Anti-alpha-synuclein monoclonal antibodies and methods of use thereof

Abstract

The present disclosure provides monoclonal antibodies that bind to α-synuclein. In certain aspects, the antibodies preferentially bind to α-synuclein fibrils over α-synuclein monomers. In other aspects, the present disclosure provides methods for treating, ameliorating, and / or preventing α-synucleinopathies in a subject, comprising administering to the subject any one of the antibodies of the present disclosure. In yet another aspect, the present disclosure provides methods for detecting α-synuclein fibrils using any one of the antibodies of the present disclosure.

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority under U.S.C. §119(e) to U.S. Provisional Patent Application No. 62 / 937,636, filed November 19, 2019, which is incorporated herein by reference in its entirety.

[0002] Federally Sponsored Research and Development Statement This invention was made with government support under Grant Nos. T32-AG000255, P30-AG10124, P50-NS053488 and R01-NS088322 awarded by the National Institutes of Health (NIH). The government has certain rights in this invention. [Background technology]

[0003] Background of the Disclosure Parkinson's disease (PD) is a progressive neurodegenerative disorder with no disease-modifying treatment that affects 1% of the world's population. Even the most effective dopamine replacement therapies fail to halt disease progression, including the development of dementia in up to 80% of cases. The disease's characteristic motor symptoms are often preceded by non-motor symptoms, including constipation, sleep disturbances, and olfactory impairment. The motor symptoms are often followed by cognitive decline, which allows for the diagnosis of PD dementia (PDD). Dementia with Lewy bodies (DLB) shares symptoms and pathology with PD and PDD, suggesting that these three disorders belong to a family of neurodegenerative disorders collectively known as α-synucleinopathies, which are caused by the abnormal accumulation of normal synaptic α-synuclein protein in neuronal Lewy bodies (LBs) and axonal Lewy neurites (LNs). Alpha-synuclein is not just a bystander in these diseases, as rare mutations, double duplications, and triple duplications of alpha-synuclein lead to familial PD.

[0004] The progression from manageable to debilitating symptoms corresponds to the burden of α-synuclein pathology in higher cortical regions of the brain. Overexpression of α-synuclein in mice with or without familial mutations is sufficient to drive the formation of LB-like inclusions and neurodegeneration. Furthermore, reduction of α-synuclein levels has beneficial effects in neurotoxin-induced models of PD. Collectively, these studies suggest that reducing α-synuclein, particularly misfolded forms of α-synuclein, may be a therapeutic strategy for treating PD and related α-synucleinopathies. Because α-synuclein is primarily localized in the cytoplasm of neurons, it has been thought that therapeutic molecules must cross not only the blood-brain barrier (BBB) ​​but also the plasma membrane of neurons to interact with α-synuclein. However, numerous recent in vitro and in vivo studies suggest that misfolded α-synuclein species can be released by neurons and taken up by nearby neurons, triggering transcellular transmission of pathogenic α-synuclein. Therefore, minimizing the effects of this reservoir of extracellular pathogenic α-synuclein offers a unique and more accessible therapeutic opportunity for the treatment of PD, especially when pathological α-synuclein is also present in peripheral areas such as the enteric nervous system.

[0005] There are numerous promising approaches to inhibiting the transmission of α-synuclein pathology. For example, extracellular α-synuclein could be targeted for vascular or glymphatic clearance, its uptake could be blocked, or glial cells could be modified to promote the clearance of extracellular α-synuclein. At least some of these potential mechanisms could be facilitated through antibody-mediated immunotherapy. For example, antibodies could block neuronal α-synuclein uptake while promoting glymphatic clearance to peripheral or glial clearance through binding to surface Fc receptors. Passive immunotherapy (direct antibody treatment instead of injecting immunogens) is a particularly attractive option because therapeutic antibodies have been demonstrated to be relatively safe and immunotherapy has been shown to promote the clearance of extracellular targets. Administered antibodies must achieve sufficient brain levels to affect disease biology, but antibodies are known to have poor BBB penetration.

[0006] There is a need to develop anti-α-Syn antibodies that can be used to investigate the neuropathological features of PD and related disorders. In certain embodiments, these antibodies can be used in the treatment of these diseases. The present disclosure addresses and meets these needs. Summary of the Invention

[0007] BRIEF SUMMARY OF THE DISCLOSURE The present disclosure provides certain monoclonal antibodies comprising a light chain variable region (VL) and a heavy chain variable region (VH) as defined elsewhere herein. The present disclosure further provides pharmaceutical compositions comprising at least one monoclonal antibody contemplated herein and at least one pharmaceutical excipient. The present disclosure further provides certain isolated polynucleotides comprising at least one of the nucleic acid sequences contemplated herein.

[0008] The present disclosure further provides an autonomously replicating or integrating mammalian cell vector comprising at least one recombinant nucleic acid encoding at least one antibody comprising a light chain variable region (VL) and a heavy chain variable region (VH) as defined elsewhere herein. The present disclosure further provides an isolated host cell comprising any of the vectors contemplated herein.

