Polyomavirus neutralizing antibodies

Abstract

To provide anti-VP1 antibodies, antibody fragments, and their uses for the prevention and treatment of polyoma virus infection and associated diseases.SOLUTION: Antibodies that specifically bind to VP1 of BK virus serotypes I to IV are provided. A method for treating or reducing likelihood of BK virus or JC virus associated disorders is provided. The method comprises administering via injection or infusion to a patient in need an effective amount of the antibody or a pharmaceutical composition, wherein the disorders are nephropathy, BKVAN, hemorrhagic cystitis (HC), progressive multifocal leukoencephalopathy (PML), granule cell neuronopathy (GCN), interstitial kidney disease, ureteral stenosis, vasculitis, colitis, retinitis, meningitis, and immune reconstitution inflammatory syndrome (IRIS).SELECTED DRAWING: None

Description

[Technical Field]

[0001] FIELD OF THE INVENTION The present disclosure relates to anti-VP1 antibodies, antibody fragments, and methods for reducing the likelihood of polyomavirus infection. or their use for treatment.

[0002] Background of the Invention Among the human polyomaviruses, BK virus (BKV) and JC virus (JCV) ) were the first two polyomaviruses identified. These two polyomaviruses affect immunosuppressed patients. isolated from a human and published in the same issue of The Lancet in 1971 (Gardner et al., La Lancet 1971 1:1253-1527, and Padgett et al., Lancet 1971 1:1257-1260). Maviruses are icosahedral, non-enveloped, double-stranded DNA viruses. They have a diameter of They are 40-45 nm in size and contain 88% protein and 12% DNA.

[0003] The BKV genome is a circular double-stranded DNA approximately 5 Kb in length, and is divided into three major sections: the early core The early coding region contains a 3′-coding region, a late coding region, and a non-coding regulatory region. Three regulatory proteins (large tumor antigen [TAg], small tumor antigen [tAg], and It encodes a truncated tumor antigen (truncTAg) and is expressed in newly infected cells. It is the first viral protein expressed and facilitates viral DNA replication, favoring The late coding region is responsible for establishing the cellular environment. The structural proteins (VP1, VP2, and VP3) and agnoproteins are encoded by However, its role during viral replication is less clear. and early and late promoters that drive expression of viral gene products. .

[0004] BKV infects epithelial cells of the kidney, bladder, and ureter (typical persistence sites), tonsillar tissue, and in many different cell types, including lymphocytes (the proposed site of primary infection and dissemination) have been detected (Chatterjee et al., J. Med. Virol. 2000; 60:353-362, Goudsmit et al., J. Med. Virol. 1982; 10:91-99, Heritage et al., J. Med. Virol. 1981; 8:143- 150, Shinohara et al., J. Med. Virol. 1993; 41(4):301-305). The primary cell The surface receptors are gangliosides GT1b, GD1b, and GD3, all of which It has terminal α2,8-linked sialic acid, making it fairly ubiquitous and enabling infection of a wide variety of cell types (Neu et al., PLos Patholog. 2013; 9(10):e1003714 and e1003688, O'Hara et al. (See also, et al., Virus Res. 2014; 189:208-285). Non-enveloped forms of BKV20 The octahedral virion contains 360 proteins arranged in three different viral proteins: 72 pentamers 1 copy of the major viral capsid protein VP1 and associated with each VP1 pentamer 72 copies of minor virus capsid combined with VP2 or VP3 molecules It is composed of the proteins VP2 and VP3. Only VP1 is exposed on the virion surface during entry. Each pentamer contains five low-affinity binding sites for ganglioside receptors. Binding of VP1 pentamers to ganglioside receptors on the surface mediates caveolae-mediated endocytic transport. This initiates internalization via the endoplasmic reticulum pathway, followed by transport of the virus to the endoplasmic reticulum and ultimately to the nucleus. (Tsai and Qian, J. Virol 2010;84(19):9840-9852).

[0005] Infection with human polyomavirus BK (BKV) is ubiquitous in nature and widespread. It is estimated that 80-90% of the population is infected (Knowles WA, Adv. Exp. Med. Biol. 2006;577:19-45). Primary infection most often occurs in childhood (i.e., before age 10). and cause mild, nonspecific, self-limited illness or no symptoms at all. Persistent infection is established in the epithelial cells of the renal tubules, ureters, and bladder and is effectively treated by the immune system. Transient asymptomatic viral shedding in the urine of immunocompetent adults has been reported sporadically. However, immune dysfunction, especially renal metastasis, may occur. Immunosuppression after hematopoietic stem cell transplantation or transplantation can lead to uncontrolled BKV replication and ultimately to pain. A bladder disease called BKV-associated nephropathy (BKVAN) or hemorrhagic cystitis (HC) There is no effective antiviral therapy for BKV, and the current standard of care is immunotherapy. This reduces immune control and increases the risk of acute rejection. Despite the use of preventative measures, up to 10% of kidney transplant recipients develop BKVAN. BKVAN occurs, and 15-30% of these patients suffer graft loss due to BKVAN. Of these experiencing reductions in immunosuppressive regimens upon detection of KV viremia , up to 30% experience an acute rejection episode as a result.

[0006] BKV was first described in 1971 (see above), but until the 1990s, BK-related Nephropathia Purigh's disease (BKVAN) has not been reported in the literature as a cause of kidney transplant injury. alla et al., Am. J. Kidney Dis. 1995; 26:671-673 and Randhawa et al., Transplan tation 1999; 67:103-109). In the initial management of BKVAN, a positive BK test is considered a serious consequence. The results were favorable, with over 50% of patients experiencing graft dysfunction and graft loss (Hirsch et al., 2004). (Eds. et al., New Engl. J. Med. 2002; 347:488-496). BK virus reactivation occurs after transplantation. It can begin in approximately 30% to 50% of patients by 3 months after transplantation ( Bressollette-Bodin et al., Am J. Transplant. 2005; 5(8):1926-1933 and Brennan et al. (t al., Am. J. Transplant. 2004;4(12):2132-2134). BK virus reactivation initially is detected by the virus and viral DNA in the urine, then in the plasma, and finally in the kidneys. (Brennan et al., Am. J. Transplant. 2005;5(3):582-594 and Hirsch et al., N Eng. J. Med. 2002;347(7):488-496). Approximately 80% of kidney transplant patients have BK virus in their urine. (BK viruria), and 5-10% of these patients progress to BKVAN (Bine t et al., Transplantation 1999;67(6):918-922 and Bressollette-Bodin et al., Am J. Transplant. 2005; 5(8):1926-1933). BKV attacks the renal tubular epithelial cells, exposing the basement membrane. It causes necrosis and lytic destruction of the alveoli, leading to accumulation of luminal fluid in the interstitium and interstitial fibrosis and This leads to tubular atrophy (Nickeleit et al., J. Am. Soc. Neprol. 1999; 10(5):1080-108 9), all of which can affect the condition of the graft. Patients may experience decreased renal function, tubulo-interstitial Nephritis and ureteral strictures may occur (Garner et al., Lancet 1971; 1(7712):1253-1257 and and Hirsch Am. J. Transplant 2002; 2(1)25-30).

[0007] BKV can also cause pneumonia, retinitis, and meningoencephalitis in immunocompromised hosts (Repl oeg et al., Clin. Infect. Dis. 2001;33(2):191-202). Hematopoietic stem cell transplantation (HSCT) BKV disease in recipients typically manifests as hemorrhagic cystitis (HC), which can be severe. May vary in severity. Viruria (but not viremia) and painful Hematuria is associated with the clinical presentation of HC. The current standard of care is primarily forced fluid replacement / diuresis and In the most severe cases, treatment is supportive in nature, including blood transfusions, clot removal, and pain management measures. may require medical attention and in some cases may result in death. HC of rhesus (HV) is relatively common among HSCT recipients, but is associated with BKV. HC usually occurs in approximately 10-12% of patients within 6 months of transplantation. viral etiology, and adenoviruses are associated with a higher incidence of pediatric HSCT compared with adult HSCT recipients. It is a more common cause of HC among CT recipients. BK virus also causes systemic erythropoietin. in other immunocompromised states such as erythroderma, other solid organ transplants, and in patients with HIV / AIDS This has also been observed in other pathogenic bacteria (Jiang et al., Virol. 2009; 384:266-273).

[0008] In this regard, treatment of BK nephropathy associated with organ transplantation is important for preventing graft dysfunction and transplant failure. The reduction of immunosuppression in an attempt to prevent fragment loss (Wiseman et al., Am. J. Kidney Dis. 2009; 54(1): 131-142 and Hirsch et al., Transplantation 2005; 79(1): 1277- 1286). Reduction of immunosuppression reduces the progression from viremia to the widespread damage associated with clinical nephropathy. There is no fixed clinical regimen for reduction, which may help prevent progression However, this also increases the risk of acute organ rejection (Brennan et al., Am. J. Transplant 20 05; 5(3):582-594). Physicians recommend the use of cidofovir, leflunomide, and fluconazole in combination with reduced immunosuppressant medication. reported the use of therapeutic agents such as phenom- ilamides or quinolones, but this report has led to the invalidation of this approach. The effectiveness of these drugs is high, and the burden of managing additional side effects is increased (Randhawa and Brennan Am. J. Transplant 2006; 6(9):2000-2005). The present invention relates to therapeutics that neutralize human HIV and can be used in immunocompromised hosts. There is a useful unmet need.

[0009] JC virus is also a highly prevalent (80%) polyomavirus in the population However, JC virus is generally acquired later than BK virus (Padgett et al. al., J. Infect. Dis. 1973;127(4):467-470 and Sabath et al., J. Infect. Dis. 20 02; 186 Suppl. 2:5180-5186). After initial infection, JC virus infects lymphoid organs and kidneys. establish latency and, if reactivated, enter the central nervous system via infected B lymphocytes Once in the CNS, the JC virus causes progressive demyelinating central nervous system disease, known as progressive cerebrospinal fluid (JC). PML is a common disease in HIV / AIDS patients. It is most commonly found as a casual infection and has also been reported in immunosuppressed patients (Angs trom et al., Brain 1958; 81(1):93-111 and Garcia-Suarez et al., Am. J. Hematol. 2005; 80(4):271-281). Patients with PML may experience confusion, altered mental status, gait ataxia, and hemiparesis. It presents with focal neurological abnormalities such as total paralysis and quadriparesis, as well as visual changes (Richardson EP ., N. Eng. J. Med. 1961; 265:815-823). Patients with PML have a poor prognosis, and H This is particularly poor in patients with IV / AIDS (Antinori et al., J. Neurovir ol. 2003;9 suppl.1:47-53). This is a therapeutic approach to neutralize polyomaviruses such as JC. This further highlights the valuable unmet need in the field.

[0010] Summary of the Invention The present disclosure provides neutralizing antibodies against human polyomavirus and / or fragments thereof, including BK virus. Antibodies that recognize the VP1 pentamers and / or JC virus and their corresponding VP1 pentamers and and fragments thereof.

[0011] An antibody or an antigen-binding fragment thereof, wherein the antibody or the antigen-binding fragment thereof specifically binds to VP1. .

[0012] The antibody or antigen-binding fragment thereof is specific to VP1 of BK virus serotypes I to IV. In one embodiment, the antibody or antigen-binding fragment thereof binds to 5.0 pM binds to VP1 of BKV serotype I with a binding affinity of 29.0 pM or less It binds to VP1 of BKV serotype II with a binding affinity of 6.0 pM or less and binds to VP1 of BKV serotype III with a binding affinity of 6.0 pM or less. VP1 of BKV serotype IV with a binding affinity of 185.0 pM or less. In another embodiment, the antibody or antigen-binding fragment thereof further binds to VP1 of to JCV VP1 and specific JCV VP1 mutants with binding affinities in the monomolar concentration range Combine.

[0013] An antibody, wherein the antibody or antigen-binding fragment specifically binds to a VP1 of Table 1. In one embodiment, the antibody or antigen-binding fragment thereof binds to two or more VP1s in Table 1. In this embodiment, the antibody or antigen-binding fragment thereof is capable of binding to BKV VP1 serotype I and BKV VP In another embodiment, the antibody or antigen-binding fragment thereof binds to BKV serotype II. In another embodiment, the antibody binds to BKV VP1 serotype I and BKV VP1 serotype III. The antibody or antigen-binding fragment thereof is a BKV VP1 serotype I antibody or a BKV VP1 serotype II antibody. In another embodiment, the antibody or antigen-binding fragment thereof binds to BKV VP1. In another embodiment, the antibody or The antigen-binding fragment binds to BKV VP1 serotype II and BKV VP1 serotype IV. In another embodiment, the antibody or antigen-binding fragment thereof binds to BKV VP1 serotype I. and JCV VP1. In a preferred embodiment, the antibody or antigen-binding fragment thereof The fragment binds to BKV VP1 serotypes I, II, III, and IV. The antigen-binding fragments are directed against BKV VP1 serotypes I, II, III, and IV, as well as J. CV binds to VP1.

[0014] The antibody or antigen-binding fragment binds to the VP1 epitope (SEQ ID NO: 500 or SEQ ID NO: 5 01). In one embodiment, the antibody The antibody or antigen-binding fragment may contain one or more of the amino acids Y169, R170, and K172. or more than one, e.g., by alanine scanning mutagenesis as described herein. For example, it binds to Y169 and R170 as determined by

[0015] the antibody or antigen-binding fragment has the sequence GFTFXNYWMT (SEQ ID NO: 507) wherein X may be any amino acid (Xaa). X may be N (Asn), S (Ser), K (Lys) or Q (Gln).

[0016] An antibody, wherein the antibody or antigen-binding fragment thereof comprises: (i) (a) HCDR1 (CDR-complementarity determining region) of SEQ ID NO: 6, (b) SEQ ID NO: 7 (c) a heavy chain variable region comprising an HCDR2 of SEQ ID NO: 8, an HCDR3 of SEQ ID NO: 1, and (e) LCDR1 of SEQ ID NO: 17, and (f) LCDR2 of SEQ ID NO: 18 the light chain variable region, including R3; (ii) (a) HCDR1 of SEQ ID NO: 26, (b) HCDR2 of SEQ ID NO: 27, (c) (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 28; and (e) an LCDR1 of SEQ ID NO: 36. (e) a light chain variable domain comprising an LCDR2 of SEQ ID NO: 37, and (f) an LCDR3 of SEQ ID NO: 38. region; (iii) (a) HCDR1 of SEQ ID NO: 46, (b) HCDR2 of SEQ ID NO: 47, (c (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 48; and (e) an LCDR1 of SEQ ID NO: 56; (e) an LCDR2 of SEQ ID NO: 57, and (f) an LCDR3 of SEQ ID NO: 58. variable region; (iv) (a) HCDR1 of SEQ ID NO: 66, (b) HCDR2 of SEQ ID NO: 67, (c) (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 68; and (e) an LCDR1 of SEQ ID NO: 76; (e) a light chain variable domain comprising an LCDR2 of SEQ ID NO: 77, and (f) an LCDR3 of SEQ ID NO: 78. region; (v) (a) HCDR1 of SEQ ID NO: 86, (b) HCDR2 of SEQ ID NO: 87, (c) HCDR3 of SEQ ID NO: 88, (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 88; (e) an LCDR1 of SEQ ID NO: 96; (f) a light chain variable region comprising an LCDR2 of SEQ ID NO: 97, and (f) an LCDR3 of SEQ ID NO: 98. area; (vi) (a) HCDR1 of SEQ ID NO: 106, (b) HCDR2 of SEQ ID NO: 107, ( (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 108; and (d) an LCD of SEQ ID NO: 116. R1, (e) LCDR2 of SEQ ID NO: 117, and (f) LCDR3 of SEQ ID NO: 118 a light chain variable region comprising: (vii) (a) HCDR1 of SEQ ID NO: 126, (b) HCDR2 of SEQ ID NO: 127, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 128; and (d) an LC of SEQ ID NO: 136. DR1, (e) LCDR2 of SEQ ID NO: 137, and (f) LCDR3 of SEQ ID NO: 138 a light chain variable region comprising: (viii) (a) HCDR1 of SEQ ID NO: 146, (b) HCDR2 of SEQ ID NO: 147 (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 148; and (d) an L of SEQ ID NO: 156. (e) CDR1 of SEQ ID NO: 157, and (f) LCDR of SEQ ID NO: 158 a light chain variable region comprising 3; (ix) (a) HCDR1 of SEQ ID NO: 166, (b) HCDR2 of SEQ ID NO: 167, ( (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 168; and (d) an LCD of SEQ ID NO: 176. R1, (e) LCDR2 of SEQ ID NO: 177, and (f) LCDR3 of SEQ ID NO: 178 a light chain variable region comprising: (x) (a) HCDR1 of SEQ ID NO: 186, (b) HCDR2 of SEQ ID NO: 187, (c (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 188; and (e) an LCDR of SEQ ID NO: 196. 1, (e) LCDR2 of SEQ ID NO: 197, and (f) LCDR3 of SEQ ID NO: 198. light chain variable region; (xi) (a) HCDR1 of SEQ ID NO: 206, (b) HCDR2 of SEQ ID NO: 207, ( (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 208; and (d) an LCD of SEQ ID NO: 216. R1, (e) LCDR2 of SEQ ID NO: 217, and (f) LCDR3 of SEQ ID NO: 218 a light chain variable region comprising: (xii) (a) HCDR1 of SEQ ID NO: 226, (b) HCDR2 of SEQ ID NO: 227, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 228; and (d) an LC of SEQ ID NO: 236. (e) LCDR2 of SEQ ID NO: 237, and (f) LCDR3 of SEQ ID NO: 238 a light chain variable region comprising: (xiii) (a) HCDR1 of SEQ ID NO: 246, (b) HCDR2 of SEQ ID NO: 247 (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 248; and (d) an L of SEQ ID NO: 256. (e) CDR1 of SEQ ID NO: 257, and (f) LCDR of SEQ ID NO: 258 a light chain variable region comprising 3; (xiv) (a) HCDR1 of SEQ ID NO: 266, (b) HCDR2 of SEQ ID NO: 267, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 268; and (d) an LC of SEQ ID NO: 276. (e) LCDR2 of SEQ ID NO: 277, and (f) LCDR3 of SEQ ID NO: 278 a light chain variable region comprising: (xv) (a) HCDR1 of SEQ ID NO: 286, (b) HCDR2 of SEQ ID NO: 287, ( (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 288; and (d) an LCD of SEQ ID NO: 296. R1, (e) LCDR2 of SEQ ID NO: 297, and (f) LCDR3 of SEQ ID NO: 298 a light chain variable region comprising: (xvi) (a) HCDR1 of SEQ ID NO: 306, (b) HCDR2 of SEQ ID NO: 307, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 308; and (d) an LC of SEQ ID NO: 314. (e) LCDR2 of SEQ ID NO: 315, and (f) LCDR3 of SEQ ID NO: 316 a light chain variable region comprising: (xvii) (a) HCDR1 of SEQ ID NO: 322, (b) HCDR2 of SEQ ID NO: 323 (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 324; and (d) an L of SEQ ID NO: 332. (e) CDR1 of SEQ ID NO: 333, and (f) LCDR of SEQ ID NO: 334 a light chain variable region comprising 3; (xviii) (a) HCDR1 of SEQ ID NO: 342, (b) HCDR of SEQ ID NO: 343 2, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 344; and (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 349. (e) LCDR1 of SEQ ID NO: 350, and (f) LCDR2 of SEQ ID NO: 351 the light chain variable region, including R3; (xix) (a) HCDR1 of SEQ ID NO: 356, (b) HCDR2 of SEQ ID NO: 357, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 358; and (d) an LC of SEQ ID NO: 363. (e) LCDR2 of SEQ ID NO: 364, and (f) LCDR3 of SEQ ID NO: 365 a light chain variable region comprising: (xx) (a) HCDR1 of SEQ ID NO: 370, (b) HCDR2 of SEQ ID NO: 371, ( (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 372; and (d) an LCD of SEQ ID NO: 377. R1, (e) LCDR2 of SEQ ID NO: 378, and (f) LCDR3 of SEQ ID NO: 379 a light chain variable region comprising: (xxi) (a) HCDR1 of SEQ ID NO: 384, (b) HCDR2 of SEQ ID NO: 385, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 386; and (d) an LC of SEQ ID NO: 391. (e) LCDR2 of SEQ ID NO: 392, and (f) LCDR3 of SEQ ID NO: 393 a light chain variable region comprising: (xxii) (a) HCDR1 of SEQ ID NO: 398, (b) HCDR2 of SEQ ID NO: 399 (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 400; and (d) an L of SEQ ID NO: 405. (e) CDR1 of SEQ ID NO: 406, and (f) LCDR of SEQ ID NO: 407 a light chain variable region comprising 3; (xxiii) (a) HCDR1 of SEQ ID NO: 412, (b) HCDR of SEQ ID NO: 413 2, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 414; and (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 419. (e) LCDR1 of SEQ ID NO: 420, and (f) LCDR2 of SEQ ID NO: 421 the light chain variable region, including R3; (xxiv) (a) HCDR1 of SEQ ID NO: 426, (b) HCDR2 of SEQ ID NO: 427 (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 428; and (d) an L of SEQ ID NO: 433. (e) CDR1 of SEQ ID NO: 434, and (f) LCDR of SEQ ID NO: 435 a light chain variable region comprising 3; (xxv) (a) HCDR1 of SEQ ID NO: 440, (b) HCDR2 of SEQ ID NO: 441, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 442; and (d) an LC of SEQ ID NO: 447. (e) LCDR2 of SEQ ID NO: 448, and (f) LCDR3 of SEQ ID NO: 449 a light chain variable region comprising: (xxvi) (a) HCDR1 of SEQ ID NO: 454, (b) HCDR2 of SEQ ID NO: 455 (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 456; and (d) an L of SEQ ID NO: 461. (e) CDR1 of SEQ ID NO: 462, and (f) LCDR of SEQ ID NO: 463 a light chain variable region comprising 3; (xxvii) (a) HCDR1 of SEQ ID NO: 468, (b) HCDR of SEQ ID NO: 469 2, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 470; and (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 475. (e) LCDR2 of SEQ ID NO: 476, and (f) LCDR of SEQ ID NO: 477 the light chain variable region, including R3; (xxviii) (a) HCDR1 of SEQ ID NO: 482, (b) HCD of SEQ ID NO: 483 R2, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 484; and (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 489. (e) LCDR1 of SEQ ID NO: 490, and (f) LCDR2 of SEQ ID NO: 491. Light chain variable region containing DR3 an antibody,

[0017] An antibody, wherein the antibody or antigen-binding fragment thereof comprises: (i) (a) HCDR1 (CDR-complementarity determining region) of SEQ ID NO: 508; (b) SEQ ID NO: (c) a heavy chain variable region comprising an HCDR2 of SEQ ID NO: 509, (d) an HCDR3 of SEQ ID NO: 510, and (e) LCDR1 of SEQ ID NO: 511, (f) LCDR2 of SEQ ID NO: 512, and (g) the sequence a light chain variable region containing LCDR3 at number 513; (ii) (a) HCDR1 of SEQ ID NO: 514, (b) HCDR2 of SEQ ID NO: 515, ( (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 516; and (d) an LCD of SEQ ID NO: 517. R1, (e) LCDR2 of SEQ ID NO: 518, and (f) LCDR3 of SEQ ID NO: 519 a light chain variable region comprising: (iii) (a) HCDR1 of SEQ ID NO: 520, (b) HCDR2 of SEQ ID NO: 521, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 522; and (d) an LC of SEQ ID NO: 523. (e) LCDR2 of SEQ ID NO: 524, and (f) LCDR3 of SEQ ID NO: 525 a light chain variable region comprising: (iv) (a) HCDR1 of SEQ ID NO: 526, (b) HCDR2 of SEQ ID NO: 527, ( (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 528; and (d) an LCD of SEQ ID NO: 529. R1, (e) LCDR2 of SEQ ID NO: 530, and (f) LCDR3 of SEQ ID NO: 531 a light chain variable region comprising: (v) (a) HCDR1 of SEQ ID NO: 532, (b) HCDR2 of SEQ ID NO: 533, (c (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 534; and (e) an LCDR of SEQ ID NO: 535. 1, (e) LCDR2 of SEQ ID NO: 536, and (f) LCDR3 of SEQ ID NO: 537. light chain variable region; (vi) (a) HCDR1 of SEQ ID NO: 538, (b) HCDR2 of SEQ ID NO: 539, ( (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 540; and (d) an LCD of SEQ ID NO: 541. R1, (e) LCDR2 of SEQ ID NO: 542, and (f) LCDR3 of SEQ ID NO: 543 Light chain variable region containing an antibody,

[0018] At least one amino acid in a CDR is identical to the corresponding CDR of another anti-VP1 antibody in Table 2. The antibody is substituted by the corresponding residue.

[0019] An antibody in which one or two amino acids within the CDRs have been modified, deleted or substituted.

[0020] at least 90, 91, 92 across either the variable heavy chain region or the variable light chain region; An antibody that retains 93, 94, 95, 96, 97, 98 or 99% identity.

[0021] An antibody containing the modifications in Table 3.

[0022] Monoclonal antibodies, chimeric antibodies, humanized antibodies, human engineered antibodies, human antibodies, single-chain antibodies (scFv) or antibody fragment.

