Anti-par2 antibodies and uses thereof

Abstract

The present disclosure provides antibodies and antigen-binding fragments capable of binding PAR2. In some embodiments, the anti-PAR2 antibodies or antigen-binding fragments thereof bind PAR2 in a pH-dependent manner. The disclosure further provides methods for making and using the antibodies and antigen-binding fragments.

What is claimed is: 
     
         1 . An antibody or antigen-binding fragment thereof comprising a heavy chain variable domain (VH) and a light chain variable domain (VL), wherein the VH comprises:
 i) a VH-CDR1 having the amino acid sequence of SEQ ID NO: 3, but wherein 1, 2, 3, 4, or 5 amino acid substitutions, deletions or insertions are optionally present in the sequence of SEQ ID NO: 3;   ii) a VH-CDR2 having the amino acid sequence of SEQ ID NO: 4, but wherein 1, 2, 3, 4, or 5 amino acid substitutions, deletions or insertions are optionally present in the sequence of SEQ ID NO: 4; and   iii) a VH-CDR3 having the amino acid sequence of SEQ ID NO: 5, but wherein 1, 2, 3, 4, or 5 amino acid substitutions, deletions or insertions are optionally present in the sequence of SEQ ID NO: 5;   and wherein the VL comprises:   i) a VL-CDR1 having the amino acid sequence of SEQ ID NO: 8; but wherein 1, 2, 3, 4, or 5 amino acid substitutions, deletions or insertions are optionally present in the sequence of SEQ ID NO: 8;   ii) a VL-CDR2 having the amino acid sequence of SEQ ID NO: 9, but wherein 1, 2, 3, 4, or 5 amino acid substitutions, deletions or insertions are optionally present in the sequence of SEQ ID NO: 9; and   iii) a VL-CDR3 having the amino acid sequence of SEQ ID NO: 10; but wherein 1, 2, 3, 4, or 5 amino acid substitutions, deletions or insertions are optionally present in the sequence of SEQ ID NO: 10;   wherein the amino acid substitutions, deletions or insertions reduce the binding affinity of the antibody or antigen-binding fragment thereof for human PAR2 by no more than 1000, 800, 700, 500, 400, 300, 200, 100, 50 or 10-fold as compared to an antibody or antigen-binding fragment having a VH with an amino acid sequence of SEQ ID NO: 2 and VL with an amino acid sequence of SEQ ID NO: 7 when tested at a pH of 7.4 in a PAR2 binding assay.   
     
     
         2 . An antibody or antigen-binding fragment thereof comprising a heavy chain variable domain (VH) and a light chain variable domain (VL), wherein the VH comprises:
 i) a VH-CDR1 having the amino acid sequence of SEQ ID NO: 3;   ii) a VH-CDR2 having the amino acid sequence of SEQ ID NO: 4, but wherein a histidine is optionally present at any one or more of the amino acid positions corresponding to positions 1-17 of SEQ ID NO: 4; and   iii) a VH-CDR3 having the amino acid sequence of SEQ ID NO: 5, but wherein a histidine is optionally present at any one or more of the amino acid positions corresponding to positions 1-8 of SEQ ID NO: 5;   and wherein the VL comprises:   i) a VL-CDR1 having the amino acid sequence of SEQ ID NO: 8;   ii) a VL-CDR2 having the amino acid sequence of SEQ ID NO: 9; and   iii) a VL-CDR3 having the amino acid sequence of SEQ ID NO: 10; but wherein a histidine is optionally present at any one or more of the amino acid positions corresponding to positions 1-14 of SEQ ID NO: 10.   
     
     
         3 . The antibody or antigen-binding fragment thereof of  claim 2 , wherein the VH comprises:
 i) a VH-CDR1 having the amino acid sequence of SEQ ID NO: 3,   ii) a VH-CDR2 having the amino acid sequence of SEQ ID NO: 4,   iii) a VH-CDR3 having the amino acid sequence of SEQ ID NO: 5,   and wherein the VL comprises:   i) a VL-CDR1 having the amino acid sequence of SEQ ID NO: 8,   ii) a VL-CDR2 having the amino acid sequence of SEQ ID NO: 9,   iii) a VL-CDR3 having the amino acid sequence of SEQ ID NO: 10.   
     
