Polyoma virus-neutralizing antibody

Abstract

To provide an anti-polyoma virus antibody and an antibody fragment for treatment of polyoma virus infection or reduction of probability thereof, and a method of using the antibody.SOLUTION: Provided is an antibody, where the antibody or an antigen-binding fragment thereof specifically binds to BK virus and / or JC virus. In one embodiment, the antibody or the antigen-binding fragment thereof binds to BKV serotype I, BKV serotype II, BKV serotype III or BKV serotype IV, or combination of serotypes I to IV. In another embodiment, the antibody or antigen-binding fragment thereof further binds to JC virus.SELECTED DRAWING: None

Description

Technical Field

[0001] The present disclosure relates to anti-polyomavirus antibodies, antibody fragments, and their use for the treatment or reduction of the likelihood of polyomavirus infection.

Background Art

[0002] Among human polyomaviruses, the first to be identified were BK virus (BKV) and JC virus (JCV). These two types of polyomaviruses were isolated from immunosuppressed patients and published in the same issue of the Lancet in 1971 (Gardner et al., Lancet 1971 1:1253-1527, and Padgett et al., Lancet 1971 1:1257-1260). Polyomaviruses are icosahedral non-enveloped double-stranded DNA viruses. The virus has a diameter of 40-45 nm and is composed of 88% protein and 12% DNA. et al., Lancet 1971 1:1253-1527, and Padgett tt et al., Lancet 1971 1:1257-1260). Polyoma viruses are icosahedral non-enveloped double-stranded DNA viruses. The virus has a diameter of 40-45 nm and is composed of 88% protein and 12% DNA.

[0003] The BKV genome is a circular double-stranded DNA approximately 5000 base pairs in length and includes three major regions: an early coding region, a late coding region, and a non-coding control region. The early coding region encodes three regulatory proteins (large tumor antigen [TAg], small tumor antigen [tAg], and truncated tumor antigen [truncTAg]), which are the first viral proteins expressed in newly infected cells and play a role in promoting viral DNA replication and establishing a favorable cellular environment. The late coding region encodes three structural proteins (VP1, VP2, and VP3) that make up the viral capsid, as well as an agnoprotein. ​​​​​​​​​It codes, but its role during virus replication is not very clear. The non-coding control region contains , the origin of genome replication, as well as early and late promoters that drive the expression of viral gene products. It is included.

[0004] BKV has been detected in many different cell types, such as epithelial cells of the kidney, bladder and ureter (typical sites of persistence), tonsillar tissue and lymphocytes (proposed primary infection and seeding sites) (Chatterjee et al., J. Med. Virol. 2000; 60:353 - 362, Goudsmit et al., J. Med. Virol. 1 982;10:91 - 99, Heritage et al., J. Med. Virol .1981;8:143 - 150, Shinohara et al., J. Med. V irol.1993;41(4):301 - 305). The main cell surface receptors for BKV are gangliosides GT1b, GD1b, GD3, all of which have terminal α2,8-linked sialic acid and are quite widely distributed, enabling infection of various cell types (Neu et al., PLos Patholog. 2013;9(l0):el10371 4 and e1003688, and also see O’Hara et al., Virus Res. 2014;189:208 - 285). The non-enveloped icosahedral virion of BKV is composed of three different viral proteins, with 360 copies of the major viral capsid protein VP1 arranged in 72 pentamers and 72 copies of the non-major viral capsid proteins VP2 and VP3 combined, with one VP2 or VP3 molecule bound to each VP1 pentamer. During translocation, only VP1 is exposed on the virion surface. , Each pentamer has five low-affinity binding sites for ganglioside receptors. VP1 When the pentamer binds to the ganglioside receptor on the cell surface, internalization is initiated through the caveolae-mediated endocytosis pathway, followed by the virus being transported to the endoplasmic reticulum and finally to the nucleus (Tsai and Qian, J. Virol 2010;84(19): 9840-9852).

[0005] Infection with BKV is essentially ubiquitous, and the estimated range of the infected population worldwide is 80-90 % (Knowles W.A., Adv. Exp. Med. Biol. 2006; 577:19-45). Primary infection is most commonly seen in childhood (i.e., before the age of 10) and leads to a mild, non-specific, self-limiting disease or is asymptomatic. Persistent infection is established in the epithelial cells of the renal tubules, ureters, and bladder and is effectively controlled by the immune system. Transient and asymptomatic viral shedding in the urine of immunocompetent adults occurs sporadically but does not result in disease or sequelae. However, immunosuppression, especially during renal or hematopoietic stem cell transplantation, can lead to uncontrolled replication of BKV and ultimately to BK virus-associated nephropathy (BKVAN) or hemorrhagic cystitis (HC), a painful disease of the bladder. There is no effective antiviral therapy for BKV, and current treatment guidelines are to reduce immunosuppression, but this increases the risk of acute rejection. Even with a more aggressive current approach to monitoring and prevention, up to 10% of renal transplant patients develop BKVA N, and 15-30% of those patients suffer graft loss due to BKVAN . Up to 30% of patients who had their immunosuppressive therapy reduced at the time of detection of BK viremia ​​​​​​​​As a result, it experiences the onset of acute rejection.

[0006] BKV was first described in 1971 (as above), but until the 1990s, BK virus-associated nephropathy (BKVAN) had not been reported in the literature as a cause of kidney transplant injury (Purighalla et al., Am. J. Kidney Dis. 1995;26:671-6 73, and Randhawa et al., Transplantation 19 99;67:103-109). In the early management of BKVAN, patients who tested positive for BK had a serious outcome, with graft dysfunction and graft loss seen in more than 50% of them (Hirsch et al., New Engl. J. Med. 2002;3 47:488-496). Reactivation and replication of BKV are associated with an established clinical course in kidney transplant patients, first with detection of the virus and viral DNA in urine (viremia), followed by detection of the virus in the bloodstream (viremia), and finally evidenced by the development of nephropathy and subsequent decline in renal function as a result of viral replication. Usually, within the first 3 months after transplantation, approximately 30-40% of all kidney transplant patients develop viremia, and 10-20% develop BK virus viremia (Sawinski and Goral, Nephrol Dial Transplant. 2015;30:209-217; Hirsch et a l., Am J Transplant. 2013;13:136-145; Dharn idharka et al., Pediatr Nephrol. 2011;26:1 763-1774; Babel et al., Transplantation. 20 ​​​09;88:89-95). Usually, within one year after transplantation, about 1-10% of all kidney transplant patients progress to BKVAN (Bohl and Brennan, Clin J Am Soc Nephrol. 2007;2(Suppl 1):S36-46, Sawin ski and Goral, Nephrol Dial Transplant. 20 15;30:209-217). BKV replication in renal tubular epithelial cells causes necrotic and lytic destruction, detachment of the basement membrane, accumulation of tubular fluid in the stroma, and ultimately interstitial fibrosis and tubular atrophy (Nickeleit et al., J. Am. Soc. Ne prol. 1999;10(5):1080-1089). Patients may present with deterioration of renal function, tubulointerstitial nephritis, and ureteral stricture (Garner et al., Lancet 1 971;1(7712):1253-1257, and Hirsch Am. J. Tra nsplant 2002;2(1)25-30).

[0007] In addition, BKV can cause interstitial pneumonia, retinitis, and meningoencephalitis in immunocompromised hosts (Reploeg et al., Clin. Infect. Dis. 20 01;33(2):191-202). BKV disease in hematopoietic stem cell transplant (HSCT) patients typically manifests as hemorrhagic cystitis (HC), although the severity varies. Viraluria (but not necessarily viremia) and painful hematuria are associated with the clinical symptoms of HC . Current treatment guidelines are essentially supportive and mainly include forced hydration / diuresis and pain management. In the most severe cases, blood transfusion and hematoma removal are required, and in some cases is fatal. Among HSCT patients, HC of all causes (e.g., drugs, radiation, vi ruses) is relatively common, but HC associated with BKV usually occurs in about 10-12% of patients within 6 months after transplantation. There are also other viral etiologies of HC. In pediatric HSCT patients, adenovirus is a more common cause of HC compared to adult HSCT patients. Also, BK virus has been observed in other immunocompromised states such as organ transplantation and in HIV / AIDS patients (Jiang et al., Virol. 20 09;384:266-273). At present, the standard treatment for BKVAN is reduction of immunosuppression to prevent graft dysfunction and graft injury (Wiseman et al., Am. J. Ki dney Dis. 2009;54(1):131-142, and Hirsch et

[0008] al., Transplantation 2005;79(1):1277-128 6). Although reduction of immunosuppression helps prevent progression from viremia to severe injury associated with clinical nephropathy, this increases the risk of acute organ rejection, so there is no stable clinical treatment plan for its reduction (Brennan et al., A m. J. Transplant 2005;5(3):582- 594). Clinicians have reported the use of therapeutic agents such as cidofovir, leflunomide, and quinolones in combination with reduction of immunosuppressive agents, but this method has been ineffective and has increased the burden of managing additional side effects (R andhawa and Brennan Am. J. Transplant 2006 ;6(9):2000-2005). Therefore, neutralizing polyomaviruses such as BKV ​​​​​​​ , in the field of therapeutic methods that can be used in immunocompromised hosts, there are unaddressed and useful needs .

[0009] The JC virus (JCV) is another human polyomavirus that is widely found in the population (80%), but JCV generally infects later than BKV (Padgett e t al., J. Infect. Dis. 1973;127(4):467 - 470, and Sabath et al., J. Infect. Dis. 2002;186 Su ppl.2:5180 - 5186). After primary infection, JCV establishes a latent infection in lymphoid organs and the kidney, and upon reactivation, it invades the central nervous system (CNS) via infected B lymphocytes . Once in the CNS, JCV causes progressive multifocal leukoencephalopathy (PML), a progressive demyelinating CNS disorder. Most cases of PML are associated with immunomodulatory therapies used in the treatment of multiple sclerosis (natalizumab, fingolimod, etc.) or rheumatoid arthritis (rituximab, etc.), and usually, the progression of the disease stops upon discontinuation of the treatment. Given the progressive nature of PML, in patients who have received JCV neutralizing antibodies over several months, improvement may be documented over several months by clinical criteria or by MRI, which is already routinely used for monitoring multiple sclerosis, in patients who have received JCV neutralizing antibodies. PML also manifests in patients with HIV / AIDS and has been reported in immunocompromised patients (Angstrom et al., Brain 1 958;81(1):93 - 111, and Garcia - Suarez et al. , Am. J. Hematol. 2005;80(4):271 - 281). In PML patients Considering the progressive nature of PML, in patients who have received JCV neutralizing antibodies over several months, by clinical criteria or by MRI, which is already routinely used for monitoring multiple sclerosis, in patients who have received JCV neutralizing antibodies over several months, significant improvement may be documented. PML also manifests in patients with HIV / AIDS and has been reported in immunocompromised patients (Angstrom et al., Brain 1 958;81(1):93 - 111, and Garcia - Suarez et al. , Am. J. Hematol. 2005;80(4):271 - 281). In PML patients 958;81(1):93 - 111, and Garcia - Suarez et al. , Am. J. Hematol. 2005;80(4):271 - 281). In PML patients Local nerves such as confusion, changes in mental state, ataxia, hemiparesis, quadriplegia, and visual changes are recognized (Richardson E.P., N.Eng.J.Med.196 1;265:815-823). The prognosis of PML patients is poor, especially in HIV / AIDS patients (Antinori et al., J.Neurovirol. 2003;9 suppl.1:47-53). This further emphasizes unaddressed and useful needs in the field of therapeutic methods that neutralize polyomaviruses such as JCV . SUMMARY OF THE INVENTION

[0010] The present disclosure is directed to neutralizing antibodies against human polyomavirus and / or fragments thereof, and antibodies that recognize BK virus and / or JC virus.

[0011] An antibody, wherein the antibody or an antigen-binding fragment thereof specifically binds to BK virus and / or JC virus.

[0012] An antibody, wherein the antibody or an antigen-binding fragment thereof specifically binds to BK virus and / or JC virus. In one embodiment, the antibody or an antigen-binding fragment thereof binds to BKV serotype I, BKV serotype II, BKV serotype III, or BKV serotype IV, or a combination of serotypes I-IV. In another embodiment, the antibody or an antigen-binding fragment thereof further binds to JC virus.

[0013] An antibody, wherein the antibody or antigen-binding fragment specifically binds to BK virus and / or JC virus. In one embodiment, the antibody or an antigen-binding fragment thereof is a BK serum type Bind to and neutralize Clearotype I. In one embodiment, the antibody or antigen-binding fragment thereof binds to BKV blood Bind to and neutralize Clearotype I and BKV serotype II. In another embodiment, the antibody or its Antigen-binding fragment binds to and neutralizes BKV serotype I and BKV serotype III. In another Embodiment, the antibody or antigen-binding fragment thereof binds to and neutralizes BKV serotype I and BKV serotype IV Bind to and neutralize. In another embodiment, the antibody or antigen-binding fragment thereof binds to BKV serotype I I and BKV serotype III bind to and neutralize. In another embodiment, the antibody or its antigen The binding fragment binds to and neutralizes BKV serotype II and BKV serotype IV. In another embodiment In the form, the antibody or antigen-binding fragment thereof binds to and neutralizes BKV serotype I and JCV To neutralize. In a preferred embodiment, the antibody or antigen-binding fragment thereof binds to BKV serotypes I, II, III and IV bind to and neutralize. Further, the antibody or antigen-binding fragment thereof binds to BKV Serotypes I, II, 234III and IV, and JCV bind to and neutralize.

[0014] An isolated antibody or antigen-binding fragment thereof, comprising (i) a heavy chain region and (ii) a light chain region as defined in Table 2, an isolated antibody or antigen-binding fragment thereof.

[0015] An isolated antibody, wherein the antibody or antigen-binding fragment thereof is (i) (a) HCDR1 (CDR - Complementary Determining Region) of SEQ ID NO: 9, (b) SEQ ID NO: 10 Of HCDR2, (c) HCDR3 of SEQ ID NO: 11, a heavy chain variable region comprising, and (d) LCDR1 of SEQ ID NO: 25, (e) LCDR2 of SEQ ID NO: 26, and (f) SEQ ID NO: 2 7 of LCDR3, a light chain variable region comprising, and (ii)(a) The HCDR1 of SEQ ID NO: 41, (b) the HCDR2 of SEQ ID NO: 42, (c) the HCDR3 of SEQ ID NO: 43, a heavy chain variable region, and (d) the LCDR 1 of SEQ ID NO: 57, (e) the LCDR2 of SEQ ID NO: 58, and (f) the LCDR3 of SEQ ID NO: 59, a light chain variable region, and (iii)(a) The HCDR1 of SEQ ID NO: 73, (b) the HCDR2 of SEQ ID NO: 74, (c) the HCDR3 of SEQ ID NO: 75, a heavy chain variable region, and (d) the LCD R1 of SEQ ID NO: 89, (e) the LCDR2 of SEQ ID NO: 90, and (f) the LCDR3 of SEQ ID NO: 91, a light chain variable region, and (iv)(a) The HCDR1 of SEQ ID NO: 105, (b) the HCDR2 of SEQ ID NO: 106, (c ) the HCDR3 of SEQ ID NO: 107, a heavy chain variable region, and (d) the LCD of SEQ ID NO: 121 , (e) the LCDR2 of SEQ ID NO: 122, and (f) the LCD R3 of SEQ ID NO: 123, a light chain variable region, and (v)(a) The HCDR1 of SEQ ID NO: 137, (b) the HCDR2 of SEQ ID NO: 138, (c the HCDR3 of SEQ ID NO: 139, a heavy chain variable region, and (d) the L of SEQ ID NO: 153 CDR1, (e) the LCDR2 of SEQ ID NO: 154, and (f) the LCDR 3 of SEQ ID NO: 155, a light chain variable region, and (vi)(a) The HCDR1 of SEQ ID NO: 169, (b) the HCDR2 of SEQ ID NO: 170, (c ) the HCDR3 of SEQ ID NO: 171, a heavy chain variable region, and (d) the LCD of SEQ ID NO: 185 , (e) the LCDR2 of SEQ ID NO: 186, and (f) the LCD R3 of SEQ ID NO: 187, a light chain variable region, and (vii)(a) The HCDR1 of SEQ ID NO: 201, (b) the HCDR2 of SEQ ID NO: 202, ( (c) A heavy chain variable region comprising the HCDR3 of SEQ ID NO: 203, and (d) SEQ ID NO: 217 of LCDR1, (e) LCDR2 of SEQ ID NO: 218, and (f) LC of SEQ ID NO: 219 A light chain variable region comprising CDR3, and (viii) (a) HCDR1 of SEQ ID NO: 233, (b) HCDR2 of SEQ ID NO: 234, (c) A heavy chain variable region comprising the HCDR3 of SEQ ID NO: 235, and (d) SEQ ID NO: 24 9 of LCDR1, (e) LCDR2 of SEQ ID NO: 250, and (f) L of SEQ ID NO: 251 A light chain variable region comprising CDR3, and (xi) (a) HCDR1 of SEQ ID NO: 265, (b) HCDR2 of SEQ ID NO: 266, (c ) A heavy chain variable region comprising the HCDR3 of SEQ ID NO: 267, and (d) SEQ ID NO: 281 of LCDR1, (e) LCDR2 of SEQ ID NO: 282, and (f) LCD of SEQ ID NO: 283 R3, and an isolated antibody comprising a light chain variable region.

[0016] An antibody in which one or two amino acids within the CDR are modified, deleted, or substituted.

[0017] An antibody that retains at least 90, 91, 92 , 93, 94, 95, 96, 97, 98, or 99% identity over either the variable heavy chain region or the variable light chain region.

[0018] An antibody, wherein the antibody is a monoclonal antibody, chimeric antibody, humanized antibody, human engineered antibody, human antibody, single-chain antibody (scFv), or antibody fragment.

[0019] An isolated antibody or antigen-binding fragment thereof, (i) A heavy chain variable region (vH) comprising SEQ ID NO: 18, and a light chain variable region (vL) comprising SEQ ID NO: 34, and (ii) A heavy chain variable region (vH) containing SEQ ID NO: 50, and a light chain variable region (vL) containing SEQ ID NO: 66, (iii) A heavy chain variable region (vH) containing SEQ ID NO: 82, and a light chain variable region (vL) containing SEQ ID NO: 98, (iv) A heavy chain variable region (vH) containing SEQ ID NO: 114, and a light chain variable region (vL) containing SEQ ID NO: 130, (v) A heavy chain variable region (vH) containing SEQ ID NO: 146, and a light chain variable region (vL) containing SEQ ID NO: 162, (vi) A heavy chain variable region (vH) containing SEQ ID NO: 178, and a light chain variable region (vL) containing SEQ ID NO: 194, (vii) A heavy chain variable region (vH) containing SEQ ID NO: 210, and a light chain variable region (vL) containing SEQ ID NO: 226, (viii) A heavy chain variable region (vH) containing SEQ ID NO: 242, and a light chain variable region (vL) containing SEQ ID NO: 258, (ix) A heavy chain variable region (vH) containing SEQ ID NO: 274, and a light chain variable region (vL) containing SEQ ID NO: 290, comprising an isolated antibody or an antigen-binding fragment thereof.

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

[0021] An antibody in which 1, 2, 3, 4, or 5, but less than 10, amino acids within the variable light chain or variable heavy chain region are modified, deleted, or substituted.

[0022] An antibody, wherein the antibody is a monoclonal antibody, chimeric antibody, humanized antibody, humanized modified antibody, human antibody, single-chain antibody (scFv), or antibody fragment.

[0023] Natural signal / leader pairs that fit the appropriate VH and / or VL gene segments Methods for isolating and producing antibodies using peptide sequences.

[0024] Use synthetic and / or optimized signal / leader peptide sequences to enhance expression and and a method for isolating and producing antibodies, the method improving the efficiency and yield of the antibody.

[0025] The antibody or fragment thereof is reduced or aglycosylated. or hypofucosylated.

[0026] A pharmaceutical composition comprising an antibody or fragment thereof, and further comprising a pharma- ceutically acceptable carrier.

[0027] A pharmaceutical composition, wherein the pharma- ceutically acceptable carrier comprises a histidine or a carbohydrate.

[0028] A pharmaceutical composition, wherein the carbohydrate is sucrose.

[0029] A pharmaceutical composition comprising a plurality of antibodies or antigen-binding fragments, wherein at least one of the antibodies in the composition α2,3-bonded sialic acid of 0.05%, 0.1%, 0.5%, 1%, 2%, 3%, 5% or more A pharmaceutical composition having a carboxylic acid residue.