[0023] the antibody or antigen-binding fragment thereof (i) a heavy chain variable region (vH) comprising SEQ ID NO: 12 and a light chain variable region comprising SEQ ID NO: 22 (vL); (ii) a heavy chain variable region (vH) comprising SEQ ID NO: 32 and a light chain variable region comprising SEQ ID NO: 42 area(vL); (iii) a heavy chain variable region (vH) comprising SEQ ID NO: 52 and a light chain variable region (vH) comprising SEQ ID NO: 62 area(vL); (iv) a heavy chain variable region (vH) comprising SEQ ID NO: 72 and a light chain variable region comprising SEQ ID NO: 82 area(vL); (v) a heavy chain variable region (vH) comprising SEQ ID NO: 92 and a light chain variable region (vH) comprising SEQ ID NO: 102 area(vL); (vi) a heavy chain variable region (vH) comprising SEQ ID NO: 112 and a light chain variable region (vL) comprising SEQ ID NO: 122; variable region (vL); (vii) a heavy chain variable region (vH) comprising SEQ ID NO: 132 and a light chain comprising SEQ ID NO: 142 variable region (vL); (viii) a heavy chain variable region (vH) comprising SEQ ID NO: 152 and a light chain variable region (vH) comprising SEQ ID NO: 162 chain variable region (vL); (ix) a heavy chain variable region (vH) comprising SEQ ID NO: 172 and a light chain variable region (vH) comprising SEQ ID NO: 182 variable region (vL); (x) a heavy chain variable region (vH) comprising SEQ ID NO: 192 and a light chain variable region (vL) comprising SEQ ID NO: 202 area(vL); (xi) a heavy chain variable region (vH) comprising SEQ ID NO: 212 and a light chain variable region (vH) comprising SEQ ID NO: 222 variable region (vL); (xii) a heavy chain variable region (vH) comprising SEQ ID NO: 232 and a light chain comprising SEQ ID NO: 242 variable region (vL); (xiii) a heavy chain variable region (vH) comprising SEQ ID NO: 252 and a light chain variable region (vH) comprising SEQ ID NO: 262 chain variable region (vL); (xiv) a heavy chain variable region (vH) comprising SEQ ID NO: 272 and a light chain comprising SEQ ID NO: 282 variable region (vL); (xv) a heavy chain variable region (vH) comprising SEQ ID NO: 292 and a light chain variable region (vL) comprising SEQ ID NO: 302 variable region (vL); (xvi) a heavy chain variable region (vH) comprising SEQ ID NO: 312 and a light chain comprising SEQ ID NO: 320 variable region (vL); (xvii) a heavy chain variable region (vH) comprising SEQ ID NO: 328 and a light chain variable region (vH) comprising SEQ ID NO: 338 chain variable region (vL); (xviii) a heavy chain variable region (vH) comprising SEQ ID NO: 348 and a heavy chain variable region (vH) comprising SEQ ID NO: 355 Light chain variable region (vL); (xix) a heavy chain variable region (vH) comprising SEQ ID NO: 362 and a light chain comprising SEQ ID NO: 369 variable region (vL); (xx) a heavy chain variable region (vH) comprising SEQ ID NO: 376 and a light chain variable region (vH) comprising SEQ ID NO: 383 variable region (vL); (xxi) a heavy chain variable region (vH) comprising SEQ ID NO: 390 and a light chain comprising SEQ ID NO: 397 variable region (vL); (xxii) a heavy chain variable region (vH) comprising SEQ ID NO: 404 and a light chain variable region (vH) comprising SEQ ID NO: 411 chain variable region (vL); (xxiii) a heavy chain variable region (vH) comprising SEQ ID NO: 418 and a heavy chain variable region (vH) comprising SEQ ID NO: 425 Light chain variable region (vL); (xxiv) a heavy chain variable region (vH) comprising SEQ ID NO: 432 and a light chain variable region (vH) comprising SEQ ID NO: 439 chain variable region (vL); (xxv) a heavy chain variable region (vH) comprising SEQ ID NO: 446 and a light chain comprising SEQ ID NO: 453 variable region (vL); (xxvi) a heavy chain variable region (vH) comprising SEQ ID NO: 460 and a light chain variable region (vH) comprising SEQ ID NO: 467 chain variable region (vL); (xxvii) a heavy chain variable region (vH) comprising SEQ ID NO: 474 and a heavy chain variable region (vH) comprising SEQ ID NO: 481 a light chain variable region (vL); or (xxviii) a heavy chain variable region (vH) comprising SEQ ID NO: 488 and a heavy chain variable region (vH) comprising SEQ ID NO: 495 Light chain variable region (vL) an antibody,

[0024] at least 90, 91, 92 across either the variable light chain region or the variable heavy chain region; An antibody that retains 93, 94, 95, 96, 97, 98 or 99% identity.

[0025] 1, 2, 3, 4, or 5, but less than 10, in the variable light chain region or variable heavy chain region an antibody in which amino acids of the

[0026] Monoclonal antibodies, chimeric antibodies, humanized antibodies, human engineered antibodies, human antibodies, single-chain antibodies (scFv) or antibody fragment.

[0027] Antibodies or fragments thereof may have reduced or no glycosylation. The antibody of any preceding embodiment, wherein the antibody is absent or hypofucosylated.

[0028] A composition comprising a plurality of antibodies or antigen-binding fragments thereof according to any of the preceding embodiments. Therefore, at least 0.05%, 0.1%, 0.5%, 1%, 2%, 3% of the antibody in the composition 5% or more of the sialic acid residues are α2,3-linked, (i) (a) HCDR1 (CDR - complementarity determining region) of SEQ ID NO: 6; (b) SEQ ID NO: (c) a heavy chain variable region comprising an HCDR2 of SEQ ID NO: 7, (d) an HCDR3 of SEQ ID NO: 8, and (e) LCDR1 of SEQ ID NO: 16, (f) LCDR2 of SEQ ID NO: 17, and (g) LCDR3 of SEQ ID NO: 18. the light chain variable region including CDR3; (ii) (a) HCDR1 of SEQ ID NO: 26, (b) HCDR2 of SEQ ID NO: 27, (c) (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 28; and (e) an LCDR1 of SEQ ID NO: 36. (e) a light chain variable domain comprising an LCDR2 of SEQ ID NO: 37, and (f) an LCDR3 of SEQ ID NO: 38. region; (iii) (a) HCDR1 of SEQ ID NO: 46, (b) HCDR2 of SEQ ID NO: 47, (c (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 48; and (e) an LCDR1 of SEQ ID NO: 56; (e) an LCDR2 of SEQ ID NO: 57, and (f) an LCDR3 of SEQ ID NO: 58. variable region; (iv) (a) HCDR1 of SEQ ID NO: 66, (b) HCDR2 of SEQ ID NO: 67, (c) (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 68; and (e) an LCDR1 of SEQ ID NO: 76; (e) a light chain variable domain comprising an LCDR2 of SEQ ID NO: 77, and (f) an LCDR3 of SEQ ID NO: 78. region; (v) (a) HCDR1 of SEQ ID NO: 86, (b) HCDR2 of SEQ ID NO: 87, (c) HCDR3 of SEQ ID NO: 88, (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 88; (e) an LCDR1 of SEQ ID NO: 96; (f) a light chain variable region comprising an LCDR2 of SEQ ID NO: 97, and (f) an LCDR3 of SEQ ID NO: 98. area; (vi) (a) HCDR1 of SEQ ID NO: 106, (b) HCDR2 of SEQ ID NO: 107, ( (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 108; and (d) an LCD of SEQ ID NO: 116. R1, (e) LCDR2 of SEQ ID NO: 117, and (f) LCDR3 of SEQ ID NO: 118 a light chain variable region comprising: (vii) (a) HCDR1 of SEQ ID NO: 126, (b) HCDR2 of SEQ ID NO: 127, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 128; and (d) an LC of SEQ ID NO: 136. DR1, (e) LCDR2 of SEQ ID NO: 137, and (f) LCDR3 of SEQ ID NO: 138 a light chain variable region comprising: (viii) (a) HCDR1 of SEQ ID NO: 146, (b) HCDR2 of SEQ ID NO: 147 (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 148; and (d) an L of SEQ ID NO: 156. (e) CDR1 of SEQ ID NO: 157, and (f) LCDR of SEQ ID NO: 158 a light chain variable region comprising 3; (ix) (a) HCDR1 of SEQ ID NO: 166, (b) HCDR2 of SEQ ID NO: 167, ( (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 168; and (d) an LCD of SEQ ID NO: 176. R1, (e) LCDR2 of SEQ ID NO: 177, and (f) LCDR3 of SEQ ID NO: 178 a light chain variable region comprising: (x) (a) HCDR1 of SEQ ID NO: 186, (b) HCDR2 of SEQ ID NO: 187, (c (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 188; and (e) an LCDR of SEQ ID NO: 196. 1, (e) LCDR2 of SEQ ID NO: 197, and (f) LCDR3 of SEQ ID NO: 198. light chain variable region; (xi) (a) HCDR1 of SEQ ID NO: 206, (b) HCDR2 of SEQ ID NO: 207, ( (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 208; and (d) an LCD of SEQ ID NO: 216. R1, (e) LCDR2 of SEQ ID NO: 217, and (f) LCDR3 of SEQ ID NO: 218 a light chain variable region comprising: (xii) (a) HCDR1 of SEQ ID NO: 226, (b) HCDR2 of SEQ ID NO: 227, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 228; and (d) an LC of SEQ ID NO: 236. (e) LCDR2 of SEQ ID NO: 237, and (f) LCDR3 of SEQ ID NO: 238 a light chain variable region comprising: (xiii) (a) HCDR1 of SEQ ID NO: 246, (b) HCDR2 of SEQ ID NO: 247 (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 248; and (d) an L of SEQ ID NO: 256. (e) CDR1 of SEQ ID NO: 257, and (f) LCDR of SEQ ID NO: 258 a light chain variable region comprising 3; (xiv) (a) HCDR1 of SEQ ID NO: 266, (b) HCDR2 of SEQ ID NO: 267, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 268; and (d) an LC of SEQ ID NO: 276. (e) LCDR2 of SEQ ID NO: 277, and (f) LCDR3 of SEQ ID NO: 278 a light chain variable region comprising: (xv) (a) HCDR1 of SEQ ID NO: 286, (b) HCDR2 of SEQ ID NO: 287, ( (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 288; and (d) an LCD of SEQ ID NO: 296. R1, (e) LCDR2 of SEQ ID NO: 297, and (f) LCDR3 of SEQ ID NO: 298 a light chain variable region comprising: (xvi) (a) HCDR1 of SEQ ID NO: 306, (b) HCDR2 of SEQ ID NO: 307, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 308; and (d) an LC of SEQ ID NO: 314. (e) LCDR2 of SEQ ID NO: 315, and (f) LCDR3 of SEQ ID NO: 316 a light chain variable region comprising: (xvii) (a) HCDR1 of SEQ ID NO: 322, (b) HCDR2 of SEQ ID NO: 323 (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 324; and (d) an L of SEQ ID NO: 332. (e) CDR1 of SEQ ID NO: 333, and (f) LCDR of SEQ ID NO: 334 a light chain variable region comprising 3; (xviii) (a) HCDR1 of SEQ ID NO: 342, (b) HCDR of SEQ ID NO: 343 2, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 344; and (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 349. (e) LCDR1 of SEQ ID NO: 350, and (f) LCDR2 of SEQ ID NO: 351 the light chain variable region, including R3; (xix) (a) HCDR1 of SEQ ID NO: 356, (b) HCDR2 of SEQ ID NO: 357, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 358; and (d) an LC of SEQ ID NO: 363. (e) LCDR2 of SEQ ID NO: 364, and (f) LCDR3 of SEQ ID NO: 365 a light chain variable region comprising: (xx) (a) HCDR1 of SEQ ID NO: 370, (b) HCDR2 of SEQ ID NO: 371, ( (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 372; and (d) an LCD of SEQ ID NO: 377. R1, (e) LCDR2 of SEQ ID NO: 378, and (f) LCDR3 of SEQ ID NO: 379 a light chain variable region comprising: (xxi) (a) HCDR1 of SEQ ID NO: 384, (b) HCDR2 of SEQ ID NO: 385, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 386; and (d) an LC of SEQ ID NO: 391. (e) LCDR2 of SEQ ID NO: 392, and (f) LCDR3 of SEQ ID NO: 393 a light chain variable region comprising: (xxii) (a) HCDR1 of SEQ ID NO: 398, (b) HCDR2 of SEQ ID NO: 399 (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 400; and (d) an L of SEQ ID NO: 405. (e) CDR1 of SEQ ID NO: 406, and (f) LCDR of SEQ ID NO: 407 a light chain variable region comprising 3; (xxiii) (a) HCDR1 of SEQ ID NO: 412, (b) HCDR of SEQ ID NO: 413 2, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 414; and (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 419. (e) LCDR1 of SEQ ID NO: 420, and (f) LCDR2 of SEQ ID NO: 421 the light chain variable region, including R3; (xxiv) (a) HCDR1 of SEQ ID NO: 426, (b) HCDR2 of SEQ ID NO: 427 (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 428; and (d) an L of SEQ ID NO: 433. (e) CDR1 of SEQ ID NO: 434, and (f) LCDR of SEQ ID NO: 435 a light chain variable region comprising 3; (xxv) (a) HCDR1 of SEQ ID NO: 440, (b) HCDR2 of SEQ ID NO: 441, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 442; and (d) an LC of SEQ ID NO: 447. (e) LCDR2 of SEQ ID NO: 448, and (f) LCDR3 of SEQ ID NO: 449 a light chain variable region comprising: (xxvi) (a) HCDR1 of SEQ ID NO: 454, (b) HCDR2 of SEQ ID NO: 455 (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 456; and (d) an L of SEQ ID NO: 461. (e) CDR1 of SEQ ID NO: 462, and (f) LCDR of SEQ ID NO: 463 a light chain variable region comprising 3; (xxvii) (a) HCDR1 of SEQ ID NO: 468, (b) HCDR of SEQ ID NO: 469 2, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 470; and (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 475. (e) LCDR2 of SEQ ID NO: 476, and (f) LCDR of SEQ ID NO: 477 the light chain variable region, including R3; (xxviii) (a) HCDR1 of SEQ ID NO: 482, (b) HCD of SEQ ID NO: 483 R2, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 484; and (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 489. (e) LCDR1 of SEQ ID NO: 490, and (f) LCDR2 of SEQ ID NO: 491. Light chain variable region containing DR3 A composition comprising:

[0029] A composition comprising a plurality of antibodies or antigen-binding fragments according to any of the preceding embodiments. , wherein none of the antibodies contains a bisecting GlcNAc.

[0030] The antibody or composition according to any of the preceding embodiments, wherein the composition is prepared as a lyophilizate. A pharmaceutical composition comprising a fragment of

[0031] The antibody or fragment thereof of any of the preceding embodiments and a pharmaceutically acceptable carrier. In one embodiment, the carrier is a histidine buffer. In some cases, the pharmaceutical composition contains a sugar (e.g., sucrose).

[0032] An effective amount of the antibody or pharmaceutical composition is administered to a patient in need thereof by injection or infusion. and (c) neutralizing BK virus or JC virus infection. In another embodiment, the synthetic fragment neutralizes BKV serotype I and BKV serotype II. The antibody or antigen-binding fragment thereof neutralizes BKV serotype I and BKV serotype III. In another embodiment, the antibody or antigen-binding fragment thereof is BKV serotype I and BKV In another embodiment, the antibody or antigen-binding fragment thereof neutralizes BKV serotype IV. In another embodiment, the antibody or its antigen neutralizes BKV serotype II and BKV serotype III. The binding fragment neutralizes BKV serotype II and BKV serotype IV. The antibody or antigen-binding fragment thereof neutralizes BKV serotype I and JCV. In embodiments, the antibody or antigen-binding fragment thereof is directed against BKV serotypes I, II, III and I. Furthermore, the antibody or antigen-binding fragment thereof neutralizes BKV serotypes I, II, and II. In a preferred embodiment, the anti-VP1 antibody neutralizes BK, I, IV, and JCV. These anti-VP1 antibodies neutralized infection with all four serotypes (I-IV) of V. In particular, these include P8D11, modifications of P8D11, and EBB-C1975-B5.

[0033] An effective amount of the antibody or pharmaceutical composition is administered to a patient in need thereof by injection or infusion. and (iii) treating or potentially treating disorders associated with BK virus or JC virus, including A method for reducing the risk of urinary tract infection, wherein the disorder is nephropathy, BKVAN, hemorrhagic cystitis (HC), , progressive multifocal leukoencephalopathy (PML), granular cell neuropathy (GCN), interstitial kidney disease , ureteral stenosis, vasculitis, colitis, retinitis, meningitis, and immune reconstitution inflammatory syndrome (IRIS). There is a way.

[0034] The method wherein the antibody or composition is reconstituted prior to injection or infusion.

[0035] The method wherein the antibody or pharmaceutical composition is administered in combination with another therapeutic agent.

[0036] The method wherein said therapeutic agent is an immunosuppressant.

[0037] The immunosuppressant is selected from the group consisting of a monophosphate dehydrogenase inhibitor, a purine synthesis inhibitor, and a carboxylase inhibitor. The method is a lucineurin inhibitor or an mTOR inhibitor.

[0038] Immunosuppressants include mycophenolate mofetil (MMF), mycophenolate sodium, azathioprine, tacrolimus, sirolimus or cyclosporine.

[0039] The method wherein the therapeutic agent is an additional anti-VP1 antibody.

[0040] 2. The antibody or fragment thereof of any of the preceding embodiments for use as a medicament.

[0041] Antibodies or their derivatives for use in neutralizing BK virus or JC virus infection Fragments or pharmaceutical compositions.

[0042] Nephropathy, BKVAN, hemorrhagic cystitis (HC), progressive multifocal leukoencephalopathy (PML) , granular cell neuropathy (GCN), interstitial kidney disease, ureteral stenosis, vasculitis, colitis, retina in the treatment or reduction of the likelihood of inflammatory bowel disease, meningitis, and immune reconstitution inflammatory syndrome (IRIS) An antibody or fragment thereof or a pharmaceutical composition for use in

[0043] The use of an antibody or fragment thereof administered in combination with another therapeutic agent.

[0044] The use of an antibody or fragment thereof, wherein said therapeutic agent is an immunosuppressant.

[0045] The immunosuppressant is a monophosphate dehydrogenase inhibitor, a purine synthesis inhibitor, a calcitonin inhibitor, The use of an antibody or fragment thereof that is a syneurin inhibitor or an mTOR inhibitor.

[0046] The immunosuppressant is mycophenolate mofetil (MMF), mycophenolate sodium antibody or cyclosporine, which is cyclosporine, azathioprine, tacrolimus, sirolimus, or cyclosporine is the use of that fragment.

[0047] A nucleic acid encoding the antibody or antigen-binding fragment of any of the preceding embodiments.

[0048] A vector containing a nucleic acid.

[0049] A host cell containing a vector.

[0050] Culturing the host cells and recovering the antibody from the culture. A method for generating

[0051] A diagnostic agent comprising a labeled antibody or an antigen-binding fragment thereof.

[0052] Labels consist of radiolabels, fluorophores, chromophores, imaging agents, and metal ions A diagnostic agent selected from the group:

[0053] definition Unless otherwise stated, the following terms and phrases used herein have the following meanings: It is intended to.

[0054] As used herein, the term "antibody" refers to a molecule that binds to a target molecule in a non-covalent, reversible, and specific manner. It refers to a polypeptide of the immunoglobulin family that is capable of binding to a corresponding antigen. For example, a natural IgG antibody contains at least two amino acids interconnected by disulfide bonds. It is a tetramer containing a heavy chain (H) and two light chains (L). Each heavy chain contains a heavy chain variable region ( The heavy chain constant region consists of three domains (abbreviated as VH in the literature) and a heavy chain constant region. Each light chain is composed of a light chain variable region (see the description). The light chain constant region consists of one domain (abbreviated as VL in the literature) and a light chain constant region. The VH and VL regions are called framework regions (FR). hypervariable regions called complementarity-determining regions (CDRs) interspersed with more conserved regions Each VH and VL can be further subdivided into domains from the amino terminus to the carboxy terminus. In the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, and The heavy and light chains are composed of three CDRs and four FRs, which are located in FR1 and FR2. The variable region contains the binding domain that interacts with an antigen. The constant region of an antibody is responsible for the functions of the immune system. Various cells (e.g., effector cells) and the first component of the classical complement system (C1q) These antibodies can mediate the binding of immunoglobulins to host tissues or factors.

[0055] The term "antibody" includes, but is not limited to, monoclonal antibodies, human antibodies, humanized antibodies, and the like. antibodies, camelid antibodies, chimeric antibodies, and anti-idiotypic (anti-Id) antibodies (e.g., Antibodies of any isotype / class (e.g., anti-Id antibodies to the disclosed antibodies) may be used. IgG, IgE, IgM, IgD, IgA and IgY), or subclass (e.g. For example, IgG1, IgG2, IgG3, IgG4, IgA1 and IgA2). It is also possible.

[0056] "Complementarity-determining domains" or "complementarity-determining regions ("CDRs")" refer to the VL and VH The CDRs are interchangeably referred to as the hypervariable regions of a target protein. Each human VL or VH contains three CDs. CDRs (CDRs 1 to 3, numbered consecutively from the N-terminus) are present in approximately 1 CDRs can be described by their region and order. For example, "VHCDR1" or "HCDR1" both refer to the first CDR of a heavy chain variable region. The CDRs are structurally complementary to the epitope of the target protein and thus provide binding specificity. The remaining stretches of the VL or VH, the so-called framework regions, are directly responsible for the isomerism. They show little change in amino acid sequence (Kuby, Immunology, 4th ed., Chapter 4. WH F Reeman & Co., New York, 2000).

[0057] The positions of the CDRs and framework regions may be determined according to various well-known definitions in the art, for example, abat, Chothia, and AbM (e.g., Johnson et al., Nucleic Acids Re s., 29:205-206 (2001); Chothia and Lesk, J. Mol. Biol., 196:901-917 (1987); Chot hia et al., Nature, 342:877-883 (1989); Chothia et al., J. Mol. Biol., 227:799-8 17 (1992); Al-Lazikani et al., J. Mol. Biol., 273:927-748 (1997)). The definition of an antigen-binding site can be found in Ruiz et al. et al., Nucleic Acids Res., 28:219-221 (2000); and Lefranc, MP, Nucleic Acids Res. ds Res., 29:207-209 (2001); MacCallum et al., J. Mol. Biol., 262:732-745 (1996); and Martin et al., Proc. Natl. Acad. Sci. USA, 86:9268-9272 (1989); Martin et al., Methods Enzymol., 203:121-153 (1991); and Rees et al., In Sternberg MJ E. (ed.), Protein Structure Prediction, Oxford University Press, Oxford, 141-172. (1996). In some embodiments, a combined Kabat and Chothia numbering scheme is used. In this group, the CDRs may be the Kabat CDRs, the Chothia CDRs, or both. For example, in some embodiments, the CDRs correspond to amino acid residues that are part of the VH For example, amino acid residues 26 to 35 (HC CDRs) in a mammalian VH, for example, a human VH 1), 50-65 (HC CDR2), and 95-102 (HC CDR3); and to amino acid residues 24 to 34 (LC CDR1), 50-56 (LC CDR2), and 89-97 (LC CDR3) Respond.

[0058] Both the light and heavy chains are divided into regions of structural and functional homology. and "variable" is used functionally. In this context, the light chain (VL) and heavy chain (VH) portions It is understood that both variable domain portions of the Conversely, the constant domains of the light chain (CL) and heavy chain (CH1, CH2, or CH3) are They confer important biological properties such as secretion, transplacental transfer, Fc receptor binding, and complement fixation. The numbering of constant region domains determines whether they are located at the antigen binding site or the amino terminus of the antibody. The N-terminus is the variable region, and the C-terminus is the constant region. CH3 and CL domains are the carboxy-terminal domains of the heavy and light chains, respectively. Actually includes.

[0059] As used herein, the term "antigen-binding fragment" refers to a fragment that specifically interacts with an epitope of an antigen. retain the ability to act (e.g., by binding, steric hindrance, stabilization / destabilization, spatial distribution) A binding fragment refers to one or more portions of an antibody that binds to the antibody. Examples of binding fragments include, but are not limited to, However, single-chain Fv (scFv), disulfide-linked Fv (sdFv), Fab fragments, F (ab') fragment, a monovalent fragment consisting of the VL, VH, CL, and CH1 domains; F(ab) 2 fragments, bivalent fragments containing two Fab fragments linked by a disulfide bridge at the hinge region a fragment consisting of the VH and CH1 domains; a fragment consisting of the VL and VH domains of a single arm of an antibody; Fv fragment consisting of the main domain; dAb fragment consisting of the VH domain (Ward et al., Nature 341: 544-546, 1989); and isolated complementarity determining regions (CDRs), or other end portions of antibodies. Examples include pitope-binding fragments.

