Single Domain Antibodies to Programmed Cell Death (PD-1)

Abstract

The invention relates to multifunctional PD-1 binding agents and the use of such binding agents in the treatment, prevention and detection of disease.

1 . An isolated binding agent comprising a) a first single domain antibody directed against a first epitope of human PD-1 and b) a second single domain antibody directed against a second epitope of human PD-1 wherein said first single domain antibody comprises one of the following CDR combinations: SEQ ID NOs: 1, 2, 3; SEQ ID NOs: 5, 6, 7; SEQ ID NOs: 9, 10, 11; SEQ ID NOs: 13, 14, 15; SEQ ID NOs: 17, 18, 19; SEQ ID NOs: 21, 22, 23; SEQ ID NOs: 25, 26, 27; SEQ ID NOs: 29, 30, 31; SEQ ID NOs: 33, 34, 35; SEQ ID NOs: 37, 38, 39; SEQ ID NOs: 41, 42, 43; SEQ ID NOs: 45, 46, 47; SEQ ID NOs: 49, 50, 51; SEQ ID NOs: 53, 54, 55; SEQ ID NOs: 57, 58, 59; SEQ ID NOs: 61, 62, 63; SEQ ID NOs: 65, 66, 67; SEQ ID NOs: 69, 70, 71; SEQ ID NOs: 73, 74, 75; SEQ ID NOs: 77, 78, 79; SEQ ID NOs: 101, 102, 103; SEQ ID NOs: 105, 106, 107; SEQ ID NOs: 109, 110, 111; SEQ ID NOs: 113, 114, 115; SEQ ID NOs: 117, 118, 119; SEQ ID NOs: 121, 122, 123; SEQ ID NOs: 125, 126, 127; SEQ ID NOs: 129; 130; 131; SEQ ID NOs: 133, 134, 135; SEQ ID NOs: 137, 138, 139; SEQ ID NOs: 141, 142, 143; SEQ ID NOs: 145, 146, 147; SEQ ID NOs: 149, 150, 151; SEQ ID NOs: 153, 154, 155; SEQ ID NOs: 157, 158, 159; SEQ ID NOs: 161, 162, 163; SEQ ID NOs: 165, 166, 167; SEQ ID NOs: 169, 170, 171; SEQ ID NOs: 173, 174, 175; SEQ ID NOs: 177, 178, 179; SEQ ID NOs: 181, 182, 183; SEQ ID NOs: 185, 186, 187; SEQ ID NOs: 189, 190, 191; SEQ ID NOs: 193, 194, 195; SEQ ID NOs: 197, 198, 199; SEQ ID NOs: 201, 202, 203; SEQ ID NOs: 205, 206, 207; SEQ ID NOs: 209, 210, 211; SEQ ID NOs: 213, 214, 215 or SEQ ID NOs: 217, 218, 219. 
     
     
         2 . A binding agent according to  claim 1 , wherein said domain of the single domain antibody is a human heavy chain variable domain (V H ). 
     
     
         3 . An isolated binding agent according to  claim 1 , wherein said first single domain antibody does not block the interaction of human PD-1 with human PD-L1 and/or PD-L2 and wherein said second single domain antibody blocks the interaction of human PD-1 with human PD-L1 and/or PD-L2. 
     
     
         4 - 9 . (canceled) 
     
     
         10 . An isolated binding agent according to  claim 1 , wherein said first single domain antibody comprises a sequence selected from SEQ ID NOs: 4, 8, 12, 16, 20, 24, 28, 32, 36, 40, 44, 48, 52, 56, 60, 64, 68, 72, 76, 80, 104, 108, 112, 116, 120, 124, 128, 132, 136, 140, 144, 148, 152, 156, 160, 164, 168, 172, 176, 180, 184, 188, 192, 196, 200, 204, 208, 212, 216 or 220 or a sequence with at least 90% homology thereto. 
     