[0030] A pharmaceutical composition comprising a plurality of antibodies or antigen-binding fragments, each of the antibodies being bisecting. A pharmaceutical composition comprising:

[0031] A pharmaceutical composition comprising an antibody or a fragment thereof, the composition being prepared as a lyophilizate. , pharmaceutical compositions.

[0032] A method for neutralizing BK virus or JC virus infection that is effective in a patient in need thereof. A method comprising administering an amount of an antibody via injection or infusion.

[0033] A method wherein the patient in need is diagnosed with BK virusuria or BK viremia.

[0034] A method wherein the patient in need is diagnosed with JC virusuria or JC viremia.

[0035] A method for treating or reducing the likelihood of a BK virus or JC virus-related disorder, comprising administering an effective amount of an antibody via injection or infusion to a patient in need, wherein the disorder is nephropathy, BKVAN, hemorrhagic cystitis (HC), progressive multifocal leukoencephalopathy (PML), granulocyte neuronopathy (GCN), interstitial kidney disease, ureteral stricture, vasculitis, colitis, retinitis, meningitis, and immune reconstitution inflammatory syndrome (IRIS).

[0036] A method wherein the antibody or composition is reconstituted prior to injection or infusion.

[0037] A method wherein the antibody or pharmaceutical composition is administered in combination with another therapeutic agent.

[0038] A method wherein the therapeutic agent is an immunosuppressive agent.

[0039] A method wherein the immunosuppressive agent is a dehydrogenase inhibitor, a purine synthesis inhibitor, a calcineurin inhibitor or an mTOR inhibitor.

[0040] A method wherein the immunosuppressive agent is mycophenolate mofetil (MMF), sodium mycophenolate, azathioprine, tacrolimus, sirolimus or cyclosporine.

[0041] A method wherein the therapeutic agent is an additional anti-VP1 antibody.

[0042] A method in which PML is related to the treatment of multiple sclerosis, rheumatoid arthritis, or psoriasis.

[0043] A method in which the treatment of multiple sclerosis is natalizumab, fingolimod, or dimethyl fumarate, fuma Dimethyl fumarate, fumaric acid, or alemtuzumab.

[0044] A method in which the treatment of rheumatoid arthritis is rituximab.

[0045] A method in which the treatment of psoriasis is efalizumab.

[0046] An antibody or a fragment thereof for use as a medicament.

[0047] An antibody or a fragment thereof for use in neutralizing BK virus or JC virus infection.

[0048] For the treatment of nephropathy, BKVAN, hemorrhagic cystitis (HC), progressive multifocal leukoencephalopathy (PML), granulocytic Neuronopathy (GCN), interstitial kidney disease, ureteral stricture, vasculitis, colitis, retinitis, myelitis And immune reconstitution inflammatory syndrome (IRIS), or for reducing the possibility thereof, an antibody or a fragment thereof. For use.

[0049] Use of an antibody or a fragment thereof administered in combination with another therapeutic agent.

[0050] Use of an antibody or a fragment thereof, wherein the therapeutic agent is an immunosuppressant.

[0051] Use of an antibody or a fragment thereof, wherein the immunosuppressant is a dehydrogenase inhibitor, a purine synthesis inhibitor, a calcineurin inhibitor Or an mTor inhibitor.

[0052] Use of an antibody or a fragment thereof, wherein the immunosuppressant is mycophenolate mofetil (MMF), mycophenolate sodium , the use of an antibody or a fragment thereof, which is azathioprine, tacrolimus, sirolimus or cyclosporine.

[0053] The use of an antibody or a fragment thereof, wherein the therapeutic agent is an additional anti-BK antibody.

[0054] The use of an antibody or a fragment thereof, wherein PML is related to the treatment of multiple sclerosis, rheumatoid arthritis, or psoriasis.

[0055] The use, wherein the treatment of multiple sclerosis is natalizumab, fingolimod, or dimethyl fumarate, fumaric acid esters, or alemtuzumab.

[0056] The use, wherein the treatment of rheumatoid arthritis is rituximab.

[0057] The use, wherein the treatment of psoriasis is efalizumab.

[0058] A nucleic acid encoding an antibody or an antigen-binding fragment.

[0059] A vector containing the nucleic acid.

[0060] A host cell containing the vector.

[0061] A diagnostic agent comprising a labeled antibody or an antigen-binding fragment thereof.

[0062] The diagnostic agent, wherein the label is selected from the group consisting of a radioactive label, a fluorophore, a chromophore, an imaging agent, and a metal ion and.

[0063] Definitions Unless otherwise specified, the following terms and phrases used herein are intended to have the following meanings. be.

[0064] As used herein, the term "antibody" refers to a non-covalent, reversible and a polypeptide of the immunoglobulin family that can bind in a specific manner Means. For example, a naturally occurring IgG antibody is a tetramer containing at least two heavy chains (H) and two light chains (L) interconnected by disulfide bonds. Each heavy chain is composed of a heavy chain variable region (abbreviated as VH herein) and a heavy chain constant region. The heavy chain constant The region is composed of three domains, CH1, CH2 and CH3. Each light chain is composed of a light chain variable Region (abbreviated as VL herein) and a light chain constant region. The light chain constant region Is composed of one domain CL. The VH region and the VL region are further divided into regions of hypervariability named complementarity Determining regions (CDRs), which are interspersed with more conserved regions named framework regions (FRs). Each VH and VL Are composed of three CDRs and four FRs arranged in the following order from the amino terminus to the carboxy terminus: FR1, CDR1, FR2, CDR2, FR3, CDR3 And FR4. The variable regions of the heavy and light chains contain a binding domain that interacts with the antigen . The constant region of the antibody can mediate the binding of the immunoglobulin to host tissues or factors, and these include various cells of the immune system (e.g., effector cells) and the first component of the classical complement System (Clq). The term "antibody" includes, but is not limited to, monoclonal antibodies, human antibodies, humanized antibodies , camel antibodies, chimeric antibodies, and anti-idiotype (anti-Id) antibodies (e.g., including anti-Id antibodies against the antibodies of the present disclosure) . Antibodies can be of any isotype / class (e.g.,

[0065] IgG, IgE, IgM, IgD, IgA and IgY), or subclasses (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, IgA2, etc.). IgG1, IgG2, IgG3, IgG4, IgA1, IgA2, etc.). Including IgG, IgE, IgM, IgD, IgA and IgY), or subclasses (e.g., (such as IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2).

[0066] "Complementary determining domain" or "complementary determining region" ("CDR") interchangeably refers to the hypervariable regions of VL and VH. CDRs are the target protein binding sites of antibody chains that have specificity for such target proteins. Each human VL or VH that constitutes about 15 - 20% of the variable domain has three CDRs (CDR1 - 3, numbered sequentially from the N - terminus ). CDRs can be referred to by those regions and order. For example, both "VHCDR1" or "HCDR1" mean the first CDR of the heavy chain variable region . CDRs are structurally complementary to the epitopes of the target protein and thus are directly involved in binding specificity. The remaining segments of VL or VH, the so - called framework regions, have less variation in amino acid sequence (Kuby, Immunology, 4th ed., Chapter 4. W.H. Freeman & Co., New York , 2000).

[0067] ).

[0067] The positions of CDRs and framework regions can be determined using various definitions known in the art, such as Kabat, Chothia, IMGT, and AbM ( for example, Johnson et al., Nucleic Acids Res., 29 :205 - 206(2001), Chothia and Lesk, J. Mol. Biol., 196:901 - 917(1987), Chothia et al., Nature, 342:877 - 883(1989), Chothia et al., J. M ol., 196:901 - 917(1987), Chothia et al., Nature, 342:877 - 883(1989), Chothia et al., J. Mol. Biol., 196:901 - 917(1987), Chothia et al., Nature, 342:877 - 883(1989), Chothia et al., J. M ol., 196:901 - 917(1987), Chothia et al., Nature, 342:877 - 883(1989), Chothia et al., J. Mol. Biol., 196:901 - 917(1987), Chothia et al., Nature, 342:877 - 883(1989), Chothia et al., J. M ol., 196:901 - 917(1987), Chothia et al., Nature, 342:877 - 883(1989), Chothia et al., J. Mol. Biol., 196:901 - 917(1987), Chothia et al., Nature, 342:877 - 883(1989), Chothia et al., J. M ol. Biol., 227:799 - 817 (1992), Lefranc, M.P., Nucleic Acids Res., 29:207 - 209 (2001), Al - L azikani et al., J. Mol. Biol., 273:927 - 748 (1 997). (See also). Also, the definition of the antigen - binding site is described below: Rui z et al., Nucleic Acids Res., 28:219 - 221 (2 000), MacCallum et al., J. Mol. Biol., 262:73 2 - 745 (1996), 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 M.J.E. (ed.), Pr otein Structure Prediction, Oxford Univer sity Press, Oxford, 141 - 172 (1996). In the combined K abat and Chothia numbering schemes, in some embodiments, C DR corresponds to amino acid residues that are part of the Kabat CDR, Chothia CDR, or both. For example, in some embodiments, the CDR corresponds to amino acid residues 26 - 35 (HC CDR1), 50 - 65 (HC CDR2), and 95 - 10 2 (HC CDR3) in VH, for example, mammalian VH, such as human VH; and corresponds to amino acid residues 24 - 34 (LC CDR1), 50 - 56 (LC CDR2 ), and 89 - 97 (LC CDR3) in VL, for example, mammalian VL, such as human V L It is L. Under IMGT, the CDR amino acid residues in VH are numbered approximately 26 - 35 (CDR1 ), 51 - 57 (CDR2), and 93 - 102 (CDR3), and the CDR amino acid residues in VL are numbered approximately 27 - 32 (CDR1), 50 - 52 (CDR2), and 89 - 97 (CDR3). Under IMGT, the CDR regions of an antibody can be determined using the program IMGT / DomainGap Align .

[0068] Both the light and heavy chains are divided into regions of structural and functional homology. The terms "constant" and "variable" are used functionally. In this regard, it will be understood that the variable domains of both the light chain (VL) and heavy chain (VH) portions determine antigen recognition and specificity. In contrast, the constant domains of the light chain (CL) and heavy chain (CH1, CH2, or CH3) confer important biological properties such as secretion, transplacental mobility, Fc receptor binding, complement binding, etc. By convention, the numbering of the constant region domains increases as they become more distal from the antigen-binding site or amino terminus of the antibody. The N-terminus is the variable region and the C-terminus is the constant region, and in fact, the CH3 and CL domains include the carboxy-terminal domains of the heavy and light chains, respectively.

[0069] As used herein, the term "antigen-binding fragment" means one or more portions of an antibody that retain the ability to specifically interact with an epitope of an antigen (e.g., by binding, steric hindrance, stabilization / destabilization, spatial distribution). Examples of binding fragments include single-chain Fv (scFv ), disulfide-linked Fv (sdFv), Fab fragments, F(ab’) fragments, VL, VH, C . ) ​​​​​​A monovalent fragment consisting of L and the CH1 domain; a bivalent fragment containing two Fab fragments linked by a disulfide bridge in the hinge region; an Fd fragment consisting of VH and CH1 domains; an Fv fragment consisting of the VL and VH domains of a single arm of an antibody ; a dAb fragment consisting of the VH domain (Ward et al., Nature 341: 544 - 546, 1989); and isolated complementarity-determining regions (CDRs), or other epitope-binding fragments of an antibody, although not limited to these. Furthermore, the two domains of VL and VH of the Fv fragment are encoded by separate genes, but these can be joined by a synthetic linker using recombinant methods

[0070] to create a single protein chain where the VL and VH regions pair to form a monovalent molecule (known as single-chain Fv, see, for example, Bird et al., Science 242:423 - 426, 1988, and Huston et al., Proc. Natl. Acad. Sci. 85:5879 - 5883, 1988). Such single-chain antibodies are also intended to be encompassed by the term "antigen-binding fragment". These antigen-binding fragments are obtained using conventional techniques known to those skilled in the art, and the fragments are screened for utility in the same manner as the intact antibodies. Antigen-binding fragments can also be incorporated into single-domain antibodies, maxibodies, minibodies, nanobodies, intrabodies, diabodies, triabodies, tetrabodies, v-NAR, and bis- scFv (for example, Hollinger and Hudson

[0071] ​​​​, refer to Nature Biotechnology 23:1126-1136, 2005 (see also). Antigen-binding fragments can be grafted onto scaffolds based on polypeptides such as fibronectin type III (Fn3) (see U.S. Patent No. 6,703,199 which describes monomers of fibronectin polypeptides). (see also).

[0072] Antigen-binding fragments can be incorporated into single-chain molecules, which consist of a pair of tandem Fv segments (VH-CH1-VH-CH1) that together with a complementary light-chain polypeptide form a pair of antigen-binding regions (Zapata et al., Protein Eng. 8:1057-1062, 1995, and U.S. Patent No. 5,641,870). (see also).

[0073] As used herein, the terms "monoclonal antibody" or "monoclonal antibody composition" mean polypeptides that include antibodies and antigen-binding fragments having substantially the same amino acid sequence or derived from the same genetic source. Also, the term includes preparations of antibody molecules of single-molecule compositions. Monoclonal antibody compositions exhibit a single binding specificity and affinity for a particular epitope. (see also). (see also).

[0074] As used herein, the term "human antibody" includes antibodies having variable regions in which both the framework and CDR regions are derived from human-derived sequences. Further, when the antibody includes a constant region, the constant region is derived from such human sequences, such as human germline sequences or variant forms of human germline sequences, or from consensus framework sequences derived from antibodies that include human framework sequences analysis (e.g., Knappik et (see also). (see also). al. ​​​​​​(described in al., J. Mol. Biol. 296:57-86, 2000).

[0075] The human antibodies of the present disclosure contain amino acid residues not encoded by human sequences (e.g., mutations introduced by random or site-directed mutagenesis in vitro or somatic mutations in vivo, or conservative substitutions to promote stability or production).

[0076] As used herein, the term "recognize" means an antibody or antigen-binding fragment thereof that finds and interacts (e.g., binds) to its epitope, and the epitope can be linear or conformational. The term "epitope" means the site on an antigen to which the antibody or antigen-binding fragment of the present disclosure specifically binds. An epitope can be formed from both adjacent amino acids or non-adjacent amino acids juxtaposed by the tertiary folding of a protein. Epitopes formed from adjacent amino acids are typically retained even when exposed to a denaturing solvent, while epitopes formed by three-dimensional folding are typically lost upon treatment with a denaturing solvent. An epitope typically contains at least 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14 or 15 amino acids in a particular spatial conformation. Methods for determining the spatial conformation of an epitope include techniques in the art, e.g., X-ray crystallography and two-dimensional nuclear magnetic resonance (see, e.g., Epitope Mapping Protocols in Methods in Molecular Biology, Vol. 66, G.E. Morris, Ed. (1996)). niques, e.g., X-ray crystallography and two-dimensional nuclear magnetic resonance (see, e.g., Epitope Mapping Protocols in Methods in Molecular Biology, Vol. 66, G.E. Morris, Ed. (1996)). g Protocols in Methods in Molecular Biol ogy, Vol. 66, G.E. Morris, Ed. (1996)). or electron microscopy. A "paratope" is a part of an antibody that recognizes an epitope of an antigen. A part.

[0077] The phrases "specifically binds" or "selectively binds" are used in the context of describing the interaction between an antigen (e.g., a protein) and an antibody, antibody fragment, or binding agent derived from an antibody, and mean a binding reaction that determines the presence of the antigen in a heterogeneous collection of proteins or biological substances (e.g., in a biological sample such as blood, serum, plasma, or tissue sample). Thus, under specific designated immunoassay conditions, an antibody or binding agent having a specific binding specificity binds to a specific antigen at least 2-fold above background and does not bind in a significant amount to other antigens present in the sample. In the context of describing the interaction between an antigen (e.g., a protein) and an antibody, antibody fragment, or binding agent derived from an antibody, and mean a binding reaction that determines the presence of the antigen in a heterogeneous collection of proteins or biological substances (e.g., in a biological sample such as blood, serum, plasma, or tissue sample). In the context of describing the interaction between an antigen (e.g., a protein) and an antibody, antibody fragment, or binding agent derived from an antibody, and mean a binding reaction that determines the presence of the antigen in a heterogeneous collection of proteins or biological substances (e.g., in a biological sample such as blood, serum, plasma, or tissue sample). In the context of describing the interaction between an antigen (e.g., a protein) and an antibody, antibody fragment, or binding agent derived from an antibody, and mean a binding reaction that determines the presence of the antigen in a heterogeneous collection of proteins or biological substances (e.g., in a biological sample such as blood, serum, plasma, or tissue sample). In the context of describing the interaction between an antigen (e.g., a protein) and an antibody, antibody fragment, or binding agent derived from an antibody, and mean a binding reaction that determines the presence of the antigen in a heterogeneous collection of proteins or biological substances (e.g., in a biological sample such as blood, serum, plasma, or tissue sample). In the context of describing the interaction between an antigen (e.g., a protein) and an antibody, antibody fragment, or binding agent derived from an antibody, and mean a binding reaction that determines the presence of the antigen in a heterogeneous collection of proteins or biological substances (e.g., in a biological sample such as blood, serum, plasma, or tissue sample). In the context of describing the interaction between an antigen (e.g., a protein) and an antibody, antibody fragment, or binding agent derived from an antibody, and mean a binding reaction that determines the presence of the antigen in a heterogeneous collection of proteins or biological substances (e.g., in a biological sample such as blood, serum, plasma, or tissue sample). In one aspect, under specific designated immunoassay conditions, an antibody or binding agent having a specific binding specificity binds to a specific antigen at least 10-fold above background and does not bind in a significant amount to other antigens present in the sample. Specific binding of an antibody or binding agent under such conditions may require that the antibody or agent be selected for its specificity for a particular protein. This selection can be achieved by reducing cross-reacting antibodies with molecules from other species (e.g., mouse or rat) or other subtypes, as desired or appropriate. Alternatively, in some aspects, an antibody or antibody fragment is selected to cross-react with a particular desired molecule. In one aspect, under specific designated immunoassay conditions, an antibody or binding agent having a specific binding specificity binds to a specific antigen at least 10-fold above background and does not bind in a significant amount to other antigens present in the sample. In the context of describing the interaction between an antigen (e.g., a protein) and an antibody, antibody fragment, or binding agent derived from an antibody, and mean a binding reaction that determines the presence of the antigen in a heterogeneous collection of proteins or biological substances (e.g., in a biological sample such as blood, serum, plasma, or tissue sample). In the context of describing the interaction between an antigen (e.g., a protein) and an antibody, antibody fragment, or binding agent derived from an antibody, and mean a binding reaction that determines the presence of the antigen in a heterogeneous collection of proteins or biological substances (e.g., in a biological sample such as blood, serum, plasma, or tissue sample). In the context of describing the interaction between an antigen (e.g., a protein) and an antibody, antibody fragment, or binding agent derived from an antibody, and mean a binding reaction that determines the presence of the antigen in a heterogeneous collection of proteins or biological substances (e.g., in a biological sample such as blood, serum, plasma, or tissue sample). In the context of describing the interaction between an antigen (e.g., a protein) and an antibody, antibody fragment, or binding agent derived from an antibody, and mean a binding reaction that determines the presence of the antigen in a heterogeneous collection of proteins or biological substances (e.g., in a biological sample such as blood, serum, plasma, or tissue sample). In the context of describing the interaction between an antigen (e.g., a protein) and an antibody, antibody fragment, or binding agent derived from an antibody, and mean a binding reaction that determines the presence of the antigen in a heterogeneous collection of proteins or biological substances (e.g., in a biological sample such as blood, serum, plasma, or tissue sample). In the context of describing the interaction between an antigen (e.g., a protein) and an antibody, antibody fragment, or binding agent derived from an antibody, and mean a binding reaction that determines the presence of the antigen in a heterogeneous collection of proteins or biological substances (e.g., in a biological sample such as blood, serum, plasma, or tissue sample).

[0078] The term "affinity" as used herein refers to the strength of the interaction between an antibody and an antigen at a single antigenic site. means the strength of the interaction between. Within each antigenic site, the variable regions of the antibody "arms" interact with the antigen through weak non-covalent binding forces at multiple sites, and the more interactions there are, the stronger the affinity is shown. A number of sites interact with the antigen through weak non-covalent binding forces, and the more interactions there are, the stronger the affinity. is shown.

[0079] The term "isolated antibody" substantially means an antibody that does not contain other antibodies having different antigen specificities. However, an isolated antibody that specifically binds to one antigen may have cross-reactivity with other antigens. Furthermore, an isolated antibody may not substantially contain other cellular materials and / or chemical substances. An antibody that specifically binds to one antigen may have cross-reactivity with other antigens. Furthermore, an isolated antibody may not substantially contain other cellular materials and / or chemical substances.

[0080] The term "corresponding human germline sequence" means a nucleic acid sequence encoding a human variable region amino acid sequence or a partial sequence, which, when compared to all other known or putative variable region amino acid sequences encoded by the human germline immunoglobulin variable region sequences, has the highest determined amino acid sequence identity with the reference variable region amino acid sequence or partial sequence. Also, the corresponding human germline sequence means a human variable region amino acid sequence or partial sequence that has the highest amino acid sequence identity with the reference variable region amino acid sequence or partial sequence when compared to all other evaluated variable region amino acid sequences. The corresponding human germline sequence can be a sequence containing only the framework region, only the complementarity-determining region, the framework and complementarity-determining regions, variable segments (as defined above), or other combinations of sequences or partial sequences containing the variable region. Sequence identity can be determined by aligning two sequences using the methods described herein, for example, using BLAST, ALIGN, or another alignment algorithm known in the art. The corresponding human germline sequence sequence can be determined by aligning two sequences using the methods described herein, for example, using BLAST, ALIGN, or another alignment algorithm known in the art. The corresponding human germline​​​​​​​​​​ The series of nucleic acid or amino acid sequences can have at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99 %, or 100% sequence identity with a reference variable region nucleic acid sequence or amino acid sequence.

[0081] Using various immunoassay formats, antibodies that are specifically immunoreactive with a particular protein can be selected. For example, solid-phase ELISA immunoassays are routinely used to select antibodies that are specifically immunoreactive with a protein (for a description of immunoassay formats and conditions that can be used to determine specific immunoreactivity, see, for example, Harlow & Lane, Using Antibodies, A Labor atory Manual (1998)). Typically, specific or selective binding reactions produce a signal that is at least 2-fold, more typically at least 10- to 100-fold, that of the background signal. For a description of immunoassay formats and conditions that can be used to determine specific immunoreactivity, see, for example, Harlow & Lane, Using Antibodies, A Labor atory Manual (1998)). Typically, specific or selective binding reactions produce a signal that is at least 2-fold, more typically at least 10- to 100-fold, that of the background signal. binding reactions produce a signal that is at least 2-fold, more typically at least 10- to 100-fold, that of the background signal. binding reactions produce a signal that is at least 2-fold, more typically at least 10- to 100-fold, that of the background signal.

[0082] The term "equilibrium dissociation constant (KD, M)" means the dissociation rate constant (Kd, time -1 , M -1 ) divided by the association rate constant (Ka, time -1 ). The equilibrium dissociation constant can be measured using methods known in the art. The antibodies of the present disclosure generally have an equilibrium dissociation constant of about 10 or less than 10 -7 or less than 10 -8 M, for example, about 10 -9 M or less than 10 -10 M, and in some embodiments, less than 10 -11 M, 10 -12 M or less than 10 -13 -13 M.

[0083] The term "biological availability" means the systemic availability (i.e., blood / plasma levels) of a given dose of a drug administered to a patient. Biological availability is an absolute term and indicates measurements of both the time (rate) and the total amount (extent) of drug reaching the general circulation from the administered dosage form.

[0084] As used in the specification, the phrase "consisting essentially of" means the genus or species of active agent(s) included in a method or composition, as well as any excipients that are inactive for the intended purpose of the method or composition. In some embodiments, the phrase "consisting essentially of" expressly excludes the inclusion of one or more additional active agents other than the anti-BK or JC antibodies of the present disclosure. In some embodiments, the phrase "consisting essentially of" expressly excludes the inclusion of one or more additional active agents other than the anti-BK or JC antibodies of the present disclosure and a second co-administered agent.

[0085] The term "amino acid" refers to amino acids of natural origin, synthetic and non-natural amino acids, as well as amino acid analogs and mimetics that function in a manner similar to amino acids of natural origin. Amino acids of natural origin are those encoded by the genetic code, as well as amino acids that are subsequently modified, such as hydroxyproline, γ-carboxyglutamic acid and O-phosphoserine. Amino acid analogs mean compounds having the same basic chemical structure as amino acids of natural origin, i.e., an α-carbon bonded to a hydrogen, a carboxyl group, an amino group, and an R group, and include, for example, homoserine, norleucine, methionine sulfoxide, methionine methylsulfonium. Such analogs are modified ... has an R group (e.g., norleucine) or a modified peptide backbone, but retains the same basic chemical structure as amino acids of natural origin and retains the same basic chemical structure as the amino acids of natural origin. An amino acid mimetic means a chemical compound having a structure different from the general chemical structure of an amino acid, but which functions in a manner similar to an amino acid of natural origin .

[0086] The term "conservatively modified variant" applies to both amino acid and nucleic acid sequences. With respect to a particular nucleic acid sequence, a conservatively modified variant is a nucleic acid that encodes the same or essentially the same amino acid sequence, or, if the nucleic acid does not encode an amino acid sequence, a nucleic acid that encodes an essentially the same sequence. Because of the degeneracy of the genetic code, a large number of functionally identical nucleic acids encode any given protein. For example, the codons GCA, GCC, GCG, and GCU all encode the amino acid alanine. Thus, at every position where an alanine is specified by a codon, the codon can be altered to any of the corresponding codons described without changing the encoded polypeptide. Such nucleic acid variants are "silent variants" and are one type of conservatively modified variant. Also, all nucleic acid sequences herein that encode polypeptides describe every possible silent variant of the nucleic acid. One of ordinary skill in the art will recognize that each codon in a nucleic acid (except for the ATG which is ordinarily the only codon for methionine, and TGG which is ordinarily the only codon for tryptophan) can be modified to provide a functionally identical molecule . Thus, each silent variant of a nucleic acid that encodes a polypeptide is implicit in each described sequence . Such nucleic acid variants are "silent variants" and are one type of conservatively modified variant. Also, all nucleic acid sequences herein that encode polypeptides describe every possible silent variant of the nucleic acid. One of ordinary skill in the art will recognize that each codon in a nucleic acid (except for the ATG which is ordinarily the only codon for methionine, and TGG which is ordinarily the only codon for tryptophan) can be modified to provide a functionally identical molecule . Thus, each silent variant of a nucleic acid that encodes a polypeptide is implicit in each described sequence . will recognize that each codon in a nucleic acid (except for the ATG which is ordinarily the only codon for methionine, and TGG which is ordinarily the only codon for tryptophan) can be modified to provide a functionally identical molecule . Thus, each silent variant of a nucleic acid that encodes a polypeptide is implicit in each described sequence .

[0087] For a polypeptide sequence, a "conservatively modified variant" is an individual substitution, deletion, or addition to the polypeptide sequence that results in a substitution of an amino acid with a chemically similar amino acid. Tables of conserved substitutions that give functionally similar amino acids are known in the art . Such conservatively modified variants are polymorphic variants, interspecies homologs, and alleles and are additional to, and do not exclude, the following 8 groups include amino acids that are conservative substituents of each other : 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 ); 8) cysteine (C), methionine (M) (see, e.g., Creighton, Proteins (1984)). In some embodiments, the term "conservative sequence modification" is used to mean an amino acid modification that does not significantly affect or change the binding properties of an antibody comprising the amino acid sequence .

[0088] As used herein, the term "optimized" means a nucleotide sequence that has been modified to encode an amino acid sequence using codons that are preferred in a production cell or organism (generally, eukaryotic cells, such as yeast cells, Pichia cells, fungal cells, Trichoderma cells, Chinese hamster ovary cells (CHO) or human cells). An optimized nucleotide sequence is designed to retain, as completely as possible, the amino acid sequence originally encoded by the first nucleotide sequence, also known as the "parent sequence" . ​

[0089] The terms "percent identity" or "percent identical" in the context of two or more nucleic acids or polypeptide sequences mean the degree to which two or more sequences or subsequences are identical. Two sequences are "identical" if they have the same amino acid or nuc leotide sequence over the region being compared. Two sequences are "substantially the same" if, when compared using one of the following sequence comparison algorithms or by measurement using manual alignment and visual inspection, the two sequences have a specified percentage of amino acid residues or nucleotides that are identical over a comparison window or specified region (i.e., 60% identity, optionally over a specified region or, if not specified, over the entire sequence, 6 5%, 70%, 75%, 80%, 85%, 90%, 95%, or 99% identity). Optionally, the identity exists over a region that is at least about 30 nucleotides (or 10 amino acids) in length, more preferably over a region that is 100 - 500 or 1000 or more nucleotides (or 20, 50, 200 or more amino acids) in length.

[0090]

[0090] For sequence comparison, typically one sequence is made to act as a reference sequence and the test sequence is compared. When using a sequence comparison algorithm, the test and reference sequences are input into a computer, subsequence coordinates are specified if necessary, and the program parameters of the sequence algorithm are specified. Default program parameters can be used, or other parameters A tag can be specified. Then, the array comparison algorithm calculates the percent sequence identity of the test array to the reference array based on the program parameter and.

[0091] As used herein, "comparison window" refers to any segment of a number of contiguous positions selected from the group consisting of 20 to 600, usually about 50 to about 200, more usually about 100 to about 150, wherein the sequences can be compared to a reference sequence of the same number of adjacent positions after the two sequences are optimally aligned. The methods of aligning sequences for comparison are known in the art. Optimal alignment of sequences for comparison can be conducted, for example, by the local homology algorithm of Smith and Waterman, Adv. Appl. Math. 2:482 c(1970), by the homology alignment algorithm of Needleman and Wunsch, J. Mol. Biol. 48:443 (1970), by the similarity search method of Pearson and Lipman, Proc. Natl. Acad. Sci. USA 85:2444 (1988), by computerized implementations of these algorithms (GAP, BESTFIT, FASTA, and TFASTA in the Wisconsin Genetics Software Package, Genetics Computer Group, 575 Science Dr., Madison, WI), or by manual alignment and visual inspection (see, e.g., Brent et al., Current Protocols in Molecular Biology, 2003). and can be performed. algorithm, by the similarity search method of Pearson and Lipman, Proc. Natl. Acad. Sci. USA 85:2444 (1988), by computerized implementations of these algorithms (GAP, BESTFIT, FASTA, and TFASTA in the Wisconsin Genetics Software Package, Genetics Computer Group, 575 Science Dr., Madison, WI), or by manual alignment and visual inspection (see, e.g., Brent et al., Current Protocols in Molecular Biology, 2003). inspection (see, e.g., Brent et al., Current Protocols in Molecular Biology, 2003) (see also). This can be done.

[0092] Two examples of algorithms suitable for determining sequence identity and sequence similarity are BLA The ST and BLAST2.0 algorithms are based on the Altschul et al. l., Nuc. Acids Res. 25:3389-3402, 1977, and Al tschul et al., J. Mol. Biol. 215:403-410,199 0. Software for performing BLAST analyses is provided by National Academic Press, New York, NY. Through the Center for Biotechnology Information The algorithm first finds short sequences of length W in the query sequence. The method includes identifying high-scoring sequence pairs (HSPs) by identifying words, When aligned with a word of the same length in the database sequence, it has a positive threshold score T. T is the neighborhood word score threshold (Altschul et al., supra) These initial neighborhood word hits act as seeds for initiating searches. The word hits are then combined to find longer HSPs that contain them. The cumulative alignment score is The cumulative score is calculated by multiplying the nucleotide sequence by 100. For , the parameters M (the reward score for matching residue pairs, always > 0) and N (penalty score for mismatched residues, always <0). For a sequence, a scoring matrix is ​​used to calculate the cumulative score. The extension of a word hit in increases the cumulative alignment score by an amount X from its maximum score. When a negative score is reached, the accumulation of one or more negatively scoring residue alignments reduces the cumulative score below zero. Stop when going down or when reaching the end of any array. BLAST a The BLAST algorithm parameters W, T, and X determine the sensitivity and speed of the alignment. B The LASTN program (for nucleotide sequences) uses a word length (W) of 11, an expectation value (E) of 10, M = 5, N = 4, and two-strand comparison as the default. For amino acid sequences the BLASTP program uses a word length of 3, an expectation value (E) of 10, and the BLOSUM62 scoring matrix (Henikoff and Heniko ff, (1989) Proc. Natl. Acad. Sci. USA 89:10915 see reference), an alignment (B) of 50, an expectation value (E) of 10, M = 5, N = 4, and two-strand comparison as the default.

[0093] Also, the BLAST algorithm performs a statistical analysis of the similarity between two sequences (e.g., see Karlin and Altschul, Proc. Natl. Acad. Sci . USA 90:5873 - 5787, 1993). One measure of similarity provided by the BLAST algorithm is the minimum total probability (P(N)), which provides an indication of the likelihood that a match between two nucleotide sequences or amino acid sequences occurs by chance. For example, if the minimum total probability in the comparison of a test nucleic acid and a control nucleic acid is less than about 0.2, more preferably less than about 0.01, and most preferably less than about 0.001, the nucleic acid is considered to be similar to the control sequence.

[0094] Also, the percent identity between two amino acid sequences is determined by the algorithm of E. Meyers and W. Miller incorporated into the ALIGN program (vers ion 2.0)​​ which can be determined using (Comput.Appl.Biosci.4:11-17 ,1988), the PAM120 weight residue table, a gap length penalty of 12, and a gap penalty of 4. Additionally, it is incorporated into the GAP program of the GCG software package (available from the Univers ity of South Florida) and the Needleman and Wunsch (J.Mol.Biol.48:444- 453,1970) algorithm is used to determine the percent identity of two amino acid sequences, with the BLOSUM 62 matrix or PAM250 matrix and gap weights of 16, 14, 12, 10, 8, 6, or 4 and length weights of 1, 2, 3, 4, 5, or 6.

[0095] Except for the percent sequence identity described above, another indication that two nucleic acid sequences or polypeptides are substantially identical is that the polypeptide encoded by the first nucleic acid is immunologically cross-reactive with an antibody produced against the polypeptide encoded by the second nucleic acid, as follows. Thus, a polypeptide is typically, for example, substantially identical to a second polypeptide when the two polypeptides differ only by conservative substitutions. Another indication that two nucleic acid sequences are substantially identical is that, as shown below, the two molecules or their complements hybridize to each other under stringent conditions. Additionally, another indication that two nucleic acid sequences are substantially identical is that they can be used to amplify sequences using the same primers.

[0096] ​​​​​The term "nucleic acid" is used interchangeably herein with the term "polynucleotide" and means deoxyribonucleotides or ribonucleotides and polymers thereof, which can be in either single-stranded or double-stranded form. The term includes nucleic acids containing known nucleotide analogs or modified backbone residues or linkages, which are synthetic, naturally occurring, and non-naturally occurring, have similar binding properties as reference nucleic acids, and are metabolized in a manner similar to reference nucleotides. Examples of such analogs include, but are not limited to, phosphorothioates, phosphoramidates, methylphosphonates, chiral-methylphosphonates, 2-O-methylribonucleotides, peptide-nucleic acids (PNAs), etc. Unless otherwise specified, a particular nucleic acid sequence implicitly encompasses its conservatively modified variants (e.g., degenerate codon substitutions) and complementary sequences, as well as the explicitly recited sequences. Specifically, as detailed below, degenerate codon substitutions can be achieved by generating sequences in which the third position of one or more selected (or all) codons is substituted with a mixture of bases and / or deoxyinosine residues (Batzer et al., (1991) Nucleic Acid Res. 19:5081, Ohtsuka et al., (1985) J. Biol. Chem. 260:2605-2608,

[0097] and Rossolini et al., (1994) Mol. Cell Probes 8:91-98). In the context of nucleic acids, the term "operably linked" means that two or more polynucleotides are ligated together such that the resulting nucleic acid molecule can be transcribed and / or translated into a functional product, wherein the transcription and / or translation of the nucleic acid molecule is regulated by the presence of a regulatory sequence(s). For example, a promoter sequence and a coding sequence are operably linked if the promoter sequence can drive the transcription of the coding sequence into mRNA, which can then be translated into a polypeptide. Similarly, a regulatory sequence and a coding sequence are operably linked if the regulatory sequence can modulate the expression of the coding sequence, either by enhancing or repressing its transcription and / or translation.

[0098] In the context of nucleic acids, the term "functionally linked" means that two or more polynucleotides ​It refers to the functional relationship between two or more sequences (e.g., DNA) of a gene. Typically, transcriptional regulation Refers to the functional relationship between a sequence and a transcribed sequence, e.g., a promoter or enhancer. The sequence may be used to stimulate or regulate transcription of a coding sequence in an appropriate host cell or other expression system. A coding sequence is operably linked when it is operably linked to a coding sequence. Generally, it is operably linked to a transcribed sequence. The ligated promoter transcriptional regulatory sequence is physically adjacent to the transcribed sequence, i.e. However, some transcriptional regulatory sequences, such as enhancers, act in a cis-acting manner. It is not necessary that the gene be physically adjacent or located contiguous to the coding sequence it enhances.

[0099] The terms "polypeptide" and "protein" are used herein to refer to a group consisting of amino acid residues. The term is used interchangeably to refer to a polymer in which one or more amino acid residues correspond Amino acid polymers that are artificial chemical mimics of naturally occurring amino acids that exhibit applies to amino acid polymers of any of the above and to amino acid polymers of non-naturally occurring origins unless otherwise specified. To the extent possible, a particular polypeptide sequence also implicitly embraces conservatively modified variants thereof.

[0100] The term "subject" includes humans and non-human animals. Non-human animals include all vertebrate motion. animals, such as non-human primates, sheep, dogs, cows, chickens, amphibians, and reptiles. Except where noted, the term "patient" or "subject" may be used interchangeably with "subject." The terms "elephant" and "elephant" are used interchangeably herein.

[0101] The term "BKV" or "BK virus" refers to the Polyomaviridae, Orthopolypeptide Polyomaviruses are members of the genus Polyomavirus. They are icosahedral, non-enveloped, double-stranded DNA viruses with a genome. They have a diameter of approximately 40 - 45 nm (Bennett et al., Microbes and Infection. 2012:14(9):672 - 683).

[0102] The terms "JCV" or "JC virus" refer to members of the Polyomaviridae family, Orthopolyomavirus genus. JCV is related to BKV and is also an icosahedral, non-enveloped, double-stranded DNA virus with a genome of approximately 5,000 base pairs. They have a diameter of approximately 40 - 45 nm (Johne et al., Arch. V irol. 2011;156(9):1627 - 1634).

[0103] The terms "BKV nephropathy" or "BKV-associated nephropathy" or "BKVAN" refer to inflammatory interstitial nephritis resulting from lytic infection by BKV, characterized mainly by viral cytopathic changes and expression of viral genes

[0104] in renal tubular epithelium. The term "VP1" refers to the major polyomavirus capsid subunit protein. [Table 1-1] [Table 1-2]

[0105] A "virus-like particle" or "VLP" is an assembly of VP1 pentamers into the viral capsid. A VLP is composed of 72 VP1 pentamers. A VLP has a structure structurally very similar, but lack the minor capsid proteins (VP2, VP3) as well as the viral DNA genome and are therefore non-infectious. VLPs are useful as viral epitopes and are presented in a form similar to that of the actual virus. and are therefore non-infectious. VLPs are useful as viral epitopes and are presented in a form similar to that of the actual virus. and are presented in a form similar to that of the actual virus.

[0106] The "IC50" (half-maximal inhibitory concentration) refers to the concentration of a specific antibody that induces a signal midway (50%) between the baseline control and the maximum possible signal. For example, the IC50 is the concentration of antibody at which 50% of the available binding sites on the VP1 antigen are occupied. is the concentration of antibody at which 50% of the available binding sites on the VP1 antigen are occupied.

[0107] The "EC50" (half-maximal effective concentration) means the concentration of a specific antibody that induces a response midway (50%) between the baseline control and the maximum possible effect after a specific exposure or treatment time. For example, the EC50 is the concentration of antibody at which 50% of the viral infection is neutralized.

[0108] "EC90" means the concentration of a specific antibody that induces a response corresponding to 90% of the maximum possible effect after a specific exposure or treatment time. For example, the EC90 is the concentration of antibody at which 90% of the viral infection is neutralized. is the concentration of antibody at which 90% of the viral infection is neutralized.