[0060] Furthermore, the two domains of the Fv fragment, VL and VH, are encoded by separate genes. However, they can be engineered using recombinant methods to pair the VL and VH regions into monovalent molecules (monovalent molecules). Synthetic Fvs can be made as a single protein chain to form single-chain Fvs ("scFvs"). They can be linked by a linker; see, for example, Bird et al., Science 242:423-426, 1999. 88; and Huston et al., Proc. Natl. Acad. Sci. 85:5879-5883, 1988. Such single chain antibodies are also intended to be encompassed within the term "antigen-binding fragment." These antigen-binding fragments can be obtained using conventional techniques known to those skilled in the art, and the fragments can be They are screened for utility in the same manner as intact antibodies.

[0061] Antigen-binding fragments can be expressed as single domain antibodies, maxibodies, minibodies, nanobodies, Tribodies, diabodies, triabodies, tetrabodies, v-NAR and bis-sc Fv (see, e.g., Hollinger and Hudson, Nature Biotechnology 23:1126-1136, 2005) The antigen-binding fragment can also be incorporated into a type III fibronectin. (Fn3) can be implanted into a scaffold based on polypeptides such as fibronectin. See U.S. Patent No. 6,703,199, which describes polypeptide monobodies. sea ​​bream).

[0062] The antigen-binding fragment is combined with a complementary light chain polypeptide to form a pair of antigen-binding regions. It can be incorporated into a single chain molecule containing a pair of Fv segments (VH-CH1-VH-CH1). (Zapata et al., Protein Eng. 8:1057-1062, 1995; and U.S. Pat. No. 5,644,644. 1,870 specification).

[0063] As used herein, the term "monoclonal antibody" or "monoclonal antibody composition" refers to a " refers to antibodies that have substantially identical amino acid sequences or are derived from the same genetic source. The term also refers to polypeptides such as antibodies and antigen-binding fragments of a single molecule. Monoclonal antibody compositions also include preparations of antibody molecules. Binding specificity and affinity are shown.

[0064] As used herein, the term "human antibody" refers to an antibody in which both the framework and CDR regions are humanized. The present invention also includes antibodies having variable regions derived from sequences of human origin. If contained, the constant region may also contain such human sequences, e.g., human germline sequences, or are variants of the human germline sequence or, for example, Knappik et al., J. Mol. Biol. 296:57-86, Consensus framework sequences derived from human framework sequence analysis, as described in 2000. It is derived from an antibody containing a framework sequence.

[0065] The human antibodies of the present disclosure may include amino acid residues not encoded by human sequences (e.g., For example, by random or site-directed mutagenesis in vitro or in Conservative substitutions to enhance stability or production, or by somatic mutation in vivo Mutations introduced by recombination).

[0066] As used herein, the term "recognize" refers to an epitope, whether linear or conformational. Also, antibodies or other molecules that discover and interact with (e.g., bind to) the epitope may be used. The term "epitope" refers to an antigen-binding fragment of an antibody or antigen-binding fragment of the present disclosure. The term "epitope" refers to the site on an antigen that specifically binds to a protein. It can be formed both from juxtaposed contiguous amino acids or from non-contiguous amino acids. Epitopes formed from consecutive amino acids are typically preserved upon exposure to denaturing solvents. epitopes formed by tertiary folding are typically denatured by treatment with denaturing solvents. Epitopes are typically found in unique spatial conformations. At least 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14 or 15 Methods for determining the spatial conformation of epitopes are well known in the art and include amino acids. techniques, such as x-ray crystallography and two-dimensional nuclear magnetic resonance (e.g., Epitope Mapping) g Protocols in Methods in Molecular Biology, Vol. 66, GE Morris, Ed. (1996). (See, e.g., ). A "paratope" is the portion of an antibody that recognizes an epitope of an antigen.

[0067] Interaction between an antigen (e.g., a protein) and an antibody, antibody fragment, or antibody-derived binding agent The phrases "specifically bind" or "selectively bind" when used in a context describing a use "To identify" refers to the identification of, for example, a biological sample, e.g., blood, in a heterogeneous population of proteins and other biologics. It refers to a binding reaction that determines the presence of an antigen in a fluid, serum, plasma, or tissue sample. Under certain specified immunoassay conditions, an antibody or The binding agent binds to the specific antigen at least twice as much as background and is present in the sample. does not substantially bind to other antigens in any significant amount. In one aspect, under designated immunoassay conditions , an antibody or binder with a particular binding specificity is at least 10-fold higher than background The antibody binds to a specific antigen and does not substantially bind in significant amounts to other antigens present in the sample. Specific binding to an antibody or binder under such conditions indicates its specificity for a particular protein. It may be necessary to select antibodies or agents for specificity, if desired or appropriate. If necessary, this selection can be extended to other species (e.g., mouse or rat) or other subtypes. This can be achieved by removing antibodies that cross-react with the molecule from which it is derived. In some embodiments, antibodies or antibody fragments that cross-react with a particular desired molecule are selected. do.

[0068] As used herein, the term "affinity" refers to the interaction of an antibody with an antigen at a single antigenic site. Within each antigen site, the variable regions of the antibody "arms" bind to each other through weak non-covalent bonds. interacts with the antigen at several sites via Become stronger.

[0069] The term "isolated antibody" refers to an antibody that is substantially free of other antibodies having different antigen specificities. However, an isolated antibody that specifically binds to one antigen may not bind to other antigens. Additionally, the isolated antibody may have cross-reactivity with other cellular material and / or may be substantially free of chemicals.

[0070] The term "corresponding human germline sequence" refers to a human germline immunoglobulin variable region sequence. The reference variable region amino acid sequence is compared to all other known variable region amino acid sequences encoded by The human variable region with the highest determined amino acid sequence identity to the amino acid sequence or subsequence refers to a nucleic acid sequence that encodes an amino acid sequence or subsequence. The corresponding human germline sequence is Additionally, the reference variable region amino acid sequence was compared to all other evaluated variable region amino acid sequences. the human variable region amino acid sequence or subsequence with the highest amino acid sequence identity The corresponding human germline sequence may refer to a subsequence of the framework region only. Complementarity determining regions only, framework regions and complementarity determining regions, variable segments (as defined above) or other combinations of sequences or subsequences comprising the variable regions. For example, BLAST, ALIGN, or another alignment algorithm known in the art. The two sequences are aligned using the algorithm described herein, and sequence identity is determined using the methods described herein. The corresponding human germline nucleic acid or amino acid sequence can be determined by the reference variable region. At least about 90%, 91%, 92%, 93%, 94%, or more of the nucleic acid or amino acid sequence of the target gene may have 95%, 96%, 97%, 98%, 99%, or 100% sequence identity .

[0071] Antibodies that specifically immunoreact with a particular protein using a variety of immunoassay formats For example, to select antibodies that are specifically immunoreactive with a protein, Solid-phase ELISA immunoassays are routinely used to detect specific immune reactions (e.g., For a description of immunoassay formats and conditions that can be used to determine sex, see , Harlow & Lane, Using Antibodies, A Laboratory Manual (1998). Typically, a specific or selective binding reaction will result in a signal that is at least 2x higher than the background signal. fold, more typically yielding a signal at least 10-100 times greater than background vinegar.

[0072] The term "equilibrium dissociation constant (KD, M)" refers to the equilibrium dissociation rate constant (kd, time-1) relative to the binding rate. It refers to division by a constant (ka, time-1, M-1). The equilibrium dissociation constant can be measured using the method. Antibodies of the present disclosure generally have an equilibrium dissociation constant of about 10 - 7 or 10 -8 Less than m, e.g., about 10 -9 M or 10 -10 Less than M, some states In this case, about 10 -11 M, 10 -12 M or 10 -13 have an equilibrium dissociation constant less than M .

[0073] The term "bioavailability" refers to the systemic availability of a given amount of drug administered to a patient. (i.e., blood / plasma levels). Bioavailability refers to the amount of a drug that is absorbed from the administered dosage form. is an absolute term that measures both the time (rate) and total amount (extent) of drug reaching the systemic circulation from .

[0074] As used herein, the phrase "consisting essentially of" refers to a method or composition that The genus or species of the active agent, as well as any inactive agent for the intended purpose of the method or composition. In some embodiments, the phrase "consisting essentially of" refers to any excipient of the present disclosure. The inclusion of one or more additional active agents other than the antibody is expressly excluded. The phrase "consisting essentially of" refers to the inclusion of one or more additional antibodies other than the anti-VP1 antibodies of the present disclosure. The inclusion of an active agent and a second, co-administered agent is expressly excluded.

[0075] The term "amino acid" refers to natural, synthetic, and unnatural amino acids, as well as naturally occurring amino acids. This refers to amino acid analogs and amino acid mimetics that function in a manner similar to natural amino acids. The amino acids include those encoded by the genetic code as well as those that are later modified, e.g. Examples are hydroxyproline, γ-carboxyglutamic acid, and O-phosphoserine. Amino acid analogs are compounds that have the same basic chemical structure as natural amino acids, i.e., A carboxyl group, an amino group, and an R group, such as homoserine, norleucine, methyl It refers to the α-carbon bonded to methionine sulfoxide or methionine methyl sulfonium. Such analogs have modified R groups (e.g., norleucine) or modified peptide backbones. Amino acid mimetics are compounds that mimic the general structure of amino acids but retain the same basic chemical structure as natural amino acids. compounds that have a different chemical structure from the natural amino acids but function in a manner similar to natural amino acids. Point.

[0076] The term "conservatively modified variants" applies to both amino acid and nucleic acid sequences. With respect to a particular nucleic acid sequence, conservatively modified variants are identical or Nucleic acids encoding a qualitatively identical amino acid sequence, or nucleic acids that do not encode an amino acid sequence Because of the degeneracy of the genetic code, there are many functionally identical sequences. The nucleic acid encodes any given protein, e.g., the codons GCA, GCC, GCG and GCU all encode the amino acid alanine. Thus, the codons with alanine At every position specified by, the codon is changed to alter the encoded polypeptide. The nucleic acid may be changed to any of the corresponding codons described without modification. Polypeptides are "silent mutations" and are a type of conservatively modified variant. Every nucleic acid sequence encoding a nucleic acid also describes every possible silent variation of the nucleic acid. Those skilled in the art will recognize each codon in a nucleic acid (usually the only codon for methionine) and TGG, which is usually the only codon for tryptophan. It is recognized that one can obtain functionally identical molecules by altering the polypeptide. Each silent variation of a nucleic acid encoding a peptide is implicit in each described sequence. It is silent.

[0077] For polypeptide sequences, "conservatively modified variants" refer to those variants of an amino acid sequence that: Individual substitutions, deletions, etc. in a polypeptide sequence that result in substitution with chemically similar amino acids. Conservative substitution tables providing functionally similar amino acids are well known in the art. Such conservatively modified variants include polymorphic variants, interspecies homologs, The following eight groups are conservative substitutions for each other. Contains the amino acids: 1) alanine (A), glycine (G); 2) aspartic acid (D) ), glutamic acid (E); 3) asparagine (N), glutamine (Q); 4) arginine (R), lysine (K); 5) isoleucine (I), leucine (L), methionine (M), valine (V); 6) phenylalanine (F), tyrosine (Y), tryptophan (W); 7) serine (S), threonine (T); and 8) cysteine ​​(C), methionine (M). (See, e.g., Creighton, Proteins (1984)). In some embodiments, the term " "Conservative sequence modifications" do not significantly affect or affect the binding characteristics of the antibody containing the amino acid sequence. The term "amino acid modification" is used to refer to an amino acid modification that does not alter the amino acid sequence.

[0078] As used herein, the term "optimized" refers to a generative cell or organism, generally a gene that is optimized for the Nucleic cells, such as yeast cells, Pichia cells, fungal cells, Trichoderma rma cells, Chinese hamster ovary cells (CHO) or human cells. refers to a nucleotide sequence that has been altered to encode an amino acid sequence using new codons. The optimized nucleotide sequence is the starting nucleotide sequence, also known as the "parent" sequence. to retain the entire amino acid sequence originally encoded by the sequence, or as much as possible. Being manipulated.

[0079] The term "percent identical" in the context of two or more nucleic acid or polypeptide sequences Alternatively, "percent identity" refers to the degree to which two or more sequences or subsequences are the same. The two sequences have identical amino acid or nucleotide sequences over the region where they are compared. Two sequences are "identical" if they have the same sequence. Ratios measured using a meter or by manual alignment and visual inspection Compared and aligned for maximum match over a comparison window, or specified region. In this case, a certain percentage of amino acid residues or nucleotides are the same (i.e., a certain percentage of 60% identity over a specified region, or if not specified, over the entire sequence; 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 99% at your discretion. Two sequences are "substantially identical" if they share a common sequence identity. Optionally, the identity can be determined by over a region that is at least about 30 nucleotides (or 10 amino acids) in length, and More preferably, the length is 100 to 500 or 1000 or more nucleotides (or 2 The amino acid sequence is present over a region of 0, 50, 200 or more amino acids.

[0080] For sequence comparison, typically one sequence serves as a reference sequence, to which test sequences are compared. When using a sequence comparison algorithm, test and reference sequences are compared by computer. Enter the subarray coordinates and, if necessary, the array algorithm program parameters. You can use the default program parameters or specify alternative parameters. The sequence comparison algorithm then runs based on the program parameters. The percent sequence identity for the test sequence relative to the reference sequence is then calculated.

[0081] As used herein, a "comparison window" refers to the time period after optimal alignment of two sequences. The sequence can then be compared to a reference sequence at the same number of consecutive positions, typically 20 to 600. Usually, the number of consecutive numbers is about 50 to about 200, more usually about 100 to about 150. The alignment of sequences for comparison includes reference to any one segment of the sequence. Methods for optimal alignment of sequences for comparison are well known in the art. For example, the partial homology algorithm of Smith and Waterman, Adv. Appl. Math. 2:482c (1970) Based on the homology alignment of Needleman and Wunsch, J. Mol. Biol. 48:443 (1970) Algorithm: Pearson and Lipman, Proc. Natl. Acad. Sci. USA 85:2444 (1988) The similarity search method of Wisco nsin Genetics Software Package, Genetics Computer Group, 575 Science Dr., Madison, W. by GAP, BESTFIT, FASTA, and TFASTA in I; or Manual alignment and visual inspection (e.g., Brent et al., Current Protocol This can be done by the method described in Molecular Biology, 2003.

[0082] Two algorithms that are suitable for determining percent sequence identity and sequence similarity are: Two examples are the BLAST and BLAST 2.0 algorithms, which are , Altschul et al., Nuc. Acids Res. 25:3389-3402, 1977; and Altschul et al., J. Mol. Biol. 215:403-410, 1990. Software for performing BLAST analysis. The software is supported by the National Center for Biotechnology This algorithm is publicly available under the Data Protection Act 2000. When aligned with words of the same length in the base sequence, a threshold score T Identifying a short word length W in the query sequence that matches or satisfies This involves first identifying high-scoring sequence pairs (HSPs) by These initial neighborhood word hits are called the score threshold (Altschul et al., supra). Act as seeds to initiate searches to discover longer HSPs that contain them Each word hit is added as far as possible to increase the cumulative alignment score. The cumulative score is calculated as follows for a nucleotide sequence: , parameters M (reward score for a pair of matching residues; always >0) and N (non-matching) The penalty score for matching residues (always <0) is used for calculation. For each direction, a scoring matrix is ​​used to calculate the cumulative score. An extension of a word hit in If there are too few alignments, the alignment is stopped; due to the accumulation of one or more negatively scored residues. The cumulative score falls below zero; or the end of either sequence is reached. The algorithm parameters W, T, and X determine the sensitivity and speed of the alignment. The BLASTN program (for nucleotide sequences) uses 11 Using a word length (W), an expectation (E) of 10, M=5, N=-4, and a comparison of both strands. For amino acid sequences, the BLASTP program defaults to a word count of 3. The expected length (E) of 10 and the BLOSUM62 scoring matrix ( See Henikoff and Henikoff, (1989) Proc. Natl. Acad. Sci. USA 89:10915 ), 50 alignments (B), 10 expectations (E), M=5, N=-4, and both Use a chain comparison.

[0083] The BLAST algorithm also performs a statistical analysis of the similarity between two sequences (e.g., , Karlin and Altschul, Proc. Natl. Acad. Sci. USA 90:5873-5787, 1993 One measure of similarity provided by the BLAST algorithm is the ratio of two nucleotides The minimum sum probability (P (N)). For example, the minimum sum probability in the comparison of the test nucleic acid with the reference nucleic acid is about 0.2 less than about 0.01, more preferably less than about 0.001, and most preferably less than about 0.001. A nucleic acid is considered to be similar to a reference sequence if it is found to be similar to a reference sequence.

[0084] The percent identity between two amino acid sequences is calculated using the PAM120 weight residue table (weig ht residue table), using a gap length penalty of 12 and a gap penalty of 4 and incorporated into the ALIGN program (version 2.0) by E. Meyers and W. Mil. It can also be determined using the algorithm of [Lee, Comput. Appl. Biosci. 4:11-17, 1988]. Furthermore, the percent identity between two amino acid sequences can be calculated using the BLOSUM 62 matrix. PAM250 matrix and 16, 14, 12, 10, 8, 6, or GCG with a gap weight of 1, 2, 3, 4, 5, or 6 and a length weight of 1, 2, 3, 4, 5, or 6 The GAP program (University of Southampton) in the software package h Florida) incorporated into Needleman and Wunsch, (J. Mol. Biol. The determination can be made using the algorithm of [Paragraph 1], [Paragraph 2], [Paragraph 3], [Paragraph 4], [Paragraph 5], [Paragraph 6], [Paragraph 7], [Paragraph 8], [Paragraph 9], [Paragraph 10], [Paragraph 11], [Paragraph 12], [Paragraph 13], [Paragraph 14], [

[0085] In addition to the percentage of sequence identity noted above, it is possible to determine whether two nucleic acid sequences or polypeptides are substantially identical. Another indication of genetic identity is the sequence encoded by the first nucleic acid, as described below. The polypeptide is immunoreactive with antibodies raised against a polypeptide encoded by a second nucleic acid. Thus, for example, if two peptides are identical by conservative substitutions, A polypeptide is typically substantially identical to a second polypeptide if it differs only in one amino acid sequence. Another indication that two nucleic acid sequences are substantially identical is the identification of two sequences, as described below. The molecules or their complements hybridize to each other under stringent conditions. Further indication that two nucleic acid sequences are substantially identical is the use of the same primers. The advantage is that it can be used to amplify the sequence.

[0086] The term "nucleic acid" is used interchangeably herein with the term "polynucleotide" and refers to a single Deoxyribonucleotides or ribonucleotides and their polynucleotides in either single or double stranded form This term refers to a nucleic acid analog that is synthetic, natural, and non-naturally occurring. known nucleotides that have similar binding properties and are metabolized in a manner similar to the reference nucleotides Nucleic acids containing nucleotide analogs or modified backbone residues or linkages of Examples of such analogs include, but are not limited to, phosphorothioates (ph osphorothioate, phosphoramidate, methylphosphonic acid, chiral These include 2-O-methylphosphonic acid, 2-O-methylribonucleotide, and peptide nucleic acid (PNA). It can be obtained.

[0087] Unless otherwise indicated, a particular nucleic acid sequence may include conservatively modified variants thereof, e.g. , degenerate codon substitutions) and complementary sequences, as well as sequences explicitly indicated. In particular, as detailed below, one or more selected (or all) codons Creating sequences where the third position is substituted with mixed base and / or deoxyinosine residues Degenerate codon substitution can be achieved by using the Acid Res. 19:5081; Ohtsuka et al., (1985) J. Biol. Chem. 260:2605-2608; and Ro ssolini et al., (1994) Mol. Cell. Probes 8:91-98).

[0088] The term "operably linked" in the context of nucleic acids refers to two or more polynucleotides Refers to the functional relationship between segments of (e.g., DNA). Typically, it is transcribed Refers to the functional relationship of a transcriptional regulatory sequence to a sequence, e.g., a promoter or enhancer. - sequences that stimulate or modulate transcription of a coding sequence in an appropriate host cell or other expression system It is operably linked to a coding sequence if it transcribed the sequence. The promoter transcriptional regulatory sequence operably linked to the sequence is physically related to the sequence to be transcribed. However, enhancers are often contiguous with one another, i.e., they are cis-acting. Some transcriptional regulatory sequences, such as They need not be contiguous or located adjacent to each other.

[0089] The terms "polypeptide" and "protein" are used herein to refer to a polymer of amino acid residues. The terms are used interchangeably herein. The term refers to a nucleotide sequence corresponding to one or more amino acid residues. Amino acid polymers that are artificial chemical mimics of natural amino acids, as well as natural amino acid polymers and and unnatural amino acid polymers. Unless otherwise indicated, the term "natural amino acid" refers to a particular polypeptide sequence. A sequence also implicitly encompasses conservatively modified variants thereof.

[0090] The term "subject" includes human and non-human animals. Non-human animals include all vertebrates, e.g., For example, mammals and non-mammals, e.g., non-human primates, sheep, dogs, cows, chickens Unless otherwise noted, the terms "patient" and "subject" are used interchangeably. As used herein generically.

[0091] The term "BKV" or "BK virus" refers to a polyomavirus (Polyomaviridae) It refers to a member of the Orthopolyomavirus family and the Orthopolyomavirus genus. Rus has a genome of approximately 5,000 base pairs, consisting of an icosahedral, non-enveloped double-stranded DNA. They are viruses. They measure approximately 40-45 nM in diameter (Bennett et al., Microb es and Infection. 2012:14(9):672-683).

[0092] "JCV" or "JC virus" refers to a virus belonging to the Polyomaviridae family, It refers to a member of the Orthopolyomavirus genus. JCV is a member of the Orthopolyomavirus genus. A related, icosahedral, non-enveloped duplex also has a genome of approximately 5,000 base pairs. They are DNA viruses. They measure approximately 40-45 nM in diameter (Johne et al., Arch. Virol. 2011;156(9):1627-1634).

[0093] The terms "BKV nephropathy" or "BKV-associated nephropathy" or "BKVAN" This refers to viral cytopathological changes and viral gene expression mainly in the renal tubular epithelium. It refers to an inflammatory interstitial nephropathy resulting from lytic infection with BKV, characterized by:

[0094] The term "VP1" refers to the major polyomavirus capsid subunit protein. A "VP1 pentamer" is composed of five VP1 monomers.

[0095] [Table 1]

[0096] "Virus-like particles" or "VLPs" are aggregates of VP1 pentamers within the viral capsid. VLPs are composed of 72 VP1 pentamers. VLPs are the actual virus It is structurally very similar to the VP1, but contains minor capsid proteins (VP2 and VP3). VLPs lack the viral DNA genome and are therefore non-infectious. This is useful because the epitopes are presented in a conformation similar to that of the actual virus. do.

[0097] "IC50" (half maximal inhibitory concentration) is the median value between the baseline control and the maximum possible signal. IC50 refers to the concentration of a particular antibody that induces a signal between VP and VP. It is the concentration of antibody at which 50% of the available binding sites on an antigen are occupied.

[0098] "EC50" (median effective concentration) is the concentration at which a given concentration is reduced relative to baseline after a specific exposure or treatment period. It refers to the concentration of a particular antibody that induces a response midway (50%) between the control and the maximum possible effect. For example, EC50 is the concentration of antibody that neutralizes 50% of viral infection.

[0099] "EC90" is the response corresponding to 90% of the maximum possible effect after a specific exposure or treatment period. For example, EC90 refers to the concentration of a specific antibody that induces a 90% neutralization of viral infection. is the concentration of antibody used.

[0100] "Neutralization" refers to the reduction of host cell susceptibility to a virus, as indicated by the absence of viral gene expression. Without being bound by any theory, it is believed that the neutralization by specific antibodies The mechanism is blocking the interaction of viral capsid proteins with cell surface receptors or host Disruption of any stage of the entry and transport process prior to delivery of the viral genome to the nucleus of the host cell It can be induced.

[0101] As used herein, the term "treat" or "treatment of any disease or disorder" refers to "Treatment" or "treatment" refers, in one embodiment, to ameliorating a disease or disorder (i.e., preventing or reducing the disease). or slowing or arresting the development of at least one of its clinical symptoms, and In another embodiment, "treat," "treating," or "treatment" refers to " means a reduction in at least one physical parameter, including one that is not discernible by the patient. In yet another embodiment, "treat," "treating," or "improving" refers to "Treatment" means the physical (e.g., stabilization of discernible symptoms) or therapeutic improvement of a disease or disorder. This refers to regulating the physiological (e.g., stabilization of physical parameters), or both.

[0102] The phrase "reducing the likelihood" means reducing the likelihood of the onset or occurrence of a disease, infection, or disorder. It means to slow down the progression.

[0103] The term "therapeutically acceptable amount" or "therapeutically effective dose" refers to a dose that is sufficient to achieve the desired result (i.e., tumor suppression). reduction of tumor size, inhibition of tumor growth, prevention of metastasis, viral, bacterial, fungal or parasitic infection In some embodiments, therapeutically effective means refers to an amount sufficient to achieve a therapeutically effective effect (inhibition or prevention of a disease or condition). An acceptable amount is one that does not induce or cause undesirable side effects. The amount tolerated is initially administered at a low dose, and then increased until the desired effect is achieved. "Prophylactically effective doses" and "therapeutically effective doses" can be determined by gradually increasing the dose. The molecules of the present disclosure may be used to treat or alleviate disease symptoms, such as those associated with polyomavirus infection. These may prevent its onset or result in a reduction in its severity, respectively.