[0009] The present disclosure further provides methods for treating, ameliorating, and / or preventing a synucleopathic disease in a subject, hi certain embodiments, the methods comprising administering to the subject a therapeutically effective amount of at least one isolated monoclonal antibody contemplated herein.

[0010] The present disclosure further provides methods for detecting synucleinopathy in a subject. In certain embodiments, the methods include administering to the subject at least one labeled isolated monoclonal antibody as contemplated herein. In certain embodiments, the methods include detecting the presence or absence of a complex between the labeled isolated monoclonal antibody and any α-Syn fibrils, oligomers, and / or other misfolded α-Syn species present in the subject. In certain embodiments, if a complex is detected, the subject has a synucleinopathy.

[0011] The present disclosure further provides methods for detecting total α-syn, α-syn fibrils, and / or α-syn oligomer species in a sample. In certain embodiments, the methods include contacting the sample with at least one labeled isolated monoclonal antibody as contemplated herein. In certain embodiments, the methods include detecting the presence or absence of a complex between the labeled isolated monoclonal antibody and total α-syn, α-syn monomers, α-syn fibrils, and / or α-syn oligomer species present in the sample. In certain embodiments, detection of a complex indicates the presence of total α-syn, α-syn monomers, α-syn fibrils, and / or α-syn oligomer species in the sample. [The present invention 1001] An isolated monoclonal antibody comprising a light chain variable region (VL) and a heavy chain variable region (VH), The VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO: 27, 46, 70, 114, 126, 185, 213, or 240; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29, 72, 116, 214, or 242; and a CDR3 region comprising the amino acid sequence of SEQ ID NO: 31, 85, 118, 129, 152, 205, 216, or 244 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO: 11, 58, 92, 101, 144, 160, or 226; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 13, 40, 60, 93, 102, 134, 145, 161, 200, or 228; and a CDR3 region comprising the amino acid sequence of SEQ ID NO: 15, 62, 81, 147, 163, 194, 201, or 230 Including, The isolated monoclonal antibody. [The present invention 1002] The VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO: 27, 46, 70, 27, 27, 70, 70, 114, 126, 46, 46, 46, 70, or 46; A CDR2 region comprising the amino acid sequence of SEQ ID NOs: 29, 29, 72, 29, 29, 72, 72, 116, 72, 29, 29, 29, 29, or 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 31, 31, 85, 31, 31, 85, 85, 118, 129, 31, 31, 31, 152, or 31 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO: 11, 58, 92, 101, or 144; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 13, 40, 60, 93, 102, 134, or 145; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 15, 62, 81, or 147 Including, The monoclonal antibody of the present invention. [The present invention 1003] The VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:70, 185, 213, or 240; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29, 72, 214, or 242; and a CDR3 region comprising the amino acid sequence of SEQ ID NO: 152, 129, 205, 216, or 244 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO: 160, 58, or 226; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 161, 145, 200, or 228; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 163, 194, 201, or 230 Including, The monoclonal antibody of the present invention. [The present invention 1004] The monoclonal antibody of the present invention 1001 is adapted for at least one of the following: (a) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:27; A CDR2 region comprising the amino acid sequence of SEQ ID NO:29; and CDR3 region comprising the amino acid sequence of SEQ ID NO:31 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:11; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 13; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 15 including; (b) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:46; A CDR2 region comprising the amino acid sequence of SEQ ID NO:29; and CDR3 region comprising the amino acid sequence of SEQ ID NO:31 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:11; A CDR2 region comprising the amino acid sequence of SEQ ID NO:40; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 15 including; (c) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:70; A CDR2 region comprising the amino acid sequence of SEQ ID NO:72; and CDR3 region comprising the amino acid sequence of SEQ ID NO:85 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:58; A CDR2 region comprising the amino acid sequence of SEQ ID NO:60; and CDR3 region comprising the amino acid sequence of SEQ ID NO:62 including; (d) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:70; A CDR2 region comprising the amino acid sequence