[0109] "Neutralization" means the inhibition of viral infection of host cells as indicated by the absence of viral gene expression. Without being bound by any theory, the mechanism of neutralization by a specific antibody may involve preventing the interaction between the viral capsid protein and the cell surface receptor before the viral genome is delivered to the nucleus of the host cell, or preventing the disruption of any stage of the entry and transport processes. Without being bound by any theory, the mechanism of neutralization by a specific antibody may involve preventing the interaction between the viral capsid protein and the cell surface receptor before the viral genome is delivered to the nucleus of the host cell, or preventing the disruption of any stage of the entry and transport processes. Without being bound by any theory, the mechanism of neutralization by a specific antibody may involve preventing the interaction between the viral capsid protein and the cell surface receptor before the viral genome is delivered to the nucleus of the host cell, or preventing the disruption of any stage of the entry and transport processes. Without being bound by any theory, the mechanism of neutralization by a specific antibody may involve preventing the interaction between the viral capsid protein and the cell surface receptor before the viral genome is delivered to the nucleus of the host cell, or preventing the disruption of any stage of the entry and transport processes. Without being bound by any theory, the mechanism of neutralization by a specific antibody may involve preventing the interaction between the viral capsid protein and the cell surface receptor before the viral genome is delivered to the nucleus of the host cell, or preventing the disruption of any stage of the entry and transport processes.

[0110] As used herein, the terms "treating," "treatment of," or "treatment" of any disease or disorder, in one aspect, means alleviating (i.e., delaying, or arresting, or reducing) at least one of the disease or its clinical symptoms. In another aspect, it means reducing or alleviating at least one physical parameter, including those not identifiable in the patient. In yet another aspect, "treating," "treatment of," or "treatment" means modulating the disease or disorder, either physically (e.g., stabilization of identifiable symptoms), physiologically (e.g., stabilization of physical parameters), or both. The phrase "reducing the likelihood of" means delaying the onset or progression of a disease, infection, or disorder. The terms "therapeutically acceptable amount" or "therapeutically effective amount" are used interchangeably to mean an amount sufficient to produce the desired result (i.e., reduction in tumor size, inhibition of tumor growth, prevention of metastasis, inhibition or prevention of viral, bacterial, fungal, or parasitic infection). In some aspects, a therapeutically acceptable amount does not induce or cause undesirable side effects. A therapeutically acceptable amount can be administered at a low dose initially determined and then increased incrementally until the desired effect is achieved. The "prophylactically effective dosage" and "therapeutically effective dosage" of the molecules of the present disclosure can each prevent the onset of disease symptoms, including those associated with polyomavirus infection, or result in a reduction in severity.

[0111]

[0112]

[0113] ​​​​​​​​​​​​​ The term "co - administration" means the simultaneous presence of two active agents in the blood of an individual. Co - administered active agents can be delivered simultaneously or sequentially.

Brief Description of the Drawings

[0114]

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Figure 3H

[0115] The present disclosure provides antibodies, antibody fragments (e.g., antigen-binding fragments) that bind to and neutralize BKV. Further, the present disclosure provides antibodies having desired pharmacokinetic properties and other desirable attributes, and thus can be used to treat or reduce the likelihood of BK virus-associated nephropathy (e.g., BKVAN) and / or JC virus-associated progressive multifocal leukoencephalopathy (PML). The present disclosure further provides pharmaceutical compositions comprising the antibodies, and for the prevention and treatment of polyomavirus infections and related disorders, such pharmaceutical compositions and and / or and / or JC virus-associated progressive multifocal leukoencephalopathy (PML). The present disclosure further provides pharmaceutical compositions comprising the antibodies, and for the prevention and treatment of polyomavirus infections and related disorders, such pharmaceutical compositions can be used. The present disclosure further provides pharmaceutical compositions comprising the antibodies, and for the prevention and treatment of polyomavirus infections and related disorders, such pharmaceutical compositions can be used for the prevention and treatment of polyomavirus infections and related disorders. A method of making and using an article is provided.

[0116] Anti-polyomavirus antibody The present disclosure provides antibodies and antibody fragments (e.g., antigen-binding fragments) that specifically bind to BK or JC virus. The antibodies or antibody fragments (e.g., antigen-binding fragments) of the present disclosure include, but are not limited to, the isolated human monoclonal antibodies or fragments thereof described in the following examples. These are not limited thereto.

[0117] One aspect of the present disclosure provides an antibody or antibody fragment (e.g., antigen-binding fragment) that specifically binds to BK or JC virus, wherein the antibody or antibody fragment (e.g., antigen-binding fragment) includes a VH domain having the amino acid sequence of SEQ ID NOs: 18, 50, 82, 114, 146, 178, 210, 242, and 274 (Table 2). The present disclosure also provides an antibody or antibody fragment (e.g., antigen-binding fragment) that specifically binds to BK or J C virus, and the antibody or antibody fragment (e.g., antigen-binding fragment) includes a VH CDR having any of the amino acid sequences of the VH CDRs listed in Table 2. In certain aspects, the present disclosure provides an antibody or antibody fragment (e.g., antigen-binding fragment) that specifically binds to BK or JC virus, and the antibody includes, or alternatively consists of, 1, 2, 3 or more of the VH CDRs having any of the amino acid sequences of the VH CDRs listed in Table 2.

[0118] The present disclosure provides an antibody or antibody fragment (e.g., antigen-binding fragment) that specifically binds to BK or JC virus, and the antibody or antibody fragment (e.g., antigen-binding fragment) has a SEQ ID NO. ​​​Numbers 34, 66, 98, 130, 162, 194, 226, 258, and 290 (Table 2) comprises a VL domain having the amino acid sequence. The present disclosure also provides an antibody or antibody fragment (e.g., antigen-binding fragment) that specifically binds to BK or JC virus, and said antibody or antibody fragment (e.g., antigen-binding fragment) comprises a VL CDR having the amino acid sequence of any of the VL CDRs listed in Table 2. In particular, the present disclosure provides an antibody or antibody fragment (e.g., antigen-binding fragment) that specifically binds to BK or JC virus, and said antibody or antibody fragment (e.g., antigen-binding fragment) comprises one, two, three or more VL CDRs having the amino acid sequence of any of the VL CDRs listed in Table 2 (or, equivalently, consists of them). antibody fragment (e.g., antigen-binding fragment) comprises one, two, three or more VL CDRs having the amino acid sequence of any of the VL CDRs listed in Table 2 (or, equivalently, consists of them). consists of them).

[0119] Other antibodies or antibody fragments (e.g., antigen-binding fragments) of the present disclosure contain mutated amino acids, but have at least 60, 70, 8 0, 90 or 95% identity in the CDR regions shown in the sequences listed in Table 2. In some embodiments, when compared to the CDR regions shown in the sequences listed in Table 2, the CDR regions contain an amino acid sequence in which 1, 2, 3, 4 or 5 or more amino acids are not mutated. are not mutated. are not mutated.

[0120] The present disclosure also provides nucleic acid sequences encoding VH, VL, full-length heavy chains, and full-length light chains of antibodies that specifically bind to BK or JC virus. Such nucleic acid sequences can be optimized for expression in mammalian cells

Table 2-1

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[0121] Other antibodies of the present disclosure include those in which the amino acids or the nucleic acids encoding the amino acids are mutated, but are at least 60, 70, 80, 90 or 95% It has identity. In some embodiments, when compared to the variable regions shown in the sequences listed in Table 2, one, two, three, four, or five or more amino acids in the variable regions are not mutated and contain a mutant amino acid sequence that retains substantially the same therapeutic activity.

[0122] Since these antibodies can each bind to VP1, VH, VL, full-length light chain, and full-length heavy chain sequences (amino acid sequences and nucleotide sequences encoding the amino acid sequences) can be "mixed and paired" to generate VP1-binding antibodies. Such " mixed and paired" VP1-binding antibodies can be tested using binding assays known in the art (e.g., ELISA and other assays described in the Examples section). When these chains are mixed and paired, the VH sequence from a particular VH / VL pairing should be replaced with a structurally similar VH sequence. Similarly, the full-length heavy chain sequence from a particular full-length heavy chain / full-length light chain pairing must be replaced with a structurally similar full-length heavy chain sequence. Similarly, the VL sequence from a particular VH / VL pairing must be replaced with a structurally similar VL sequence. Similarly, the full-length light chain sequence from a particular full-length heavy chain / full-length light chain pairing must be replaced with a structurally similar full-length light chain sequence. Thus, in one embodiment, the present disclosure provides an isolated monoclonal having a heavy chain variable region comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 18, 50, 82, 114, 146, 178, 210, 242, and 27 4 (Table 2), and a light chain variable region comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 34, 66, 98, 130, 162, 194, 226, 258, and 290 (Table 2). ​​​​​​​​​​and providing a monoclonal antibody or antigen-binding region thereof, the antibody being capable of binding to BK or JC virus. Binds specifically.

[0123] In another aspect, the disclosure provides a method for the production of an antibody comprising: (i) a full-length heavy chain comprising an amino acid sequence selected from Table 2; and and a full length light chain comprising an amino acid sequence selected from Table 2, The present invention provides isolated monoclonal antibodies having sequences optimized for the expression of the same. In this regard, the present disclosure provides: (i) SEQ ID NOs: 20, 52, 84, 116, 148, 180, 2 12, 244, and 276 (Table 2), and expression in mammalian cells. and full-length heavy chains comprising amino acid sequences optimized for SEQ ID NOs: 36, 68, 100, , 132, 164, 196, 228, 260, and 292 (Table 2) and a full-length light chain comprising an amino acid sequence optimized for expression in mammalian cells. or (ii) a functional antibody comprising an antigen-binding portion thereof. Provides quality.

[0124] In another embodiment, the present disclosure provides heavy and light chain CDR1, CDR2, and CDR3, or a combination thereof. The amino acid sequences of the VH CDR1 of the antibodies are set forth in SEQ ID NOs: 9, 41, 73, 105, 137, , 169, 201, 233, and 265. The sequences are SEQ ID NOs: 10, 42, 74, 106, 138, 170, 202, 234, and The amino acid sequences of the VH CDR3 of the antibodies are shown in SEQ ID NOs: 11, 43, 75 , 107, 139, 171, 203, 235, and 267. The amino acid sequences of DR1 are shown in SEQ ID NOs: 25, 57, 89, 121, 153, 185, 217 , 249, and 281. The amino acid sequences of the VL CDR2 of the antibodies are shown in SEQ ID NOs: 26, 58, 90, 122, 154, 186, 218, 250, and 282 . The amino acid sequences of the VL CDR3 of the antibodies are shown in SEQ ID NOs: 27, 59, 91, 123, 155 , 187, 219, 251, and 283.

[0125] If each of these antibodies can bind to BK or JC virus and its antigen-binding specificity is mainly provided by the CDR1, 2, and 3 regions, assuming that the VH CDR1, 2 , and 3 sequences, or the VL CDR1, 2, and 3 sequences, can be "mixed and aligned" (i.e., each antibody must contain VH CDR1, 2 , and 3, as well as VL CDR1, 2, and 3 to generate other VP1-binding molecules, but the CDRs from different antibodies can be mixed and aligned). Such "mixed and aligned" V P1-binding antibodies can be tested using binding assays known in the art described in the examples (e.g., E LISA). When the VH CDR sequences are mixed and aligned , the CDR1, CDR2, and / or CDR3 sequences from a particular VH sequence must be replaced with structurally similar CDR sequences. Similarly, when the VL CDR sequences are mixed and aligned , the CDR1, CDR2, and / or CDR3 sequences from a particular VL sequence must be replaced with structurally similar CDR sequences. For the monoclonal antibodies of the present disclosure, one or more VH and / or VL CDR region sequences are shown herein , and can be used to generate "mixed and aligned" antibodies with altered antigen-binding specificities, such as by replacing the CDR sequences from a particular VH or VL sequence with structurally similar CDR sequences from another VH or VL sequence. Replacing with structurally similar sequences from the CDR sequences that can be obtained allows for the generation of novel VH and VL sequences, which will be readily apparent to those skilled in the art.

[0126] Accordingly, the present disclosure provides a heavy chain CDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 9, 41, 73, 105, 137, 169, 201, 233, and 265, a heavy chain CDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 10, 42, 74, 106, 138, 170, 202, 234, and 266, a heavy chain CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 11, 43, 75, 107, 139, 171, 203, 235, and 267, a light chain CDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 25, 57, 89, 121, 153, 185, 217, 249, and 281, a light chain CDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 26, 58, 90, 122, 154, 186, 218, 250, and 2 82, and a light chain CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 27, 59 91, 123, 155, 187, 219, 251, and 283, and provides an isolated monoclonal antibody or its antigen-binding region, wherein the antibody specifically binds to BK or JC virus. In certain embodiments, an antibody that specifically binds to BK or JC virus is an antibody or antibody fragment (e.g., antigen-binding fragment) described in Table 2. In one aspect, an antibody that specifically binds to BK or JC virus is an antibody or antibody fragment (e.g., antigen-binding fragment) described in Table 2. antigen-binding region, wherein the antibody specifically binds to BK or JC virus. The antibody specifically binds to BK or JC virus.

[0127] In certain embodiments, an antibody that specifically binds to BK or JC virus is an antibody or antibody fragment (e.g., antigen-binding fragment) described in Table 2. antibody or antibody fragment (e.g., antigen-binding fragment) described in Table 2.

[0128] 1. Identification of Antibodies The present disclosure provides antibodies and antibody fragments (e.g., antigen-binding to provide fragments). In certain embodiments, the antibodies and antibody fragments are capable of binding to the same epitope among all four BKV serotypes and / or JCV.

[0129] In addition, the present disclosure provides antibodies and antibody fragments (e.g., antigen-binding fragments) that bind to the same epitope as the anti-BK or JC antibodies described in Table 2. Thus, additional antibodies and antibody fragments (e.g., antigen-binding fragments) can be identified based on their ability to cross-compete with other antibodies in a binding assay (e.g., competitively inhibit binding in a statistically significant manner). The ability of a test antibody to inhibit the binding of the antibodies and antibody fragments (e.g., antibody fragments) of the present disclosure to BK or JC virus can demonstrate that the test antibody can compete with an antibody or antibody fragment (e.g., antigen-binding fragment) for binding to BK or JC virus, and such an antibody, according to non-limiting theory, can bind to the same or a related (e.g., structurally similar or spatially proximal) epitope on BK or JC virus as the antibody or antibody fragment (e.g., antigen-binding fragment) with which it competes. In certain embodiments, the antibody or antibody fragment (e.g., antigen-binding fragment) of the present disclosure that binds to the same epitope on BK or JC virus is a human or humanized monoclonal antibody. Such human or humanized monoclonal antibodies can be prepared and isolated as described in the present specification. 2. Further modification of the framework of the Fc region The present disclosure discloses specific anti-BK or JC virus antibodies. These antibodies have a VH region that binds to the same epitope on BK or JC virus as the antibody or antibody fragment (e.g., antigen-binding fragment) with which it competes. In certain embodiments, the antibody or antibody fragment (e.g., antigen-binding fragment) of the present disclosure that binds to the same epitope on BK or JC virus is a human or humanized monoclonal antibody. Such human or humanized monoclonal antibodies can be prepared and isolated as described in the present specification. region that binds to the same epitope on BK or JC virus as the antibody or antibody fragment (e.g., antigen-binding fragment) with which it competes. In certain embodiments, the antibody or antibody fragment (e.g., antigen-binding fragment) of the present disclosure that binds to the same epitope on BK or JC virus is a human or humanized monoclonal antibody. Such human or humanized monoclonal antibodies can be prepared and isolated as described in the present specification. region that binds to the same epitope on BK or JC virus as the antibody or antibody fragment (e.g., antigen-binding fragment) with which it competes. In certain embodiments, the antibody or antibody fragment (e.g., antigen-binding fragment) of the present disclosure that binds to the same epitope on BK or JC virus is a human or humanized monoclonal antibody. Such human or humanized monoclonal antibodies can be prepared and isolated as described in the present specification. region that binds to the same epitope on BK or JC virus as the antibody or antibody fragment (e.g., antigen-binding fragment) with which it competes. In certain embodiments, the antibody or antibody fragment (e.g., antigen-binding fragment) of the present disclosure that binds to the same epitope on BK or JC virus is a human or humanized monoclonal antibody. Such human or humanized monoclonal antibodies can be prepared and isolated as described in the present specification. region that binds to the same epitope on BK or JC virus as the antibody or antibody fragment (e.g., antigen-binding fragment) with which it competes. In certain embodiments, the antibody or antibody fragment (e.g., antigen-binding fragment) of the present disclosure that binds to the same epitope on BK or JC virus is a human or humanized monoclonal antibody. Such human or humanized monoclonal antibodies can be prepared and isolated as described in the present specification. region that binds to the same epitope on BK or JC virus as the antibody or antibody fragment (e.g., antigen-binding fragment) with which it competes. In certain embodiments, the antibody or antibody fragment (e.g., antigen-binding fragment) of the present disclosure that binds to the same epitope on BK or JC virus is a human or humanized monoclonal antibody. Such human or humanized monoclonal antibodies can be prepared and isolated as described in the present specification. region that binds to the same epitope on BK or JC virus as the antibody or antibody fragment (e.g., antigen-binding fragment) with which it competes. In certain embodiments, the antibody or antibody fragment (e.g., antigen-binding fragment) of the present disclosure that binds to the same epitope on BK or JC virus is a human or humanized monoclonal antibody. Such human or humanized monoclonal antibodies can be prepared and isolated as described in the present specification. region that binds to the same epitope on BK or JC virus as the antibody or antibody fragment (e.g., antigen-binding fragment) with which it competes. In certain embodiments, the antibody or antibody fragment (e.g., antigen-binding fragment) of the present disclosure that binds to the same epitope on BK or JC virus is a human or humanized monoclonal antibody. Such human or humanized monoclonal antibodies can be prepared and isolated as described in the present specification.

[0130] 2. Further modification of the framework of the Fc region The present disclosure discloses specific anti-BK or JC virus antibodies. These antibodies have a VH and / or further comprising modifications to residues of the framework within VL, the modified anti- body or antigen-binding fragment thereof, e.g., to improve antibody properties. Typically, such framework modifications are made to reduce the immunogenicity of the antibody. For example, one approach is to "revert mutate" one or more framework residues to their corresponding germline sequences. More specifically, an antibody that has undergone somatic mutations may contain framework residues that differ from the germline sequence from which the antibody is derived. Such residues can be identified by comparing the antibody framework sequence to the germline sequence from which the antibody is derived. To return the framework region sequences to their germline configuration, for example, by site-directed mutagenesis, somatic mutations can be "reverted mutated" to the germline sequence. Such "reverted mutated" antibodies are

[0131] also intended to be encompassed. Another type of framework modification involves mutating one or more residues within the framework region or even within one or more CDR regions, removing T cell epitopes, thereby reducing the potential immunogenicity of the antibody. This approach is also referred to as "deimmunization" and is described in more detail in U.S. Patent Publication No. 2003 / 0153043 by Carr et al.

[0132] In addition to, or alternatively to, modifications made within the framework or CDR regions, the antibody can be designed to include modifications within the Fc region, typically one or more functional properties of the antibody, e.g., serum half-life, complement binding, Fc Modify cytotoxicity. Furthermore, the antibody may be chemically modified (e.g., one or more chemical moieties can be attached to the antibody), or modified to alter its glycosylation to change one or more functional properties of the antibody. Each of these embodiments is described in more detail below.

[0133] In one embodiment, the hinge region of CH1 is modified by changing the number of cysteine residues in the hinge region, for example, increasing or decreasing it. This approach is further described in U.S. Patent No. 5,677,425 by Bode et al. The number of cysteine residues in the hinge region of CH1 is changed, for example, to facilitate the construction of light and heavy chains, or to increase or decrease the stability of the antibody.

[0134] In another embodiment, the fc hinge region of the antibody is mutated to reduce the biological half-life of the antibody. More specifically, one or more amino acid mutations are introduced into the CH2-CH3 domain interface region of the Fc-hinge fragment such that the SpA binding of the antibody is impaired relative to the binding of the native Fc-hinge domain to Staphylococcus aureus protein A ( SpA). This approach is further described in detail in U.S. Patent No. 6,165,745 by Ward et al.

[0135] In yet another embodiment, the Fc region is modified by substituting at least one amino acid residue with a different amino acid residue to change the effector function of the antibody. For example, one or more amino acids can be substituted with different amino acid residues such that the antibody has a changed affinity for effector ligands while retaining the antigen-binding ability of the parental antibody. The affinity is changed while the antigen-binding ability of the parental antibody is retained, so that the antibody has a changed affinity for effector ligands. The affinity is changed The effector ligand can be, for example, an Fc receptor or the C1 component of complement. This approach is described, for example, in U.S. Patent Nos. 5,624,821 and 5,648,260 by Winter et al.

[0136] In another aspect, one or more amino acids selected from amino acid residues can be used to change the C 1q binding property and / or the complement-dependent cytotoxicity (CDC) that has decreased or disappeared of the antibody, and can be substituted with different amino acid residues. This approach is described, for example, in U.S. Patent No. 6,194,551 by Idusogie et al.