[0104] The term "co-administer" refers to the simultaneous presence of two active agents in the blood of an individual. The administered active agents can be delivered simultaneously or sequentially. [Brief explanation of the drawings]

[0105] [Figure 1-1]1A-1D illustrate the affinity measurements of anti-VP1 antibodies on VP1 pentamers for BKV serotypes I-IV by SET assay. [Figure 1-2] As stated above. [Figure 1-3] As stated above. [Figure 1-4] As stated above. [Figure 2] FIG. 2 is a table of SET affinity values ​​(KD) of anti-VP1 antibodies on VP1 pentamers for BKV serotypes I-IV. [Figure 3-1] 3A-3E illustrate the affinity measurements of anti-VP1 antibodies on VP1 pentamers or VLPs for BKV serotypes I-IV by Biacore. [Figure 3-2] As stated above. [Figure 3-3] As stated above. [Figure 3-4] As stated above. [Figure 3-5] As stated above. [Figure 4] FIG. 4 is a graph of anti-VP1 antibody binding to BKV serotype I VLPs as measured by ELISA. [Figure 5] FIG. 5 is a graph of anti-VP1 antibody binding to BKV serotype IV VLPs as measured by ELISA. [Figure 6] FIG. 6 is a graph of anti-VP1 antibody binding to BKV serotype IV VP1 pentamers as measured by ELISA. [Figure 7] FIG. 7 is a table of IC50 values ​​generated by ELISA for anti-VP1 antibody binding to BKV serotype I and IV VLPs or VP1 pentamers. [Figure 8] FIG. 8 is a graph of anti-VP1 antibody binding to BKV serotype I VLPs as measured by ELISA. [Figure 9] FIG. 9 is a table of IC50 values ​​generated by ELISA for anti-VP1 antibody binding to BKV serotype I VLPs. [Figure 10] FIG. 10 is a graph of anti-VP1 antibody binding to JC virus VLPs as measured by ELISA. [Figure 11] FIG. 11 is a table of IC50 values ​​generated by ELISA for anti-VP1 antibody binding to JCV VLPs. [Figure 12] Figures 12A-B show two blots. The top panel (Figure 12A) is a Western blot showing no binding of anti-VP1 antibody to denatured BKV VP1. The bottom panel (Figure 12B) is a dot blot of non-denatured BKV VP1 pentamer showing binding of anti-VP1 antibody to non-denatured VP1 pentamer. [Figure 13-1] 13A-13F illustrate the binding of anti-VP1 antibodies to wild-type BKV serotype I VP1 pentamers by Biacore, and that point mutations in VP1 can abolish binding. [Figure 13-2] As stated above. [Figure 13-3] As stated above. [Figure 13-4] As stated above. [Figure 13-5] As stated above. [Figure 13-6] As stated above. [Figure 14] FIG. 14 is a table summarizing the critical residues for binding identified in the epitope of the anti-VP1 antibody. [Figure 15] FIG. 15 is a graph of anti-VP1 antibodies neutralizing BKV serotype I infection. [Figure 16] FIG. 16 is a graph of anti-VP1 antibodies neutralizing BKV serotype II infection. [Figure 17] FIG. 17 is a graph of anti-VP1 antibodies neutralizing BKV serotype III infection. [Figure 18] FIG. 18 is a graph of anti-VP1 antibodies neutralizing BKV serotype IV infection. [Figure 19] FIG. 19 is a table summarizing the neutralizing activity (EC50 and EC90) of anti-VP1 antibodies against BKV serotypes I to IV and JC virus. [Figure 20] FIG. 20 is a graph of anti-VP1 antibodies neutralizing infection by BKV serotypes I, II, and IV. [Figure 21]FIG. 21 is a table summarizing the neutralizing activity (EC50 and EC90) of anti-VP1 antibodies against BKV serotypes I to IV. [Figure 22] FIG. 22 is a graph of anti-VP1 antibodies neutralizing infection by BKV serotype I. [Figure 23] FIG. 23 is a table summarizing the neutralizing activity (EC50 and EC90) of anti-VP1 antibodies against BKV serotype I. [Figure 24] FIG. 24 is a graph of anti-VP1 antibodies neutralizing infection by JCV. [Figure 25] FIG. 25 is a graph of anti-VP1 antibodies neutralizing infection by JCV. [Figure 26] FIG. 26 is a table summarizing the neutralizing activity (EC50 and EC90) of anti-VP1 antibodies against JCV infection. [Figure 27] FIG. 27 is a table of the affinity of antibody P8D11 for JC virus VLPs and VLPs containing point mutations. [Figure 28] FIG. 28 is a deuterium exchange epitope mapping of P8D11 Fab bound to BKV VP1 pentamer. [Figure 29-1] Figure 29A is a table showing anti-BKV antibody contact residues in the EF loop when certain mutations are introduced by alanine scanning, and Figures 29B-29C show SPR graphs of anti-BKV antibody binding to wild-type and mutant residues in VP1. [Figure 29-2] As stated above. [Figure 29-3] As stated above. [Figure 30] FIG. 30 is the X-ray crystal structure of P8D11 in complex with the BKV VP1 pentamer. [Figure 31] Figures 31A-B illustrate how P8D11 contacts residues of the VP1 pentamer.

[0106] Detailed Description The present disclosure provides antibodies, antibody fragments (e.g., antigen-binding fragments) that bind to and neutralize BKV. In particular, the present disclosure provides a method for detecting viral vectors that bind to VP1 protein and, upon such binding, inhibit viral replication. The present invention relates to antibodies and antibody fragments (e.g., antigen-binding fragments) that neutralize virulence infection. The present disclosure has desirable pharmacokinetic properties and other desirable attributes, and thus is Reduce the likelihood of or treat cancer-related nephropathy (e.g., BKVAN) The present disclosure further provides pharmaceutical compositions comprising the antibodies. and such for the prevention and treatment of polyomavirus infections and related disorders. Methods of making and using the pharmaceutical compositions are provided.

[0107] Anti-VP1 antibody The present disclosure provides antibodies or antibody fragments (e.g., antigen-binding fragments) that specifically bind to VP1. The antibodies or antibody fragments (e.g., antigen-binding fragments) of the present disclosure include, but are not limited to, Human monoclonal antibodies isolated as described in the Examples below, but not limited to: or a fragment thereof.

[0108] In certain aspects, the present disclosure provides an antibody or antibody fragment (e.g., For example, antigen-binding fragments) and SEQ ID NOs: 12, 32, 52, 72, 92, 112, 13 2, 152, 172, 192, 212, 232, 252, 272, 292, 312, 32 8, 348, 362, 376, 390, 404, 418, 432, 446, 460, 47 4, and 488. The present disclosure also provides fragments (e.g., antigen-binding fragments) specific for VP1 (Table 2). and antibodies or antibody fragments (e.g., antigen-binding fragments) that bind to any of the antibodies listed in Table 2. The antibody or The present disclosure also provides antibody fragments (e.g., antigen-binding fragments). In certain aspects, the present disclosure provides antibodies against VP1 an antibody or antibody fragment (e.g., an antigen-binding fragment) that specifically binds to 1, 2, 3 or more VHs having any of the amino acid sequences of the VH CDRs listed in 2. The antibodies described above comprise (or alternatively consist of) the CDRs.

[0109] The present disclosure provides antibodies or antibody fragments (e.g., antigen-binding fragments) that specifically bind to VP1. SEQ ID NOs: 22, 42, 62, 82, 102, 122, 142, 162, 182 , 202, 222, 242, 262, 282, 302, 320, 338, 355, 369 , 383, 397, 411, 425, 439, 453, 467, 481, and 495 the antibody or antibody fragment (e.g., antigen-binding The present disclosure also provides antibodies or antibody fragments that specifically bind to VP1. a fragment (e.g., an antigen-binding fragment) comprising any of the VL CDRs listed in Table 2 below. the antibody or antibody fragment (e.g., In particular, antibodies or antibody fragments (e.g., antigen-binding fragments) that specifically bind to VP1 are also provided. and a fragment thereof (e.g., an antigen-binding fragment) comprising any of the amino acids of the VL CDRs listed in Table 2. The antibody comprises (or alternatively consists of) one, two, three or more VL CDRs having the sequence The antibody or antibody fragment (e.g., antigen-binding fragment) is provided.

[0110] Other antibodies or antibody fragments (e.g., antigen-binding fragments) of the present disclosure may be mutated but still contain the antibody fragments shown in Table 1. 2 and at least 60, 70, 80 or more CDR regions In some embodiments, the amino acid sequence contains amino acids with 90 or 95 percent identity. This may be 1, 2, 3, 4 or 5 CDR regions as compared to the CDR regions set forth in the sequences set forth in Table 2. The following amino acid sequence variants are included in which the amino acids are mutated in the CDR regions:

[0111] The present disclosure also provides VH, VL, full-length heavy chain, and full-length heavy chain antibodies that specifically bind to VP1. Nucleic acid sequences encoding long light chains are also provided. Such nucleic acid sequences can be used for expression in mammalian cells. can be optimized for the present.

[0112] [Table 2-1]

[0113] [Table 2-2]

[0114] [Table 2-3]

[0115] [Table 2-4]

[0116] [Table 2-5]

[0117] [Table 2-6]

[0118] [Table 2-7]

[0119]

Table 2-8

[0120]

Table 2-9

[0121]

Table 2-10

[0122]

Table 2-11

[0123]

Table 2-12

[0124]

Table 2-13

[0125]

Table 2-14

[0126]

Table 2-15

[0127]

Table 2-16

[0128]

Table 2-17

[0129]

Table 2-18

[0130]

Table 2-19

[0131]

Table 2-20

[0132]

Table 2-21

[0133]

Table 2-22

[0134]

Table 2-23

[0135]

Table 2-24

[0136]

Table 2-25

[0137]

Table 2-26

[0138]

Table 2-27

[0139]

Table 2-28

[0140]

Table 2-29

[0141]

Table 2-30

[0142]

Table 2-31

[0143]

Table 2-32

[0144]

Table 2-33

[0145]

Table 2-34

[0146]

Table 2-35

[0147]

Table 2-36

[0148]

Table 2-37

[0149]

Table 2-38

[0150]

Table 2-39

[0151]

Table 2-40

[0152]

Table 2-41

[0153]

Table 2-42

[0154]

Table 2-43

[0155]

Table 2-44

[0156]

Table 2-45

[0157]

Table 2-46

[0158]

Table 2-47

[0159]

Table 2-48

[0160]

Table 2-49

[0161]

Table 2-50

[0162]

Table 2-51

[0163]

Table 2-52

[0164]

Table 2-53

[0165]

Table 2-54

[0166]

Table 2-55

[0167]

Table 2-56

[0168]

Table 2-57

[0169] [Table 2-58]

[0170] Other antibodies of the present disclosure include antibodies in which the amino acid or the nucleic acid encoding the amino acid has been mutated. or at least 60, 70, 80, 90, or 95 percent of the sequences listed in Table 2 In some embodiments, it has an identity to a sequence listed in Table 2. No more than 1, 2, 3, 4, or 5 amino acids in the variable region when compared to the variable region described and amino acid sequence variants which are mutated in the nucleotide sequence but retain substantially the same therapeutic activity.

[0171] These antibodies each bind to VP1, and therefore VH, VL, and full-length light chains and the full-length heavy chain sequence (amino acid sequence and nucleotide sequence encoding the amino acid sequence). The antibodies (columns) can be "mixed and matched" to generate other VP1-binding antibodies. Such "mixed and matched" VP1-binding antibodies can be assayed using binding assays known in the art (e.g., Test using ELISA, and other assays described in the Examples section. When these chains are mixed and matched, the VH from a particular VH / VL pairing can be The sequence should be replaced with a structurally similar VH sequence. / Replacing a full-length heavy chain sequence derived from a full-length light chain pair with a structurally similar full-length heavy chain sequence Similarly, VL sequences from a particular VH / VL pair should be compared with structurally similar sequences. Similarly, VL sequences derived from a particular full-length heavy chain / full-length light chain pair should be replaced with The full-length light chain sequence should be replaced with a structurally similar full-length light chain sequence. In one aspect, the present disclosure provides antibodies against SEQ ID NOs: 12, 32, 5, and 6, wherein the antibodies specifically bind to VP1. 2, 72, 92, 112, 132, 152, 172, 192, 212, 232, 252, 272, 292, 312, 328, 348, 362, 376, 390, 404, 418, 432, 446, 460, 474, and 488 (Table 2). heavy chain variable region comprising the amino acid sequence of SEQ ID NOs: 22, 42, 62, 82, 102, 122; , 142, 162, 182, 202, 222, 242, 262, 282, 302, 320 , 338, 355, 369, 383, 397, 411, 425, 439, 453, 467 a light chain variable region comprising an amino acid sequence selected from the group consisting of: 481, 482, and 495 (Table 2) The present invention provides an isolated monoclonal antibody or antigen-binding region thereof having a target region.

[0172] In another aspect, the present disclosure provides a method for the preparation of a nucleic acid sequence comprising: (i) SEQ ID NOs: 14, 34, 54, 74, 94, 114, 1 34, 154, 174, 194, 214, 234, 254, 274, 294, 313 and and 330; an amino acid sequence optimized for expression in mammalian cells selected from the group consisting of full-length heavy chains containing the amino acid sequences of SEQ ID NOs: 24, 44, 64, 84, 104, 124, 144, 164, 184, 204, 224, 244, 264, 284, 304, 321, 340. An amino acid optimized for expression in mammalian cells selected from the group consisting of an isolated monoclonal antibody having a full-length light chain comprising the sequence A functional protein comprising the binding moiety is provided.

[0173] In another aspect, the disclosure provides heavy and light chain CDR1, CDR2 and CDR3 sequences as set forth in Table 2. R3, or combinations thereof. The amino acid sequences are SEQ ID NOs: 6, 26, 46, 66, 86, 106, 126, 146, 166, 186, 206, 226, 246, 266, 286, 306, 322, 342, 356, 370, 384, 398, 412, 426, 440, 454, 468, and 482 The amino acid sequences of the VH CDR2 of the antibodies are shown in SEQ ID NOs: 7, 27, 47, 67, 87, and 1. 07, 127, 147, 167, 187, 207, 227, 247, 267, 287, 3 07, 323, 343, 357, 371, 385, 399, 413, 427, 441, 4 The amino acid sequences of the VH CDR3 of the antibodies are shown in SEQ ID NOs: 55, 469, and 483. 8, 28, 48, 68, 88, 108, 128, 148, 168, 188, 208, 22 8, 248, 268, 288, 308, 324, 344, 358, 372, 386, 40 0, 414, 428, 442, 456, 470, and 484. The amino acid sequences of R1 are shown in SEQ ID NOs: 16, 36, 56, 76, 96, 116, 136, 15 6, 176, 196, 216, 236, 256, 276, 296, 314, 332, 34 9, 363, 377, 391, 405, 419, 433, 447, 461, 475 and The amino acid sequences of the VL CDR2 of the antibodies are shown in SEQ ID NOs: 17, 37, 57, 7 7, 97, 117, 137, 157, 177, 197, 217, 237, 257, 277 , 297, 315, 333, 350, 364, 378, 392, 406, 420, 434 The amino acid sequences of the VL CDR3 of the antibodies are shown in , SEQ ID NOs: 18, 38, 58, 78, 98, 118, 138, 158, 178, 198, 218, 238, 258, 278, 298, 316, 334, 351, 365, 379, 393, 407, 421, 435, 449, 463, 477 and 491.

[0174] Each of these antibodies can bind to VP1, and their antigen-binding specificity is mainly CDR1 VH CDR1, 2 and 3 sequences, taking into account that the VH CDR1, 2 and 3 regions are provided by and VL CDR1, 2 and 3 sequences can be "mixed and matched" (all That is, CDRs from different antibodies can be mixed and matched, but each The antibody contains VH CDR1, 2 and 3 and VL CDR1, 2 and 3, and other (These "mixed and matched" V P1-binding antibodies were assayed using binding assays known in the art and those described in the Examples (e.g., Mix and match VH CDR sequences and test using ELISA. In this case, the CDR1, CDR2 and / or CDR3 sequences derived from a particular VH sequence are The VL CDRs should be replaced with structurally similar CDR sequence(s). When mixing and matching sequences, the CDR1, CDR2 and CDR3 sequences from a particular VL sequence may be and / or the CDR3 sequence should be replaced with a structurally similar CDR sequence(s). The novel VH and VL sequences are synthesized by combining one or more VH and / or VL CDR region sequences. The sequences are constructed from the CDR sequences shown herein for the monoclonal antibodies of the disclosure. New VH and VL sequences can be generated by substituting structurally similar sequences. It will be readily apparent to those skilled in the art that this is possible.

[0175] Thus, the present disclosure provides antibodies against SEQ ID NOs: 6, 26, 46, 66, 86, 106, 126, 146, 166, 186, 206, 226, 246, 26 6, 286, 306, 322, 342, 356, 370, 384, 398, 412, 42 6, 440, 454, 468, and 482; Heavy chain CDR1 comprising: SEQ ID NOs: 7, 27, 47, 67, 87, 107, 127, 147, 1 67, 187, 207, 227, 247, 267, 287, 307, 323, 343, 3 57, 371, 385, 399, 413, 427, 441, 455, 469, and 48 heavy chain CDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 8, 28, 4; 8, 68, 88, 108, 128, 148, 168, 188, 208, 228, 248, 268, 288, 308, 324, 344, 358, 372, 386, 400, 414, an amino acid sequence selected from the group consisting of: 428, 442, 456, 470, and 484; Heavy chain CDR3 comprising the sequences: SEQ ID NOs: 16, 36, 56, 76, 96, 116, 136, 15 6, 176, 196, 216, 236, 256, 276, 296, 314, 332, 34 9, 363, 377, 391, 405, 419, 433, 447, 461, 475 and 489; light chain CDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 17, 3 7, 57, 77, 97, 117, 137, 157, 177, 197, 217, 237, 2 57, 277, 297, 315, 333, 350, 364, 378, 392, 406, 4 20, 434, 448, 462, 476 and 490; light chain CDR2 comprising the amino acid sequence of SEQ ID NOs: 18, 38, 58, 78, 98, 118, 138, 158, 178, 198, 218, 238, 258, 278, 298, 316, 334, 351, 365, 379, 393, 407, 421, 435, 449, 463, 477 and 491; and a light chain CDR3 comprising an amino acid sequence selected from the group consisting of The present invention provides an isolated monoclonal antibody or antigen-binding region thereof.

[0176] In certain embodiments, the antibody that specifically binds to VP1 is an antibody described in Table 2 or An antibody fragment (eg, an antigen-binding fragment).

[0177] 1. Identification of epitopes and antibodies that bind to the epitopes The present disclosure provides antibodies and antibody fragments (e.g., antigen-binding fragments) that bind to epitopes of VP1. In certain embodiments, the antibodies and antibody fragments are directed against all four BKV serotypes. and / or capable of binding to the same epitope in JCV.

[0178] The present disclosure also provides antibodies and antibodies that bind to the same epitope as the anti-VP1 antibodies listed in Table 2. Fragments (e.g., antigen-binding fragments) are also provided. Accordingly, additional antibodies and antibody fragments are provided. (e.g., antigen-binding fragments) cross-compete with other antibodies in binding assays (e.g., and identifying the compound based on its ability to competitively inhibit the binding of the compound in a statistically significant manner. The antibodies of the present disclosure directed to VP1 (e.g., human BKV or JCV VP1) and The ability of the test antibody to inhibit binding of the antibody fragment (e.g., antigen-binding fragment) to the target protein is determined by the test antibody's ability to inhibit binding of the target protein to the target protein. Competing with an antibody or antibody fragment (e.g., an antigen-binding fragment) for binding to VP1 Such antibodies, according to non-limiting theory, may be capable of competing with or against antibodies or antibody fragment (e.g., antigen-binding fragment) and the same or related (e.g., For example, it can bind to epitopes that are structurally similar or spatially close. In certain embodiments, the antibody or antibody fragment (e.g., antigen-binding fragment) of the present disclosure may have the same VP The antibody that binds to the epitope on 1 is a human or humanized monoclonal antibody. Such human or humanized monoclonal antibodies can be prepared and isolated as described herein. It is possible.

[0179] 2. Further changes in the Fc region framework The present disclosure discloses specific anti-VP1 antibodies. These antibodies can be used to, for example, improve the properties of antibodies. These modifications may further include modifications to framework residues within VH and / or VL to improve The antibody or antigen-binding fragment thereof may include a modified antibody or antigen-binding fragment thereof. Framework modifications are performed to reduce the immunogenicity of antibodies. For example, one approach is to by "backmutating" one or more framework residues to the corresponding germline sequence. More particularly, antibodies that have undergone somatic mutations differ from the germline sequence from which they are derived. Antibody framework sequences may contain framework residues. Comparison with the germline sequence can identify such residues. To restore the gene region sequence to its germline configuration, e.g., by site-directed mutagenesis, In this way, somatic mutations can be "reverted" to the germline sequence. Antibodies that are "antibodies" are also intended to be encompassed.

[0180] Another type of framework modification is within the framework region, or even within one or more Mutation of one or more residues within multiple CDR regions to remove T cell epitopes This approach involves reducing the immunogenic potential of antibodies by Also known as "Pesticide-Free" and U.S. Patent Application Publication No. 2003 / 0153043 by Carr et al. Further details are provided in the specification.

[0181] In addition to, or instead of, modifications made within the framework or CDR regions, Contains modifications within the Fc region, typically affecting serum half-life, complement fixation, Fc receptor binding, and and / or antigen-dependent cellular cytotoxicity. Additionally, antibodies can be chemically modified (e.g., one or more chemical moieties can be attached to the antibody) or modified to The glycosylation can be altered, again altering one or more functional properties of the antibody. Each of these aspects is described in further detail below.

[0182] In one aspect, the number of cysteine ​​residues in the hinge region is altered, e.g., increased or The hinge region of CH1 is modified to increase or decrease the binding affinity. This technique was previously described by Bodmer et al. The hinge region of CH1 is further described in U.S. Patent No. 5,677,425. The number of cysteine ​​residues in the or increasing or decreasing the stability of the antibody.

[0183] In another embodiment, the Fc hinge region of the antibody is mutated to decrease the biological half-life of the antibody. More particularly, antibodies that bind weaker staphylococci to the native Fc-hinge domain of SpA compared to native SpA. Staphylococcal protein A (SpA) binding A number of amino acid mutations are introduced into the CH2-CH3 domain interface region of the Fc-hinge fragment. This approach is described in more detail in US Pat. No. 6,165,745 to Ward et al. It is written.

[0184] In yet another embodiment, at least one amino acid residue is replaced with a different amino acid residue. In addition, the Fc region is altered by changing the effector functions of the antibody. , where the antibody has an altered affinity for the effector ligand but retains the antigen-binding capacity of the parent antibody. Replacing one or more amino acids with different amino acid residues to retain potency The effector ligand that alters affinity can be, for example, an Fc receptor or a complement receptor. This approach is described, for example, in US Pat. No. 6,229,493, both by Winter et al. As described in Patent Nos. 5,624,821 and 5,648,260 .

[0185] In another embodiment, the antibody has altered C1q binding and / or reduced or abrogated C1q binding. The amino acid residues selected from the group consisting of 1 or 2 are selected to have complement-dependent cytotoxicity (CDC) activity. can replace multiple amino acids with different amino acid residues.

[0186] In another embodiment, one or more amino acid residues are altered to modify the polypeptide that fixes complement. This technique is described, for example, in PCT International Publication No. 2003 / 0100900 by Bodmer et al. In certain embodiments, the antibodies or or an antigen-binding fragment thereof, by substituting one or more amino acids of one or more allotypic amino acids. Acid residues are substituted for the IgG1 subclass and kappa isotype. The amino acid residues include, but are not limited to, IgG1, IgG2, and IgG3 The constant region of the heavy chain of each subclass, as well as the Also included are constant regions of the light chain of the kappa isotype as described in

[0187] In yet another embodiment, the Fc region is modified to enhance antibody-dependent cellular cytotoxicity (ADCC). Increase the ability of the antibody to mediate and / or modify one or more amino acids This increases the affinity of the antibody for the Fcγ receptor. ta, PCT International Publication No. 00 / 42072. Bindings on human IgG1 to FcγRI, FcγRII, FcγRIII, and FcRn The binding site has been mapped and variants with improved binding have been described (Shield (See, e.g., S. et al., J. Biol. Chem. 276:6591-6604, 2001).

[0188] In yet another embodiment, the glycosylation of the antibody is modified. For example, an aglycosylated antibody is The antibody can be made to have altered glycosylation (i.e., the antibody lacks glycosylation). Such carbon can increase the affinity of an antibody for an "antigen," for example. Hydrate modifications can be made, for example, by altering one or more glycosylation sites within the antibody sequence. For example, one or more variable region framework glycosylation sites can be by making one or more amino acid substitutions that result in the removal of the ligation site. Glycosylation at positions 1 and 2 can be removed. Such aglycosylation can enhance the binding of the antigen to the target molecule. This can increase the affinity of antibodies to the target molecule. Such techniques are described, for example, by Co et al. Nos. 5,714,350 and 6,350,861. It is being done.