     
         11 - 14 . (canceled) 
     
     
         15 . An isolated binding agent according to  claim 1 , wherein said second single domain antibody comprises a CDR1, CDR2 and CDR3 selected from CDRs1-3 of SEQ ID NOs: 519, 523, 527, 531, 535, 539, 543, 547, 551, 555, 559, 563, 567, 571, 575, 579, 583, 587, 591, 595, 599, 603, 607, 611, 615, 619, 623, 627, 631, 635, 639, 643, 647, 651, 655, 659, 663, 667, 671, 675, 679, 683, 687, 691, 695, 699, 703, 707, 711, 715, 719, 723, 727, 731, 735, 739, 743, 747, 751, 755, 759, 763, 767, 771, 775, 779, 783, 787, 791, 795, 799, 803, 807, 884, 888, 892, 896, 900, 904, 908, 912, 916, 920, 924, 928, 932, 936, 940, 944, 948, 952, 956, 960, 964, 968, 972, 976, 980, 984, 988, 992, 996, 1000, 1004, 1008, 1012, 1016, 1020, 1024, 1028, 1032, 1036, 1040, 1044 or 1048. 
     
     
         16 . An isolated binding agent according to  claim 1 , wherein said second single domain antibody comprises a sequence selected from SEQ ID NOs: 519, 523, 527, 531, 535, 539, 543, 547, 551, 555, 559, 563, 567, 571, 575, 579, 583, 587, 591, 595, 599, 603, 607, 611, 615, 619, 623, 627, 631, 635, 639, 643, 647, 651, 655, 659, 663, 667, 671, 675, 679, 683, 687, 691, 695, 699, 703, 707, 711, 715, 719, 723, 727, 731, 735, 739, 743, 747, 751, 755, 759, 763, 767, 771, 775, 779, 783, 787, 791, 795, 799, 803, 807, 884, 888, 892, 896, 900, 904, 908, 912, 916, 920, 924, 928, 932, 936, 940, 944, 948, 952, 956, 960, 964, 968, 972, 976, 980, 984, 988, 992, 996, 1000, 1004, 1008, 1012, 1016, 1020, 1024, 1028, 1032, 1036, 1040, 1044 or 1048 or a sequence with at least 90% homology thereto. 
     
     
         17 . An isolated binding agent according to  claim 1 , wherein said first and said second single domain antibody are covalently linked via a peptide, optionally wherein said peptide is between 3 and 50 amino acids in length, optionally wherein the linker comprises glycine and/or serine amino acid residues, optionally wherein the peptide has the formula (Gly4Ser)n, where n=from 1 to 20. 
     
     
         18 - 20 . (canceled) 
     
     
         21 . An isolated binding agent according to  claim 1 , comprising one or more further binding molecule,
 optionally wherein said binding molecule is an antibody or fragment thereof,   optionally wherein said binding molecule is a single domain antibody,   optionally wherein said binding molecule binds to human PD-1 or a different antigen,   optionally an immunooncology target.   
     
     
         22 - 26 . (canceled) 
     
     
         27 . An isolated binding agent according to  claim 1 , wherein at least one single domain antibody is conjugated to a toxin, enzyme, radioisotope, half-life extending moiety, therapeutic molecule or other chemical moiety, optionally wherein said half-life extending moiety is selected from the group consisting of an albumin binding moiety, a transferrin binding moiety, a polyethylene glycol molecule, a recombinant polyethylene glycol molecule, human serum albumin, a fragment of human serum albumin, and an albumin binding peptide or single domain antibody that binds to human serum albumin. 
     
     
         28 . (canceled) 
     
     
         29 . An isolated binding agent according to  claim 1 , wherein said first single domain antibody is located N-terminally and said second single domain antibody is located C-terminally or, wherein said first single domain antibody is located C-terminally and said second single domain antibody is located N-terminally. 
     
     
         30 - 31 . (canceled) 
     
     
         32 . A pharmaceutical composition comprising a binding agent according to  claim 1  and a pharmaceutical carrier. 
     