[0137] In another aspect, by modifying one or more amino acid residues, the ability of the antibody to fix complement is changed. This approach is described, for example, in International Patent Application No. WO94 / 29351 by Bodmer et al. In certain aspects, one or more amino acids of the antibody or its antigen-binding fragment are substituted with one or more allotype amino acid residues for IgG1 subclass and κ isotype. Also, the allotype amino acid residues include, but are not limited to, the constant regions of the heavy chains of IgG1, IgG2, and IgG3 subclasses, as described in Jefferis et al., MAbs. 1:332 - 338 (2009), and the constant region of the light chain of the κ isotype.

[0138] In yet another aspect, the Fc region is modified to increase the ability of the antibody to mediate antibody-dependent cell cytotoxicity (ADCC) and / or to increase the affinity of the antibody for the Fcγ receptor by modifying one or more amino acids. This approach , for example, as described in International Patent Application No. WO00 / 42072 by Presta . Furthermore, the binding sites on human Ig G1 for FcγRl, FcγRII, FcγRIII and FcRn have been mapped and mutants with improved binding are described (see Shields et al., J. Biol. Chem. 276:6591-66 04, 2001).

[0139] In yet another aspect, the glycosylation of the antibody is modified. For example, non-glycosylated antibodies can be produced (i.e., the antibody lacks glycosylation). Glycosylation can be altered, for example, to increase the affinity of the antibody for the "antigen". Such carbohydrate modifications can be achieved, for example, by changing one or more glycosylation sites within the antibody sequence . For example, one or more amino acid substitutions can be made, resulting in the removal of glycosylation sites in one or more variable region frameworks, thereby eliminating glycosylation at those sites. Such glycosylation can increase the affinity of the antibody for the antigen. Such an approach is described, for example, in U.S. Pat. Nos. 5,714,350 and 6,350,861 by Co et al. . .

[0140] Additionally, or alternatively, antibodies with reduced amounts of fucosyl residues, hypofucosylated antibodies, or antibodies with an increased bisecting GlcNac structure, antibodies with modified forms of glycosylation, can be produced. Such modified glycosylation patterns have been demonstrated to increase the ADCC ability of the antibody. Such carbohydrate modifications are , for example, it can be achieved by expressing an antibody in a host cell having a modified glycosylation mechanism. Cells having a modified glycosylation mechanism are described in the art, and by using them as host cells to express a recombinant antibody, an antibody having modified glycosylation can be produced. For example, European Patent No. 1,176,195 by Hang et al. describes a cell line having an FUT8 gene encoding a functionally disrupted fucosyltransferase, and antibodies expressed in such cell lines exhibit hypofucosylation. International Patent Application No. WO03 / 035835 by Presta describes mutant CHO cell lines, Lecl3 cells, with reduced ability to bind fucose to Asn(297)-linked carbohydrates, resulting in hypofucosylation of antibodies expressed in those host cells (see Shields et al., (2002) J. Biol. Chem. 277:26733-26740). International Patent Application No. WO99 / 54342 by Umana et al. describes a cell line designed to express a glycoprotein-modifying glycosyltransferase (e.g., β(1,4)-N-acetylglucosaminyltransferase III (GnTIII)), and antibodies expressed in the modified cell line exhibit an increased bisecting GlcNAc structure, resulting in increased ADCC activity of the antibody (see Umana et al., Nat. Biotech. 17:176-180, 1999). In another aspect, the antibody is modified to increase its biological half-life. Various approaches are possible. For example, U.S. Patent No. 6,277,375 by Ward describes

[0141] ​​​​​​​​​​​​​​​​As described, one or more of the following mutations: T252L, T254S, T256F, can be introduced. Alternatively, to increase the biological half-life, Presta et al. described in U.S. Patent Nos. 5,869,046 and 6,121,022, the antibody can be modified within the CH1 or CL region to include a salvage receptor binding epitope obtained from two loops of the CH2 domain of the Fc region of IgG.

[0142] In minimizing the ADCC activity of the antibody, specific mutations in the Fc region result in an "Fc silent" antibody with minimal interaction with effector cells. Generally, the "IgG Fc region" is used to define the C-terminal region of the immunoglobulin heavy chain, including the native sequence Fc region and variant Fc regions. The human IgG heavy chain Fc region is generally defined as the amino acid residues from position C226 or P230 to the carboxyl terminus of the IgG antibody. The numbering of residues in the Fc region is that of the Kabat EU index. The C-terminal lysine (residue K447) of the Fc region can be removed, for example,

[0143] during antibody production or purification. Suppressed effector functions are obtained by mutations in the Fc region of the antibody 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., 2008, J. Immu ​See U.S. Patent No. 12 / 065,950. Examples of silent Fc IgG1 antibodies are LALA mutants that contain the L234A and L235A mutations in the Fc amino acid sequence and there are some. Other examples of silent IgG1 antibodies are the DAPA (D265A, P329A) mutations (U.S. Patent No. US6,737,056). Another silent IgG1 antibody contains the N 297A mutation, and a non-glycosylated / unglycosylated antibody is obtained.

[0144] Fc silent antibodies have no or low ADCC activity, which means that Fc silent antibodies exhibit ADCC activity where specific cell lysis is less than 50% (low ADCC activity) or specific cell lysis is less than 1% (no ADCC activity).

[0145] 3. Production of Antibodies Anti-BK or JC virus antibodies and antibody fragments (e.g., antigen-binding fragments) can be produced by any means known in the art, including but not limited to recombinant expression, chemical synthesis, and enzymatic digestion of antibody tetramers, while full-length monoclonal antibodies can be obtained, for example, by hybridoma or recombinant production. Recombinant expression can be carried out in any suitable host cell known in the art, such as mammalian host cells, bacterial host cells, yeast host cells, insect host cells, etc.

[0146] The present disclosure provides polynucleotides encoding the antibodies described herein, for example, polynucleotides encoding heavy or light chain variable regions or segments containing the complementarity-determining regions described herein or segments. In some embodiments, the polynucleotide encoding the heavy chain variable region is set forth in SEQ ID NOs: 19, 51, 83, 115, 147, 179, 211, 243, and 27​​​​ A polynucleotide selected from the group consisting of 5., and at least 85%, 89%, 90% , 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% nucleic acid sequence identity. In some embodiments, the polynucleotide encoding the light chain variable region is a polynucleotide selected from the group consisting of SEQ ID NOs: 35, 67, 99, 131, 163, 195, 227, 259, and 291, and has at least 85%, 89% , 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% , or 100% nucleic acid sequence identity.

[0147] In some embodiments, the polynucleotide encoding the heavy chain variable region is a polynucleotide selected from the group consisting of SEQ ID NOs: 21, 53 , 85, 117, 149, 181, 213, 245, 277 and has at least 85%, 89%, 90%, 91%, 92%, 93%, 94 %, 95%, 96%, 97%, 98%, 99%, or 100% nucleic acid sequence identity. In some embodiments, the polynucleotide encoding the light chain variable region is a polynucleotide selected from the group consisting of SEQ ID NOs: 37, 6 9, 101, 133, 165, 197, 229, 261, and 293 and has at least 85%, 89%, 90%, 91%, 92%, 9 3%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% nucleic acid sequence identity. The polynucleotides of the present disclosure can encode only the variable region sequences of anti-BK or JC virus antibodies. They can also encode both the variable and constant regions of the antibody .

[0148] ​​Some of the polynucleotide sequences can be used to identify exemplified anti-BK or JC viruses. The polypeptide encodes a polypeptide containing both the heavy and light chain variable regions of a single antibody. Several other nucleotide sequences correspond to the heavy and light chain variable regions of one mouse antibody. , each encoding two substantially identical polypeptide segments.

[0149] The polynucleotide sequences can be synthesized by de novo solid-phase DNA synthesis or by using anti-BK or JC Generated by PCR mutagenesis of existing sequences encoding viral antibodies or their binding fragments. Direct chemical synthesis of nucleic acids can be achieved using the method described by Narang et al. The phosphate diele method (Meth. Enzymol. 68:90, 1979), Brown et al. Stell method (Meth. Enzymol. 68:109, 1979), Beaucage et al. The diethyl phosphoramidite method (Tetra. Lett., 22:1859, 1981 ), and the solid support method of U.S. Pat. No. 4,458,066. This can be achieved by the method of PCR, which involves the introduction of mutations into the polynucleotide sequence. For example, PCR Technology Principles and Applications for DNA Amplification,HA Erlich (Ed.), Freeman Press, NY, NY, 1992, PCR Protocols: A Guide to Methods and Applic ations, Innis et al. (Ed.), Academic Press, San Diego, CA, 1990, Mattila et al., Nucleic Acids Res. 19:967, 1991, and Eckert et al., as described in PCR Methods and Applications 1:17, 1991, can be performed.

[0150] In addition, the present disclosure provides expression vectors and host cells for producing the above-mentioned anti-BK or JC virus antibodies. Using various expression vectors, polynucleotides encoding anti-BK or JC virus antibody chains or binding fragments can be expressed. Both virus-based expression vectors and non-virus expression vectors can be used to produce antibodies in mammalian host cells. Non-viral vectors and systems include plasmids, episomal vectors, typically expression cassettes that express proteins or RNA, and human artificial chromosomes (see, for example, Harrington et al., Nat Genet 15:345, 1997). For example, non-viral vectors useful for the expression of anti-BK or JC virus polynucleotides and polypeptides in mammalian (e.g., human to) cells include pThioHis A, B & C, pcDNA3. 1 / His, pEBVHis A, B & C (Invitrogen, San Diego , CA), MPSV vectors, and numerous other vectors known in the art for expressing other proteins, including. Useful viral vectors include retrovirus-based vectors, adenoviruses, adeno-associated viruses, herpesviruses, SV4 0-based vectors, papillomaviruses, HBP Epstein-Barr virus, vaccinia virus vectors, and Semliki Forest virus (SFV). Bre ​​​​ nt et al., supra, Smith, Annu. Rev. Microbiol. 49 :807, 1995, and Rosenfeld et al., Cell 68:14 3, 1992, see also.

[0151] The choice of expression vector depends on the intended host cell in which the vector is to be expressed. Typ ically, an expression vector includes a promoter and other regulatory sequences (e.g., an enhancer) oper ably linked to a polynucleotide encoding an anti-BK or JC virus antibody chain or fragment. In some as pects, an inducible promoter is used to prevent expression of the inserted sequence under inducing cond itions. Examples of inducible promoters include, for example, arabinose, lacZ, metallothionein promo ter or heat shock promoter. Cultures of transformed organisms can be grown under non-inducing conditi ons without biasing the population of codon sequences for which the expression product is better tolera ted by the host cell. In addition to the promoter, other regulatory elements may also be necessary or desirable for the effi cient expression of an anti-VP1 antibody chain or fragment. These elements typically include an ATG st art codon and an adjacent ribosome binding site or other sequences. Furthermore, the efficiency of ex pression can be enhanced by including an enhancer appropriate for the cell line in use (e.g., Scharf et al., Results Probl. Cell Differ. 20:125 , 1994; and Bittner et al., Meth Enzymol., 15 3:516, 1987, see also). For example, the SV40 enhancer or CMV enhancer can be used to increase ex pression in mammalian host cells.

[0152] The expression vector also provides a secretion signal sequence position to form a fusion protein with the polypeptide encoded by the inserted anti-BK antibody sequence. In more cases, the inserted anti-BK antibody sequence is ligated to the signal sequence before being incorporated into the vector. The vectors used to receive the sequences encoding the anti-BK antibody light and heavy chain variable domains may also encode a constant region or a portion thereof. Such vectors enable the expression of the variable region as a fusion protein with a constant region, thereby resulting in the production of intact antibodies or fragments thereof. Typically, such constant regions are human.

[0153] Host cells expressing and carrying anti-BK or JC antibody chains may be either prokaryotic or eukaryotic cells. E. coli is a useful prokaryotic host for cloning and expressing the polynucleotides of the present disclosure. Other microbial hosts suitable for use include Bacilli such as Bacillus subtilis, Salmonella, Serrati a, and other Enterobacteriaceae such as Pseudomonas. In these prokaryotic hosts, expression vectors can also be prepared, typically containing expression control sequences (e.g., origin of replication) compatible with the host cell. Furthermore, any number of known promoters such as the lactose promoter system, the tryptophan (trp) promoter system, the β-lactamase promoter system, or the promoter system from phage λ exist. The promoter typically optionally controls expression with an operator sequence and initiates transcription and translation. ​​​​​​​It has ribosome binding site sequences for starting and ending, etc. Other microorganisms such as yeast can also be used to express the anti-VP1 polypeptide. Insect cells combined with baculovirus vectors can also be used.

[0154] In other embodiments, mammalian host cells are used to express and produce the anti-VP1 polypeptide of the present disclosure. For example, they can be either hybridoma cell lines that express endogenous immunoglobulin genes (e.g., the myeloma hybridoma clones described in the examples) or mammalian cell lines having an exogenous expression vector. These include any normal cell or normal or abnormal immortalized animal or human cell. For example, many suitable host cell lines capable of secreting intact immunoglobulins have been developed and include CHO cell lines, various COS cell lines, HeLa cells, myeloma cell lines, transformed B-cells and hybridomas. The use of mammalian tissue cell culture to express polypeptides is generally described, for example, in Winnacker, From Genes t o Clones, VCH Publishers, N.Y., N.Y., 1987 The expression vectors for mammalian host cells can include an origin of replication, a promoter, and expression control sequences such as enhancers (see, for example, Queen et al., Immunol. Rev. 89:49-68, 1986), as well as necessary processing information sites such as ribosome binding sites, RNA splice sites, polyadenylation sites, and transcription terminator sequences. These expression vectors usually contain mammalian genes For example, Winnacker, From Genes t o Clones, VCH Publishers, N.Y., N.Y., 1987 described. The expression vectors for mammalian host cells can include an origin of replication, a promoter, and expression control sequences such as enhancers (see, for example, Queen et al., Immunol. Rev. 89:49-68, 1986), as well as necessary processing information sites such as ribosome binding sites, RNA splice sites, polyadenylation sites, and transcription terminator sequences. These expression vectors usually contain mammalian genes processing information sites such as ribosome binding sites, RNA splice sites, polyadenylation sites, and transcription terminator sequences. These expression vectors usually contain mammalian genes processing information sites such as ribosome binding sites, RNA splice sites, polyadenylation sites, and transcription terminator sequences. These expression vectors usually contain mammalian genes It includes a promoter derived from a gene or a promoter derived from a mammalian virus. A suitable pro moter may be constitutive, cell-type specific, stage-specific, and / or regulatable or controllable. Useful promoters include, but are not limited to, the metallothione in promoter, the constitutive adenovirus major late promoter, the dexamethasone-inducible MMTV promoter, the SV40 promoter, the MRP polIII promoter, the constitutive MPSV promoter, the tetracycline-inducible CMV promoter (such as the human immediate-early C MV promoter, etc.), the constitutive CMV promoter, and promoter-enhancer combinations known in the art.

[0155] The method of introducing an expression vector containing the target polynucleotide sequence varies depending on the type of cell host. For example, calcium chloride transfection is generally used for prokaryotic cells, while calcium phosphate treatment or electroporation can be used for other cell hosts (generally, Sambrook et al., supra). Other methods include, for example electroporation, calcium phosphate treatment, liposome-mediated transformation, injection, micro injection, microinjection, ballistics, virosomes, immunoliposomes, polycation nucleic acid conjugate complexes, naked DNA, artificial virions, fusion to the herpes virus structural protein V P22 (Elliot and O’Hare, Cell 88:223, 19 97), drug-enhanced uptake of DNA, and ex vivo transduction. For the long-term, high-yield production of recombinant proteins, stable expression is often desired. For example ​​​, expression containing a virus-origin replication or endogenous expression element and a selectable marker gene Using a vector, a cell line that stably expresses an anti-BK or JC virus antibody chain or binding fragment can be prepared . After introduction of the vector, the cells can be grown in enriched medium for 1-2 days before switching to selective medium . The purpose of the selectable marker is to confer resistance to selection, and its presence allows the growth of cells that express the introduced sequence well in selective medium . Cells stably transfected with resistance can be grown using tissue culture techniques suitable for the cell type

[0156] Therapeutic and diagnostic uses The antibodies, antibody fragments (e.g., antigen-binding fragments) of the present disclosure are useful in a variety of applications including, but not limited to, polyomavirus infections and diseases . In one aspect, the antibodies, antibody fragments (e.g., antigen-binding fragments) are useful for neutralizing BKV or JCV infection and are useful for the prevention or treatment of BK virus nephropathy, e.g., BKVAN . The methods of use can be in vitro, ex vivo, or in vivo methods

[0157] In one aspect, the antibodies, antibody fragments (e.g., antigen-binding fragments) are useful for detecting the presence of BKV in a biological sample . As used herein, the term "detecting" encompasses quantitative or qualitative detection . In one aspect, the biological sample includes cells or tissues . In one aspect, such tissues include normal and / or cancerous tissues that express BKV at higher levels than other tissues

[0158] In one aspect, the present disclosure provides a method for detecting the presence of BK or JC virus in a biological sample​​​​​ provides a method. In one aspect, the method comprises, under conditions that permit binding of an antibody to an antigen, contacting a biological sample with an anti-BK or anti-JC virus antibody and detecting whether a complex is formed between the antibody and the antigen. The biological sample can include, but is not limited to, a urine or blood sample. Also included is a method of diagnosing a disorder associated with the expression of BK or JC virus. In one

[0159] aspect, the method comprises contacting test cells with an anti-BK or anti-JC virus antibody, detecting binding of the antibody to BK or JC virus, thereby determining the level (quantitative or qualitative) of expression of BK or JC virus in the test cells, and comparing the level of infection in the test cells to the level of infection of BK or JC virus in control cells (e.g., normal cells of the same tissue origin as the test cells or non-virus-infected cells). A higher level of presence of BK or JC virus in the test cells compared to the control cells indicates the presence of a disorder associated with infection with BK or JC virus. In one aspect, the test cells are obtained from an individual suspected of having an infection with BK or JC virus. In one aspect, the diagnostic or detection method as described above comprises detecting binding of an anti-BK or anti-JC virus antibody to virus-infected cells. An exemplary assay for detecting binding of an anti-BK or anti-JC virus antibody to BK or JC virus-infected cells is the " FACS" assay. In one aspect, the test cells are obtained from an individual suspected of having an infection with BK or JC virus. In one aspect, the test cells are obtained from an individual suspected of having an infection with BK or JC virus.

[0160] In one aspect, the diagnostic or detection method as described above comprises detecting binding of an anti-BK or anti-JC virus antibody to virus-infected cells. The BK or JC virus-infected cells An exemplary assay for detecting binding of an anti-BK or anti-JC virus antibody to BK or JC virus-infected cells is the " FACS" assay.

[0161] Alternatively, other methods can be used to detect binding of the anti-BK or anti-JC virus antibody. Such methods include, but are not limited to, antigen-binding assays known in the art, for example, Western blot, radioimmunoassay, ELISA (enzyme-linked immunosorbent assay), "sandwich" immunoassay, immunoprecipitation assay, fluorescence immunoassay, protein A immunoassay, and immunohistochemistry (IHC).

[0162] In certain embodiments, the anti-BK or JC virus antibody is labeled. The label can be a label or moiety that is directly detectable (e.g., fluorescence, chromophore, high electron density, chemiluminescence, and radiolabel ), as well as a moiety such as an enzyme or ligand that is indirectly detectable (e.g., via an enzymatic reaction or molecular interaction), but is not limited thereto. )

[0163] In certain embodiments, the anti-BK or JC virus antibody is immobilized on an insoluble matrix. Immobilization involves separating the anti-BK or JC virus antibody from any BKV or JCV protein remaining in solution. This can be accomplished by insolubilizing the anti-BK antibody or JC antibody, such as by adsorption to a water-insoluble matrix or surface (U.S. Patent No. 3,720,760 to Benn ich et al.), by covalent bonding (e.g., using glutaraldehyde cross-linking), or by forming a complex between the anti-BK or JC antibody and the BKV or JCV protein and then insolubilizing the anti-BK or JC antibody (e.g., by immunoprecipitation).

[0164] Diagnosis or detection according to any of the above embodiments can be performed using the anti-BK antibody or JC antibody of the present disclosure, instead of or in addition to, another anti-BK or JC antibody. .