     
         4 . The antibody or antigen-binding fragment thereof of  claim 2 , wherein the VH comprises:
 i) a VH-CDR1 having the amino acid sequence of SEQ ID NO: 13,   ii) a VH-CDR2 having the amino acid sequence of SEQ ID NO: 14,   iii) a VH-CDR3 having the amino acid sequence of SEQ ID NO: 15,   and wherein the VL comprises:   i) a VL-CDR1 having the amino acid sequence of SEQ ID NO: 18,   ii) a VL-CDR2 having the amino acid sequence of SEQ ID NO: 19,   iii) a VL-CDR3 having the amino acid sequence of SEQ ID NO: 20.   
     
     
         5 . The antibody or antigen-binding fragment thereof of  claim 2 , wherein a histidine is present at any one or more of the amino acid positions corresponding to:
 a) positions 5, 7, 8, 12, 15, 16 or 17 of SEQ ID NO: 4;   b) positions 2 or 3 of SEQ ID NO: 5; and/or   c) positions 1, 5, 6 or 14 of SEQ ID NO: 10.   
     
     
         6 . The antibody or antigen-binding fragment thereof of  claim 2 , wherein a histidine is present at the amino acid positions corresponding to positions 5 and 8 of SEQ ID NO: 4. 
     
     
         7 . The antibody or antigen-binding fragment thereof of  claim 6 , wherein a histidine is present at the amino acid positions corresponding to positions 2 and 3 of SEQ ID NO: 5. 
     
     
         8 . The antibody or antigen-binding fragment thereof of  claim 2 , wherein:
 a) the VH-CDR2 comprises an amino acid sequence selected from the group consisting of: SEQ ID NOs: 4, 14, 24, 34, 44, 54, 64, 74, 84, 94, 104, 114, 124, 134, 144, 154, 164, 174, 184, 194, 204, 214, 224, 234, 244, 254, 264, 274, 284, 294, 304, 314, 324, 334, 344, 354, 364, 374, 384, 394, 404, 414, 424, 434, 444, 454, 464, 474, 484, 494, 504, 514, 524, 534, 544, 554, 564, 574, 584, 594, 604, 614, 624, 634, 644, 654, 664, 674, 684, 694, 704, 714, 724, 734, 744, 754, 764, 774, 784, 794, and 811-818;   b) the VH-CDR3 comprises an amino acid sequence selected from the group consisting of: SEQ ID NOs: 5, 15, 25, 35, 45, 55, 65, 75, 85, 95, 105, 115, 125, 135, 145, 155, 165, 175, 185, 195, 205, 215, 225, 235, 245, 255, 265, 275, 285, 295, 305, 315, 325, 335, 345, 355, 365, 375, 385, 395, 405, 415, 425, 435, 445, 455, 465, 475, 485, 495, 505, 515, 525, 535, 545, 555, 565, 575, 585, 595, 605, 615, 625, 635, 645, 655, 665, 675, 685, 695, 705, 715, 725, 735, 745, 755, 765, 775, 785, 795, and 819-820; and/or   c) the VL-CDR3 comprises an amino acid sequence selected from the group consisting of: SEQ ID NOs: 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, 270, 280, 290, 300, 310, 320, 330, 340, 350, 360, 370, 380, 390, 400, 410, 420, 430, 440, 450, 460, 470, 480, 490, 500, 510, 520, 530, 540, 550, 560, 570, 580, 590, 600, 610, 620, 630, 640, 650, 660, 670, 680, 690, 700, 710, 720, 730, 740, 750, 760, 770, 780, 790 and 800.   
     
     
         9 . The antibody or antigen-binding fragment thereof of  claim 2 , wherein the VH-CDR2 comprises an amino acid sequence corresponding to SEQ ID NO: 14; wherein the VH-CDR3 comprises an amino acid sequence corresponding to SEQ ID NO: 15, and wherein the VL-CDR3 comprises an amino acid sequence corresponding to SEQ ID NO: 20. 
     