     
         33 - 45 . (canceled) 
     
     
         46 . A combination comprising a) a first single domain antibody directed against a first epitope of human PD-1 and b) a second single domain antibody directed against a second epitope of human PD-1 wherein said first single domain antibody comprises one of the following CDR combinations: SEQ ID NOs: 1, 2, 3; SEQ ID NOs: 5, 6, 7; SEQ ID NOs: 9, 10, 11; SEQ ID NOs: 13, 14, 15; SEQ ID NOs: 17, 18, 19; SEQ ID NOs: 21, 22, 23; SEQ ID NOs: 25, 26, 27; SEQ ID NOs: 29, 30, 31; SEQ ID NOs: 33, 34, 35; SEQ ID NOs: 37, 38, 39; SEQ ID NOs: 41, 42, 43, SEQ ID NOs: 45, 46, 47; SEQ ID NOs: 49, 50, 51; SEQ ID NOs: 53, 54, 55; SEQ ID NOs: 57, 58, 59; SEQ ID NOs: 61, 62, 63; SEQ ID NOs: 65, 66, 67; SEQ ID NOs: 69, 70, 71; SEQ ID NOs: 73, 74, 75; SEQ ID NOs: 77, 78, 79; SEQ ID NOs: 101, 102, 103; SEQ ID NOs: 105, 106, 107; SEQ ID NOs: 109, 110, 111; SEQ ID NOs: 113, 114, 115; SEQ ID NOs: 117, 118, 119; SEQ ID NOs: 121, 122, 123; SEQ ID NOs: 125, 126, 127; SEQ ID NOs: 129; 130; 131; SEQ ID NOs: 133, 134, 15; SEQ ID NOs: 137, 138, 139; SEQ ID NOs: 141, 142, 143; SEQ ID NOs: 145, 146, 147; SEQ ID NOs: 149, 150, 151; SEQ ID NOs: 153, 154, 155; SEQ ID NOs: 157, 158, 159; SEQ ID NOs: 161, 162, 163; SEQ ID NOs: 165, 166, 167; SEQ ID NOs: 169, 170, 171; SEQ ID NOs: 173, 174, 175; SEQ ID NOs: 177, 178, 179; SEQ ID NOs: 181, 182, 183; SEQ ID NOs: 185, 186, 187; SEQ ID NOs: 189, 190, 191; SEQ ID NOs: 193, 194, 195; SEQ ID NOs: 197, 198, 199; SEQ ID NOs: 201, 202, 203; SEQ ID NOs: 205, 206, 207; SEQ ID NOs: 209, 210, 211; SEQ ID NOs: 213, 214, 215 or SEQ ID NOs: 217, 218, 219. 
     
     
         47 - 49 . (canceled) 
     
     
         50 . An isolated binding agent comprising a) a first single domain antibody directed against a first epitope of human PD-1 and b) a second single domain antibody directed against a second epitope of human PD-1 wherein said first single domain antibody comprises one of the following CDR combinations: SEQ ID NOs: 251, 252, 253; SEQ ID NOs: 255, 256, 257; SEQ ID NOs: 259, 260, 261; SEQ ID NOs: 263, 264, 265; SEQ ID NOs: 267, 268, 269; SEQ ID NOs: 271, 272, 273; SEQ ID NOs: 275, 276, 277; SEQ ID NOs: 279, 280, 281; SEQ ID NOs: 283, 284, 285; SEQ ID NOs: 287, 288, 289; SEQ ID NOs: 291, 292, 293; SEQ ID NOs: 295, 296, 297; SEQ ID NOs: 299, 300, 301; SEQ ID NOs: 303, 304, 305; SEQ ID NOs: 307, 308, 309; SEQ ID NOs: 311, 312, 313; SEQ ID NOs: 315, 316, 317; SEQ ID NOs: 319, 320, 321; SEQ ID NOs: 323, 324, 325; SEQ ID NOs: 327, 328, 329; SEQ ID NOs: 331, 332, 333; SEQ ID NOs: 335, 336, 337; SEQ ID NOs: 339, 340, 341; SEQ ID NOs: 343, 344, 345; SEQ ID NOs: 347, 348, 349; SEQ ID NOs: 351, 352, 353; SEQ ID NOs: 355, 356, 357; SEQ ID NOs: 359, 360, 361; SEQ ID NOs: 363, 364, 365; SEQ ID NOs: 367, 368, 369; SEQ ID NOs: 371, 372, 373; SEQ ID NOs: 375, 376, 377; SEQ ID NOs: 379, 380, 381; SEQ ID NOs: 383, 384, 385; SEQ ID NOs: 387, 388, 389; SEQ ID NOs: 391, 392, 393; SEQ ID NOs: 395, 396, 397; SEQ ID NOs: 399, 400, 401; SEQ ID NOs: 403, 404, 405; SEQ ID NOs: 407, 408, 409; SEQ ID NOs: 411, 412, 413; SEQ ID NOs: 415, 416, 417; SEQ ID NOs: 419, 420, 421; SEQ ID NOs: 423, 424, 425; SEQ ID NOs: 427, 428, 429; SEQ ID NOs: 431, 432, 433; SEQ ID NOs: 435, 436, 437; SEQ ID NOs: 439, 440, 441; SEQ ID NOs: 443, 444, 445; SEQ ID NOs: 447, 448, 449; SEQ ID NOs: 451, 452, 453; SEQ ID NOs: 455, 456, 457 or SEQ ID NOs: 459, 460, 461. 
     