[0165] In one aspect, the disclosure provides a method for administering an antibody, antibody fragment (e.g., an antigen-binding fragment) to a patient. and providing a method for treating, reducing the likelihood of, or ameliorating a disease, thereby In some embodiments, the disease is treated with an antibody, antibody fragment (e.g., an antigen-binding fragment). The disease is infection with the BK virus or JC virus. Examples of BKV and JCV diseases that can be treated include nephropathy, hemorrhagic cystitis, and progressive multifocal leukocytosis. Plasmid encephalopathy (PML), interstitial kidney disease, ureteral stenosis, granule cell neuronopathy (GCN), blood These include ductitis, colitis, retinitis, meningitis, and immune reconstitution inflammatory response syndrome (IRIS). In one embodiment, the infection is caused by the expression of anti-BK antibodies, antibody fragments (e.g., or antigen-binding fragment) or a BKV- or JCV-expressing cell to which the JC antibody can specifically bind. It can be characterized as follows.

[0166] The present disclosure provides a method of treating a BK virus infection, comprising administering to a subject a therapeutically effective amount of BKVAN. In one embodiment, the subject is administered an antibody, antibody fragment (e.g., an antigen-binding fragment) of is a human.

[0167] In one embodiment, the method of reducing BK virus infection comprises administering a therapeutically effective amount of an antibody or antibody In one embodiment, the subject is a human. In one embodiment, the subject is immunosuppressed. Due to the therapeutic effect of the K antibodies, the amount of immunosuppression can be increased or decreased.

[0168] In one embodiment, the transplanted tissue is infected with a BK virus to which the anti-BK antibody binds. Due to the high incidence of BK infection in the general population, in the case of kidney transplants, The patient is BK virus positive, the kidney donor is BK virus positive, or There is a high probability that either or both of the donors are BK virus positive. - or before and / or after the kidney transplant procedure, depending on the seropositivity of the transplant recipient. Anti-BK antibodies can be administered to kidney transplant recipients. is when the virus is found in the urine (viruria) or in the blood. It may be administered to patients when they develop viremia.

[0169] For the treatment of BK or JC viral infection, an antibody or antibody fragment (e.g., an antigen-binding The appropriate dosage of the compound (fragment) will depend on the type of infection being treated, the severity and course of the infection, the response to the infection, and the severity of the infection. The disease depends on a number of factors, including the patient's response to treatment, the development of viral resistance to treatment, previous treatments, and the patient's medical history. Antibodies may be administered once or over a series of treatments lasting from a few days to several months. until a cure is effected or a reduction in infection is achieved (e.g., viral infection of the kidneys) reduction in urinary incontinence or viral disorders) can be administered. , a measure of drug accumulation in the patient's body is calculated from the individual antibodies or antibody fragments (e.g. In some embodiments, the dosage will vary depending on the relative potency of the antibody (e.g., antigen-binding fragment). 0.01mg to 10mg (e.g., 0.01mg, 0.05mg, 0. lmg, 0.5mg, 1mg, 2mg, 3mg, 4mg, 5mg, 7mg, 8mg, 9m g, or 10 mg) administered one or more times daily, weekly, monthly, or yearly. It is possible. In certain embodiments, the antibodies or antibody fragments (e.g., antigen-binding fragments) of the present disclosure are administered once every two weeks or once every three weeks. The treating physician can estimate the dosing frequency for administration and the concentration of the antibody in body fluids or tissues based on the measured half-life. In certain examples, the antibodies or antibody fragments (e.g., antigen-binding fragments) of the present disclosure are combined with other therapeutic agents such as other antiviral agents, anti-allergy agents, anti-nausea agents (or anti-emetics), analgesics, cytoprotective agents, immunosuppressive agents, and combinations thereof. As used herein, the term "pharmaceutical combination" means either a defined combination in a single dosage unit form, or an undefined combination or kit of parts of combined administrations, wherein two or more therapeutic agents may be administered independently simultaneously or separately within a time interval, and in particular, such time intervals are provided such that the combination partners exhibit a synergistic effect (e.g., a synergistic effect).

[0170] Combination Therapy In certain examples, the antibodies or antibody fragments (e.g., antigen-binding fragments) of the present disclosure are combined with other therapeutic agents such as other antiviral agents, anti-allergy agents, anti-nausea agents (or anti-emetics), analgesics, cytoprotective agents, immunosuppressive agents, and combinations thereof. As used herein, the term "pharmaceutical combination" means either a defined combination in a single dosage unit form, or an undefined combination or kit of parts of combined administrations, wherein two or more therapeutic agents may be administered independently simultaneously or separately within a time interval, and in particular, such time intervals are provided such that the combination partners exhibit a synergistic effect (e.g., a synergistic effect). The term "combination therapy" means the administration of two or more therapeutic agents for treating the therapeutic situations or infections described in the present disclosure. Such administration includes the simultaneous administration of these therapeutic agents in a substantially simultaneous manner, such as in a single capsule where the active ingredients have a certain ratio.

[0171] Alternatively, such administration includes the simultaneous administration of each active ingredient in a plurality of, or separate containers (e.g., capsules, powders, and liquids). The powders and / or liquids can be reconstituted or diluted to the desired dosage before administration. Further, such administration also includes the use of each type of therapeutic agent in a sequential manner, almost simultaneously or The term "combination therapy" means the administration of two or more therapeutic agents for treating the therapeutic situations or infections described in the present disclosure. Such administration includes the simultaneous administration of these therapeutic agents in a substantially simultaneous manner, such as in a single capsule where the active ingredients have a certain ratio. Alternatively, such administration includes the simultaneous administration of each active ingredient in a plurality of, or separate containers (e.g., capsules, powders, and liquids). The powders and / or liquids can be reconstituted or diluted to the desired dosage before administration. Further, such administration also includes the use of each type of therapeutic agent in a sequential manner, almost simultaneously or The term "combination therapy" means the administration of two or more therapeutic agents for treating the therapeutic situations or infections described in the present disclosure. Such administration includes the simultaneous administration of these therapeutic agents in a substantially simultaneous manner, such as in a single capsule where the active ingredients have a certain ratio. Alternatively, such administration includes the simultaneous administration of each active ingredient in a plurality of, or separate containers (e.g., capsules, powders, and liquids). The powders and / or liquids can be reconstituted or diluted to the desired dosage before administration. Further, such administration also includes the use of each type of therapeutic agent in a sequential manner, almost simultaneously or

[0172] The term "combination therapy" means the administration of two or more therapeutic agents for treating the therapeutic situations or infections described in the present disclosure. Such administration includes the simultaneous administration of these therapeutic agents in a substantially simultaneous manner, such as in a single capsule where the active ingredients have a certain ratio. Alternatively, such administration includes the simultaneous administration of each active ingredient in a plurality of, or separate containers (e.g., capsules, powders, and liquids). The powders and / or liquids can be reconstituted or diluted to the desired dosage before administration. Further, such administration also includes the use of each type of therapeutic agent in a sequential manner, almost simultaneously or The term "combination therapy" means the administration of two or more therapeutic agents for treating the therapeutic situations or infections described in the present disclosure. Such administration includes the simultaneous administration of these therapeutic agents in a substantially simultaneous manner, such as in a single capsule where the active ingredients have a certain ratio. Alternatively, such administration includes the simultaneous administration of each active ingredient in a plurality of, or separate containers (e.g., capsules, powders, and liquids). The powders and / or liquids can be reconstituted or diluted to the desired dosage before administration. Further, such administration also includes the use of each type of therapeutic agent in a sequential manner, almost simultaneously or The term "combination therapy" means the administration of two or more therapeutic agents for treating the therapeutic situations or infections described in the present disclosure. Such administration includes the simultaneous administration of these therapeutic agents in a substantially simultaneous manner, such as in a single capsule where the active ingredients have a certain ratio. Alternatively, such administration includes the simultaneous administration of each active ingredient in a plurality of, or separate containers (e.g., capsules, powders, and liquids). The powders and / or liquids can be reconstituted or diluted to the desired dosage before administration. Further, such administration also includes the use of each type of therapeutic agent in a sequential manner, almost simultaneously or The term "combination therapy" means the administration of two or more therapeutic agents for treating the therapeutic situations or infections described in the present disclosure. Such administration includes the simultaneous administration of these therapeutic agents in a substantially simultaneous manner, such as in a single capsule where the active ingredients have a certain ratio. which includes using at different times. In either case, the treatment plan provides a beneficial effect with a combination of agents in treating the conditions or disorders described herein.

[0173] Combination therapy can provide a "synergistic effect", i.e., when the active ingredients used together produce a greater effect than the sum of the effects obtained by using the compounds separately, it can be seen that the resulting effect is "synergistic". The synergistic effect can be achieved when the active ingredients are (1) co-formulated and administered, or simultaneously delivered in a combined unit dosage formulation, (2) delivered alternately or in parallel as separate formulations, or (3) delivered by other dosing regimens. When delivered by alternating therapy, a synergistic effect can be achieved when the compounds are sequentially administered or delivered by different injections, for example, by different syringes. Generally, during alternating therapy, the effective dosages of each active ingredient are administered sequentially, i.e., continuously, while in combination therapy, the effective dosages of two or more active ingredients are administered together.

[0174] In one aspect, the present disclosure provides an anti - BK antibody or JC antibody that treats BKV or JCV infection by administering to a subject in need of an antibody in combination with immunosuppressive therapy. The anti - BK antibody or JC antibody acts prophylactically to neutralize BKV or JCV primary infection or viral reactivation resulting from immunosuppressive therapy before or after transplantation. Examples of immunosuppressive therapy include, but are not limited to, dehydrogenase inhibitor, purine synthesis inhibitor, calcineurin inhibitor or mTOR inhibitor. Specific examples of immunosuppressive therapeutic agents include mycophenolate mofetil (MMF), mycophenolate sodium, azathioprine ​ Examples include, but are not limited to, tacrolimus, sirolimus, and cyclosporine.

[0175] Pharmaceutical composition To prepare a pharmaceutical or sterile composition comprising an anti-BK antibody or an anti-JC antibody, the antibodies of the present disclosure are mixed with a pharmaceutically acceptable carrier or excipient. The composition may further comprise one or more other therapeutic agents suitable for neutralizing BKV or JCV infection.

[0176] Formulations of therapeutic agents and diagnostic agents are prepared by mixing a physiologically acceptable carrier, excipient, or stabilizer in the form of, for example, a lyophilized powder, slurry, aqueous solution, lotion, or suspension (e.g., Hardman et al., Good man and Gilman’s The Pharmacological Bas is of Therapeutics, McGraw-Hill, New York, N.Y., 2001, Gennaro, Remington: The Science and Practice of Pharmacy, Lippincott, Wil liams, and Wilkins, New York, N.Y., 2000; Avi s, et al. (eds.), Pharmaceutical Dosage For ms: Parenteral Medications, Marcel Dekker, NY, 1993, Lieberman, et al. (eds.), Pharmaceu tical Dosage Forms: Tablets, Marcel Dekker NY, 1990, Lieberman, et al. (eds.) Pharmaceu , NY, 1990, Lieberman, et al. (eds.) Pharmaceu ​tical Dosage Forms:Disperse Systems,Marc el Dekker, NY, 1990, Weiner and Kotkoskie, E xcipient Toxicity and Safety,Marcel Dekk (See, er, Inc., New York, NY, 2000).

[0177] In certain embodiments, the anti-BK or JC antibody is lyophilized in a vial containing the antibody. The lyophilized product can be reconstituted with water or a pharmaceutical carrier suitable for injection. For subsequent intravenous administration, the resulting solution is usually further diluted in a carrier solution. .

[0178] The antibodies disclosed herein are useful for preventing and treating BKV or JC in immunosuppressed tissue transplant patients. V neutralization and in bone marrow transplant patients receiving CytoGam® The previously used pharmaceutical carriers of sucrose and human albumin can be used. (DeRienzo et al.Pharmacotherapy 2000;20 Alternatively, anti-BK or JC antibodies may be associated with other antiviral antibodies (International As described in patent application WO2003 / 105894 under the trademark Synagis® The drug may be introduced into the transplant patient via a pharmaceutical carrier, such as those described in the publication. The body is made up of histidine and / or glycine, sugars (e.g., sucrose) and polyols ( It is composed of polysorbates (e.g., polysorbates).

[0179] Choosing a treatment regimen depends on the severity of the infection, the level of symptoms, and the biology of the disease. depends on several factors, including the accessibility of the target cells in the matrix. In one aspect the dosing schedule maximizes the therapeutic amount delivered to the patient that is consistent with an acceptable level of side effects Accordingly, the amount of biological delivery administered depends in part on the particular entity and the severity of the condition being treated Guidance is available for selecting appropriate dosages of antibodies, cytokines, and small molecules (see, for example, Wawrzynczak, Antibody Therapy, Bios Scientific Pub. Ltd, Oxfordshire, UK, 1996, Kresina (ed.), Monoclonal Antibodies, Cytokines and Arthritis, Marcel Dekker, New York, N.Y., 1991, Bach (ed.), Monoclonal Antibodies and Peptide Therapy in Autoimmune Diseases, Marcel Dekker, New York, N.Y., 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. 344:783-792, 2001, Beniaminovitz et al., New Engl. J. Med. 342:613-619, 2000, Ghosh et al., New Engl. J. Med. 348:24-32, 2003, Lipsky et al., New Engl. J. Med. 343:1594-1602, 2000). ​​​​​​​​​​​​​​​

[0180] Determination of appropriate dosages is a matter for the clinician, and should be understood by those skilled in the art to affect, for example, treatment. parameters known or suspected to affect treatment, or that are expected to affect Generally, doses are started at somewhat less than the optimal dose. and then increasing in small increments until the desired or optimal effect is achieved relative to any negative side effects. Important diagnostic measures include, for example, symptoms of infusion reactions.

[0181] The actual dosage level of the active ingredient in the pharmaceutical composition having an anti-BK antibody will depend on the particular patient, group, or tissue. The amount of active ingredient that is effective to achieve the desired therapeutic response, given the composition, and the mode of administration. can be varied to obtain a dose that is consistent with the intended outcome without causing toxicity to the patient. The levels depend on a variety of pharmacokinetic factors, including the neutralizing activity of the antibody, the route and time of administration, and the patient's The half-life of the antibody in the setting, the duration of treatment, the duration of treatment, and the specific composition and combination used. Other drugs, compounds and / or substances used in combination, the age and sex of the patient being treated These include weight, condition, general health and previous medical history, and similar factors known in the medical field. do.

[0182] The composition comprising the antibody or fragment thereof can be administered by continuous infusion or, for example, once a day. Doses may be given intravenously for one week or at intervals of one to seven times per week. It can be delivered subcutaneously, topically, orally, nasally, rectally, intramuscularly, intracerebrally, or by inhalation. Specific dosing protocols may be used to determine the maximum or lowest dose that avoids significant undesirable side effects. This includes frequency of administration.

[0183] For the antibodies described in this specification, the dosage administered to a patient may be from 0.000 1 mg / kg to 100 mg / kg. The dosage may be, per patient weight, 0. 0001 mg / kg to 20 mg / kg, 0.0001 mg / kg to 10 mg / kg, 0. 0001 mg / kg to 5 mg / kg, 0.0001 to 2 mg / kg, 0.0001 to 1 mg / kg, 0.0001 mg / kg to 0.75 mg / kg, 0.0001 mg / kg to 0.5 mg / kg, 0.0001 mg / kg to 0.25 mg / kg, 0.0001 to 0. 15 mg / kg, 0.0001 to 0.10 mg / kg, 0.001 to 0.5 mg / kg, 0.01 to 0.25 mg / kg, or between 0.01 and 0.10 mg / kg may be. The dosage of the antibody or its fragment can be calculated using the patient's weight (in kilograms kg) multiplied by the dosage (mg / kg) administered.

[0184] Subsequently, the dosage of the antibody can be repeated, and the administration can be at least once a day, every 2 days, 3 days, 5 days, 10 days, 15 days, 30 days, 45 days, 2 months, 75 days, 3 months, or at least every 6 months.

[0185] The effective amount for a particular patient can vary depending on factors such as the condition being treated, the patient's overall health, the method, route and dosage of administration and the severity of side effects (for example, Mayna rd et al., A Handbook of SOPs for Good Cl inical Practice, Interpharm Press, Boca Ra ton, Fla., 1996, Dent, Good Laboratory and G ood Clinical Practice, Urch Publ., London, See UK, 2001).

[0186] Routes of administration include, for example, topical or transdermal application, intravenous, intraperitoneal, intracerebral, intramuscular, intraocular, intraarterial, intrathecal, intralesional injection or infusion, or via a sustained release system or implant (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, Huang et al., Proc. Natl. Acad. Sci. USA 77:4030-4034, 1980, U.S. Pat. Nos. 6,350,466 and 6,316,024). Optionally, the composition may contain a solubilizing agent or a local anesthetic (e.g., lidocaine to relieve pain at the injection site), or both. Additionally, pulmonary administration can be utilized using an inhaler, nebulizer, etc., and by using the composition with an aerosolizing agent. For example, U.S. Pat. Nos. 6,019,968, 5,985,320, 5,985,309, 5,934,272, 5,874,064, 5,855,913, 5,290,540, and 4,880,078, as well as International Patent Publications WO92 / 19244, WO97 / 32572, WO97 / 44013, WO98 / 31346, and etc. See, for example, 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, Hw ang et al., Proc. Natl. Acad. Sci. USA 77:403 0-4034, 1980, U.S. Pat. Nos. 6,350,466 and 6, 316,024). Optionally, the composition may contain a solubilizing agent or a local anesthetic (e.g., lidocaine to relieve pain at the injection site), or both. Additionally, pulmonary administration can be utilized using an inhaler, nebulizer, etc., and by using the composition with an aerosolizing agent. For example, U.S. Pat. Nos. 6,019,968, 5, 985,320, 5,985,309, 5,934,272, 5,8 74,064, 5,855,913, 5,290,540, and 4 880,078, as well as International Patent Publications WO92 / 19244, WO97 / 32572, WO97 / 44013, WO98 / 31346, and ,880,078, and International Patent Publications WO92 / 19244, WO97 / 32572, WO97 / 44013, WO98 / 31346, and WO99 / 66903, which is incorporated herein by reference in its entirety for each. is incorporated herein by reference.

[0187] In addition, the compositions of the present disclosure can be administered via one or more routes of administration using various methods known in the art. As will be appreciated by those skilled in the art, the route and / or mode of administration will vary depending on the desired result. Routes of administration selected for antibodies include intravenous, intramuscular, intradermal, intraperitoneal, subcutaneous, intraspinal or other parenteral routes, such as administration by injection or infusion. Parenteral administration may represent a mode of administration other than enteral and topical administration and is usually by injection, but not limited to, intravenous, intramuscular, intraarterial, intrathecal, intra-articular, intra-orbital, intracardiac, intradermal, intraperitoneal, transtracheal, subepidermal, subclavian, intra-articular, subcapsular, intrathecal, intraspinal, epidural, and intrasternal injection and infusion. Alternatively, the compositions of the present disclosure can be administered via parenteral routes, such as topical, epithelial, or mucosal routes, such as intranasal, oral, vaginal, rectal, sublingual, or locally. In one aspect, the antibodies of the present disclosure are administered by infusion. In another aspect, the antibody is administered subcutaneously. The route and / or mode of administration will vary depending on the desired result. Routes of administration selected for antibodies include intravenous, intramuscular, intradermal, intraperitoneal, subcutaneous, intraspinal or other parenteral routes, such as administration by injection or infusion. Parenteral administration may represent a mode of administration other than enteral and topical administration and is usually by injection, but not limited to, intravenous, intramuscular, intraarterial, intrathecal, intra-articular, intra-orbital, intracardiac, intradermal, intraperitoneal, transtracheal, subepidermal, subclavian, intra-articular, subcapsular, intrathecal, intraspinal, epidural, and intrasternal injection and infusion. Alternatively, the compositions of the present disclosure can be administered via parenteral routes, such as topical, epithelial, or mucosal routes, such as intranasal, oral, vaginal, rectal, sublingual, or locally. In one aspect, the antibodies of the present disclosure are administered by infusion. In another aspect, the antibody is administered subcutaneously. When the antibodies of the present disclosure are administered in a controlled release or sustained release system, pumps can be used to achieve controlled or sustained release (see Langer, supra, Sefton , CRC Ref Biomed.Eng. 14:20, 1987, Buchwald et al., Surgery 88:507, 1980, Saudek et al. , N.Engl.J.Med. 321:574, 1989). Polymer materials can be used.

