabat. 
     
     
         4 . The antibody of  claim 1 , wherein said altered binding affinity for said one or more FcγRs is increased as compared to the same antibody not comprising at least the high effector function amino acid residue 332E. 
     
     
         5 . The antibody of  claim 4 , wherein said FcγR is FcγRIIIA. 
     
     
         6 . The antibody of  claim 4 , wherein said FcγR is FcγRIIB. 
     
     
         7 . The antibody of  claim 1 , wherein said altered binding affinity for said one or more FcγRs is decreased as compared to the same antibody not comprising at least the high effector function amino acid residue 332E. 
     
     
         8 . The antibody of  claim 5 , wherein the equilibrium dissociation constant (K D ) of binding for FcγRIIIA is decreased at least 2 fold as compared to the same antibody not comprising at least the high effector function amino acid residue 332E. 
     
     
         9 . The antibody of  claim 8 , wherein the equilibrium dissociation constant (K D ) of binding for FcγRIIIA is decreased at least 70 fold as compared to the same antibody not comprising at least the high effector function amino acid residue 332E. 
     
     
         10 . The antibody of  claim 5 , wherein said increased affinity for FcγRIIIA results in an enhanced ADCC activity relative to a comparable molecule not comprising at least the high effector function amino acid residue 332E. 
     
     
         11 . The antibody of  claim 10 , wherein said enhanced ADCC activity is at least 2 fold greater relative to a comparable molecule not comprising at least the high effector function amino acid residue 332E. 
     
     
         12 . The antibody of  claim 1 , wherein said antibody is humanized, fully human, CDR-grafted, or chimeric. 
     
     
         13 . The antibody of  claim 12 , wherein said antibody binds at least one Eph receptor selected from the group consisting of: EphA1, EphA2, EphA3a, EphA3b, EphA4, EphA5a, EphA5b, EphA6, EphA7, EphA8, EphB1, EphB2a, EphB2b, EphB3, EphB4 and EphB6. 
     
     
         14 . The antibody of  claim 12 , wherein said antibody binds at least the Eph receptor EphA2. 
     
     
         15 . The antibody of  claim 12 , wherein said antibody binds at least the Eph receptor EphA4. 
     
     
         16 . The antibody of  claim 12 , wherein said antibody variable sequences comprise SEQ ID Nos. 68 and 69. 
     
     
         17 . The antibody of  claim 12 , wherein said antibody is conjugated to a detectable agent, therapeutic agent or drug. 
     
     
         18 . A method of generating the antibody of  claim 1 , comprising (a) isolating antibody coding regions; and (b) making one or more desired substitutions in said Fc region of said isolated antibody coding region. 
     
     
         19 . A method of generating the antibody of  claim 1 , comprising subcloning variable regions into a vector encoding said Fc region comprising at least one or more high effector function amino acid residues. 
     
     
         20 . A formulation comprising the antibody of  claim 1  in a pharmaceutically-acceptable excipient.