of SEQ ID NO:72; and CDR3 region comprising the amino acid sequence of SEQ ID NO:85 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:58; A CDR2 region comprising the amino acid sequence of SEQ ID NO:60; and CDR3 region comprising the amino acid sequence of SEQ ID NO:81 including; (e) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:70; A CDR2 region comprising the amino acid sequence of SEQ ID NO:72; and CDR3 region comprising the amino acid sequence of SEQ ID NO:85 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:92; a CDR2 region comprising the amino acid sequence of SEQ ID NO:93; and CDR3 region comprising the amino acid sequence of SEQ ID NO:62 including; (f) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:114; A CDR2 region comprising the amino acid sequence of SEQ ID NO:116; and CDR3 region comprising the amino acid sequence of SEQ ID NO:118 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:101; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 102; and CDR3 region comprising the amino acid sequence of SEQ ID NO:62 including; (g) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:126; A CDR2 region comprising the amino acid sequence of SEQ ID NO:72; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 129 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:101; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 102; and CDR3 region comprising the amino acid sequence of SEQ ID NO:62 including; (h) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:46; A CDR2 region comprising the amino acid sequence of SEQ ID NO:29; and CDR3 region comprising the amino acid sequence of SEQ ID NO:31 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:11; A CDR2 region comprising the amino acid sequence of SEQ ID NO:134; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 15 including; (i) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:70; A CDR2 region comprising the amino acid sequence of SEQ ID NO:29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 152 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:144; A CDR2 region comprising the amino acid sequence of SEQ ID NO:145; and CDR3 region comprising the amino acid sequence of SEQ ID NO:147 including; (j) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:70; A CDR2 region comprising the amino acid sequence of SEQ ID NO:29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 152 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:160; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 161; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 163 including; (k) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:185; A CDR2 region comprising the amino acid sequence of SEQ ID NO:29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 129 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:160; A CDR2 region comprising the amino acid sequence of SEQ ID NO:145; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 163 including; (l) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:70; A CDR2 region comprising the amino acid sequence of SEQ ID NO:29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 152 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:160; A CDR2 region comprising the amino acid sequence of SEQ ID NO:145; and CDR3 region comprising the amino acid sequence of SEQ ID NO:194 including; (m) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:70; A CDR2 region comprising the amino acid sequence of SEQ ID NO:72; and CDR3 region comprising the amino acid sequence of SEQ ID NO:205 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:58; A CDR2 region comprising the amino acid sequence of SEQ ID NO:200; and CDR3 region comprising the amino acid sequence of SEQ ID NO:201 including; (n) The VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:213; A CDR2 region comprising the amino acid sequence of SEQ ID NO:214; and CDR3 region comprising the amino acid sequence of SEQ ID NO:216 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:58; A CDR2 region comprising the amino acid sequence of SEQ ID NO:200; and CDR3 region comprising the amino acid sequence of SEQ ID NO:201 including; (o) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:240; A CDR2 region comprising the amino acid sequence of SEQ ID NO:242; and CDR3 region comprising the amino acid sequence of SEQ ID NO:244 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:226; A CDR2 region comprising the amino acid sequence of SEQ ID NO:228; and CDR3 region comprising the amino acid sequence of SEQ ID NO:230 Including. [The present invention 1005] The VL is SEQ ID NO:27-SEQ ID NO:28-SEQ ID NO:29-SEQ ID NO:30-SEQ ID NO:31, SEQ ID NO:46-SEQ ID NO:28-SEQ ID NO:29-SEQ ID NO:30-SEQ ID NO:31, SEQ ID NO:70-SEQ ID NO:71-SEQ ID NO:72-SEQ ID NO:73-SEQ ID NO:85, SEQ ID NO:70-SEQ ID NO:127-SEQ ID NO:29-SEQ ID NO:151-SEQ ID NO:152, SEQ ID NO:70-SEQ ID NO:127-SEQ ID NO:29-SEQ ID NO:172-SEQ ID NO:152, SEQ ID NO:70-SEQ ID NO:127-SEQ ID NO:72-SEQ ID NO:204-SEQ ID NO:205, SEQ ID NO:114-SEQ ID NO:115-SEQ ID NO:116-SEQ ID NO:117-SEQ ID NO:118, SEQ ID NO:126-SEQ ID NO:127-SEQ ID NO:72-SEQ ID NO:128-SEQ ID NO:129, SEQ ID NO:185-SEQ ID NO:186-SEQ ID NO:29-SEQ ID NO:187-SEQ ID NO:129, SEQ ID NO:213-SEQ ID NO:115-SEQ ID NO:214-SEQ ID NO:215-SEQ ID NO:216, or SEQ ID NO:240-SEQ ID NO:241-SEQ ID NO:242-SEQ ID NO:243-SEQ ID NO:244 comprising the amino acid sequence and the VH is SEQ ID NO:11-SEQ ID NO:12-SEQ ID NO:13-SEQ ID NO:14-SEQ ID NO:15, SEQ ID NO:11-SEQ ID NO:12-SEQ ID NO:40-SEQ ID NO:41-SEQ ID NO:15, SEQ ID NO:11-SEQ ID NO:12-SEQ ID NO:134-SEQ ID NO:135-SEQ ID NO:15, SEQ ID NO:58-SEQ ID NO:59-SEQ ID NO:60-SEQ ID NO:61-SEQ ID NO:62, SEQ ID NO:58-SEQ ID NO:59-SEQ ID NO:60-SEQ ID NO:80-SEQ ID NO:81, SEQ ID NO:58-SEQ ID NO:59-SEQ ID NO:200-SEQ ID NO:94-SEQ ID NO:201, SEQ ID NO:92-SEQ ID NO:59-SEQ ID NO:93-SEQ ID NO:94-SEQ ID NO:62, SEQ ID NO:101-SEQ ID NO:59-SEQ ID NO:102-SEQ ID NO:103-SEQ ID NO:62, SEQ ID NO:144-SEQ ID NO:59-SEQ ID NO:145-SEQ ID NO:146-SEQ ID NO:147, SEQ ID NO:160-SEQ ID NO:59-SEQ ID NO:145-SEQ ID NO:177-SEQ ID NO:163, SEQ ID NO:160-SEQ ID NO:59-SEQ ID NO:145-SEQ ID NO:193-SEQ ID NO:194, SEQ ID NO:160-SEQ ID NO:59-SEQ ID NO:161-SEQ ID NO:162-SEQ ID NO:163, or SEQ ID NO:226-SEQ ID NO:227-SEQ ID NO:228-SEQ ID NO:229-SEQ ID NO:230 comprising the amino acid sequence The monoclonal antibody of the present invention. [The present invention 1006] The VL is SEQ ID NO:27-SEQ ID NO:28-SEQ ID NO:29-SEQ ID NO:30-SEQ ID NO:31, SEQ ID NO:46-SEQ ID NO:28-SEQ ID NO:29-SEQ ID NO:30-SEQ ID NO:31, SEQ ID NO:70-SEQ ID NO:71-SEQ ID NO:72-SEQ ID NO:73-SEQ ID NO:85, SEQ ID NO:70-SEQ ID NO:127-SEQ ID NO:29-SEQ ID NO:151-SEQ ID NO:152, SEQ ID NO:114-SEQ ID NO:115-SEQ ID NO:116-SEQ ID NO:117-SEQ ID NO:118, or SEQ ID NO:126-SEQ ID NO:127-SEQ ID NO:72-SEQ ID NO:128-SEQ ID NO:129 comprising the amino acid sequence and the VH is SEQ ID NO:11-SEQ ID NO:12-SEQ ID NO:13-SEQ ID NO:14-SEQ ID NO:15, SEQ ID NO:11-SEQ ID NO:12-SEQ ID NO:40-SEQ ID NO:41-SEQ ID NO:15, SEQ ID NO:11-SEQ ID NO:12-SEQ ID NO:134-SEQ ID NO:135-SEQ ID NO:15, SEQ ID NO:58-SEQ ID NO:59-SEQ ID NO:60-SEQ ID NO:61-SEQ ID NO:62, SEQ ID NO:58-SEQ ID NO:59-SEQ ID NO:60-SEQ ID NO:80-SEQ ID NO:81, SEQ ID NO:92-SEQ ID NO:59-SEQ ID NO:93-SEQ ID NO:94-SEQ ID NO:62, SEQ ID NO:101-SEQ ID NO:59-SEQ ID NO:102-SEQ ID NO:103-SEQ ID NO:62, or SEQ ID NO:144-SEQ ID NO:59-SEQ ID NO:145-SEQ ID NO:146-SEQ ID NO:147 comprising the amino acid sequence The monoclonal antibody of the present invention. [The present invention 1007] The VL is SEQ ID NO:70-SEQ ID NO:127-SEQ ID NO:29-SEQ ID NO:172-SEQ ID NO:152, SEQ ID NO:70-SEQ ID NO:127-SEQ ID NO:72-SEQ ID NO:204-SEQ ID NO:205, SEQ ID NO:185-SEQ ID NO:186-SEQ ID NO:29-SEQ ID NO:187-SEQ ID NO:129, SEQ ID NO:213-SEQ ID NO:115-SEQ ID NO:214-SEQ ID NO:215-SEQ ID NO:216, or SEQ ID NO:240-SEQ ID NO:241-SEQ ID NO:242-SEQ ID NO:243-SEQ ID NO:244 comprising the amino acid sequence and the VH is SEQ ID NO:58-SEQ ID NO:59-SEQ ID NO:200-SEQ ID NO:94-SEQ ID NO:201, SEQ ID NO:160-SEQ ID NO:59-SEQ ID NO:145-SEQ ID NO:177-SEQ ID NO:163, SEQ ID NO:160-SEQ ID NO:59-SEQ ID NO:145-SEQ ID NO:193-SEQ ID NO:194, SEQ ID NO:160-SEQ ID NO:59-SEQ ID NO:161-SEQ ID NO:162-SEQ ID NO:163, or SEQ ID NO:226-SEQ ID NO:227-SEQ ID NO:228-SEQ ID NO:229-SEQ ID NO:230 comprising the amino acid sequence The monoclonal antibody of the present invention. [The present invention 1008] 1001. A monoclonal antibody of the present invention which is humanized. [The present invention 1009] A labeled monoclonal antibody of the present invention. [The present invention 1010] A pharmaceutical composition comprising a monoclonal antibody of the present invention and at least one pharmaceutical excipient. [The present invention 1011] 1. An isolated polynucleotide comprising at least one of the nucleic acid sequences of SEQ ID NOs:3, 5, 7, 19, 21, 23, 35, 43, 44, 50, 52, 54, 66, 68, 75, 77, 88, 89, 96, 97, 99, 106, 108, 110, 121, 132, 138, 139, 141, 148, 150, 155, 157, 167, 168, 170, 175, 180, 191, 195, 197, 198, 203, 207, 209, 211, 219, 221, 223, 233, 235, and 237. [The present invention 1012] At least one nucleic acid sequence selected from the group consisting of SEQ ID NOs: 3, 5, 7, 35, 50, 52, 54, 75, 77, 88, 89, 97, 99, 132, 138, 139, 141, 155, 157, 175, 191, 197, 198, 219, 221, and 223; and At least one nucleic acid sequence selected from the group consisting of SEQ ID NOs: 19, 21, 23, 43, 44, 66, 68, 96, 106, 108, 110, 121, 148, 150, 167, 168, 170, 180, 195, 203, 207, 209, 211, 233, 235, and 237. 1011. An isolated polynucleotide of the present invention comprising: [The present invention 1013] At least one nucleic acid sequence selected from the group consisting of SEQ ID NOs:3, 5, 7; SEQ ID NOs:3, 35, 7; SEQ ID NOs:3, 132, 7; SEQ ID NOs:50, 52, 54; SEQ ID NOs:50, 75, 77; SEQ ID NOs:50, 139, 175; SEQ ID NOs:50, 139, 191; SEQ ID NOs:50, 155, 157; SEQ ID NOs:50, 197, 198; SEQ ID NOs:88, 89, 54; SEQ ID NOs:97, 99, 54; SEQ ID NOs:138, 139, 141; and SEQ ID NOs:219, 221, 223; and At least one nucleic acid sequence selected from the group consisting of SEQ ID NOs: 19, 21, 23; SEQ ID NOs: 19, 43, 44; SEQ ID NOs: 19, 66, 68; SEQ ID NOs: 19, 66, 96; SEQ ID NOs: 19, 66, 203; SEQ ID NOs: 19, 148, 150; SEQ ID NOs: 106, 108, 110; SEQ ID NOs: 121, 66, 68; SEQ ID NOs: 167, 168, 170; SEQ ID NOs: 167, 168, 195; SEQ ID NOs: 180, 168, 68; SEQ ID NOs: 213, 214, 216; and SEQ ID NOs: 233, 235, 237. 