[0188] When the antibodies of the present disclosure are administered in a controlled release or sustained release system, pumps can be used to achieve controlled or sustained release (see Langer, supra, Sefton , CRC Ref Biomed.Eng. 14:20, 1987, Buchwald , Surgery 88:507, 1980, Saudek et al. , N.Engl.J.Med. 321:574, 1989). Polymer materials can be used. can be used to achieve controlled or sustained release of antibody therapy (e.g., , Medical Applications of Controlled Rele ase, Langer and Wise (eds.), CRC Pres., Boca Raton, Fla., 1974, Controlled Drug Bioavai lability, Drug Product Design and Perform ance, Smolen and Ball (eds.), Wiley, New Yor k, 1984, Ranger and Peppas, J. Macromol. Sci. Rev. Macromol. Chem. 23:61, 1983, and Levy et al., Science 228:190, 1985, During et al., A nn. Neurol. 25:351, 1989, Howard et al., J. Ne urosurg. 71:105, 1989, U.S. Patent Nos. 5,679,377, 5 ,916,597, 5,912,015, 5,989,463, 5, 128,326, International Patent Publication Nos. WO99 / 15154, and WO99 / 20 253). Examples of polymers used in sustained release formulations include poly(2- hydroxyethyl methacrylate), poly(methyl methacrylate), poly(acrylic acid ), poly(ethylene-co-vinyl acetate), poly(methacrylic acid), polyglycolide (PL G), polyanhydrides, poly(N-vinylpyrrolidone), poly(vinyl alcohol), poly acrylamide, poly(ethylene glycol), polylactic acid (PLA), poly(lactide- co-glycolide) (PLGA), and polyorthoesters, but are not limited to these. It is not determined. In one aspect, the polymer used in the sustained release formulation is inert, leachable free of impurities, stable in storage, sterile, and biodegradable. A controlled or sustained release system can be placed in proximity to the prophylactic or therapeutic target and thus requires only a fraction of the systemic dose (see, e.g., Goodson, in Medical Applications of Controlled Release, supra, vol. 2, pp. 115-138, 1984).

[0189] Controlled release systems are described in Langer, Science 249:1527-1533, (1990). Sustained release formulations containing one or more antibodies of the present disclosure can be manufactured using any technique known to those skilled in the art. For example, U.S. Patent No. 4,526, 938, International Patent Application Nos. WO91 / 05548, WO96 / 20698, Ni ng et al., Radiotherapy & Oncology 39:179 -189, 1996, Song et al., PDA Journal of Pharmaceutical Science & Technology 50:372- 397, 1995, Cleek et al., Pro. 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, each of which is hereby incorporated by reference in its entirety.

[0190] When the antibodies of the present disclosure are administered locally, they can be formulated in the form of ointments, creams, transdermal patches, lo tions, gels, sprays, aerosols, solutions, emulsions, or other forms well known to those skilled in the art. For example, see Remington’s Pharma ceutical Sciences and Introduction to Ph armaceutical Dosage Forms, 19th ed., Mack Pub.Co., Easton, Pa. (1995). For non-sprayable topical administration forms, semi-solid or solid forms that contain a carrier or one or more excipients compatible with topical application and have a viscous consistency and, in some cases, a dynamic viscosity greater than that of water are typically used. Suitable formulations include solutions, suspensions, emulsions, creams, ointments, powders, liniments, plasters, etc., and may include, but are not limited to, mixing with sterilizing or adjuvants (e.g., preservatives, stabilizers, wetting agents, buffers, or salts) that affect various properties such as osmotic pressure, as required. Other suitable topical administration forms include sprayable aerosol preparations, which may, in some cases, combine the active ingredient with a solid or liquid inert carrier and package the mixture in a compressed volatile (e.g., a gaseous propellant such as Freon) bottle or squeeze bottle. Moisturizers or humectants can also be added to the pharmaceutical compositions and dosage forms as needed. Examples of such additional ingredients are well known in the art. When compositions containing antibodies are administered intranasally, they can be formulated in aerosol form, sprays, mists, or droplets. In particular, prophylactic or therapeutic agents for use according to the present disclosure are

[0191] ​​​​​​​​​​​Using a suitable propellant (e.g., dichlorofluoromethane, trichlorofluoromethane, dichloro tetrafluoroethane, carbon dioxide, or other suitable gas), it can be conveniently delivered in the form of an aerosol spray presentation from a pressurized package or nebulizer. In the case of a pressurized aerosol, the dosage unit can be determined by providing a valve that supplies a measured amount. Capsules and cartridges for use in inhalers or injectors (e.g., made of gelatin) can be formulated to contain a powder mixture of the compound and a suitable powder base such as lactose or dextrin. Methods for the co-administration or treatment with a second therapeutic agent, e.g., an immunosuppressant, cytokine, steroid, chemotherapeutic agent, antibiotic

[0192] or radiation, are known in the art (e.g., Hardman et al., (eds.)(2001) Goodman and Gilman’s The Pharmacological Basis o f Therapeutics, 10th ed., McGraw-Hill, New York, N.Y, Poole and Peterson (eds.)(2001) P harmacotherapeutics for Advanced Practic e: A Practical Approach, Lippincott, Willia ms & Wilkins, Phila., Pa, Chabner and Longo (eds.)(2001) Cancer Chemotherapy and Biot (eds.)(2001) Cancer Chemotherapy and Biot herapy, Lippincott, Williams & Wilkins, Phi See la., Pa. The effective amount of the therapeutic agent can reduce the symptoms by at least 10%, at least 20%, at least about 30%, at least 40%, or at least 50%.

[0193] Additional therapies (e.g., prophylactic or therapeutic agents) that can be administered in combination with the anti-BK antibody are from the anti-VP1 antibodies of the present disclosure, at intervals of less than 5 minutes, less than 30 minutes, 1 hour intervals, about 1 hour intervals, about 1 hour to about 2 hour intervals, about 2 hours to about 3 hour intervals, about 3 hours to about 4 hour intervals, about 4 hours to about 5 hour intervals, about 5 hours to about 6 hour intervals, about 6 hours to about 7 hour intervals, about 7 hours to about 8 hour intervals, about 8 hours to about 9 hour intervals, about 9 hours to about 10 hour intervals, about 10 hours to about 11 hour intervals, about 11 hours to about 12 hour intervals, about 12 hours to about 18 hour intervals, 18 hours to 24 hour intervals, 24 hours to 36 hour intervals, 36 hours to 48 hour intervals, 48 hours to 52 hour intervals , 52 hours to 60 hour intervals, 60 hours to 72 hour intervals, 72 hours to 84 hour intervals, 84 hours to 96 hour intervals, 96 hours to 120 hour intervals, and may be administered. Two or more therapies may be administered during the same patient's hospital stay.

[0194] In certain embodiments, the anti-BK antibodies can be formulated to ensure proper distribution in vivo. For example, the blood-brain barrier (BBB) excludes many highly hydrophilic compounds. To ensure that the anti-BK antibodies cross the BBB (if necessary), they can be formulated, for example, with liposomes. For methods of manufacturing liposomes, see, for example, U.S. Pat. Nos. 4,522,811, 5,374,548, and 5,399,331 for reference. These liposomes are selectively transported to specific cells or organs, and thus ​can include one or more moieties that enhance targeted drug delivery ( see, for example, Ranade, (1989) J.Clin.Pharmacol.29:685 ). Exemplary targeting moieties include folic acid or biotin (see, for example, U.S. Patent No. 5,416,016 to Low et al.), mannoside (Ume zawa et al., (1988) Biochem.Biophys.Res.Co mmun.153:1038), antibody (Bloeman et al., (1995) F EBS Lett.357:140, Owais et al., (1995) Anti microb.Agents Chemother.39:180), surfactant protein A receptor (Briscoe et al., (1995) Am.J.Phys iol.1233:134); p120 (Schreier et al,(1994 ) J.Biol.Chem.269:9090), also, K.Keinanen; M.L .Laukkanen(1994) FEBS Lett.346:123, J.J.Ki llion; I.J.Fidler(1994) Immunomethods 4:27 3. See also.

[0195] The present disclosure provides a protocol for administering to a patient in need thereof a pharmaceutical composition comprising only an antibody or a combination with other therapies. Combination therapies (e.g., prophylactic or therapeutic agents) can be administered to a subject simultaneously or sequentially. Combination therapies (e.g., prophylactic or therapeutic agents ) can also be administered periodically. Cycling therapy involves administering a first therapy (e.g., a first prophylactic or therapeutic agent) for a period of time, followed by a second therapy (e.g., a second pro phylactic or therapeutic agent) for a period of time, followed by a second therapy (e.g., a second pro Administration for a period of a prophylactic or therapeutic agent), and this sequential administration, i.e., repeating the cycle including avoiding or reducing one side effect of a treatment (e.g., an agent) and reducing the development of resistance to one of the treatments (e.g., an agent), and / or improving the effectiveness of the treatment to do.

[0196] The treatments (e.g., prophylactic or therapeutic agents) of the combination therapies of the present disclosure can be administered to a subject simultaneously The term "simultaneously" is not limited to the administration of a treatment (e.g., prophylactic or therapeutic agent) exactly simultaneously Rather, the pharmaceutical composition containing the antibody or a fragment thereof means that the antibody can act with other treatments such that the pharmaceutical composition provides an increased benefit over the way they are administered otherwise and is administered to the subject sequentially at time intervals. For example, each treatment may be administered to the subject simultaneously or sequentially in any order at different times, but if not administered simultaneously they must be administered at a time close enough to provide the desired therapeutic or prophylactic effect Each treatment can be administered to the subject separately in any suitable form and by any suitable route In various embodiments, the treatment (e.g., prophylactic or therapeutic agent) is administered to the subject at intervals of less than 15 minutes, less than 30 minutes, less than 1 hour, about 1 hour interval, about 1 hour to about 2 hour interval about 2 hours to about 3 hours interval, about 3 hours to about 4 hours interval, about 4 hours to about 5 hours interval, about 5 hours to about 6 hours interval, about 6 hours to about 7 hours interval, about 7 hours to about 8 hours interval, about 8 hours to about 9 hours interval, about 9 hours to about 10 hours interval, about 10 hours to about 11 hours interval, about 11 hours to about 1 2 hours interval, 24 hours interval, 48 hours interval, 72 hours interval, or 1 week interval In other embodiments, two or more treatments (e.g., prophylactic or therapeutic agents) are administered to the same patient ​​​​It is administered during hospitalization.

[0197] The prophylactic or therapeutic agent for combination therapy can be administered to the subject for the same pharmaceutical composition. Or the prophylactic or therapeutic agent for combination therapy can be administered simultaneously to the subject for a separate pharmaceutical composition. The prophylactic or therapeutic agent can be administered to the subject by the same or different routes of administration.

Examples

[0198] Example 1: Generation of anti-BK or JC virus antibodies B cells expressing anti-BKV and / or anti-JCV antibodies were lysed, and the VH (heavy) and V L (light) chains were amplified by RT-PCR and subsequently sequenced and analyzed to identify important post-translational modification (PTM) sites. Then, plasmids of the VH chain and VL chain were transfected into a CHO mammalian cell line as an IgG1 backbone vector for the expression of full IgG1 antibodies.

[0199] Example 2: Conjugate of anti-BKV antibody to VLP (ELISA) The binding of the antibody to VLP was analyzed by ELISA. Briefly, Nunc MaxiSo rp 384-well plates (Thermo scientific) were coated overnight with BKV VLP (100 ng / well) against BK serotype I (ST1) or serotype IV (ST4). The antibody was serially diluted in PBS containing 0.5% BSA and bound to the antigen coated plate for 2 hours. The plates were washed with PBS and then incubated for 1 hour with a secondary antibody (HRP-conjugated anti-human IgG goat antibody, Southern Biotech #2040-01) diluted 1:6000 in PBS containing 0.5% BSA. It was washed. The plate was washed with PBS, and the reaction was developed using tetramethylbenzidine (TMB) microwell peroxidase substrate (SeramunBlau Fast, Seramun, Germany). The results of ELISA binding are shown in Figure 1. For example, antibody NOV530 binds to both BKV ST1 and BKV ST4. Antibody NOV638 binds only to BKV ST1. peroxidase substrate (SeramunBlau Fast, Seramun, Germa ny). The results of ELISA binding are shown in Figure 1. For example, the antibody NOV530 binds to both BKV ST1 and BKV ST4. Antibody NO V638 binds only to BKV ST1.

[0200] Example 3: Neutralization of virus infection by anti-BKV antibodies Infectious BKV serotype I (ST1) and chimeric virus expressed serotype II (ST2), III (ST3), and IV (ST4) were pre-incubated with the purified antibody for 1 hour and subjected to binding and neutralization. Then, primary renal proximal tubular epithelial (RPTE) cells (ATCC, catalog number PCS-400-010) were exposed to the virus-antibody mixture for 4 hours, replaced with fresh medium, and incubated for 48 hours for virus entry and gene expression. The cells were fixed with 4% paraformaldehyde and analyzed by immunofluorescence to detect TAg expression (Calbiochem DP02, pAb416 mouse anti-SV40 TAg antibody ). Immunofluorescence was analyzed by high-content image analysis using Cellomics ArrayScan® VTI HCS Reader, and the percentage of BKV-infected cells (TAg -positive, DAPI-positive) was quantified, and the data were presented as the percentage inhibition of infection relative to untreated control wells. The data are presented as the concentration of antibody at which virus infection is 50% neutralized relative to untreated control wells, the EC50. )). Immunofluorescence was analyzed by high-content image analysis using Cellomics ArrayScan® VTI HCS Reader, and the percentage of BKV-infected cells (TAg -positive, DAPI-positive) was quantified, and the data were presented as the percentage inhibition of infection relative to untreated control wells. The data are presented as the concentration of antibody at which virus infection is 50% neutralized relative to untreated control wells, the EC50. -positive, DAPI-positive) was quantified, and the data were presented as the percentage inhibition of infection relative to untreated control wells. The data are presented as the concentration of antibody at which virus infection is 50% neutralized relative to untreated control wells, the EC50. The data are presented as the percentage inhibition of infection relative to untreated control wells. The data are presented as the concentration of antibody at which virus infection is 50% neutralized relative to untreated control wells, the EC50. The data are presented as the concentration of antibody at which virus infection is 50% neutralized relative to untreated control wells, the EC50.

[0201] Physiologically, antibodies perform several functions that help inhibit progressive pathogenic responses and one of these is to directly block the ability of the virus to bind to and / or translocate to target cells These “neutralizing” antibodies typically represent only a subset of antigen-binding Igs Most of the monoclonal IgG anti-BKV antibodies disclosed herein can neutralize at least BKV ST1 in a primary kidney cell infection assay, while some can neutralize additional BKV subtypes and / or related JC virus (Figure 1). For example, antibody NOV638 can bind to and neutralize BKV ST1 while antibody NOV530 can bind to and neutralize all four serotypes of BK virus and also showed sub-nM EC50 for JCV (Figure 1).

[0202] Example 4: Generation of BK virus and virus-like particles (VLPs) Genomic clones of BKV ST1 were obtained from ATCC (pBR322-BKV MM, catalog number 45026, pBR322-BKV Dunlop, catalog number 45025) Infectious genomic clones of chimeric viruses of ST2, ST3, and ST4 were generated using the cloning strategy described previously (Broekema et al, Virology 2010 407:368-373). Briefly, unique restriction sites (SacII, P mlI) were introduced into the BKV serotype I genome adjacent to the VP1-VP2-VP3 coding region using site-directed mutagenesis ST2 isolate SB (GenBank accession number CA A79596.1), serotype III isolate AS (GenBank accession number AAA46882 ), serotype IV isolate AA (GenBank accession number AAA46883 .1) and from ST4 isolated ITA-4 (GenBank accession number BAF75132) The coding region of VP1 was synthesized in the context of the VP2 / VP3 coding region from ST1 isolate (Genewiz, La Jolla, CA) such that the synthetic fragment containing the SacII-PmlI region could be used in the swap combinations described by Broekema et al. (supra) Next, using the resulting chimeric genomic clone, as previously described (Abend et al, J. Virology 2007 81:272-279), high-titer infectious virus stocks were generated in primary renal proximal tubule epithelial (RPTE) cells (ATCC, catalog number PCS-400-010) (Abend et al, J. Virology 2007 81:272-279). VLPs representing each of the four BKV serotypes were generated by expression of VP1 in Sf9 insect cells

[0203] and extracted from frozen cell pellets from 1 L cultures, microchip sonication (3× 45 second pulses with 5 minute rests on ice between pulses), pelleting of VLPs through a 20% sucrose cushion (116,000g, 2.5 hours), and purification by anion exchange on a 5 ml GE HiTrapQ HP column (GE Healthcare, Pittsburgh, PA) followed by purification on a 10 ml Capto™ Core700 (GE Healthcare, Pittsburgh, PA) resin-based size exclusion column and finally purification on a GE Sephacryl S500 26 / 60 (GE Healthcare, Pittsburgh, PA) size exclusion column The prepared VLPs were used in ELISA and SET-based binding assays​​ done.

[0204] Example 5: Measurement of Affinity of Anti-BK Antibodies (SET Assay) Using solution equilibrium titration (SET) assay, the interaction affinity (K ) of antibodies against BKV VLPs from all four serotypes was determined. The antibody was assayed at a concentration of 1 pM (constant), and the VLP was serially diluted from an initial concentration of 10 nM. The antibody-VLP solution was incubated overnight and then assayed for unbound antibody using an MSD array plate (Meso Scale Discovery catalog number L21XA, Rockville MD) coated with VLP. K d was determined by fitting the plot to a 1:1 fit model (according to Piehler et al. J. Immunol. Methods. 1997;201(2):189-206). The sample curve set used for the affinity determination of anti-BKV monoclonal IgG (clone NOV58 1) against BKV ST1 VLP via SET is shown in Figure 2A. The lower curve is the four-parameter fitting of the K control curve (based on the low concentration of antibody NOV581), while the upper curve is the fitting of the stoichiometry control curve (higher constant antibody concentration for estimating the effective ligand concentration). The signal intensity is normalized to the initial state without BKV VLP in the solution ("100% free antibody"). In Figure 2B, the binding affinity was determined with cross-neutralizing monoclonal anti-BKV IgG antibodies against BKV virus-like particles (VLPs). All antibodies tested were BKV d The curve is the four-parameter fitting of the K control curve (based on the low concentration of antibody NOV581), while the upper curve is the fitting of the stoichiometry control curve (higher constant antibody concentration for estimating the effective ligand concentration). The signal intensity is normalized to the initial state without BKV VLP in the solution ("100% free antibody"). (according to Piehler et al. J. Immunol. Methods. 1997;201(2):189-206).

[0205] The sample curve set used for the affinity determination of anti-BKV monoclonal IgG (clone NOV58 1) against BKV ST1 VLP via SET is shown in Figure 2A. The lower curve is the four-parameter fitting of the K control curve (based on the low concentration of antibody NOV581), while the upper curve is the fitting of the stoichiometry control curve (higher constant antibody concentration for estimating the effective ligand concentration). The signal intensity is normalized to the initial state without BKV VLP in the solution ("100% free antibody"). d control curve (based on the low concentration of antibody NOV581), while the upper curve is the fitting of the stoichiometry control curve (higher constant antibody concentration for estimating the effective ligand concentration). The signal intensity is normalized to the initial state without BKV VLP in the solution ("100% free antibody"). In Figure 2B, the binding affinity was determined with cross-neutralizing monoclonal anti-BKV IgG antibodies against BKV virus-like particles (VLPs). All antibodies tested were BKV for estimating the effective ligand concentration) of the higher constant antibody concentration. The signal intensity is normalized to the initial state without BKV VLP in the solution ("100% free antibody"). VLP-free initial state (''100% free antibody'').

[0206] In Figure 2B, the binding affinity was determined with cross-neutralizing monoclonal anti-BKV IgG antibodies against BKV virus-like particles (VLPs). All antibodies tested were BKV The sample curve set used for the affinity determination of anti-BKV monoclonal IgG (clone NOV58 had a K value of less than 50 pM against ST1. In this assay, antibody NOV581 d had significant affinity for 1, 2, and 3, excluding BKV serotype 4. In contrast, anti- body NOV530 had significant affinity for all four serotypes (Figure 2B).