     
         51 . A binding agent according to  claim 50 , wherein said domain of the single domain antibody is a human heavy chain variable domain (VH). 
     
     
         52 . An isolated binding agent according to  claim 50 , wherein said first single domain antibody does not block the interaction of human PD-1 with human PD-L1 and/or PD-L2 and wherein said second single domain antibody blocks the interaction of human PD-1 with human PD-L1 and/or PD-L2. 
     
     
         53 . An isolated binding agent according to  claim 50 , wherein said first single domain antibody comprises a sequence selected from SEQ ID NOs: 254, 258, 262, 266, 270, 274, 278, 282, 286, 290, 294, 298, 302, 306, 310, 314, 318, 322, 326, 330, 334, 338, 342, 346, 350, 354, 358, 362, 366, 370, 374, 378, 382, 386, 390, 394, 398, 402, 406, 410, 414, 418, 422, 426, 430, 434, 438, 442, 446, 450, 454, 458 or 462 or a sequence with at least 90% homology thereto. 
     
     
         54 . An isolated binding agent according to  claim 50 , wherein said second single domain antibody comprises a CDR1, CDR2 and CDR3 selected from CDRs1-3 of SEQ ID No: 519, 523, 527, 531, 535, 539, 543, 547, 551, 555, 559, 563, 567, 571, 575, 579, 583, 587, 591, 595, 599, 603, 607, 611, 615, 619, 623, 627, 631, 635, 639, 643, 647, 651, 655, 659, 663, 667, 671, 675, 679, 683, 687, 691, 695, 699, 703, 707, 711, 715, 719, 723, 727, 731, 735, 739, 743, 747, 751, 755, 759, 763, 767, 771, 775, 779, 783, 787, 791, 795, 799, 803, 807, 884, 888, 892, 896, 900, 904, 908, 912, 916, 920, 924, 928, 932, 936, 940, 944, 948, 952, 956, 960, 964, 968, 972, 976, 980, 984, 988, 992, 996, 1000, 1004, 1008, 1012, 1016, 1020, 1024, 1028, 1032, 1036, 1040, 1044 or 1048. 
     
     
         55 . An isolated binding agent according to  claim 50 , wherein said second single domain antibody comprises a sequence selected from SEQ ID NOs: 519, 523, 527, 531, 535, 539, 543, 547, 551, 555, 559, 563, 567, 571, 575, 579, 583, 587, 591, 595, 599, 603, 607, 611, 615, 619, 623, 627, 631, 635, 639, 643, 647, 651, 655, 659, 663, 667, 671, 675, 679, 683, 687, 691, 695, 699, 703, 707, 711, 715, 719, 723, 727, 731, 735, 739, 743, 747, 751, 755, 759, 763, 767, 771, 775, 779, 783, 787, 791, 795, 799, 803, 807, 884, 888, 892, 896, 900, 904, 908, 912, 916, 920, 924, 928, 932, 936, 940, 944, 948, 952, 956, 960, 964, 968, 972, 976, 980, 984, 988, 992, 996, 1000, 1004, 1008, 1012, 1016, 1020, 1024, 1028, 1032, 1036, 1040, 1044 or 1048 or a sequence with at least 90% homology thereto. 
     
     
         56 . An isolated binding agent according to  claim 50 , wherein said first and said second single domain antibody are covalently linked by via a peptide, optionally wherein said peptide is between 3 and 50 amino acids in length, optionally wherein the linker comprises glycine and/or serine amino acid residues, optionally wherein the peptide has the formula (Gly4Ser)n, where n=from 1 to 20. 
     