     
         10 . The antibody or antigen-binding fragment thereof of  claim 2 , wherein the VH-CDR2 comprises an amino acid sequence corresponding to SEQ ID NO: 818; wherein the VH-CDR3 comprises an amino acid sequence corresponding to SEQ ID NO: 15, and wherein the VL-CDR3 comprises an amino acid sequence corresponding to SEQ ID NO: 10 or 20. 
     
     
         11 . The antibody or antigen-binding fragment thereof of  claim 2 , wherein:
 a) the VH comprises an amino acid sequence that is at least 80%, 85%, 90%, 92%, 93%, 95%, 97%, 98%, 99% or 100% identical to a sequence selected from the group consisting of: SEQ ID NOs: 2, 12, 22, 32, 42, 52, 62, 72, 82, 92, 102, 112, 122, 132, 142, 152, 162, 172, 182, 192, 202, 212, 222, 232, 242, 252, 262, 272, 282, 292, 302, 312, 322, 332, 342, 352, 362, 372, 382, 392, 402, 412, 422, 432, 442, 452, 462, 472, 482, 492, 502, 512, 522, 532, 542, 552, 562, 572, 582, 592, 602, 612, 622, 632, 642, 652, 662, 672, 682, 692, 702, 712, 722, 732, 742, 752, 762, 772, 782, 792, and 821-831; and/or   b) the VL comprises an amino acid sequence that is at least 80%, 85%, 90%, 92%, 93%, 95%, 97%, 98%, 99% or 100% identical to a sequence selected from the group consisting of: SEQ ID NOs: 7, 17, 27, 37, 47, 57, 67, 77, 87, 97, 107, 117, 127, 137, 147, 157, 167, 177, 187, 197, 207, 217, 227, 237, 247, 257, 267, 277, 287, 297, 307, 317, 327, 337, 347, 357, 367, 377, 387, 397, 407, 417, 427, 437, 447, 457, 467, 477, 487, 497, 507, 517, 527, 537, 547, 557, 567, 577, 587, 597, 607, 617, 627, 637, 647, 657, 667, 677, 687, 697, 707, 717, 727, 737, 747, 757, 767, 777, 787, and 797.   
     
     
         12 . The antibody or antigen-binding fragment thereof of  claim 2 , wherein the VH comprises an amino acid sequence corresponding to SEQ ID NO: 831 and wherein the VL comprises an amino acid sequence corresponding to SEQ ID NO: 7 or 17. 
     
     
         13 . The antibody or antigen-binding fragment thereof of  claim 2 , wherein the VH comprises an amino acid sequence that is at least 80%, 85%, 90%, 92%, 93%, 95%, 97%, 99% or 100% identical to SEQ ID NO: 12 and wherein the VL comprises an amino acid sequence that is at least 80%, 85%, 90%, 92%, 93%, 95%, 97%, 99% or 100% identical to SEQ ID NO: 17. 
     
     
         14 . The antibody or antigen-binding fragment of  claim 2 , wherein the antigen-binding fragment is an scFv or a Fab′. 
     
     
         15 . The antibody or antigen-binding fragment of  claim 2 , wherein the antibody is a monoclonal antibody. 
     
     
         16 . The antibody or antigen-binding fragment of  claim 2 , wherein the antibody or antigen-binding fragment prevents trypsin, tryptase and/or matriptase from interacting with PAR2. 
     
     
         17 . The antibody or antigen-binding fragment of  claim 2 , wherein the antibody or antigen-binding fragment binds to PAR2 with greater affinity at a pH of 7.4 than at a pH of 6.0. 
     
     
         18 . A composition comprising a pharmaceutically acceptable carrier and the antibody or antigen-binding fragment of  claim 2 . 
     
     
         19 . A kit comprising the antibody or antigen-binding fragment of  claim 2 . 
     
     
         20 . A method for treating pain in a subject in need thereof, comprising administering to the subject a pharmaceutically effective amount of the antibody or antigen-binding fragment of  claim 2 .