[0189] Additionally or alternatively, hypofucosylated antibodies or biosynthetic antibodies having reduced amounts of fucosyl residues may be used. antibodies with altered glycosylation, such as antibodies with increased glycosylation GlcNac structures Such altered glycosylation patterns can be used to modify the AD of antibodies. Such carbohydrate modifications have been shown to increase CC capacity. This can be achieved by expressing the antibody in a host cell with an altered cosylation machinery. Cells with altered glycosylation machinery have been described in the art and can be used to express recombinant antibodies. By doing so, the cells can be used as host cells to produce antibodies with altered glycosylation. For example, European Patent No. 1,176,195 by Hang et al. The present invention describes a cell line in which the FUT8 gene encoding the transferase is functionally disrupted, Antibodies expressed in such cell lines exhibit hypoglycosylation. Publication No. 03 / 035835 discloses a method for the preparation of a nucleotide sequence in which fucose is linked to Asn(297). The ability to bind carbohydrates is reduced, resulting in hypofucosylation of antibodies expressed in the host cell. have described a variant CHO cell line, Lecl3 cells, which also produces HIV-1 (Shields et al. (2002) J. Biol. Chem. 277:26733-26740). PC by Umama et al. WO 99 / 54342 describes antibodies expressed in engineered cell lines. showed an increase in bisecting GlcNac structures, which resulted in increased ADCC activity of the antibody. As shown, glycoprotein-modifying glycosyltransferases (e.g., beta(1,4) -N-acetylglucosaminyltransferase III (GnTIII) have described engineered cell lines for this purpose (Umana et al., Nat. Biotech. 17:176-180, 1999). See also:

[0190] In another embodiment, the antibody is modified to increase its biological half-life. For example, a 1-phase ion exchange membrane, as described in U.S. Pat. No. 6,277,375 to Ward, may be used. Alternatively, multiple of the following mutations can be introduced: T252L, T254S, T256F. Alternatively, to increase the biological half-life, US Patent No. 5,529,299 by Presta et al. As described in US Pat. Nos. 8,669,046 and 6,121,022, Salvage receptor-binding epitopes taken from two loops of the CH2 domain of the Fc region The antibody can be altered in the CH1 or CL region to contain:

[0191] To minimize the ADCC activity of antibodies, specific mutations in the Fc region may be used to target effector cells. Generally, "IgG" is an Fc-silent antibody that has minimal interaction with the IgG "Fc region" refers to a region of an immunoglobulin that is a fusion protein, such as a native sequence Fc region and a variant Fc region. The C-terminal region of the IgG heavy chain is defined as the C-terminal region of the IgG heavy chain. It is defined as including amino acid residues from C226 or from position P230 to the carboxyl terminus. The numbering of residues in the Fc region is that of the EU index of Kabat. The C-terminal lysine (residue K447) of the c region may be removed, for example, during antibody production or purification. It is possible.

[0192] Silenced effector functions can be obtained by mutations in the Fc region of antibodies. and are described in the art: LALA and N297A (Strohl, W., 2009, Curr Opin. Biotechnol. vol. 20(6):685-691); and D265A (Baudino et al., 20 08, J. Immunol. 181: 6664-69), Heusser et al., International Publication No. 2012065950 See also the brochure. An example of a silent Fc IgG1 antibody is the IgG1 Fc antibody. It is a LALA mutant containing the L234A and L235A mutations in the amino acid sequence. Another example of a human IgG1 antibody is a human IgG1 antibody with a DAPA (D265A, P329A) mutation (U.S. Patent No. 6, Another silent IgG1 antibody contains the N297A mutation. This results in an aglycosylated / non-glycosylated antibody.

[0193] Fc-silenced antibodies have no or low ADCC activity, which is The antibody lyses less than 50% of the specific cells (low ADCC activity) or This means that the lysis is less than 1% (no ADCC activity).

[0194] 3. Generation of Anti-VP1 Antibodies including but not limited to recombinant expression, chemical synthesis, and enzymatic digestion of antibody tetramers Anti-VP1 antibodies and antibody fragments thereof (e.g., Although full-length monoclonal antibodies (antigen-binding fragments) can be generated, e.g., It can be obtained by hybridoma or recombinant production. Recombinant expression is well known in the art. Any suitable host cell known in the art, for example, a mammalian host cell, a bacterial host cell, or a yeast host cell , insect host cells, etc.

[0195] The present disclosure also provides polynucleotides encoding the antibodies described herein, e.g., Encoding a segment containing a heavy or light chain variable region or a complementarity determining region as described In some embodiments, a polynucleotide encoding a heavy chain variable region is further provided. The polynucleotides are SEQ ID NOs: 13, 33, 53, 73, 93, 113, 133, 15 Selected from the group consisting of 3, 173, 193, 213, 233, 253, 273 and 293 Polynucleotides with at least 85%, 89%, 90%, 91%, 92%, 93% %, 94%, 95%, 96%, 97%, 98%, 99%, or 100% nucleic acid sequence identity In some aspects, the polynucleotide encoding the light chain variable region has the sequence Numbers 23, 43, 63, 83, 103, 123, 143, 163, 183, 203, 22 a polynucleotide selected from the group consisting of: 3, 243, 263, 283, and 303; At least 85%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96% %, 97%, 98%, 99%, or 100% nucleic acid sequence identity.

[0196] In some embodiments, the polynucleotide encoding the heavy chain is selected from the group consisting of SEQ ID NOs: 15, 35, 5 5, 75, 95, 115, 135, 155, 175, 195, 215, 235, 255, At least 85%, 89%, 90%, 91% with 275 and 295 polynucleotides , 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% In some embodiments, the polynucleotide encoding the light chain has a nucleic acid sequence identity of , SEQ ID NOs: 25, 45, 65, 85, 105, 125, 145, 165, 185, 205 , 225, 245, 265, 285 and 305 polynucleotides and at least 85 %, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98 %, 99%, or 100% nucleic acid sequence identity.

[0197] The polynucleotides of the present disclosure may encode only the variable region sequences of an anti-VP1 antibody. They may also encode both the variable and constant regions of an antibody. The nucleotide sequence is a polynucleotide sequence containing the variable regions of both the heavy and light chains of one of the exemplary anti-VP1 antibodies. Some other polynucleotides encode the heavy chain of one of the mouse antibodies. and encoding two polypeptide segments substantially identical to the variable regions of the light and light chains, respectively. do.

[0198] The polynucleotide sequence is compared to an existing sequence encoding an anti-VP1 antibody or binding fragment thereof. (e.g., the sequences described in the Examples below) by de novo solid-phase DNA synthesis; can be generated by PCR mutagenesis. Narang et al., Meth. Enzymol. 68 :90, 1979; the phosphotriester method of Brown et al., Meth. Enzymol. 68:109, 1979 Phosphorodiester method: diethylphosphoroester of Beaucage et al., Tetra. Lett., 22:1859, 1981 midite method; and the solid support method of U.S. Pat. No. 4,458,066. Direct chemical synthesis of nucleic acids can be achieved by methods known in the art. The introduction of mutations into the nucleotide sequence can be carried out, for example, by PCR Technology: Principles and Applications tions for DNA Amplification, HA Erlich (Ed.), Freeman Press, NY, NY, 1992; PCR Protocols: A Guide to Methods and Applications, Innis et al. (Ed.), Academic Pr ess, San Diego, CA, 1990; Mattila et al., Nucleic Acids Res. 19:967, 1991; and Eckert et al., PCR Methods and Applications 1:17, 1991. This can be done.

[0199] The present disclosure also provides expression vectors and host cells for producing the above-described anti-VP1 antibodies. A variety of expression vectors can be used to encode anti-VP1 antibody chains or binding fragments. Viral-based expression vectors can be used to express polynucleotides that encode the vector. Both viral and non-viral expression vectors can be used to produce antibodies in mammalian host cells. Non-viral vectors and systems typically involve the delivery of proteins or RNA. Plasmids, episomal vectors, and human vectors containing expression cassettes for expression chromosomes (see, e.g., Harrington et al., Nat Genet 15:345, 1997) are listed. For example, the expression of anti-VP1 polynucleotides and Non-viral vectors useful for expression of polypeptides include pThioHis A, B, and C, pcDNA3.1 / His, pEBVHisA, B, and C (Invi (Trogen, San Diego, CA), MPSV vectors, and other proteins Several other vectors are known in the art for expressing genes. The vectors include retroviruses, adenoviruses, adeno-associated viruses, and herpes viruses. SV40-based vectors, papillomavirus, HBP Epstein-Barr virus, vaccinia virus vectors, and Semliki Forest virus Examples include SFV. Brent et al., supra; Smith, Annu. Rev. Microbiol. 49:8 07, 1995; and Rosenfeld et al., Cell 68:143, 1992.

[0200] The choice of expression vector will depend on the intended host cell in which the vector will be expressed. In one embodiment, the expression vector is operable to carry a polynucleotide encoding an anti-VP1 antibody chain or fragment. Contains a promoter and other regulatory sequences (e.g., enhancers) operably linked to it In some embodiments, an inducible promoter is used to inhibit the expression of the inserted gene except under inducing conditions. Inducible promoters include, for example, arabinose, lacZ, , metallothionein promoter, or heat shock promoter. Cultures of the selected organisms are grown using a coding sequence whose expression product is better tolerated by the host cell. The promoter can be expanded under non-inducing conditions without biasing the population. In addition, other regulatory elements are required for efficient expression of anti-VP1 antibody chains or fragments. These elements typically include the ATG start codon and and adjacent ribosome binding sites or other sequences. This can be enhanced by the inclusion of enhancers appropriate for the cell line (e.g., Scharf et al., Results Probl. Cell Differ. 20:125, 1994; and Bittner et al., M (See, e.g., Eth. Enzymol., 153:516, 1987). For example, the SV40 enhancer or The CMV enhancer can be used to increase expression in mammalian host cells. .

[0201] The expression vector also encodes the polypeptide encoded by the inserted anti-VP1 antibody sequence. A secretion signal sequence site can also be provided to form a fusion protein of the present invention. Frequently, the inserted anti-VP1 antibody sequence is ligated to a signal sequence prior to inclusion in the vector. to receive sequences encoding the anti-VP1 antibody light and heavy chain variable domains. The vector used may also encode the constant region or portions thereof. The vector allows expression of the variable region as a fusion protein with the constant region, which This results in the production of an intact antibody or fragment thereof. The realm is human.

[0202] Host cells for carrying and expressing anti-VP1 antibody chains can be prokaryotic or eukaryotic. E. coli is a suitable organism for cloning and expressing the polynucleotides of the present disclosure. Other microbial hosts suitable for use include Bacillus Bacillus subtilis and other bacilli, as well as Salmonella, Other intestinal bacteria, such as Serratia and various Pseudomonas species In these prokaryotic hosts, those skilled in the art will typically 1. Creating an expression vector containing expression control sequences (e.g., an origin of replication) compatible with the cell. In addition, lactose promoter system, tryptophan (trp) promoter system, beta-lactamase promoter system, or promoters derived from lambda phage There are any number of different well-known promoters, such as the α- and β-actin systems. optionally together with an operator sequence, controls expression and initiates and completes transcription and translation The ribosome binding site sequence for the purpose of the synthesis of the ribosome-binding site sequence is also included. Anti-VP1 polypeptides can also be expressed in combination with baculovirus vectors. Alternatively, cultured insect cells may be used.

[0203] In other embodiments, mammalian host cells are used to express and express the anti-VP1 polypeptides of the disclosure. For example, they can be used to generate hybrids that express endogenous immunoglobulin genes. myeloma cell lines (e.g., myeloma hybridoma clones described in the Examples) or exogenous These may be mammalian cell lines carrying expression vectors. Includes mortal, normal or abnormal immortal animal or human cells. For example, CH O cell lines, various COS cell lines, HeLa cells, myeloma cell lines, transformed B cells Some cells, such as cells and hybridomas, are capable of secreting intact immunoglobulins. Suitable host cell lines have been developed for the expression of mammalian tissue cells. The use of cell cultures is described, for example, in Winnacker, From Genes to Clones, VCH Publishers, NY, NY .Y., 1987. Expression vectors for mammalian host cells are replicable. Expression control sequences such as origins, promoters, and enhancers (e.g., Queen et al., Immunol. Rev. 89:49-68, 1986), as well as a ribosome binding site, RNA Necessary processes such as splice sites, polyadenylation sites, and transcription terminator sequences These expression vectors usually contain a mammalian gene or The promoters include those derived from mammalian viruses. developmental, cell type-specific, stage-specific, and / or modulatable Alternatively, it may be regulatable. Useful promoters include, but are not limited to: However, the metallothionein promoter, the constitutive adenovirus major late promoter, methasone-inducible MMTV promoter, SV40 promoter, MRP pol II I promoter, constitutive MPSV promoter, tetracycline-inducible CMV promoter promoters (such as the human immediate early CMV promoter), constitutive CMV promoters, and Examples of promoter-enhancer combinations include those known in the art.

[0204] Methods for introducing expression vectors containing polynucleotide sequences of interest into cells include For example, calcium chloride transfection is used for prokaryotic cells. Although calcium phosphate treatment or electroporation are commonly used for other cell hosts, Microporation may also be used (see generally Sambrook et al., supra). Other methods include electroporation, calcium phosphate treatment, Liposome-mediated transformation, injection and microinjection, ballistic method , virosomes, immunoliposomes, polycation:nucleic acid conjugates, naked DNA A, Artificial virion, fusion with herpesvirus structural protein VP22 (Elliot and O'H are, Cell 88:223, 1997), drug-enhanced uptake of DNA, and ex vivo phenotypes For long-term, high-yield production of recombinant proteins, stable expression is used. For example, cell lines stably expressing anti-VP1 antibody chains or binding fragments are often desirable. The vector is inserted into a vector containing a viral origin of replication or an endogenous expression element and a selectable marker gene. After the vector is introduced, the cells are cultured in a rich medium. After 1-2 days of growth, the cells can be switched to selective media. The presence of the introduced sequence confers resistance to selection in selective media. This allows for the growth of cells that successfully express the resistant stably transfected cells. , can be grown using tissue culture techniques appropriate to the cell type.

[0205] Therapeutic and Diagnostic Uses The antibodies and antibody fragments (e.g., antigen-binding fragments) of the present disclosure include, but are not limited to: These compounds are useful in a variety of applications, including the treatment of polyomavirus infection and disease. In such cases, antibodies, antibody fragments (e.g., antigen-binding fragments) may neutralize BKV or JCV infection. and BK virus nephropathy, e.g., BKVAN, are useful for preventing or treating the The method of use may be in vitro, ex vivo, or in vivo. may be.

[0206] In one embodiment, the antibody, antibody fragment (e.g., antigen-binding fragment) detects the presence of BKV in a biological sample. As used herein, the term "detecting" is useful for detecting the presence of a substance. In certain embodiments, the biological sample comprises cells or tissue. In certain embodiments, such tissues express BKV at higher levels compared to other tissues. This includes normal and / or cancerous tissues that express the tumor.

[0207] In one aspect, the present disclosure provides a method for detecting the presence of BKV in a biological sample. In certain embodiments, the method comprises treating a biological sample with an anti-V antibody under conditions that allow binding of the antibody to the antigen. P1 antibody and whether a complex is formed between the antibody and the antigen. Biological samples include, but are not limited to, urine or blood samples. Good too.

[0208] Also included are methods for diagnosing disorders associated with expression of the BKV or JCV virus. In certain embodiments, the method comprises contacting the test cell with an anti-VP1 antibody; The expression level of BK virus in test cells was determined by detecting antibody binding to BK virus. determining the level (quantitatively or qualitatively); and determining the level of infection in the test cells. Control cells (e.g., normal cells or non-BK virus-infected cells of the same tissue origin as the test cells) and comparing the level of BK virus infection in the test cells compared to the control cells. High levels of BK virus present in cells may be associated with BK virus infection and disorders. In certain embodiments, the test cells are those suspected of having a BK virus infection. It is obtained from individuals who are

[0209] In certain embodiments, the diagnostic or detection methods, such as those described above, involve the detection of BKV-infected cells. Detecting the binding of anti-VP1 antibodies to BKV-infected cells. An exemplary assay for determining this is the "FACS" assay.

[0210] Certain other methods can be used to detect binding of anti-VP1 antibodies. Such methods include, but are not limited to, Western blot, radioimmunoassay, and the like. ELISA (enzyme-linked immunosorbent assay), "sandwich" immunoassay, immunoassay Immunoprecipitation assay, fluorescent immunoassay, protein A immunoassay, and immunohistochemistry Antigen binding assays well known in the art, such as immunohistochemistry (IHC), are also included.

[0211] In certain embodiments, the anti-VP1 antibody is labeled. Labels include, but are not limited to, However, labels or moieties that are directly detected (fluorescent, chromogenic, electron-dense, chemiluminescent, and radioactive) and those that are indirectly detected, for example, via enzymatic reactions or molecular interactions. Examples of such moieties include enzymes or ligands.

[0212] In certain embodiments, the anti-VP1 antibody is immobilized on an insoluble matrix. Isolates anti-VP1 antibodies from any BKV or JCV proteins that remain free in the solution This requires adsorption to a water-insoluble matrix or surface (Benn ich et al., U.S. Pat. No. 3,720,760) or by covalent coupling. Anti-VP1 antibodies can be cross-linked (e.g., using glutaraldehyde cross-linking) prior to the assay procedure. Antibody insolubilization or, for example, immunoprecipitation with anti-VP1 antibody and BKV or by insolubilizing anti-VP1 antibodies after complex formation with JCV proteins. is conveniently achieved.

[0213] Any of the above aspects of diagnosis or detection may be used in place of or in addition to another anti-VP1 antibody. Additionally, it can be performed using the anti-VP1 antibodies of the present disclosure.

[0214] In one aspect, the present disclosure provides a method for administering an antibody, antibody fragment (e.g., an antigen-binding fragment) to a patient. Treating, reducing the likelihood of, or preventing a disease, including treating a disease by In certain embodiments, antibodies, antibody fragments (e.g., antibodies The diseases treated using the antibody-binding fragments are BK virus or JC virus infections. Examples of BKV and JCV diseases that can be treated and / or prevented include: Although not limited to these, nephropathy, hemorrhagic cystitis, and progressive multifocal leukoencephalopathy (PML) ), interstitial kidney disease, ureteral stenosis, granular cell neuropathy (GCN), vasculitis, colitis, omentum These include meningitis, meningitis, and immune reconstitution inflammatory syndrome (IRIS). Infection is caused by specific binding of anti-VP1 antibodies or antibody fragments (e.g., antigen-binding fragments). The present invention characterizes BKV- or JCV-expressing cells capable of

[0215] The present disclosure provides methods for treating cancer by administering a therapeutically effective amount of an antibody, antibody fragment (e.g., an antigen-binding fragment). In certain embodiments, methods for treating BK virus infection and BKVAN are provided, including: In this case, the subject is a human.

[0216] In certain embodiments, the method of reducing BK virus infection comprises administering to a subject a therapeutically effective amount of an anti-BK virus antibody. In certain embodiments, the method includes administering an antibody or antibody fragment (e.g., an antigen-binding fragment) to a subject. In certain embodiments, the subject is a human. In certain embodiments, the subject is immunosuppressed. For subjects, the amount of immunosuppression increased or decreased due to the therapeutic effect of the anti-VP1 antibody. That's fine.

[0217] In certain embodiments, the transplanted tissue is infected with BK virus to which the anti-VP1 antibody binds. Because of the high incidence of BK infection in the general population, kidney transplants require the kidney to be The patient is BK virus positive or the kidney donor is BK virus positive. There is a high probability that either the patient is BK virus positive or that both are BK virus positive. to determine the renal transplant procedure depending on the seropositivity of the kidney donor or transplant recipient. Anti-VP1 antibodies can be administered to kidney transplant recipients before and / or after In another embodiment, the virus is detected in the urine (viruria) or If detected in the blood (viremia), anti-VP1 antibodies can be administered to the patient .

[0218] For the treatment of BK or JCV viral infections, antibodies, or antibody fragments (e.g., antigen The appropriate dose of the antibody (binding fragment) will depend on the type of infection to be treated, the severity and course of the infection, Various factors, such as the response to infection, the development of viral resistance to therapy, previous therapy, and the patient's medical history, may influence the outcome. The antibody may be administered once or over a series of treatments lasting from several days to several months, depending on the factors involved. or cure or reduction of infection (e.g., viruria or kidney damage) The optimal dosing schedule is Joules can be calculated from measurements of drug accumulation in the patient's body and are used to measure the effect of individual antibodies or The relative potency of the antibody or antibody fragment (e.g., antigen-binding fragment) may vary depending on the specific antibody. In such cases, the dose is 0.01 mg to 10 mg (e.g., 0.01 mg, 0.05 mg, 0.1 mg, 0.5mg, 1mg, 2mg, 3mg, 4mg, 5mg, 7mg, 8mg, 9mg or 10 mg) / kg body weight once daily, weekly, monthly, or yearly or In certain embodiments, an antibody, or antibody fragment (e.g., For example, the antigen-binding fragment is administered once every two weeks or once every three weeks. A physician may decide to administer the drug based on the measured half-life and antibody concentration in body fluids or tissues. It is possible to estimate the repetition rate for

[0219] Combination Therapy In certain examples, an antibody, or antibody fragment (e.g., an antigen-binding fragment) of the present disclosure can be administered to other antiviral, antiallergic, antiemetic (or antiemetic), pain reliever, cytoprotective , immunosuppressants, and combinations thereof.

[0220] As used herein, the term "pharmaceutical combination" refers to a fixed combination in one unit dosage form or or two or more therapeutic agents simultaneously, independently, or, in particular, when the combination partners are synergistic, e.g. For example, non-fixed combinations that can be administered separately within a time interval that can exhibit a synergistic effect. Refers to a kit of parts for combined or combined administration.

[0221] The term "combination therapy" refers to two or more therapeutic agents for treating a therapeutic condition or infection as described in this disclosure. Such administration refers to the administration of the above therapeutic agents in a single capsule having a fixed ratio of active ingredients. Alternatively, the co-administration of these therapeutic agents in a substantially simultaneous manner, such as Such administration may involve multiple or separate containers (e.g., capsules) for each active ingredient. The powder and / or liquid may be administered simultaneously in a single dose. The compound can be reconstituted or diluted to the desired dose prior to administration. This also includes the use of each type of therapeutic agent in a sequential manner, at about the same time, or at different times. In either case, the treatment regimen may be in the treatment of a condition or disorder described herein. This provides the beneficial effects of the drug combination.

[0222] Combination therapy may provide "synergistic effects" and may be found to be "synergistic," i.e. That is, the effect achieved when the active ingredients are used together is greater than when the compounds are used separately. The effect is greater than the sum of the effects obtained from the active ingredients in the combined unit dose formulation. (2) Coformulated and administered or delivered simultaneously; (3) delivered by some other regimen When compounds are delivered in alternation therapy, a synergistic effect can be achieved. For example, when administered or delivered sequentially by different injections in separate syringes, Generally, during alternation therapy, an effective dose of each active ingredient is administered sequentially. In combination therapy, effective doses of two or more active ingredients are administered sequentially, i.e., sequentially. The minutes are administered together.

[0223] In one aspect, the present disclosure provides a method for treating a subject in need thereof by administering an antibody in conjunction with immunosuppressive therapy. The present invention provides a method for treating BKV or JCV infection by administering an anti-VP1 antibody to a patient. It acts as a preventative measure against BKV or JC infections that occur as a result of pre- or post-transplant immunosuppressive therapy. Neutralize primary infection or viral reactivation. Examples of immunosuppressive therapy include limited Although not a monophosphate dehydrogenase inhibitor, a purine synthesis inhibitor, calcium Specific examples of immunosuppressive therapeutic agents include neurin inhibitors or mTOR inhibitors. , including but not limited to, mycophenolate mofetil (MMF), mycophenolate sodium benzoate, azathioprine, tacrolimus, sirolimus, and cyclosporine. can be done.

[0224] Pharmaceutical Composition To prepare a pharmaceutical or sterile composition comprising an anti-VP1 antibody, the antibody of the present disclosure may be The composition is mixed with a pharmaceutically acceptable carrier or excipient. It may further contain one or more other therapeutic agents that are suitable for neutralizing.

[0225] Therapeutic and diagnostic agent formulations may be prepared, for example, as lyophilized powders, slurries, aqueous solutions, lotions, or the like. The compound is mixed with a physiologically acceptable carrier, excipient, or stabilizer in the form of a solution or suspension. (See, for example, Hardman et al., Goodman and Gilman's The Pharmacological Basis of Therapeutics, McGraw-Hill, New York, NY, 2001; Genna ro, Remington: The Science and Practice of Pharmacy, Lippincott, Williams, and W. ilkins, New York, NY, 2000; Avis, et al. (eds.), Pharmaceutical Dosage Forms: Parenteral Medications, Marcel Dekker, NY, 1993; Lieberman, et al. (eds.), Pharm aceutical Dosage Forms: Tablets, Marcel Dekker, NY, 1990; Lieberman, et al. (eds .) Pharmaceutical Dosage Forms: Disperse Systems, Marcel Dekker, NY, 1990; Weine r and Kotkoskie, Excipient Toxicity and Safety, Marcel Dekker, Inc., New York, N. (See .Y., 2000).