[0207] Example 6: Cryo-Electron Microscopy To understand the mechanism by which the isolated cross-neutralizing antibodies effectively inhibit infection by multiple polyomavirus strains, the inventors performed cryo-electron microscopy (cryoEM) of BKV ST1 VLP complexed with the single-chain variable fragment (scFv) form of the cross-neutralizing IgG NOV530 and obtained a class-averaged density map at a resolution of 4.24 Å (Figure 3 A). The inventors were able to model the capsid structure of the VLP containing the pentameric subunit linked together via the C-terminus of each individual VP1 monomer. Surprisingly, this quaternary structure forms the basis of a composite viral epitope bound by NOV53 0 having three VP1 subunits contributing amino acid residues (Figure 3B - C). A total of 2 0 viral residues are predicted to be within 5 Å of the antibody, and these residues are highly conserved across polyomavirus species, 3 show conserved homology, and the remaining 17 are identical in J CV (Figure 3D - F). The interaction positions from the antibody spread throughout the heavy and light chains and have contributions from both germline-encoded (CDR1 and CDR2) and somatic hypermutation (CDR3) loops (Figure 3G - H). Identifying the complex binding site of NOV530 to the BKV capsid protein is due to its quaternary structure requirements ​​​​​​​which was impossible by other methods. This binding mode raises further interesting questions about the mechanism of virus neutralization by NOV530, for example, whether antibodies can bind capsid sub-uni ts together, thereby preventing the uncoating process after virus translocation. Potential escape mutations can only occur at the expense of reduced virion stability. In fact, mutations in three amino acid residues (E61, R64, and R83) within the NOV530 epitope have been previously reported and probably dramatically reduce virus fitness by affecting receptor binding and capsid structure integrity (Dugan A.S.et al.,Identification of am ino acid residues in BK virus VP1 that a re critical for viability and growth.J V irol 81,11798-11808(2007)).

[0208] Method of CyroEM BKV ST1 VLPs were incubated with the scFv fragment of NOV530 (3 60 molecules of scFv per VLP, total protein concentration 1 mg / ml) for 1 hour at room temperature. The sample was then concentrated 10-fold. 4.4 μL of the concentrated VLP-scFv complex was applied onto a grid (R1.2 / 1.3, Cu300 mesh , Quantifoil Micro Tools GmbH, Grosslobich au, Germany) coated with an additional thin amorphous carbon layer. The grid was vitrified using a Leica EM GP plunger. Images were taken using a Quantum-LS Gatan image filter (G equipped with (IF) and operated at 300 kV, the Cs-corrected FEI Titan Krios TE was used, and recorded on a Gatan K2-Summit direct electron detector (Gatan Gm bH). Images were automatically collected in electron counting mode (nominal post-GIF magnification x105,00 0, calibrated pixel size 1.12 Å) (EPU, Thermo Fisher). An exposure of 7 s was dose-fractionated into 40 frames. The total exposure dose was about 40 e- / Å2, and the defocus value, which was varied from -0.8 to -2.5 μm.

[0209] Cryo-data were imaged using the following protocol. Stage drift and beam-induced motion during exposure were preprocessed, and whole-image drift correction was automated using UNBLUR (Grant, T and Grigorieff N. Measuring the optimal exposure f or single particle cryo-EM using a 2.6 Å reconstruction of rotavirus VP6 (eLife.4 (e06980):1-19(2015)). The images were aligned inline (StackGUI). The contrast transfer function (CTF) parameters were estimated using the program CTFFIND4 (Mindell JA , and Grigorieff N. Accurate determination of local defocus and specimen tilt in e , and Grigorieff N. Accurate determination of local defocus and specimen tilt in electron microscopy. J. Struct. Biol. 142:334 -347(2003)). Particles were automatically picked on each micrograph using GAUTOMATCH ​were picked up. A total of 1,400 microscopic photographs were obtained, and 6,000 particles were extracted for processing using the Relion software package (Scheres, S .H.RELION: implementation of a Bayesian a pproach to cryo-EM structure determinati on. J.Struct.Biol. 180, 519-530, doi:10.1016 / j.jsb.2012.09.006(2012)). Particle selection was by 2D classification without reference images in 2 cycles. 5,000 particles in the best 2D classes were used for 3D refinement. A sphere was used as the initial model for 3D refinement. The inventors performed particle-based beam-induced motion correction and radiation damage weighting (known as particle polishing, see Scheres, S.H., Beam-induced motion co rrection for sub-megadalton cryo-EM part icles. Elife 3, e03665, doi:10.7554 / eLife0. 03665((2014)) on the first 20 frames (corresponding to a total dose of about 20 e / Å rrection for sub-megadalton cryo-EM part icles.Elife 3, e03665, doi:10.7554 / eLife0. 03665((2014)). The resulting 5,000 polished particles yielded a reconstruction with an overall resolution of 4.5 Å. Automatic refinement of the polished particles with a soft mask around the BK-VLP_scFv complex gave a map with a resolution of 4.24 Å. The reported resolution values are based on the gold-standard Fourier shell correlation curve (FSC) with a criterion of 0.143 (Scheres, S.H. e - / Å 2 ). The 5,000 polished particles obtained resulted in a reconstruction with an overall resolution of 4.5 Å. Automatic refinement of the polished particles with a soft mask around the BK-VLP_scFv complex gave a map with a resolution of 4.24 Å. The reported resolution values are based on the gold-standard Fourier shell correlation curve (FSC) with a criterion of 0.143 (Scheres, S.H. ). The 5,000 polished particles obtained led to a reconstruction with an overall resolution of 4.5 Å. Automatic refinement of the polished particles with a soft mask around the BK-VLP_scFv complex yielded a map with a resolution of 4.24 Å. The reported resolution values are based on the gold-standard Fourier shell correlation curve (FSC) with a criterion of 0.143 (Scheres, S.H. ). Automatic refinement of the polished particles with a soft mask around the BK-VLP_scFv complex gave a map with a resolution of 4.24 Å. The reported resolution values are based on the gold-standard Fourier shell correlation curve (FSC) with a criterion of 0.143 (Scheres, S.H. ). The reported resolution values are based on the gold-standard Fourier shell correlation curve (FSC) with a criterion of 0.143 (Scheres, S.H. ). Automatic refinement of the polished particles with a soft mask around the BK-VLP_scFv complex gave a map with a resolution of 4.24 Å. The reported resolution values are based on the gold-standard Fourier shell correlation curve (FSC) with a criterion of 0.143 (Scheres, S.H. RELION: implementation of a Bayesian approach to cryo-EM structure determination. approach to cryo-EM structure determination. J.Struct.Biol.180,519-530,doi:10.1016 / j. jsb.2012.09.006(2012)). The cryoEM structure of BK virion and the crystal structure of scFv (each PDB ID code 5FUA and 4UT7) were manually fitted to the final cryoEM map using the program Coot (Emsley P. et al., Features and development of Coot Acta Crystallogr D Biol Cry stallogr 66:486-501(2010)). The obtained atomic model was rebuilt multiple times using the program Coot (Emsley P. et al., supra), and real-space refinement against the map was performed using the program Phenix (Adams PD, et al. PHENIX: A comprehensive Python-based system for macromolecular structure solution. Acta Crystallogr D Biol Cry stallogr 66:213-221(2010)). This process yielded atomic models of the pentamer and scFv complex that fit well to the cryo EM density. Structure diagrams were prepared using PyMOL (available from Schrodinger P. et al., supra). Multiple rounds of model rebuilding were performed using the program Coot (Emsley P. et al., supra), and real-space refinement against the map was performed using the program Phenix (Adams PD, et al. PHENIX: A comprehensive Python-based system for macromolecular structure solution. Acta Crystallogr D Biol Cry stallogr 66:213-221(2010)). This process yielded atomic models of the pentamer and scFv complex that fit well to the cryo EM density. Structure diagrams were prepared using PyMOL (available from Schrodinger ure solution. Acta Crystallogr D Biol Cry stallogr 66:213-221(2010)). This process yielded atomic models of the pentamer and scFv complex that fit well to the cryo EM density. Structure diagrams were prepared using PyMOL (available from Schrodinger of the pentamer and scFv complex that fit well to the cryo EM density. Structure diagrams were prepared using PyMOL (available from Schrodinger The atomic models of the pentamer and scFv complex that fit well to the cryo EM density were obtained. Structure diagrams were prepared using PyMOL (available from Schrodinger obtainable).

[0210] Example 7: Formulations The anti-BK or JC virus antibodies described herein are monoclonal antibodies, κ or λ It is the IgG1 isotype of the light chain and can be lyophilized. These antibodies are soluble and stable for 4 weeks in a histidine- sucrose formulation buffer. Furthermore, the anti-VP1 antibody is formulated as a minimum drug substance (e.g., in histidine buffer in the absence of a stabilizer) and is soluble at >200 mg / ml.

[0211] For subsequent intravenous administration, the resulting solution is usually further diluted in a carrier solution to obtain an antibody solution ready for injection.

[0212] Important stability indicator assays for selecting the most stable formulation include, among others, size exclusion chromatography for determining the aggregation level, the non-visible particulate matter test, and the titer test.

[0213] The examples and embodiments described in this specification are for illustrative purposes, and various modifications or changes are suggested to those skilled in the art in light of their content, and it is understood that they should be included within the spirit and scope of the appended claims. ​

Claims

1. An isolated antibody or antigen-binding fragment thereof, comprising: (i) a heavy chain region and (ii) An isolated antibody or antigen-binding fragment thereof, comprising a light chain region.

2. 1. An isolated antibody, comprising: (i) (a) HCDR1 (CDR-complementarity determining region) of SEQ ID NO: 9; (b) SEQ ID NO: 10 (c) a HCDR2 of SEQ ID NO: 11, and (d) a heavy chain variable region comprising: (e) LCDR1 of SEQ ID NO:25, (f) LCDR2 of SEQ ID NO:26, and (g) LCDR3 of SEQ ID NO:2 a light chain variable region comprising an LCDR3 of 7; (ii) (a) HCDR1 of SEQ ID NO: 41, (b) HCDR2 of SEQ ID NO: 42, (c) (d) a heavy chain variable region comprising an HCDR3 of sequence number 43, and (e) an LCDR of sequence number 57. 1, (e) an LCDR2 of SEQ ID NO:58, and (f) an LCDR3 of SEQ ID NO:

59. A light chain variable region; (iii) (a) HCDR1 of SEQ ID NO: 73, (b) HCDR2 of SEQ ID NO: 74, (c) (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 75, and (e) an LCDR of SEQ ID NO:

89. R1, (e) an LCDR2 of SEQ ID NO:90, and (f) an LCDR3 of SEQ ID NO:

91. A light chain variable region comprising (iv) (a) HCDR1 of SEQ ID NO: 105, (b) HCDR2 of SEQ ID NO: 106, (d) a heavy chain variable region comprising a HCDR3 of SEQ ID NO: 107, and (e) LCDR1 of SEQ ID NO: 122, and (f) LCDR2 of SEQ ID NO:

123. R3, and a light chain variable region comprising (v) (a) HCDR1 of SEQ ID NO: 137, (b) HCDR2 of SEQ ID NO: 138, (c) (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 139, and (e) LCDR2 of SEQ ID NO: 154, and (f) LCDR of SEQ ID NO:

155. 3, and a light chain variable region comprising (vi) (a) HCDR1 of SEQ ID NO: 169, (b) HCDR2 of SEQ ID NO: 170, (d) a heavy chain variable region comprising a HCDR3 of SEQ ID NO: 171, and (e) LCDR1 of SEQ ID NO: 186, and (f) LCDR2 of SEQ ID NO:

187. R3, and a light chain variable region comprising (vii) (a) HCDR1 of SEQ ID NO: 201, (b) HCDR2 of SEQ ID NO: 202, ( (c) a heavy chain variable region comprising the HCDR3 of SEQ ID NO: 203, and (d) a heavy chain variable region comprising the HCDR3 of SEQ ID NO: 217 (e) LCDR1 of SEQ ID NO: 218, and (f) LCDR2 of SEQ ID NO:

219. DR3, and a light chain variable region comprising (viii) (a) an HCDR1 of SEQ ID NO: 233, (b) an HCDR2 of SEQ ID NO: 234, (c) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 235, and (d) a heavy chain variable region comprising an HCDR3 of SEQ ID NO: 24 (e) LCDR1 of SEQ ID NO:250, and (f) LCDR2 of SEQ ID NO:

251. a light chain variable region comprising (ix) (a) HCDR1 of SEQ ID NO: 265, (b) HCDR2 of SEQ ID NO: 266, (d) a heavy chain variable region comprising a HCDR3 of SEQ ID NO: 267, and (e) LCDR1 of SEQ ID NO: 282, and (f) LCDR2 of SEQ ID NO:

283. R3, and a light chain variable region comprising:

3. 2. One or two amino acids in the CDR are modified, deleted or substituted. The antibody described in

4. At least 90, 91, 100, 150, 200, 250, 300, 350, 400, 450, 500, 600, 700, 800, 900, 1000, 1500, 2000, 3500, 4000, 5000, 6000, 7000, 800 , 92, 93, 94, 95, 96, 97, 98, or 99% identity. The antibody according to item 2.

5. The antibody is a monoclonal antibody, a chimeric antibody, a humanized antibody, a human modified antibody, a human antibody, The antibody of claim 1 which is a single chain antibody (scFv) or an antibody fragment.

6. 1. An isolated antibody or antigen-binding fragment thereof, comprising: (i) a heavy chain variable region (vH) comprising SEQ ID NO: 18, and a light chain variable region comprising SEQ ID NO: 34 Region (vL), (ii) a heavy chain variable region (vH) comprising SEQ ID NO: 50, and a light chain variable region (vL) comprising SEQ ID NO: 66 A region (vL); (iii) a heavy chain variable region (vH) comprising SEQ ID NO: 82, and a light chain variable region (vL) comprising SEQ ID NO:

98. A variable region (vL), (iv) a heavy chain variable region (vH) comprising SEQ ID NO: 114, and a light chain comprising SEQ ID NO:

130. A variable region (vL), (v) a heavy chain variable region (vH) comprising SEQ ID NO: 146, and a light chain variable region (vL) comprising SEQ ID NO:

162. A variable region (vL), (vi) a heavy chain variable region (vH) comprising SEQ ID NO: 178, and a light chain comprising SEQ ID NO:

194. A variable region (vL), (vii) a heavy chain variable region (vH) comprising SEQ ID NO: 210, and a light chain variable region (vL) comprising SEQ ID NO: 226 A chain variable region (vL), (viii) a heavy chain variable region (vH) comprising SEQ ID NO: 242, and comprising SEQ ID NO: 258 a light chain variable region (vL); (ix) a heavy chain variable region (vH) comprising SEQ ID NO: 274, and a light chain comprising SEQ ID NO: 290 An isolated antibody or antigen-binding fragment thereof comprising a variable region (vL).

7. At least 90, 91, 9 6. Claim 6, which retains 2, 93, 94, 95, 96, 97, 98, or 99% identity 2. The antibody or fragment thereof described in claim 1.

8. 1, 2, 3, 4 or 5, but not more than 10, of the variable light or heavy chain regions The antibody of claim 6, wherein less than one amino acid has been modified, deleted or substituted.

9. The antibody is a monoclonal antibody, a chimeric antibody, a humanized antibody, a human modified antibody, a human antibody, The antibody of claim 6 which is a single chain antibody (scFv) or an antibody fragment.

10. The antibody or fragment thereof may be reduced or fully glycosylated. The antibody of claim 1, 2 or 6, which is unfucosylated or hypofucosylated.

11. A method for treating a cancer, comprising administering to a patient an antibody or fragment thereof according to claim 1, 2 or 6, comprising administering to said patient a pharma- ceutically acceptable carrier. Further comprising a pharmaceutical composition.

12. The medicament of claim 11, wherein the pharma- ceutical acceptable carrier comprises a histidine or a carbohydrate. composition.

13. The pharmaceutical composition of claim 12, wherein the carbohydrate is sucrose.

14. A pharmaceutical composition comprising a plurality of antibodies or antigen-binding fragments according to claim 1, 2 or 6. and 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.

15. A pharmaceutical composition comprising a plurality of antibodies or antigen-binding fragments according to claim 1, 2 or 6. and wherein none of the antibodies contains a bisected GlcNAc.

16. A pharmaceutical composition comprising the antibody or fragment thereof according to claim 1, 2 or 6, The pharmaceutical composition, wherein the composition is prepared as a lyophilizate.

17. A method for neutralizing BK virus or JC virus infection, comprising administering to a patient in need thereof 10. The method of claim 10, further comprising administering to the patient an amount of the antibody of claim 1, 2 or 6 via injection or infusion. 、。

18. The patient in need thereof has been diagnosed with BK viruria or BK viremia. The method according to claim 17.

19. The patient in need thereof has been diagnosed with JC viruria or JC viremia. The method according to claim 17.

20. Methods for treating or reducing the likelihood of BK virus or JC virus associated disorders. and injecting or infusing an effective amount of the antibody of claim 1, 2 or 6 into a patient in need thereof. and administering the same via a bolus of water to a patient in need thereof, the disorder being nephropathy, BKVAN, hemorrhagic cystitis (HC), progressive bladder cancer, or the like. Progressive multifocal leukoencephalopathy (PML), granular cell neuronopathy (GCN), interstitial kidney disease, urinary Vasculitis, vasculitis, colitis, retinitis, meningitis, and immune reconstitution inflammatory syndrome (IRI) S).

21. 21. The method of claim 20, wherein the antibody or composition is reconstituted prior to injection or infusion. 。

22. 21. The method of claim 20, wherein the antibody or pharmaceutical composition is administered in combination with another therapeutic agent. The method described.

23. 23. The method of claim 22, wherein the therapeutic agent is an immunosuppressant.

24. The immunosuppressant is a monophosphate dehydrogenase inhibitor, a purine synthesis inhibitor, a calcineurin inhibitor, 24. The method of claim 23, wherein the compound is an inhibitor or an mTOR inhibitor.

25. The immunosuppressant is mycophenolate mofetil (MMF), mycophenolate sodium cyclosporine, azathioprine, tacrolimus, sirolimus or cyclosporine.

24. The method according to claim 23.

26. 23. The method of claim 22, wherein the therapeutic agent is an additional anti-BKV or JCV antibody.

27. 2. The method of claim 1, wherein the PML is associated with the treatment of multiple sclerosis, rheumatoid arthritis, or psoriasis. The method according to claim 0.

28. Treatment of multiple sclerosis includes natalizumab, fingolimod, dimethyl fumarate, and fumaric acid. ester, or alemtuzumab.

29. 28. The method of claim 27, wherein the treatment of rheumatoid arthritis is with rituximab.

30. 28. The method of claim 27, wherein the treatment of psoriasis is with efalizumab.

31. An antibody or fragment thereof according to claim 1, 2 or 6 for use as a medicament.

32. A method according to claim 1, 2 or 6 for use in neutralizing BK virus or JC virus infection.

2. The antibody or fragment thereof described in claim 1.

33. Nephropathy, BKVAN, hemorrhagic cystitis (HC), progressive multifocal leukoencephalopathy (PML), granuloma Glomerular neuronopathy (GCN), interstitial kidney disease, ureteral stenosis, vasculitis, colitis, retinitis, myelitis It is used to treat or reduce the likelihood of meningitis and immune reconstitution inflammatory syndrome (IRIS).

10. An antibody or fragment thereof according to claim 1, 2 or 6 for use in a therapeutic agent comprising the steps of:

34. Use of the antibody or fragment thereof according to claim 33, administered in combination with another therapeutic agent. 。

35. 34. The use of an antibody or fragment thereof according to claim 33, wherein the therapeutic agent is an immunosuppressant.

36. The immunosuppressant is a monophosphate dehydrogenase inhibitor, a purine synthesis inhibitor, a calcineurin inhibitor, 36. The use of an antibody or fragment thereof according to claim 35 which is an inhibitor or an mTOR inhibitor.

37. The immunosuppressant is mycophenolate mofetil (MMF), mycophenolate sodium cyclosporine, azathioprine, tacrolimus, sirolimus or cyclosporine.

35. Use of an antibody or a fragment thereof according to claim 35.

38. The use of the antibody or fragment thereof according to claim 34, wherein the therapeutic agent is an additional anti-BKV antibody. For.

39. 3. The method of claim 2, wherein the PML is associated with the treatment of multiple sclerosis, rheumatoid arthritis, or psoriasis.

4. Use of the antibody or fragment thereof according to claim 3.

40. Treatment of multiple sclerosis includes natalizumab, fingolimod, dimethyl fumarate, and fumaric acid.

40. The use of claim 39, wherein said ester or alemtuzumab is used.

41. 40. The use according to claim 39, wherein the treatment of rheumatoid arthritis is with rituximab.

42. 40. The use of claim 39, wherein the treatment of psoriasis is with efalizumab.

43. A nucleic acid encoding the antibody or antigen-binding fragment of claim 1, 2 or 6.

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

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

46. A diagnostic reagent comprising a labeled antibody or antigen-binding fragment thereof according to claim 1, 2 or 6. 。

47. The label may be a radiolabel, a fluorophore, a chromophore, an imaging agent, or a metal ion.

47. The diagnostic agent of claim 46, wherein the diagnostic agent is selected from the group consisting of:

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