1011. An isolated polynucleotide of the present invention comprising: [The present invention 1014] 1. A method of treating, ameliorating, and / or preventing a synucleopathic disease in a subject in need thereof, comprising: The method comprising the step of administering to the subject a therapeutically effective amount of at least one isolated monoclonal antibody of the present invention. [The present invention 1015] The method of the present invention, wherein the synucleinopathic disease is at least one selected from the group consisting of Parkinson's disease, Parkinson's disease with dementia, dementia with Lewy bodies, Alzheimer's disease, Down's syndrome, multiple system atrophy, prion disease, and other α-Syn-related neurodegenerative disorders. [The present invention 1016] The method of claim 1014, wherein said antibody is provided to said subject as a pharmaceutical composition. [The present invention 1017] The method of claim 1014, wherein said antibody is administered to said subject parenterally. [The present invention 1018] 1. A method for detecting a synucleinopathy in a subject, comprising: administering to the subject at least one labeled isolated monoclonal antibody of the present invention; and detecting the presence or absence of complexes between the labeled isolated monoclonal antibody and any α-Syn fibrils, oligomers, and / or other misfolded α-Syn species present in the subject; If the complex is detected, the subject has a synucleinopathy. The method. [The present invention 1019] 1. A method for detecting total α-Syn, α-Syn fibril, and / or α-Syn oligomeric species in a sample, comprising: contacting the sample with at least one labeled isolated monoclonal antibody of the present invention; and detecting the presence or absence of complexes between the labeled isolated monoclonal antibody and total α-Syn, α-Syn monomers, α-Syn fibrils, and / or α-Syn oligomeric species present in the sample; If the complex is detected, total α-Syn, α-Syn monomer, α-Syn fibril, and / or α-Syn oligomer species are present in the sample. The method. [The present invention 1020] The method of claim 1019, wherein said sample comprises an in vitro and / or ex vivo sample. [The present invention 1021] An autonomously replicating or integrating mammalian cell vector comprising a recombinant nucleic acid encoding an antibody comprising a light chain variable region (VL) and a heavy chain variable region (VH), The VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO: 27, 46, 70, 114, 126, 185, 213, or 240; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29, 72, 116, 214, or 242; and a CDR3 region comprising the amino acid sequence of SEQ ID NO: 31, 85, 118, 129, 152, 205, 216, or 244 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO: 11, 58, 92, 101, 144, 160, or 226; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 13, 40, 60, 93, 102, 134, 145, 161, 200, or 228; and a CDR3 region comprising the amino acid sequence of SEQ ID NO: 15, 62, 81, 147, 163, 194, 201, or 230 Including, The vector. [The present invention 1022] The vector of the present invention 1021, including a plasmid or virus. [The present invention 1023] The vectors of the present invention 1021, including mammalian cell expression vectors. [The present invention 1024] The vector of the present invention 1021 further comprising at least one nucleic acid sequence that directs and / or controls the expression of said antibody. [The present invention 1025] An isolated host cell comprising at least one vector of the present invention 1021. [The present invention 1026] The cell of the present invention 1025, which is a non-human cell. [The present invention 1027] The cell of the present invention 1025, which is a mammalian cell. [Brief explanation of the drawings]