[0226] In certain embodiments, the anti-VP1 antibody is a lyophilizate in a vial containing the antibody. The lyophilized product can be reconstituted with water or a suitable pharmaceutical carrier for injection. For intravenous administration, the resulting solution is usually further diluted in a carrier solution.

[0227] The antibodies disclosed herein may be used to treat BKV in tissue transplant patients who may be immunosuppressed. or useful in neutralizing JCV and therefore receiving CytoGam®. Pharmacology of previously used sucrose and human albumin in bone marrow transplant recipients undergoing steroid therapy. (DeRienzo et al. Pharmacotherapy 2000; 20:1175-8) Alternatively, the anti-VP1 antibody may be used in combination with another antiviral antibody described in WO2003 / 105894. Transplant patients via a pharmaceutical carrier described for the antibody Synagis® In this publication, the pharmaceutical carrier is a histidine and / or glutamic acid derivative. Lysine, saccharides (e.g., sucrose) and polyols (e.g., polysorbates ) was included.

[0228] The choice of dosing regimen for a therapeutic agent depends on the severity of the infection, the level of symptoms, and the biological It depends on several factors, such as the accessibility of the target cells in the matrix. In such cases, the dosing regimen is designed to provide a therapeutic effect delivered to the patient consistent with an acceptable side effect level. Therefore, the amount of biologic delivered will depend on the specific entity and the type of treatment. The choice of appropriate doses of antibodies, cytokines, and small molecules will depend in part on the severity of the condition being treated. Guidelines for selecting a suitable antibody are available (e.g., Wawrzynczak, Antibody Therapy, Biosciences, ntific Pub. Ltd, Oxfordshire, UK, 1996; Kresina (ed.), Monoclonal Antibodies, Cy tokines and Arthritis, Marcel Dekker, New York, NY, 1991; Bach (ed.), Monoclon al Antibodies and Peptide Therapy in Autoimmune Diseases, Marcel Dekker, New York k, NY, 1993; Baert et al., New Engl. J. Med. 348:601-608, 2003; Milgrom et al. , New Engl. J. Med. 341:1966-1973, 1999; Slamon et al., New Engl. J. Med. 344:78 3-792, 2001; Beniaminovitz et al., New Engl. J. Med. 342:613-619, 2000; Ghosh et al. al., New Engl. J. Med. 348:24-32, 2003; Lipsky et al., New Engl. J. Med. 343:15 94-1602, 2000).

[0229] Determining the appropriate dose may, for example, affect or be expected to affect treatment. by a physician using parameters or factors known or suspected in the art to be Generally, the dose should be started at a level somewhat lower than the optimal dose, and then increased to prevent any adverse effects. Increase doses in small increments until the desired or optimal effect is achieved compared to side effects. Important diagnostic measures include, for example, infusion response.

[0230] The acute dose level of the active ingredient in the pharmaceutical composition containing the anti-VP1 antibody was determined to be toxic to the patient. to achieve the desired therapeutic response for a particular patient, composition, and mode of administration so as not to The dosage level selected can be varied to obtain an amount of active ingredient that is effective for the treatment of Bell will consider the neutralizing activity of the antibody, the route of administration, the time of administration, the half-life of the antibody in patients, and the duration of treatment. duration, other drugs, compounds and / or materials used in conjunction with the particular composition used The age, sex, weight, condition, general health and previous medical history of the patient to be treated, and It depends on a variety of pharmacokinetic factors, including similar factors known to the scientific community.

[0231] The composition comprising the antibody or fragment thereof can be administered by continuous infusion or for a period of time, e.g., daily, weekly, Or it can be given in doses spaced 1 to 7 times per week. Can be given subcutaneously, topically, orally, nasally, rectally, intramuscularly, intracerebrally, or by inhalation A particular dosing protocol may be used to determine the maximum dose or dose that avoids significant undesirable side effects. It includes quantity and frequency.

[0232] For the antibodies described herein, the dose administered to a patient is between 0.0001 mg / kg and The dose may be 100 mg / kg of patient body weight. The dose may be 0.0001 mg / kg to 20 mg / kg, 0.0001mg / kg~10mg / kg, 0.0001mg / kg~5mg / kg, 0.0001~2mg / kg, 0.0001~1mg / kg, 0.0001mg / kg~0.75mg / kg, 0.0001mg / kg~0.5mg / kg, 0.0001 mg / kg~0.25mg / kg, 0.0001~0.15mg / kg, 0.0001~ 0.10mg / kg, 0.001-0.5mg / kg, 0.01-0.25mg / kg or 0.01 to 0.10 mg / kg of patient body weight. is calculated by multiplying the patient weight in kilograms (kg) by the dose administered in mg / kg. It can be calculated as follows.

[0233] The antibody dose is then repeated, with administration occurring at least every 1, 2, 3, 5, 10, 1 May be 5 days, 30 days, 45 days, 2 months, 75 days, 3 months, or at least 6 months apart .

[0234] The effective amount for a particular patient will depend on the condition being treated, the patient's overall health, the method of administration, and the patient's condition. The dosage may vary depending on factors such as route and dose, and the severity of side effects (see, e.g., Mayn ard et al., A Handbook of SOPs for Good Clinical Practice, Interpharm Press, Boc a Raton, Fla., 1996; Dent, Good Laboratory and Good Clinical Practice, Urch Publ. ., London, UK, 2001).

[0235] Routes of administration include, for example, topical or cutaneous application, intravenous, intraperitoneal, intracerebral, intramuscular, and intraocular. intraarterial, intracerebrospinal, intralesional injection or infusion, or sustained release system or implant It may be by the body (e.g., Sidman et al., Biopolymers 22:547-556, 1983; Langer et al., J. Biomed. Mater. Res. 15:167-277, 1981; Langer, Chem. Tech. 12: 98-105, 1982; Epstein et al., Proc. Natl. Acad. Sci. USA 82:3688-3692, 1985; Hwa ng et al., Proc. Natl. Acad. Sci. USA 77:4030-4034, 1980; U.S. Pat. No. 6,350, (See US Pat. Nos. 466 and 6,316,024). The composition may also contain a solubilizing agent or a topical agent such as lidocaine to reduce pain at the injection site. In addition, the device may include, for example, an inhaler or nebulizer, and Pulmonary administration may also be employed by use of a formulation containing an aerosolizing agent. Patent Nos. 6,019,968, 5,985,320, 5,985,3 Specification No. 09, Specification No. 5,934,272, Specification No. 5,874,064, No. 5, Nos. 855,913, 5,290,540, and 4,880,078 and PCT International Publication Nos. 92 / 19244 and 97 / 325 Brochure No. 72, Brochure No. 97 / 44013, Brochure No. 98 / 31346 and Brochure No. 99 / 66903 (each of which is incorporated herein by reference in its entirety). See, e.g., U.S. Pat. No. 6,229,629, which is incorporated herein by reference.

[0236] The compositions of the present disclosure can be prepared using one or more of a variety of methods known in the art. It may also be administered by a number of routes of administration. The route and / or mode of administration chosen for the antibody will vary depending on the desired results. Routes of administration include intravenous, intramuscular, intradermal, intraperitoneal, subcutaneous, spinal or intravenous routes, e.g., injection or infusion. Parenteral administration is usually by injection, whereas enteral administration is by other parenteral routes. and modes of administration other than topical administration, including but not limited to intravenous, intramuscular, Intra-arterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, joint These include intrathecal, subcapsular, subarachnoid, intraspinal, epidural and intrasternal injections and infusions. Alternatively, the compositions of the present disclosure can be administered parenterally, such as by topical, epidermal, or mucosal routes of administration, e.g., For example, it can be administered intranasally, orally, vaginally, rectally, sublingually or topically. In an embodiment, the antibody of the present disclosure is administered by infusion. In another embodiment, the antibody is administered subcutaneously. It is administered.

[0237] When administering the antibodies of the disclosure in a controlled or sustained release system, the antibodies may be administered using a pump. Controlled or sustained release can be achieved (Langer, supra; Sefton, CRC Crit. R ef Biomed. Eng. 14:20, 1987; Buchwald et al., Surgery 88:507, 1980; Saudek et al ., N. Engl. J. Med. 321:574, 1989). Polymeric materials can be used to bind antibodies. Controlled or sustained release of the therapeutic agent can be achieved (e.g., in medical applications). ns of Controlled Release, Langer and Wise (eds.), CRC Pres., Boca Raton, Fla., 1 974; Controlled Drug Bioavailability, Drug Product Design and Performance, Smole n and Ball (eds.), Wiley, New York, 1984; Ranger and Peppas, J. Macromol. Sci. R See ev. Macromol. Chem. 23:61, 1983; also Levy et al., Science 228:19 0, 1985; During et al., Ann. Neurol. 25:351, 1989; Howard et al., J. Neurosurg. 7 1:105, 1989; U.S. Patent No. 5,679,377; U.S. Patent No. 5,916,59 No. 7; U.S. Patent No. 5,912,015; U.S. Patent No. 5,989,463 Specification; U.S. Patent No. 5,128,326 Specification; PCT Publication No. 99 / 15154 See also FRET; and PCT Publication WO 99 / 20253). Examples of polymers used in sustained release formulations include, but are not limited to, poly(vinyl alcohol). Poly(2-hydroxyethyl methacrylate), poly(methyl methacrylate), poly(acrylate) acrylic acid), poly(ethylene-co-vinyl acetate), poly(methacrylic acid), polyglycol Poly(N-vinylpyrrolidone), poly(vinyl alcohol), polyanhydrides, poly(N-vinylpyrrolidone), poly(vinyl alcohol), Poly(ethylene glycol), polyacrylamide, poly(ethylene glycol), polylactide (PLA), Poly(lactide-co-glycolide) (PLGA), and polyorthoesters are examples. In one embodiment, the polymers used in the sustained release formulation are inert and leachable. It is free of toxic impurities, stable on storage, sterile, and biodegradable. The system or sustained release system is placed in proximity to the prophylactic or therapeutic target, thus reducing the systemic dose by only a small fraction of the total dose. It is possible to require only a portion (e.g., Goodson, in Medical Applications of Controlled Release, supra, vol. 2, pp. 115-138, 1984).

[0238] Controlled-release systems are discussed in the review by Langer, Science 249:1527-1533, 1990. Sustained release formulations comprising one or more antibodies of the present disclosure can be prepared using any technique known to those of skill in the art. For example, U.S. Pat. No. 4,526,938, PCT International Publication No. WO 2006 / 023106, and the like can be produced. PCT International Publication No. 91 / 05548 Pamphlet, PCT International Publication No. 96 / 20698 Pamphlet Lett, Ning et al., Radiotherapy & Oncology 39:179-189, 1996;Song et al., PDA Journal of Pharmaceutical Science & Technology 50:372-397, 1995;Cleek et al., P ro. Int'l. Symp. Control. Rel. Bioact. Mater. 24:853-854, 1997; and Lam et al. , Proc. Int'l. Symp. Control Rel. Bioact. Mater. 24:759-760, 1997 (respectively, (which is incorporated herein by reference in its entirety).

[0239] When the antibodies of the present disclosure are administered topically, they may be applied as an ointment, cream, transdermal patch, lotion, or the like. , gel, spray, aerosol, solution, emulsion, or other form known to those skilled in the art. For example, Remington's Pharmaceutical Sciences and Introductory uction to Pharmaceutical Dosage Forms, 19th ed., Mack Pub. Co., Easton, Pa. (199 For non-sprayable topical dosage forms, a carrier or Viscous solutions containing one or more excipients and, in some cases, having a dynamic viscosity higher than that of water. Semi-solid or solid forms are typically used. Suitable formulations include, but are not limited to: However, if necessary, they may be sterilized or may be modified to have various properties, e.g., osmolality. Auxiliary substances (e.g., preservatives, stabilizers, humectants, buffers, or salts) that affect the ) mixed with solutions, suspensions, emulsions, creams, ointments, powders, liminal Other suitable topical dosage forms include, in some cases, solid or The active ingredient in combination with a liquid inert carrier is mixed with a pressurized volatile substance (e.g., freon). sprays, which are packed in a mixture with a gaseous propellant such as ethanol or in a squeeze bottle; Possible aerosol preparations include: Optionally, a moisturizer or humectant Examples of such additional ingredients can also be added to pharmaceutical compositions and dosage forms. It is well known in

[0240] When the antibody-containing composition is administered intranasally, it may be in the form of an aerosol, spray, mist or can be formulated in the form of drops. In particular, prophylactic or The therapeutic agent is added to a suitable propellant (e.g., dichlorodifluoromethane, trichlorofluoromethane, with the use of a suitable gas such as chlorofluoroethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas Conveniently delivered in the form of an aerosol spray presentation from a pressurized pack or nebulizer In the case of pressurized aerosols, by providing a valve to deliver a metered amount The dosage unit can be determined by mixing the compound with a suitable carrier such as lactose or starch. Capsules for use in inhalers or insufflators containing a powder mixture with a powder base and cartridges (eg, composed of gelatin) can be formulated.

[0241] A second therapeutic agent, e.g., an immunosuppressant, cytokine, steroid, chemotherapeutic agent, antibiotic Methods for co-administration or treatment with steroids or radiation therapy are known in the art (e.g., For example, Hardman et al., (eds.) (2001) Goodman and Gilman's The Pharmacological Bas is of Therapeutics, 10th ed., McGraw-Hill, New York, NY; Poole and Peterson (e ds.) (2001) Pharmacotherapeutics for Advanced Practice:A Practical Approach, Lip Pincott, Williams & Wilkins, Phila., Pa.; Chabner and Longo (eds.) (2001) Cancer See Chemotherapy and Biotherapy, Lippincott, Williams & Wilkins, Phila., Pa. An effective amount of a therapeutic agent is one that alleviates symptoms by at least 10%; at least 20%; or at least It can be reduced by as little as about 30%; at least 40%, or at least 50%.

[0242] Additional therapies (e.g., prophylactic or therapeutic agents) that can be administered with anti-VP1 antibodies ) less than 5 minutes apart, less than 30 minutes apart, 1 hour apart, about 1 hour apart, or about 5 minutes apart from the anti-VP1 antibody of the present disclosure. Leave 1 hour between each, leave about 1 to 2 hours between each, leave about 2 to 3 hours between each, leave about 3 to 4 hours between each Leave for about 4 to 5 hours, leave for about 5 to 6 hours, leave for about 6 to 7 hours Then, wait about 7 to 8 hours, then about 8 to 9 hours, then about 9 to 10 hours. Then, wait about 10 to 11 hours, then wait about 11 to 12 hours, then wait about 12 to 14 hours. Leave 18 hours apart, leave 18-24 hours apart, leave 24-36 hours apart, leave 36-4 8 hours apart, 48-52 hours apart, 52-60 hours apart, 60-72 hours apart After a certain time, after 72 to 84 hours, after 84 to 96 hours, or after 96 hours Two or more therapies may be administered within the same patient visit. It may also be administered to

[0243] In certain embodiments, the anti-VP1 antibody is administered in a manner that ensures proper distribution in vivo. For example, the blood-brain barrier (BBB) ​​excludes many highly hydrophilic compounds. To ensure that anti-VP1 antibodies cross the BBB (if necessary), They can be formulated, for example, in liposomes. See, for example, U.S. Patent Nos. 4,522,811 and 5,374,548. and 5,399,331. Liposomes can be used to deliver specific cell or The drug may contain one or more moieties that are selectively transported into a target organ or organ, thus providing targeted drug delivery. (See, e.g., Ranade, (1989) J. Clin. Pharmacol. 29:685. Examples of targeting moieties include folic acid or biotin (see, e.g., U.S. Patent No. 6,229,393 to Low et al.). 5,416,016); mannosides (Umezawa et al., (1988) B iochem. Biophys. Res. Commun. 153:1038); Antibodies (Bloeman et al., (1995) FEBS Lett 357:140; Owais et al., (1995) Antimicrob. Agents Chemother. 39:180);Surfactant Protein A receptor (Briscoe et al., (1995) Am. J. Physiol. 1233:134);p120 (Schreier et al., (1994) J. Biol. Chem. 269:9090), K. Keinanen; ML Laukkanen (1994) FEBS Lett. 346:123; JJ Killion; IJ Fidler (1994) Immunom See also Methods 4:273.

[0244] The present disclosure provides pharmaceutical compositions comprising antibodies, alone or in combination with other therapies, to those in need thereof. Protocols for administering combination therapy (e.g., prophylactic or Therapeutic agents (or therapeutic agents) can be administered to a subject simultaneously or sequentially. Therapeutic agents (e.g., prophylactic or therapeutic agents) can also be administered cyclically. of a therapy (e.g., a drug) to reduce the development of resistance to one of the therapies (e.g., a drug). to avoid or reduce one side effect and / or improve the efficacy of the therapy administration of a first therapy (e.g., a first prophylactic or therapeutic agent) for a period of time, followed by administration of a second therapy (e.g., a first prophylactic or therapeutic agent) for a period of time; administering a second therapy (e.g., a second prophylactic or therapeutic agent), and repeating this sequential administration; , i.e., includes a period.

[0245] The therapies (e.g., prophylactic or therapeutic agents) of the combination therapies disclosed herein are administered concurrently to a subject The term "concurrently" refers to the administration of therapy (e.g., prophylactic or therapeutic agents) at exactly the same time. Rather, the present invention is not limited to administration of a pharmaceutical composition comprising an antibody or fragment thereof to a subject. , and that the antibody works together with other therapies(ies) to improve outcomes compared to when they are administered separately. This means that the doses are administered at intervals that provide increased benefit. The therapies can be administered simultaneously or sequentially at different times and in any order; However, if not administered simultaneously, they should be administered close enough in time to achieve the desired treatment or Each therapy may be administered in any suitable form and at any They can be administered separately to a subject by any suitable route. (e.g., prophylactic or therapeutic agents) less than 15 minutes apart, less than 30 minutes apart, less than 1 hour apart Leave it open, leave it open for about 1 hour, leave it open for about 1 to 2 hours, leave it open for about 2 to 3 hours, leave it open for about 3 hours ~ Leave about 4 hours between each, about 4 hours to about 5 hours between each, about 5 hours to about 6 hours between each, about 6 hours~ Leave about 7 hours between them, leave about 7 to 8 hours between them, leave about 8 to 9 hours between them, leave about 9 to 10 hours between them 10 hours apart, about 10 to 11 hours apart, about 11 to 12 hours apart, 24 The subjects are given the doses at intervals of 1 hour, 48 hours, 72 hours, or one week. In some embodiments, two or more therapies (e.g., prophylactic or therapeutic agents) are administered within the same patient visit. Administer to.

[0246] The prophylactic or therapeutic agents of the combination therapies can be administered to a subject in the same pharmaceutical composition. Alternatively, the prophylactic or therapeutic agents of the combination therapies are administered simultaneously to a subject in separate pharmaceutical compositions. The prophylactic or therapeutic agents may be administered to the target subject by the same or different routes of administration. It may also be administered to [Example]

[0247] Example 1: Generation of anti-VP1 antibodies B cells expressing anti-VP1 antibodies were lysed, and VH (heavy) and VL (light) chains were isolated by RT-PC. The DNA was sequenced and analyzed by R to identify important post-translational modification (PTM) sites. The VH and VL chain plasmids were then cloned into an IgG1 backbone vector for expression of a complete IgG1 antibody. The vector was transfected into a CHO mammalian cell line.

[0248] Methods for the production of monoclonal antibodies using hybridoma technology are well known in the art. (Antibody Methods and Protocols, Methods in Molecular Biology vol. 901, 2012, Chapter 7: 117). Briefly, various prime-boost strategies, immunogen doses, and and adjuvants (including but not limited to Freund's adjuvant and MF Female Balb / c mice were infected with BKV serotype I, ... immunized with VLPs derived from serotype IV, and JCV (individually or in combination). The supernatant of successfully fused (growing) hybridomas was analyzed by ELISA using anti-VP 1. After screening for the presence of antibodies, functional activity was assessed in a neutralization assay. The CDRs from the selected mouse IgG were then inserted into a human framework. Humanized by grafting onto the acceptor template, a mammalian IgG1 backbone expression vector The antibody was cloned into a CHO mammalian cell line for expression of the complete IgG1 antibody. Transfected.

[0249] Methods for the generation of monoclonal antibodies using phage display technology are known in the art. (Antibody Methods and Protocols, Methods in Molecular Biology, vol. 901, 2012, Chapter 3: 33). Briefly, a human B cell antibody in the scFv format containing Vκ was synthesized. The cellular antibody library was subjected to three rounds of selection at increasing stringency. Streptavidin-conjugated magnetic beads complexed with thiotinylated BKV serotype IV VLPs. The isolates were screened for anti-VP1 antibodies by solution panning using the 5'-dimer. was first expressed as an scFv and identified as a pentamer of BKV serotype IV VLPs by ELISA. Selected isolates were then screened for binding to both cloning The antibodies were then cloned, expressed as IgG1, and tested for binding to VP1 (serotypes I and IV) by ELISA. Reanalyzed for binding and for functional activity in neutralization assays, intact IgG 1 antibody was transfected into a CHO mammalian cell line for expression.

[0250] A summary of anti-VP1 antibodies is provided in Table 3.

[0251] [Table 3-1]

[0252] [Table 3-2]

[0253] Example 2: Affinity maturation of anti-VP1 antibodies Anti-VP1 antibodies were affinity cloned in yeast by error-prone PCR or CDR-directed mutagenesis. The VP1 proteins from each of the four serotypes of BKV (shown in Table 4) were ) were used as antigens in up to three rounds of selection by FACS analysis. Then, VH (heavy) chains and / or VH chains with enhanced binding affinity to VP1 were analyzed by FACS analysis. Alternatively, the VL (light) chain can be cloned into a mammalian IgG1 backbone expression vector to express the complete It was transfected into a CHO mammalian cell line for expression of an IgG1 antibody.

[0254] [Table 4]

[0255] Example 3: Production of BK virus and virus-like particles (VLPs) A genomic clone of BKV serotype I was obtained from ATCC (pBR322-BKV MM, catalog). pBR322-BKV Dunlop, Catalog No. 45025) The infectious genomic clones of serotypes II, III, and IV chimeric viruses were obtained from , using a previously described cloning strategy (Broekema et al, Virology 2010 407:368-373) Briefly, unique restriction sites (SacII, PmlI) were inserted into the , using site-directed mutagenesis to insert frankin into the VP1-VP2-VP3 coding region. The serotype II isolate SB (GenBank accession number SB) was introduced into the BKV serotype I genome. Accession number CAA79596.1), serotype III isolate AS (GenBank accession number A AA46882.1) and serotype IV strain ITA-4 (GenBank accession number BAF The coding region of VP1 derived from 75132) was synthesized by the method of Broekema et al., supra. to encompass the SacII-PmlI region used for the swap combination described in VP2 / VP from sea urchin, serotype I isolate (Genewiz, La Jolla, CA) The resulting chimeric genomic clone was then used to synthesize the following: As previously described (Abend et al, J. Virology 2007 81:272-279), primary proximal urinary High titer in canalicular tubular epithelial (RPTE) cells (ATCC, Cat. No. PCS-400-010) A titer infectious virus stock was generated.

[0256] VLPs representing each of the four BKV serotypes were subjected to VP1 expression in Sf9 insect cells. Generate 1000kJ / mL of lysed lysates by microtip sonication (3x 45 sec pulses, 5 min between pulses on ice). min resting), by pelleting VLPs through a 20% sucrose cushion. isolation (116,000 g for 2.5 hours) and 5 ml of GE HiTrap QH Anion chromatography using a P column (GE Healthcare, Pittsburgh, PA) After extraction from frozen cell pellets derived from 1 L cultures by purification by ion exchange, 10 ml of Capto™ Core 700 (GE Healthcare, Pittsburgh) Purification was performed using a size exclusion column based on (urgh, PA) resin, and finally GE Sep hacryl S500 26 / 60 (GE Healthcare, Pittsbur) The VLPs prepared were purified on a size exclusion column (gh, PA). It was used in ELISA and SPR-based binding assays.

[0257] Example 4: Purification of BKV VP1 pentamers The VP1 proteins from each of the four serotypes of BKV (shown in Table 5 below) The 6xHis-TEV sequence was cloned into pGEX vector using the N-terminal GST-6xHis-TEV sequence. subcloned into a target vector (GE Healthcare, Pittsburgh, PA). The GST fusion protein was expressed in E. coli and purified by microfluidics. Extract from the cell pellet using a disperser (15,000 PSI) and place in 20 ml of nickel. Lucepharose 6 Fast Flow column (GE Healthcare, Pitt Immobilized metal ion affinity chromatography (IMA) was performed using a 100-well platelet-free chromatograph (PPE) in the presence of 1000 uL ... The GST-6xHis-TEV tag was purified by ELISA using the TEV protease overnight. Incubate and cleave the fragments in 5 ml of His-Trap Fast Flow. w column (GE Healthcare, Pittsburgh, PA), followed by Sup Erdex 200 26 / 60 size exclusion column (GE Healthcare, Pi Final purification was performed using a HPLC (St. Johns Hopkins, PA).