     
         57 . An isolated binding agent according to  claim 50 , comprising one or more further binding molecule, optionally wherein said binding molecule is an antibody or fragment thereof, optionally wherein said binding molecule is a single domain antibody, optionally wherein said binding molecule binds to human PD-1 or a different antigen optionally an immunooncology target. 
     
     
         58 . An isolated binding agent according to  claim 50 , wherein at least one single domain antibody is conjugated to a toxin, enzyme, radioisotope, half-life extending moiety, therapeutic molecule or other chemical moiety, optionally wherein said half-life extending moiety is selected from the group consisting of an albumin binding moiety, a transferrin binding moiety, a polyethylene glycol molecule, a recombinant polyethylene glycol molecule, human serum albumin, a fragment of human serum albumin, and an albumin binding peptide or single domain antibody that binds to human serum albumin. 
     
     
         59 . An isolated binding agent according to  claim 50 , wherein said first single domain antibody is located N-terminally and said second single domain antibody is located C-terminally or wherein said first single domain antibody is located C-terminally and said second single domain antibody is located N-terminally. 
     
     
         60 . A pharmaceutical composition comprising a binding agent according to  claim 50  and a pharmaceutical carrier. 
     
     
         61 . An isolated nucleic acid molecule comprising a nucleotide sequence encoding a binding agent according to  claim 50 . 
     
     
         62 . A combination comprising a) a first single domain antibody directed against a first epitope of human PD-1 and b) a second single domain antibody directed against a second epitope of human PD-1, wherein said first single domain antibody comprises one of the following CDR combinations: SEQ ID NOs: 251, 252, 253; SEQ ID NOs: 255, 256, 257; SEQ ID NOs: 259, 260, 261; SEQ ID NOs: 263, 264, 265; SEQ ID NOs: 267, 268, 269; SEQ ID NOs: 271, 272, 273; SEQ ID NOs: 275, 276, 277; SEQ ID NOs: 279, 280, 281; SEQ ID NOs: 283, 284, 285; SEQ ID NOs: 287, 288, 289; SEQ ID NOs: 291, 292, 293; SEQ ID NOs: 295, 296, 297; SEQ ID NOs: 299, 300, 301; SEQ ID NOs: 303, 304, 305; SEQ ID NOs: 307, 308, 309; SEQ ID NOs: 311, 312, 313; SEQ ID NOs: 315, 316, 317; SEQ ID NOs: 319, 320, 321, SEQ ID NOs: 323, 324, 325; SEQ ID NOs: 327, 328, 329; SEQ ID NOs: 331, 332, 333; SEQ ID NOs: 335, 336, 337; SEQ ID NOs: 339, 340, 341; SEQ ID NOs: 343, 344, 345; SEQ ID NOs: 347, 348, 349; SEQ ID NOs: 351, 352, 353; SEQ ID NOs: 355, 356, 357; SEQ ID NOs: 359, 360, 361; SEQ ID NOs: 363, 364, 365; SEQ ID NOs: 367, 368, 369; SEQ ID NOs: 371, 372, 373; SEQ ID NOs: 375, 376, 377; SEQ ID NOs: 379, 380, 381; SEQ ID NOs: 383, 384, 385; SEQ ID NOs: 387, 388, 389; SEQ ID NOs: 391, 392, 393; SEQ ID NOs: 395, 396, 397; SEQ ID NOs: 399, 400, 401; SEQ ID NOs: 403, 404, 405; SEQ ID NOs: 407, 408, 409; SEQ ID NOs: 411, 412, 413; SEQ ID NOs: 415, 416, 417; SEQ ID NOs: 419, 420, 421; SEQ ID NOs: 423, 424, 425; SEQ ID NOs: 427, 428, 429; SEQ ID NOs: 431, 432, 433; SEQ ID NOs: 435, 436, 437; SEQ ID NOs: 439, 440, 441; SEQ ID NOs: 443, 444, 445; SEQ ID NOs: 447, 448, 449; SEQ ID NOs: 451, 452, 453; SEQ ID NOs: 455, 456, 457 or SEQ ID NOs: 459, 460, 461.