[0012] The following detailed description of specific embodiments of the present disclosure will be more fully understood when read in conjunction with the accompanying drawings. For the purpose of illustrating the disclosure, specific embodiments are shown in the drawings. It should be understood, however, that the disclosure is not limited to the precise arrangements and instrumentalities of the embodiments shown in the drawings. [Figure 1]Figures 1A-1F include an overview of the passive immunotherapy screen. Figure 1A: Mice were immunized with α-synuclein preformed fibrils (PFFs) to induce an immune response and antibody production against pathogenic α-synuclein. Figure 1B: Antibody-producing B cells were then harvested from the spleens of the immunized mice and fused with myeloma cells to generate hybridoma clonal cell lines expressing antibodies against α-synuclein. Figure 1C: The hybridomas were separated into individual clonal populations, and antibody-containing supernatants from each clone were passed through a primary screen to identify candidates with favorable properties. To confirm that the antibodies recognized pathogenic human α-synuclein, the antibodies were used in IHC of human tissues. In certain non-limiting embodiments, candidate antibodies also had a preference for fibrillar α-synuclein, so they were screened in an indirect ELISA format against monomeric and fibrillar α-synuclein. Each antibody was epitope mapped and tested for immunogenicity against mouse and human α-synuclein, allowing its use in mouse disease models. Finally, antibodies were screened for their ability to reduce α-synuclein pathology in cultured neurons in a primary neuron immunotherapy assay. (Figure 1D) Prioritized antibodies underwent two additional rounds of subcloning to ensure monoclonality. (Figure 1E) Final clones were subjected to confirmatory screening to ensure optimal properties were maintained and to identify final candidates for in vivo experiments. (Figure 1F) Non-limiting candidate antibodies were tested for their ability to reduce α-synuclein pathology and toxicity in a mouse model of PD. Candidates were tested alongside an IgG control and α-synuclein antibodies with proven therapeutic efficacy. [Figure 2]Figures 2A-2B illustrate non-limiting immunohistochemistry results revealing antibodies that preferentially bind to LBs. Figure 2A: Sections of amygdala tissue containing abundant Lewy body pathology were stained using undiluted or 1:3 diluted hybridoma supernatants. Representative staining of Lewy bodies is shown for each of the screened antibody supernatants. Scale bar = 50 μm. Figure 2B: The optical density within a Lewy body divided by the optical density of a 1 mm2 tissue section containing that Lewy body was defined as the Lewy body discrimination index. Antibodies that preferentially recognize Lewy bodies score highly on this metric. Most antibodies showed some enhanced Lewy body recognition, with the exception of antibodies with scores below 1.5 that showed little apparent Lewy body preference over neuropil α-synuclein staining. [Figure 3]Figures 3A-3C illustrate non-limiting epitope mapping confirming that certain antibodies recognize human and mouse α-synuclein. Figure 3A: Schematic of the recombinant α-synuclein fragments used to determine the epitopes of the 9000 antibody series. Full-length (FL) α-synuclein is 140 amino acids long with an internal non-amyloid component (NAC) domain. The constructs used to probe the epitopes had truncated N- or C-terminal human α-synuclein residues. Figure 3B: Most antibodies recognized both human and mouse α-synuclein to some extent. Furthermore, all antibodies recognized α-synuclein with truncated N-termini, suggesting that they all recognize the C-terminus of α-synuclein. The most extreme of these did not recognize α-synuclein unless it included even the last 10 amino acids, suggesting that the epitope is aa 130-140. Many other antibodies recognized 1-130 but not 1-120, suggesting that they have an epitope between 120 and 130. Figure 3C: Another large number of antibodies did not recognize 1-110, suggesting that they recognize 110-120. Finally, several antibodies recognized all constructs except 1-89, suggesting that the epitope is between amino acids 90 and 102. *Peptide 1-110 was run in the last lane of these gels, thereby shifting the lane to the left. [Figure 4]Figures 4A-4C illustrate a non-limiting sandwich ELISA to identify antibodies with a preference for misfolded α-synuclein. Antibody preference for misfolded α-synuclein was assessed by performing parallel sandwich ELISAs using either human α-synuclein monomer or α-synuclein PFF as the antigen. Syn211 was also used on each plate as a non-selective control antibody. Using this method, we distinguished three classes of antibodies: (Figure 4A) antibodies that did not bind in this assay and therefore had no signal for either monomer or PFF; (Figure 4B) antibodies that were non-selective and had similar binding to α-synuclein monomer and PFF; and (Figure 4C) antibodies that were PFF-selective and showed a significantly higher preference for α-synuclein PFF over monomer. The histograms shown are representative of the three classes, but all tested antibodies can be classified into one of these categories. Plots represent the mean from three technical replicates, and error bars represent the standard error. [Figure 5A] Figure 5 illustrates a non-limiting neuronal immunotherapy assay demonstrating the differential efficacy of α-synuclein antibodies in blocking LB-like pathology. Figure 5A: Examples of pS129 α-synuclein pathology (green) and NeuN (magenta) immunostaining after treatment of primary neurons with α-synuclein PFFs and the indicated