[0258] [Table 5]

[0259] Example 5: Affinity measurement of anti-VP1 antibodies (SET assay) Using a solution equilibrium titration (SET) assay, antibodies and BK from all four serotypes were identified. V Interaction affinity with VP1 pentamer (K D The antibody was added at a constant concentration of 1 pM. The VP1 pentamer was serially diluted from a starting concentration of 10 nM. After overnight incubation of the solution, the VP1 pentamer-coated MSD array plates ( Meso Scale Discovery, Catalog Number L21XA, Rockvil The unbound antibody was assayed using a 1:1 fit model. By adapting it to Dell, D was determined (Piehler et al. J. Immunol. Methods . 1997; 201(2):189-206).

[0260] In the SET assay, K D The values ​​ranged from 0.9 to 5.0 pM, and the BKV serotype I 5 The binding of P8D11 and its derivatives was similar to that of anti-VP1 antibodies to the VP1 dimer. Equivalent K for binding to BKV serotypes II, III, and IV pentamers D has a value, At least 3.5-fold higher affinity for serotype II pentamers compared to other antibodies and had 47-fold higher affinity for the serotype IV pentamer. Furthermore, P8D11 and derivatives of P8D11 are shown in Fig. 1D. The binding affinities for β-lactamase (β-lactamase) ranged from 2.5 to 6.0 pM, whereas other antibodies showed binding affinities for β-lactamase (β-lactamase) in the range of 2.5 to 6.0 pM under the conditions tested. These anti-VP1 antibodies did not detectably bind to serotype III pentamers within the A summary of the SET affinity data can be found in FIG.

[0261] Example 6: Binding of anti-VP1 antibodies to VP1 pentamers and VLPs (ELISA) Binding of anti-VP1 antibodies to VP1 pentamers and VLPs was analyzed by ELISA. Briefly, Immulon 2HB plates (VWR, 62402-972) were Plates were coated overnight with 100 ng / well of BKV VLP or VP1 pentamer. Antibodies were added at 0. Serial dilutions were made in PBS containing 5% BSA and allowed to bind to antigen-coated plates for 2 h. After washing the plate with PBS, a 1:6000 dilution of 0.5% BSA in PBS was added. Secondary antibody (HRP-conjugated rabbit antibody to human IgG, Southern Bio The plates were incubated with PBS. Wash and add tetramethylbenzidine (TMB) microwell peroxidase substrate (KP L, 52-00-03 1L) was used to carry out the reaction.

[0262] Anti-VP1 antibodies EBB-C1975-A3, A7, E7, and B5 were identified as BKV serotype IV. VLPs derived from VP1 (IC50 ranging from 0.044 to 0.1 nM) or VP1 pentamers (0 showed similar binding to serotype I antibodies (IC50s ranging from 0.026 to 0.078 nM). showed reduced and more variable binding activity to VLPs (ranging from 4.32 to 85.7 nM). The data are illustrated in Figures 4-6 and summarized in Figure 7. In contrast, 2081 and Anti-VP1 antibodies from the 2075 series and 2076 series showed enhanced activity against serotype I VLPs. The binding activity was observed with IC50 values ​​ranging from 0.046 to 0.267 nM. The results are shown in Figures 8 and 9. The 2077 series JCV-specific anti-VP1 antibodies showed a 0.034 The binding activity to JCV VLPs ranged from 0.651 nM, and the data are shown in Figure 10 and provide 11.

[0263] Example 7: Binding of anti-VP1 antibodies to VP1 pentamers and VLPs by SPR The binding of anti-VP1 antibodies to VP1 pentamers and VLPs was analyzed by surface plasmon resonance (SPR). Briefly, biotinylated protein A was analyzed by streptavidin. Immobilized on a DNA-coated SPR chip surface and bound to Protein A. The anti-VP1 antibody is then captured. BKV VP1 pentamers or VLPs are then washed over the surface. Binding to anti-VP1 antibodies during the binding step is followed by buffer washing during the dissociation step.

[0264] Using SPR, the virulence of four BKV serotypes was compared with a positive control (P165E2). The binding of anti-VP1 antibodies EBB-C1975-A3, A7, E7, and B5 was evaluated. All species antibodies had very similar binding profiles to the VP1 pentamer: Atypical binding to serotype II pentamers but no binding to serotype I and III pentamers (large bulk shift and no return to baseline), and similar to P165E2 It binds to serotype IV pentamers but with lower affinity (Figures 3A, 3C, and 3E). For VLPs, EBB-C1975-A3, A7, and E7 have similar binding profiles. Sharing files: Atypical binding to serotype I VLPs and serotype III VLs However, the binding profile of EBB-C1975-B5 was similar to that of serum P. Although there was no significant binding to types I and III VLPs, there were distinct epitopes on VP1. The binding of the ATP to the ATP-binding fragments was shown in Figures 3B and 3D.

[0265] We also used SPR to convert anti-VP1 to VP1 pentamers by scanning alanine mutagenesis. 1. Characterizing the Binding of Antibodies P165E2, NEG447, P7G11A, and P8D11 All of the anti-VP1 antibodies showed binding to the VP1 pentameric structure (Fig. 13A-F and Fig. 14). The overall effect of the natural mutations was a decrease in binding to the F66A and I145A VP1 mutants. Furthermore, K69A and E82A were also expressed in P165E (Fig. 13B and 13F). 2, NEG447, and P7G11A binding (Figures 13D and 13E).

[0266] Example 8: Anti-VP1 antibodies bind to conformational epitopes To determine whether anti-VP1 antibodies bind to conformational epitopes, SDS-PA was performed. Western blot of denatured proteins by GE and their native conformation Briefly, the V of BKV serotype I or IV was used for protein dot blot analysis. P1 pentamers are run on SDS-PAGE and transferred to nitrocellulose membranes (Western blot) or spotted directly onto a nitrocellulose membrane (dot blot). The membrane was then incubated with anti-VP1 antibody and then detected using the Licor Odyssey system. Incubation with an anti-human IgG secondary antibody conjugated to an infrared fluorescent dye for I did.

[0267] A commercially available positive control antibody (Abca) known to recognize a linear epitope was used. m 53977) detected both denatured and non-denatured VP1. 5E2, P7G11, and P8D11 detect denatured VP1 on Western blots. The VP1 fragments were unable to be detected by the VP1 gene and only recognized native VP1 on dot blots, which is likely due to the lack of these fragments. These results indicate that the antibody binds to a conformational (non-linear) epitope of VP1 (Fig. 12A and and 12B).

[0268] To further characterize the epitopes of anti-VP1 antibodies, we investigated the epitopes primarily exposed on the virion surface. The primary interaction site for cell surface receptors in the VP1 BC loop is known to Scanning alanine mutagenesis was performed on residues within these mutants. 1 pentamer was detected in the surface plasmon resonance (SPR) assay described above in Example 7. 1 and P7G11A. Mutations at several positions were The binding of P7G11A was affected by F66A, K69A, E82A, and I145A (Figure 13 A–F and Figure 14). However, mutations at only two sites were observed in P8D11 This resulted in decreased binding (F66A, I145A) (Figure 14). Mutations in this region resulted in loss of binding for all antibodies tested, making it difficult to support any one theory. Although not intended to be binding, these mutations result in a global disruption of VP1 pentameric structure. All other VP1 pentamers with BC loop mutations tested In contrast, hydrogen-deuterium exchange studies revealed that P8D11 A conserved region within the EF loop of VP1 during Fab fragment binding was identified. Up-scanning alanine mutagenesis studies identified the critical region for P8D11 binding within this region. The important contact residues include Y169, R170, and K172, and D / E175, K181, and N1 82, T184 and Q186 to M190 are important as determined by deuterium exchange. It is confirmed that the residue is ((YRXKXX(D / E)XXXXXKNXTXQ)( SEQ ID NO: 500) This is further described in Examples 14 to 17.

[0269] Example 9: Neutralization of BK virus by anti-VP1 antibodies Infectious BKV serotype I and chimeric viruses representing serotypes II, III, and IV The cells were pre-incubated with purified antibodies for 1 hour to allow binding and neutralization. Primary renal proximal tubule epithelial (RPTE) cells (ATCC, catalog number PCS-40 0-010) were exposed to the virus-antibody mixture for 4 hours, replaced with fresh medium, and incubated for 48 hours. The cells were then incubated in 4% paraformaldehyde to allow for viral entry and gene expression. Aldehyde fixation and analysis by immunofluorescence were performed to detect TAg expression (Calbio chem DP02, pAb416 Mouse anti-SV40 TAg antibody). Cellomic High-content fractions using the ArrayScan® VTI HCS Reader Immunofluorescence was analyzed by image analysis to identify the patterns of BKV-infected cells (TAg-positive, DAP1-positive). The concentrations were quantified and the data expressed as percent inhibition of infection compared to untreated control wells. and presented it.

[0270] As shown in Figures 15-23, infection with all four serotypes of BKV (I-IV) was Anti-VP1 antibodies, including a subset of neutralizing antibodies, neutralized infection by BKV. The anti-VP1 antibodies from these researchers are P8D11, modified P8D11, and EBB-C1975-B5. This particularly includes:

[0271] Example 10: Neutralization of JC virus by anti-VP1 virus antibodies The infectious JCV isolates Mad-1 and Mad-4 have identical VP1 sequences (Ge These JCV isolates were analyzed with purified antibodies. The antibody was pre-incubated for 1 hour to allow for binding and neutralization. V40 TAg-expressing African green monkey kidney fibroblast-like cell line, ATCC catalog The cells (labeled CRL-1651) were exposed to the virus-antibody mixture for 4 hours and then replaced with fresh medium. The cells were then incubated for 72 hours to allow for viral entry and gene expression. JCV VP1 expression was determined by fixing with 4% paraformaldehyde and analyzing by immunofluorescence. Detected (Abcam 53977, rabbit polyclonal anti-SV40 VP1 antibody). Cellomics ArrayScan® VTI HCS Reader by high-content image analysis using a fluorochrome (Thermo Fisher, Waltham MA). The assay was analyzed to determine the percentage of JCV-infected cells (VP1-positive, DAP1-positive). The data were presented as percent inhibition of infection compared to untreated control wells. As shown in Figures 24-26, antibodies including the P8D11 and 2077 series antibodies A subset of VP1 antibodies neutralizes infection by JCV.

[0272] Example 11: Viral Resistance Resistance selection experiments using the P8D11 antibody were performed on mice infected with BKV serotype I or serotype IV. The serotype I test was performed in renal proximal tubule epithelial (RPTE) cell cultures. No viral spike was observed in cultures containing P8D11 at the first passage (84 days). No resistance-associated variants (RAVs) were identified. As no further passages were performed beyond this point, no further passages were performed. A sharp rise in virus was detected at passage 3 (day 42). Sequencing of KV VP1 revealed resistance-associated 20 amino acid changes throughout VP1. Variants (RAVs) were identified, but clusters around specific amino acids in the VP1 sequence Subsequent phenotypic characterization of this pooled RAV virus revealed no change in , a complete loss of neutralizing activity (>7,692-fold EC50) compared to wild-type virus. Although a significant shift in EC50 was observed, the change in EC50 for P8D11 was small (3.9-fold). Furthermore, the VP1 mutant E82K was identified as a RAV during selection with another anti-VP1 antibody. Characterization of the E82K mutant virus identified (see Example 8) and cloned Evaluation showed that this variant had an EC50 of 15,880 compared to the wild-type virus. demonstrated a fold shift but no cross-resistance to P8D11 .

[0273] Similarly, in cultures of BKV serotype IV, P8D11 was associated with a virulence factor after six passages (84 days). Again, no resistance was detected. However, resistance to different anti-BK antibodies was not observed. Sex was selected as early as passage 1 (day 14). The L68R and E73K mutations were identified, with 600-fold and 22-fold increases in the EC50 value, respectively. It provided a 7-fold shift but showed no cross-resistance to P8D11. P8D11 has a high resistance barrier and is resistant to both serotype I and IV resistant variants. maintains neutralizing activity against

[0274] Example 12: Toxicity VP1 is an exogenous non-human target that is not expressed on the cell surface and is therefore not disclosed herein. The anti-VP1 antibody used in this study has a low risk of toxicity in humans. 1 showed no staining of 42 human tissues and blood smears, but human proteins This supports the lack of cross-reactivity of anti-VP1 antibodies with proteins. Although the antibody did not show antibody-dependent cell-mediated cytotoxicity (ADCC) in vitro, this is due to the This is consistent with the fact that the P1 protein is not expressed on the host cell surface.

[0275] Example 13: SET affinity assay of P8D11 for JCV VLPs Progressive multifocal leukoencephalopathy (PML) is a rare but lethal disease of the brain in immunocompromised patients caused by the JC virus. It is an infection that is often fatal. JC virus major capsid protein (VP1) is involved in binding to sialic acid receptors on the surface of host cells. Amino acids L55 and S2 Certain mutations in VP1, such as 69, abolish sialic acid recognition and contribute to the pathogenesis of PML. (Chen et al., mAbs 2015; 7(4), 681-692). Mutations occur frequently in PML patients (Gorelik et al., J. Infect. Dis. 2011 204: 103-114 and Reid et al., J. Infect. Dis. 2011; 204:237-244). We tested them to see if they bound to mutated JCV VLPs with mutations at these positions. The binding of anti-VP1 antibodies to these VLPs was confirmed by the presence of these common VP1s. This indicates that the JC virus carrying the mutation is not resistant to therapy.

[0276] Two series of 22-fold serial dilutions of the VLPs were prepared in sample buffer. The P8D11 antibody was added at a concentration of 9 nM or 1 pM. The JCV consensus concentration range was 105 μg / ml to 72 pg The concentration range of the JCV L55F mutant was 300 μg / ml to 143 μg / ml. The concentration range of the JCV S269F mutant was 300 μg / ml to A volume of 60 μl of each VLP:antibody mixture was added to 384 The wells were distributed in duplicate into polypropylene microtiter plates (PP MTPs). Sample buffer served as a negative control, and a sample containing no antigen served as a positive control. (Bmax). The plates were sealed and incubated overnight (o / n) at room temperature (RT). A 384-well standard MSD array plate was filled with 2 and 0.002 μg / ml BKV- Coated o / n with VP1 serotype I pentameric protein. 50 μl / well of washing buffer After washing three times with 50 μl / well of blocking buffer, the plates were incubated for 1 h at RT. After washing, 30 μl / well of each VLP:antibody mixture was added. The mixture was transferred from the PP MTP to the coated MSD plate and incubated for 20 min at RT. After a further washing step, 30 μl of detection antibody (1 A 1:2000 dilution of MSD Protease inhibitor (MSD Protease inhibitor) was added to each well and incubated at RT for 30 min. Wash the plate, add 35 μl / well of reading buffer, and incubate for 5 min. The ECL signal was measured using an MSD SECTOR Imager 6000. It was determined.

[0277] The reagent used was bovine serum albumin (BSA) (VWR catalog number 422351S ), Phosphate Buffered Saline (PBS) 10x (Teknova Catalog No. P0195) , MSD Read Buffer T 4x(Meso Scale Discove ry Catalog No. R92TC-1), Tris-buffered saline (TBS) 20x (Tek nova catalog number T1680), Tween-20 (VWR catalog number 43708 2Q). The buffer used was blocking buffer: 1x PBS + 5% ( (w / v) BSA, Coating buffer: 1x PBS, Sample buffer: 1x PBS + 0.5% (w / v) BSA + 0.02% (v / v) Tween-20, Washing buffer: 1xTB S + 0.05% (v / v) Tween-20 and reading buffer: 1x MSD Re It was an ad Buffer.

[0278] Using a solution equilibrium titration (SET) assay, P8D11 and J were analyzed as described in Example 5. Interaction affinity with CV VLP (K D The P8D11 antibody was administered at 9 nM or Assayed at a constant concentration of 1 pM, JCV VLPs were serially diluted as follows: Census VLPs ranged from 105 μg / ml to 72 pg / ml, with L55F and S2 Both 69F mutant VLPs ranged from 300 μg / ml to 143 pg / ml. After overnight incubation of the antibody:VP1 pentamer solution, the VP1 pentamer-coated MSD Array plate (Meso Scale Discovery, Cat. No. L21XA) The unbound antibody was assayed using a ELISA kit (BioSpectroscopy, Inc., Rockville, MD). By fitting the 1:1 fitting model, K D was determined (Piehler et al. J. Immunol. Methods. 1997; 201(2):189-206). Sapidyne (Boise I D) KinExA® Pro and n-Curve Analysis software from The analysis was performed using software.

[0279] Figure 27 describes the results of the SET assay in tabular form. This data is consistent with the consensus J for CV VLPs and VLPs containing VP1 mutations commonly associated with PML Determination of affinity of P8D11 antibody (K D P8D11 provides a therapeutic effect in the low nanomolar range. However, the L55F mutation The binding affinity of wild-type (consensus) and S269F mutant VLPs to the ribosomal domain was Therefore, this indicates that the P8D11 antibody binds to wild-type JC virus. either the JC virus or the JC virus with mutations commonly associated with PML This indicates that it remains an effective therapy.

[0280] Example 14: Deuterium Exchange Study for Epitope Mapping (BKV VP1 Pentamer and P8D11 Fab in complex with Deuterium exchange mass spectrometry (HDx-MS) measures the incorporation of deuterium into the amide backbone of proteins. These measurements are based on the solvent accessibility of the amide and the hydrogen bonding of the backbone amide. HDx-MS is highly sensitive to changes in the binding network. It is often used to compare two different states of a protein, such as pepsin In such experiments, the skilled artisan will be able to A region, typically 10-15 amino acids, that shows differential deuterium uptake between different states of the protein is identified. The protected regions are those directly involved in ligand binding or those that bind to the ligand. It is allosterically affected by antibody binding to the agonist.

[0281] In these experiments, deuterium incorporation of the BKV VP1 protein (SEQ ID NO: 502) was measured using Measurements were performed in the absence and presence of P8D11 Fab fragment. The region in VP1 that exhibits reduced uptake is likely to be contained within the epitope, but Due to their nature, it is also possible to detect changes distant from the direct binding site (allosteric Generally, the region with the greatest amount of protection is directly involved in binding.

[0282] Epitope mapping experiments were performed using the LEAP® robotic system, nanoAC QUITY® UPLC System, and Synapt® G2 A Waters Synapt® G2 HDx-MS platform, including a mass spectrometer In this method, three sets of control experiments are carried out as follows: VP1 pentamer of KV serotype I was dissolved in 110 μl of 95% deuterated PBS buffer (pH 7 4) and incubate at room temperature on a bench rotator for 25 minutes (%D = 8 Deuterium exchange was performed on ice for 5 min in cold quench buffer (6 M urea and 1 Quench by diluting 1:1 with 1M TCEP (pH=2.5). Afterwards, the tube was transferred onto the LEAP system (thermobox set to 2°C) and quenched. The checked samples are injected onto the UPLC system by the LEAP system for analysis. The PLC system contained an immobilized pepsin column 2.1 mm x 30 mm maintained at 12°C. (Life Technologies 2-3131-00) is incorporated. 35% acetonitrile gradient and Waters UPLC CSH C18 1.0x A 100 mm column is used for the separation. Triplicate experiments are then performed using the antibodies. The P8D11 Fab fragment was bound to protein G agarose beads using standard techniques. Immobilized on Thermo Scientific Cat#22851. Once the antibody is in the suspension, it is centrifuged to remove the storage buffer. Then, 200 μl of PBS is added. Buffer (pH 7.4) and a certain concentration of VP1 pentamer were applied to immobilized P8D11 F Add to the ab fragment and incubate at room temperature for 30 minutes. The cells are centrifuged, washed with 200 μl of PBS buffer, and centrifuged again. For the exchange, 200 μl of deuterated PBS was added for 25 min of incubation at room temperature. The deuterium buffer is then added to the antigen-antibody complex (%D = 85.5%). Remove the supernatant and immediately add 125 μl of ice-cold quench buffer. After quenching for 5 minutes, The column is centrifuged and the effluent is transferred to a pre-chilled HPLC vial. Analyze the samples using the same online pepsin digestion / LC-MS setup.

[0283] The results of these measurements are summarized in Figure 28. Figure 28 shows the results of the control and P8D11 antibody binding. The baseline-corrected difference between the samples divided by the standard error of measurement is shown. In this plot, the more negative the value, the greater the P8D binding to the VP1 pentamer of BKV serotype I. 11 Fab fragments show a greater amount of protection in a given region upon binding. 1. Upon binding of the Fab fragment, the present inventors determined that the amino acids 168 to 190 of the VP1 protein The most significant amount of protection is observed in the bolded and underlined sequences in Table 1 (NYR TKYPXGTXXPKNXTXQSQVM) (SEQ ID NO: 501) As such, this region of the EF loop is expressed in all four serotypes of BK virus and JC virus. It is highly conserved among the genus Illus.

[0284] In conclusion, the deuterium mapping data demonstrate that the P8D11 antibody inhibits the EF loop of BKV VP1. This region binds to an epitope within the BKV serotypes. It is highly conserved among BK and JC viruses, thus P8D11 is involved in all four BK This supports the finding that the neutralizing activity is across serotype V and JC viruses.

[0285] Example 15: Targeted alanine scanning and SEQ ID NO: 1 for epitope mapping of P8D11 PR Scanning alanine mutagenesis using Biacore surface plasmon resonance (SPR) Binding of anti-VP1 antibodies to VP1 pentamers generated for epitope mapping by The experiments were carried out in 0.005% Tween 20 surfactant (Calbiochem in phosphate-buffered saline (PBS) supplemented with PEG-1000 (Cat. No. 655206) at 25°C. The analysis was performed using a Biacore T-200 instrument (GE Healthcare Life Sciences Biotinylated Protein A (Sigma, catalogue number 101111111) was used for electrophoresis. (No. P2165) with approximately 1200 response units (RU) of the streptavidin series S The remaining free streptavidin sites were then immobilized on a sensor chip using biotin-PEG ( Blocking was performed with Pierce EZ-Link (catalog number PI21346). Antibodies were measured on the prepared Protein A sensor chip by a 4-second injection at a flow rate of 0 μl / min. Antibodies were immobilized at 20-40 RU on flow cells 2, 3, and 4, but the flow Cell 1 was left as a reference cell without antibody. VP1 pentamer was then added to the wells in 100 μl. Buffer to monitor dissociation after injection onto the chip at 1000 rpm for 200 sec. Between each pentamer and pentamer concentration, the sensor chip surface was inoculated with 30 μL of HCl. Regenerate by injection of 25 mM NaOH at 1 / min for 60 seconds and then resuspend for the next cycle. The antibody was removed before recapture of the antibody on the protein A surface. Double reference subtraction was applied. Data analysis was performed in BiaEvaluation software. VP1 pentamer binding Evaluation of the effect of alanine mutagenesis on β-glucan was achieved by comparing the levels of bound RU. The shape of the fitting curve was compared to that of the wild-type pentamer.

[0286] As previously discussed, the epitopes for antibodies P8D11 and P7G11A are: The EF loop of BKV VP1 is three-dimensional and discontinuous (Fig. 12A-B). Single mutations to alanine at Y169, R170, and K172 in P8D11 Mutations at Y169 and R170 also abolish binding (Figures 29A and 29B). It also abolishes the binding of the P7G11A antibody, which binds to the EF of BKV VP1. It is not affected by changes at position K172 of the loop (Figures 29A and 29C).

[0287] Example 16: Epitope mapping by x-ray crystallography sc of antibody P8D11 bound to the pentameric form of BKV major capsid protein VP1 The crystal structure of the Fv chain was determined. As detailed below, the scFv:BKV-VP1 pentamer A 5.5:1 solution of the antibody was used to generate a pentamer consisting of five scFv chains linked to each other. Protein crystallography was then used to generate a crystallographically suitable complex of the original protein. The fragment structures were generated and the epitopes were defined.

[0288] Crystallization and structure determination The P8D11 scFv / BKV-VP1 complex was concentrated to 5.2 mg / ml and subjected to crystallization. Crystals for data collection were screened by hanging drop vapor diffusion at 18 °C. The complex was grown in 1.0 μl of 25% (w / v) PEG3350, 0.2 M NaCl solution. 1.0 μl of lysate containing magnesium and 0.1 M Bis-Tris pH 7.0 Mix with reservoir solution and equilibrate the droplet against 350 μl of the same reservoir solution. Crystals were grown overnight and continued to grow for several days. Before filtration, transfer the crystals to 75% reservoir solution + 25% glycerol and briefly cool them in liquid nitrogen. I rejected it.

[0289] Diffraction data were collected in-house on a Rigaku FRE+ copper source and R-axis X-ray diffractometer. The data was collected using Autoproc (Global Phasing, LTD). The BKV-VP1 data were processed and scaled using a cell size of a = 22 4.4Å, b=224.4Å, c=144.04Å, alpha=90°, beta=90° The complex was processed to 2.66 Å in space group P42212 with gamma=90°. The structure of the combined protein was analyzed using Phaser (McCoy et al.) with the BKV-VP1 pentamer as a search model. The fragments were resolved by molecular replacement using the method (J. Appl. Cryst., (2007) 40:658-674). The model was constructed in COOT (Emsley & Cowtan (2004) Acta Cryst. D60:2126-2132). , Buster (Global Phasing, LTD., Cambridge, UK) The Rwork and Rfree values ​​were 17.1% and 21%, respectively. 0.4%; the root mean square (rms) deviations of bond lengths and bond angles are The angles are 0.010 Å and 1.18°.

[0290] Residues of the BKV-VP1 pentamer that contact the P8D11 scFv, the type of interaction, and The surface area and buried surface area were all calculated using PISA (Krissinel et al., (2007) J Mol Biol. 372:774-97). and are listed in Table 6 below. Each monomer of the VP1 pentamer is P8D It was found to contain a single isolated epitope for 11 antibodies. The five scFv domains are arranged in five chemically and sterically equivalent positions in each pentamer. The details of the interaction at each epitope are essentially the same, Only one scFv / VP1-epitope interface is analyzed here.

[0291] Epitope of P8D11-scFv on BKV-VP1 overall structure The overall folding of each polyomavirus VP1 pentameric structure is discussed at the tertiary structure level. The primary sequences are highly conserved with identities ranging from 69 to 85%. Each pentamer consists of a three-stranded β-sheet that stacks against another five-stranded β-sheet. It is composed of five monomers, each consisting of a β-sheet followed by a β-sheet. The 8D11 scFv has a VH and VL domains with a 20 amino acid linker between them. As shown in Figure 30, the VH-VL fusion protein , binds to an epitope located on the lateral outer surface of the BKV-VP1 pentamer.

[0292] Epitope of P8D11 Using the crystal structure of the BKV-VP1 / P8D11 complex, we investigated the P8D on BKV-VP1. Identifying epitopes. The interaction surface on VP1 with P8D11-scFv is Several contiguous and discontinuous (i.e., non-contiguous) sequences are available: It is formed by residues 77-80, 169-186, and 191-192, which are These residues form the three-dimensional conformational epitope recognized by P8D11-scFv. (Figures 31A-B). This epitope, defined by crystallography, was also identified by hydrogen-deuterium exchange mass spectrometry. Residues 168–190 of P8D1 are in good agreement with those defined by HDx-MS. 1-Fab is substantially protected (Figure 28). This was in good agreement with the alanine scan performed on the TYR169 and ARG169 sequences (Figures 29A-C). 170 and LYS172 are contact residues that are part of the epitope of the P8D11 antibody It was shown that...

[0293] P8D11-scFv epitope on BKV-VP1. All residues of BKV-VP1 that contact the Fv were identified by PISA and Enumerate and select by their buried surface area by scFv. Interactions, if applicable. The types are also listed.

[0294] [Table 6-1]

[0295] [Table 6-2]

[0296] Example 18: Formulation The anti-VP1 antibodies described herein are monoclonal antibodies with lambda light chains, IgG1 These antibodies are histidine-sucrose isotypes and can be lyophilized. The anti-VP1 antibody is soluble in the formulation buffer and stable for 4 weeks. Minimally formulated drug substance (e.g., in histidine buffer in the absence of stabilizers) It was soluble at over 200 mg / ml as a steroid.

[0297] For subsequent intravenous administration, the resulting solution is usually further diluted in a carrier solution to Prepare a ready-to-use antibody solution for infusion.

[0298] Important stability-indicating analytical methods for selecting the most stable formulations are, among others, those that determine the level of aggregation. size exclusion chromatography to determine the size of particles that are invisible to the naked eye; The study included clinical trials and efficacy studies.

[0299] The examples and embodiments described herein are for illustrative purposes only and various modifications or variations may be made in light thereof. Any modifications or variations will be suggested to one skilled in the art and fall within the spirit and scope of this application and the appended claims. It is understood that the term "internal" is included within the scope of the present invention.

Claims

1. An antibody or an antigen-binding fragment thereof, which specifically binds to VP1. 。

2. The antibody or antigen-binding fragment thereof binds to VP1 of BK virus serotypes I to IV. The antibody of claim 1 that specifically binds.

3. The antibody or antigen-binding fragment specifically binds to at least one VP1 of Table 1. The antibody of claim 1.

4. The antibody or antigen-binding fragment thereof binds to two or more VP1 serotypes of Table 1. The antibody according to claim 1.

5. the antibody or antigen-binding fragment thereof a) BKV VP1 serotype I and BKV VP1 serotype II; b) BKV VP1 serotype I and BKV VP1 serotype III; c) BKV VP1 serotype I and BKV VP1 serotype IV; d) BKV VP1 serotype II and BKV VP1 serotype III; and e) BKV VP1 serotype I and JCV VP1 The antibody of claim 4, which binds to

6. The antibody or antigen-binding fragment thereof binds to BKV serotype I with a binding affinity of 5.0 pM or less. or binds to BKV serotype II with a binding affinity of 29.0 pM or less, or binds to BKV serotype III with a binding affinity of 185.0 pM or less, or binds to BKV serotype IV with a binding affinity of 436 pM or less, or to JCV with a binding affinity of 436 pM or less The antibody of claim 1 which binds to

7. The antibody or antigen-binding fragment binds to the VP1 epitope (SEQ ID NO: 500 or SEQ ID NO: 501).

8. An antibody, wherein the antibody or antigen-binding fragment thereof comprises: (i) (a) HCDR1 (CDR-complementarity determining region) of SEQ ID NO: 6; (b) SEQ ID NO: 7 (c) a heavy chain variable region comprising an HCDR2 of SEQ ID NO: 8, an HCDR3 of SEQ ID NO: 1, and (e) LCDR1 of SEQ ID NO: 17, and (f) LCDR2 of SEQ ID NO:

18. a light chain variable region comprising R3; (ii) (a) HCDR1 of SEQ ID NO: 26, (b) HCDR2 of SEQ ID NO: 27, (c) (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 28; and (e) an LCDR1 of SEQ ID NO: 36; (e) a light chain variable region comprising an LCDR2 of SEQ ID NO: 37, and (f) an LCDR3 of SEQ ID NO:

38. region; (iii) (a) HCDR1 of SEQ ID NO: 46, (b) HCDR2 of SEQ ID NO: 47, (c (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 48; and (e) an LCDR1 of SEQ ID NO: 56; (e) an LCDR2 of SEQ ID NO: 57, and (f) an LCDR3 of SEQ ID NO:

58. variable region; (iv) (a) HCDR1 of SEQ ID NO: 66, (b) HCDR2 of SEQ ID NO: 67, (c) (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 68; and (e) an LCDR1 of SEQ ID NO: 76; (e) a light chain variable region comprising an LCDR2 of SEQ ID NO: 77, and (f) an LCDR3 of SEQ ID NO:

78. region; (v) (a) HCDR1 of SEQ ID NO: 86, (b) HCDR2 of SEQ ID NO: 87, (c) HCDR3 of SEQ ID NO: 88, (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 88; (e) an LCDR1 of SEQ ID NO: 96; (f) a light chain variable region comprising an LCDR2 of SEQ ID NO: 97, and (f) an LCDR3 of SEQ ID NO:

98. Area; (vi) (a) HCDR1 of SEQ ID NO: 106, (b) HCDR2 of SEQ ID NO: 107, ( (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 108; and (d) an LCDR of SEQ ID NO:

116. R1, (e) LCDR2 of SEQ ID NO: 117, and (f) LCDR3 of SEQ ID NO: 118 a light chain variable region comprising: (vii) (a) HCDR1 of SEQ ID NO: 126, (b) HCDR2 of SEQ ID NO: 127, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 128; and (d) an LC of SEQ ID NO:

136. (e) LCDR2 of SEQ ID NO: 137, and (f) LCDR3 of SEQ ID NO: 138 a light chain variable region comprising: (viii) (a) HCDR1 of SEQ ID NO: 146, (b) HCDR2 of SEQ ID NO: 147 (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 148; and (d) an L of SEQ ID NO:

156. CDR1, (e) LCDR2 of SEQ ID NO: 157, and (f) LCDR of SEQ ID NO: 158 a light chain variable region comprising: (ix) (a) HCDR1 of SEQ ID NO: 166, (b) HCDR2 of SEQ ID NO: 167, ( (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 168; and (d) an LCDR of SEQ ID NO:

176. R1, (e) LCDR2 of SEQ ID NO: 177, and (f) LCDR3 of SEQ ID NO: 178 a light chain variable region comprising: (x) (a) HCDR1 of SEQ ID NO: 186, (b) HCDR2 of SEQ ID NO: 187, (c (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 188; and (e) an LCDR of SEQ ID NO:

196. 1, (e) an LCDR2 of SEQ ID NO: 197, and (f) an LCDR3 of SEQ ID NO:

198. a light chain variable region; (xi) (a) HCDR1 of SEQ ID NO: 206, (b) HCDR2 of SEQ ID NO: 207, ( (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 208; and (d) an LCDR of SEQ ID NO:

216. R1, (e) an LCDR2 of SEQ ID NO: 217, and (f) an LCDR3 of SEQ ID NO: 218 a light chain variable region comprising: (xii) (a) HCDR1 of SEQ ID NO: 226, (b) HCDR2 of SEQ ID NO: 227, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 228; and (d) an LC of SEQ ID NO:

236. (e) LCDR2 of SEQ ID NO: 237, and (f) LCDR3 of SEQ ID NO: 238 a light chain variable region comprising: (xiii) (a) HCDR1 of SEQ ID NO: 246, (b) HCDR2 of SEQ ID NO: 247 (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 248; and (d) an L CDR1, (e) LCDR2 of SEQ ID NO: 257, and (f) LCDR of SEQ ID NO: 258 a light chain variable region comprising: (xiv) (a) an HCDR1 of SEQ ID NO: 266, (b) an HCDR2 of SEQ ID NO: 267, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 268; and (d) an LC of SEQ ID NO:

276. (e) LCDR2 of SEQ ID NO: 277, and (f) LCDR3 of SEQ ID NO: 278 a light chain variable region comprising: (xv) (a) HCDR1 of SEQ ID NO: 286, (b) HCDR2 of SEQ ID NO: 287, ( (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 288; and (d) an LCDR of SEQ ID NO:

296. R1, (e) LCDR2 of SEQ ID NO:297, and (f) LCDR3 of SEQ ID NO:298 a light chain variable region comprising: (xvi) (a) HCDR1 of SEQ ID NO: 306, (b) HCDR2 of SEQ ID NO: 307, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 308; and (d) an LC of SEQ ID NO:

314. (e) LCDR2 of SEQ ID NO: 315, and (f) LCDR3 of SEQ ID NO: 316 a light chain variable region comprising: (xvii) (a) HCDR1 of SEQ ID NO: 322, (b) HCDR2 of SEQ ID NO: 323 (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 324; and (d) an L CDR1, (e) LCDR2 of SEQ ID NO: 333, and (f) LCDR of SEQ ID NO: 334 a light chain variable region comprising: (xviii) (a) HCDR1 of SEQ ID NO: 342, (b) HCDR of SEQ ID NO: 343 2. (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 344; and (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO:

349. (e) LCDR1 of SEQ ID NO: 350, and (f) LCDR2 of SEQ ID NO:

351. a light chain variable region comprising R3; (xix) (a) an HCDR1 of SEQ ID NO: 356, (b) an HCDR2 of SEQ ID NO: 357, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 358; and (d) an LC of SEQ ID NO:

363. (e) LCDR2 of SEQ ID NO: 364, and (f) LCDR3 of SEQ ID NO: 365 a light chain variable region comprising: (xx) (a) HCDR1 of SEQ ID NO: 370, (b) HCDR2 of SEQ ID NO: 371, ( (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 372; and (d) an LCDR of SEQ ID NO:

377. R1, (e) LCDR2 of SEQ ID NO: 378, and (f) LCDR3 of SEQ ID NO: 379 a light chain variable region comprising: (xxi) (a) an HCDR1 of SEQ ID NO: 384, (b) an HCDR2 of SEQ ID NO: 385, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 386; and (d) an LC of SEQ ID NO:

391. (e) LCDR2 of SEQ ID NO: 392, and (f) LCDR3 of SEQ ID NO: 393 a light chain variable region comprising: (xxii) (a) an HCDR1 of SEQ ID NO: 398, (b) an HCDR2 of SEQ ID NO: 399 (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 400; and (d) an L CDR1, (e) LCDR2 of SEQ ID NO: 406, and (f) LCDR of SEQ ID NO: 407 a light chain variable region comprising: (xxiii) (a) HCDR1 of SEQ ID NO: 412, (b) HCDR of SEQ ID NO: 413 2. (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 414; and (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO:

419. (e) LCDR1 of SEQ ID NO: 420, and (f) LCDR2 of SEQ ID NO:

421. a light chain variable region comprising R3; (xxiv) (a) HCDR1 of SEQ ID NO: 426, (b) HCDR2 of SEQ ID NO: 427 (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 428; and (d) an L CDR1, (e) LCDR2 of SEQ ID NO: 434, and (f) LCDR of SEQ ID NO: 435 a light chain variable region comprising: (xxv) (a) an HCDR1 of SEQ ID NO: 440, (b) an HCDR2 of SEQ ID NO: 441, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 442; and (d) an LC of SEQ ID NO:

447. (e) LCDR2 of SEQ ID NO: 448, and (f) LCDR3 of SEQ ID NO: 449 a light chain variable region comprising: (xxvi) (a) HCDR1 of SEQ ID NO: 454, (b) HCDR2 of SEQ ID NO: 455 (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 456; and (d) an L (e) LCDR1 of SEQ ID NO: 462, and (f) LCDR of SEQ ID NO: 463 a light chain variable region comprising: (xxvii) (a) HCDR1 of SEQ ID NO: 468, (b) HCDR of SEQ ID NO: 469 2. (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 470; and (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO:

475. (e) LCDR1 of SEQ ID NO: 476, and (f) LCDR2 of SEQ ID NO: 477 a light chain variable region comprising R3; (xxviii) (a) HCDR1 of SEQ ID NO: 482, (b) HCDR of SEQ ID NO: 483 R2, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 484; and (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO:

489. (e) LCDR1 of SEQ ID NO: 490, and (f) LCDR2 of SEQ ID NO:

491. Light chain variable region containing DR3 an antibody,

9. At least one amino acid in the CDR is identical to that of the corresponding CDR of another anti-VP1 antibody in Table 2. The antibody of claim 8, wherein the amino acid sequence is substituted with a corresponding residue.

10. 9. The method of claim 8, wherein one or two amino acids in the CDR are modified, deleted or substituted. The antibodies listed.

11. 9. The antibody of claim 8, containing the modifications in Table 3.

12. at least 90, 91, 100, 120, 140, 160, 180, 190, 200, 210, 220, 230, 240, 250, 260, 270, 280, 300, 310, 320, 330, 340, 350, 360, 370, 3 , 92, 93, 94, 95, 96, 97, 98 or 99% identity to the sequence of 8. The antibody described in 8.

13. Monoclonal antibodies, chimeric antibodies, humanized antibodies, human engineered antibodies, human antibodies, single-chain antibodies 9. The antibody of claim 8, which is an scFv or antibody fragment.

14. the antibody or antigen-binding fragment thereof (i) a heavy chain variable region (vH) comprising SEQ ID NO: 12 and a light chain variable region comprising SEQ ID NO: 22 (vL); (ii) a heavy chain variable region (vH) comprising SEQ ID NO: 32 and a light chain variable region comprising SEQ ID NO: 42 Area (vL); (iii) a heavy chain variable region (vH) comprising SEQ ID NO: 52 and a light chain variable region (vH) comprising SEQ ID NO: 62 Area (vL); (iv) a heavy chain variable region (vH) comprising SEQ ID NO: 72 and a light chain variable region comprising SEQ ID NO: 82 Area (vL); (v) a heavy chain variable region (vH) comprising SEQ ID NO: 92 and a light chain variable region comprising SEQ ID NO: 102; Area (vL); (vi) a heavy chain variable region (vH) comprising SEQ ID NO: 112 and a light chain variable region (vL) comprising SEQ ID NO: 122; variable region (vL); (vii) a heavy chain variable region (vH) comprising SEQ ID NO: 132 and a light chain comprising SEQ ID NO: 142 variable region (vL); (viii) a heavy chain variable region (vH) comprising SEQ ID NO: 152 and a light chain variable region (vH) comprising SEQ ID NO: 162 Chain variable region (vL); (ix) a heavy chain variable region (vH) comprising SEQ ID NO: 172 and a light chain variable region (vL) comprising SEQ ID NO: 182; variable region (vL); (x) a heavy chain variable region (vH) comprising SEQ ID NO: 192 and a light chain variable region (vL) comprising SEQ ID NO: 202 Area (vL); (xi) a heavy chain variable region (vH) comprising SEQ ID NO: 212 and a light chain variable region (vL) comprising SEQ ID NO: 222 variable region (vL); (xii) a heavy chain variable region (vH) comprising SEQ ID NO: 232 and a light chain comprising SEQ ID NO: 242 variable region (vL); (xiii) a heavy chain variable region (vH) comprising SEQ ID NO: 252 and a light chain variable region (vH) comprising SEQ ID NO: 262 Chain variable region (vL); (xiv) a heavy chain variable region (vH) comprising SEQ ID NO: 272 and a light chain comprising SEQ ID NO:

282. variable region (vL); (xv) a heavy chain variable region (vH) comprising SEQ ID NO: 292 and a light chain variable region (vL) comprising SEQ ID NO:

302. variable region (vL); (xvi) a heavy chain variable region (vH) comprising SEQ ID NO: 312 and a light chain comprising SEQ ID NO: 320 variable region (vL); (xvii) a heavy chain variable region (vH) comprising SEQ ID NO: 328 and a light chain variable region (vH) comprising SEQ ID NO: 338 Chain variable region (vL); (xviii) a heavy chain variable region (vH) comprising SEQ ID NO: 348 and a heavy chain variable region (vH) comprising SEQ ID NO: 355 Light chain variable region (vL); (xix) a heavy chain variable region (vH) comprising SEQ ID NO: 362 and a light chain comprising SEQ ID NO:

369. variable region (vL); (xx) a heavy chain variable region (vH) comprising SEQ ID NO: 376 and a light chain variable region (vL) comprising SEQ ID NO: 383 variable region (vL); (xxi) a heavy chain variable region (vH) comprising SEQ ID NO: 390 and a light chain comprising SEQ ID NO:

397. variable region (vL); (xxii) a heavy chain variable region (vH) comprising SEQ ID NO: 404 and a light chain variable region (vH) comprising SEQ ID NO: 411 Chain variable region (vL); (xxiii) a heavy chain variable region (vH) comprising SEQ ID NO: 418 and a heavy chain variable region (vH) comprising SEQ ID NO: 425 Light chain variable region (vL); (xxiv) a heavy chain variable region (vH) comprising SEQ ID NO: 432 and a light chain variable region (vH) comprising SEQ ID NO: 439 Chain variable region (vL); (xxv) a heavy chain variable region (vH) comprising SEQ ID NO: 446 and a light chain comprising SEQ ID NO: 453 variable region (vL); (xxvi) a heavy chain variable region (vH) comprising SEQ ID NO: 460 and a light chain variable region (vL) comprising SEQ ID NO: 467 Chain variable region (vL); (xxvii) a heavy chain variable region (vH) comprising SEQ ID NO: 474 and a heavy chain variable region (vH) comprising SEQ ID NO: 481 a light chain variable region (vL); or (xxviii) a heavy chain variable region (vH) comprising SEQ ID NO: 488 and a heavy chain variable region (vH) comprising SEQ ID NO:

495. Light chain variable region (vL) The antibody of claim 1, comprising:

15. at least 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, 14. The sequence of claim 14, wherein the sequence retains 2, 93, 94, 95, 96, 97, 98 or 99% identity. The antibody or fragment thereof described in .

16. 1, 2, 3, 4, or 5, but not more than 10, of the variable light chain region or variable heavy chain region 15. The antibody of claim 14, wherein less than 10 amino acids have been modified, deleted or substituted.

17. Monoclonal antibodies, chimeric antibodies, humanized antibodies, human engineered antibodies, human antibodies, single-chain antibodies 15. The antibody of claim 14, which is an scFv or antibody fragment.

18. The antibody or fragment thereof has reduced or no glycosylation. The antibody according to any one of claims 1 to 17, which is free or hypofucosylated. body.

19. The antibody or antibody composition of any one of claims 1 to 18, further comprising a pharmaceutically acceptable carrier. or a fragment thereof.

20. 20. The pharmaceutical composition of claim 19, wherein the pharmaceutically acceptable carrier comprises histidine or a sugar. Pharmaceutical composition.

21. 21. The pharmaceutical composition of claim 20, wherein the sugar is sucrose.

22. A pharmaceutical composition comprising a plurality of antibodies or antigen-binding fragments according to any one of the preceding claims. At least 0.05%, 0.1%, 0.5%, 1%, 2% of the antibody in the composition , 3%, 5% or more of the pharmaceutical composition have α2,3-linked sialic acid residues.

23. A pharmaceutical composition comprising a plurality of antibodies or antigen-binding fragments according to any one of the preceding claims. wherein none of the antibodies contains a bisecting GlcNAc.

24. 10. The antibody or fragment thereof according to any one of the preceding claims, prepared as a lyophilisate. A pharmaceutical composition comprising:

25. A patient in need thereof, administering an effective amount of the antibody or pharmaceutical composition of claim 1 or claim 19. and administering the drug by injection or infusion to treat BK virus or JC virus infection. How to make peace.

26. 26. A patient in need thereof is diagnosed with BK viruria or BK viremia. The method described below.

27. A patient in need thereof, administering an effective amount of the antibody or pharmaceutical composition of claim 1 or claim 19. and administering to the patient by injection or infusion a vaccine containing a compound associated with BK virus or JC virus.

10. A method of treating or reducing the likelihood of a disorder, wherein the disorder is nephropathy, -, BKVAN, hemorrhagic cystitis (HC), progressive multifocal leukoencephalopathy (PML), granular cell encephalopathy Gastrointestinal neuropathy (GCN), interstitial kidney disease, ureteral stenosis, vasculitis, colitis, retinitis, meningitis, and immune reconstitution inflammatory syndrome (IRIS).

28. 28. The method of claim 25 or 27, wherein the antibody or composition is reconstituted prior to injection or infusion. The method described.

29. 25. The method of claim 24, wherein the antibody or pharmaceutical composition is administered in combination with another therapeutic agent.

27. The method according to claim 27.

30. 30. The method of claim 29, wherein the therapeutic agent is an immunosuppressant.

31. The immunosuppressant is selected from the group consisting of a monophosphate dehydrogenase inhibitor, a purine synthesis inhibitor, and a carboxylase inhibitor.

31. The method of claim 30, wherein the inhibitor is a lucineurin inhibitor or an mTOR inhibitor.

32. Immunosuppressants include mycophenolate mofetil (MMF), mycophenolate sodium, 32. The compound according to claim 31, which is azathioprine, tacrolimus, sirolimus or cyclosporine. The method described.

33. 30. The method of claim 29, wherein the therapeutic agent is an additional anti-VP1 antibody.

34. The antibody or fragment thereof according to any one of claims 1 to 18 for use as a pharmaceutical. Piece.

35. 10. The method of claim 1 for use in neutralizing BK virus or JC virus infection. An antibody or a fragment thereof, or a pharmaceutical composition according to claims 19 to 24.

36. Nephropathy, BKVAN, hemorrhagic cystitis (HC), progressive multifocal leukoencephalopathy (PML) , granular cell neuropathy (GCN), interstitial kidney disease, ureteral stenosis, vasculitis, colitis, retina in the treatment or reduction of the likelihood of inflammatory bowel disease, meningitis, and immune reconstitution inflammatory syndrome (IRIS); The antibody or fragment thereof according to claim 1 or any of claims 19 to 24 for use in The pharmaceutical composition according to any one of claims 1 to 4.

37. Use of the antibody or fragment thereof according to claim 35, administered in combination with another therapeutic agent. 。

38. 38. The use of an antibody or fragment thereof according to claim 37, wherein the therapeutic agent is an immunosuppressant.

39. The immunosuppressant is a monophosphate dehydrogenase inhibitor, a purine synthesis inhibitor, a calcitonin inhibitor, 39. The antibody or fragment thereof of claim 38, which is a sineurin inhibitor or an mTOR inhibitor. Use of pieces.

40. The immunosuppressant is mycophenolate mofetil (MMF), mycophenolate sodium, 3. The compound according to claim 2, wherein the compound is selected from the group consisting of cyclosporine, tacrolimus, sirolimus, azathioprine, tacrolimus, sirolimus, and cyclosporine.

10. Use of an antibody or fragment thereof according to 9.

41. 38. The antibody or fragment thereof of claim 37, wherein the therapeutic agent is a further anti-VP1 antibody. use.

42. A nucleic acid encoding the antibody or antigen-binding fragment of claim 1.

43. 43. A vector comprising the nucleic acid of claim 42.

44. 44. A host cell comprising the vector of claim 43.

45. and recovering the antibody or antibody fragment from the culture. Methods for generating original binding fragments.

46. A diagnostic agent comprising a labeled antibody or antigen-binding fragment thereof according to claim 1.

47. Labels consist of radiolabels, fluorophores, chromophores, imaging agents, and metal ions 47. The diagnostic agent of claim 46, selected from the group:

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