representative antibodies. Scale bar = 50 μm. [Figure 5B] A non-limiting neuronal immunotherapy assay is illustrated, revealing differential potency of α-synuclein antibodies in blocking LB-like pathology. Figure 5B: Quantification of pS129 α-synuclein area / neuron number normalized to IgG-treated neurons. Antibodies have varying effects on α-synuclein pathology and are ranked from least effective to most effective. Twenty-one of these antibodies demonstrated statistically significant reductions in pathology (Kruskal-Wallis test with Dunn's multiple comparisons test: *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001). N=6–20 / condition. Error bars represent standard error. [Figure 6]Figures 6A-6B include a summary of antibody properties shown as heat maps, with a scale bar at the top of each column. Most antibodies are from the IgG subclass. Most recognize mouse α-synuclein, and all recognize human α-synuclein. The LB discrimination index derived from immunohistochemistry reflects the antibody's ability to selectively bind LB α-synuclein. α-Synuclein PFF preference, derived from sandwich ELISA results and spanning over 100-fold differences, allows for the selection of antibodies with a preference for misfolded α-synuclein (NB = non-binding). Finally, the % inhibition metric derived from primary neuron assays indicates the antibody's ability to reduce pathology in cell lines. In certain embodiments, antibodies of interest recognize both mouse and human α-synuclein, distinguish LB from the surrounding neuropil, have a higher binding affinity for α-synuclein PFFs than monomers, and inhibit nearly all in vitro neuronal α-synuclein pathology. Based on these measurements, Syn9048 and Syn9063 were selected for further subcloning. [Figure 7]Figures 7A-7D illustrate non-limiting subclones that retain the properties of their parent clones. Figure 7A: Syn9063 subclones were assayed for PFF preference as previously described, confirming that all subclones exhibited the same selectivity as the parent clone. All but one subclone showed a high preference for α-synuclein PFFs. Figure 7B: All Syn9048 subclones exhibited similar selectivity for α-synuclein PFFs. Plots in panels (Figure 7A) and (Figure 7B) represent the mean from three technical replicates, and error bars represent standard error. Figure 7C: Example of pS129 α-synuclein pathology (green) and NeuN (magenta) immunostaining after treatment of primary neurons with purified Syn9048 clone #3 at the indicated molar ratios to α-synuclein PFFs and α-synuclein monomer. Scale bar = 50 μm. Figure 7D: Quantification of pS129 α-synuclein area / neuron number normalized to control IgG-treated neurons. Increasing molar ratios of Syn9048 dose-dependently reduced pS129 α-synuclein pathology, with an IC50 of 0.006 (R2 = 0.9539) as fit by a sigmoidal dose curve. N = 3 / condition. Error bars represent standard error, and shaded areas represent the 95% confidence interval of the sigmoidal fit. [Figure 8]Figures 8A-8G illustrate that in vivo immunotherapy was well tolerated and improved dopaminergic tone. Figure 8A: Wild-type mice were injected with α-synuclein PFFs into the dorsal striatum at 2-3 months of age. One week after α-synuclein PFF injection, mice were injected with 30 mg / kg of control IgG1, Syn303, or Syn9048. Mice were injected weekly for the next 6 months, after which they were sacrificed and assayed for pathology and motor behavior. Figure 8B: Mean weight of mice in each antibody treatment group, with standard error represented by the hatched band. Weight increased over time but did not differ between groups (2-way ANOVA: time p=0.0001; treatment p=0.7209). N=16 / group. Error bars represent standard error. Figure 8C: Representative images of TH-stained SN from mice treated with control IgG1, Syn303, or Syn9048. Scale bar = 500 µm. Figure 8D: Estimated TH+ neurons in the SN after manual quantification of neurons across the SN (C = contralateral; I = ipsilateral). Substantial neuronal loss was observed ipsilateral to the injection site for all groups (2-way ANOVA with Sidak's multiple comparison test, contralateral vs. ipsilateral: IgG1 p < 0.0001; Syn303 p < 0.0001; Syn9048 p < 0.0001), but did not differ between groups (2-way ANOVA with Sidak's multiple comparison test, ipsilateral: IgG1 vs. Syn303 p = 0.6266, IgG1 vs. Syn9048 p = 0.3561). N = 10–16 per group. Figure 8E: The ratio of remaining neurons in the ipsilateral SN to the contralateral SN similarly showed no difference between groups (one-way ANOVA with Dunnett's multiple comparison test: IgG1 vs. Syn303 p = 0.6323, IgG1 vs. Syn9048 p = 0.9084). Figure 8F: Dopamine levels measured in the dorsal striatum of injected mice were reduced ipsilateral to the injection site in IgG1-treated animals but maintained in Syn9048-treated mice (one-way ANOVA with Dunnett's multiple comparison test: IgG1 vs. naive p = 0.0283; IgG1 vs. Syn303 p = 0.9791; IgG1 vs. Syn9048 p = 0.0047). N = 7–12 / group.Figure 8G: DOPAC levels were also similarly maintained by Syn9048 treatment (one-way ANOVA with Dunnett's multiple comparison test: IgG1 vs. naive p=0.0469; IgG1 vs. Syn303 p=0.9969; IgG1 vs. Syn9048 p=0.0049). N=7-12 / group. Error bars represent standard error. [Figure 9] Figures 9A-9F illustrate that Syn9048 reduces α-synuclein pathology in the SN and amygdala. Figure 9A: Mice injected with α-synuclein PFFs accumulate substantial α-synuclein pathology in the ipsilateral SN, as recognized by the conformation-selective antibody Syn506. Rare pathology was also observed in the contralateral SN at this time point. Scale bar = 100 μm. Figure 9B: Syn506+ area / section was substantially, but not significantly, reduced when quantified throughout the contralateral SN (one-way ANOVA with Dunnett's multiple comparisons test: IgG1 vs. Syn303 p = 0.0827; IgG1 vs. Syn9048 p = 0.0847). N = 14-16 / group. Figure 9C: Immunotherapeutic treatment also reduced α-synuclein pathology in the ipsilateral SN (1-way ANOVA with Dunnett's multiple comparison test: IgG1 vs. Syn303 p = 0.0937; IgG1 vs. Syn9048 p = 0.0444). N = 14–16 / group. Figure 9D: Mice injected with α-synuclein PFFs accumulate substantial α-synuclein pathology in the ipsilateral and contralateral amygdala, as recognized by the conformation-selective antibody Syn506. Scale bar = 100 μm. Figure 9E: Syn506+ area / section, quantified throughout the contralateral amygdala, was substantially reduced (1-way ANOVA with Dunnett's multiple comparison test: IgG1 vs. Syn303 p = 0.0521; IgG1 vs. Syn9048 p = 0.0069). N = 14–16 / group. Figure 9F: Alpha-synuclein pathology was not reduced to the same extent in the ipsilateral amygdala (one-way ANOVA with Dunnett's multiple comparisons test: IgG1 vs. Syn303 p=0.8694; IgG1 vs. Syn9048 p=0.2464). N=14-16 / group. All points represent individual mice, and error bars represent standard error. [Figure 10]Figure 1 illustrates non-limiting recognition of β-synuclein. Most antibodies did not recognize β-synuclein by Western blot. Syn9030 and Syn9066 had slight reactivity with β-synuclein, indicating some cross-reactivity of these antibodies. Syn7015 cross-reacts with β-synuclein, so this antibody is used as a positive control. [Figure 11] Figures 11A-11B illustrate a non-limiting sandwich ELISA to identify non-binding antibodies. Antibody preference for misfolded α-synuclein was assessed by performing parallel sandwich ELISAs using either α-synuclein monomer or α-synuclein PFF as the antigen. Syn211 was also used on each plate as a non-selective control antibody. The figure displays antibodies that were non-binding in this assay. Plots represent the mean from three technical replicates, and error bars represent standard error. [Figure 12] Figures 12A-12B illustrate a non-limiting sandwich ELISA to identify non-selective antibodies. Antibody preference for misfolded α-synuclein was assessed by performing parallel sandwich ELISAs using either α-synuclein monomer or α-synuclein PFF as the antigen. Syn211 was also used on each plate as a non-selective control antibody. This figure displays antibodies that were non-selective in this assay. Plots represent the mean from three technical replicates, and error bars represent standard error. [Figure 13]Figures 13A-13B illustrate a non-limiting sandwich ELISA to identify α-synuclein PFF-selective antibodies. Antibody preference for misfolded α-synuclein was assessed by performing parallel sandwich ELISAs using either α-synuclein monomer or α-synuclein PFF as the antigen. Syn211 was also used on each plate as a non-selective control antibody. The figure displays antibodies that showed a preference for α-synuclein PFF in this assay. Plots represent the mean from three technical replicates, and error bars represent standard error. [Figure 14] Figure 1 illustrates a non-limiting second sandwich ELISA confirming α-synuclein PFF-selective antibodies. Antibody preference for misfolded α-synuclein was assessed by performing a second set of parallel sandwich ELISAs using either α-synuclein monomer or α-synuclein PFF as antigen and the polyclonal α-synuclein antibody SNL4. Plots represent the average from three technical replicates, and error bars represent standard error. [Figure 15] Figure 1 illustrates that Syn9048 blocks α-synuclein pathology induced by mouse α-synuclein PFFs. Syn9048 was assayed for its ability to block α-synuclein pathology induced by mouse α-synuclein in neurons. Primary hippocampal neurons were cultured as previously described and treated with increasing concentrations of Syn9048 and mouse α-synuclein PFFs simultaneously. Quantitation of pS129 α-synuclein area / neuron number normalized to IgG treatment is shown. Increasing molar ratios of Syn9048 dose-dependently reduced pS129 α-synuclein pathology, with a sigmoidal dose curve fit yielding an IC50 of 0.004 (R2 = 0.8026). N = 3 / condition. Error bars represent standard error, and shaded areas represent the 95% confidence interval of the sigmoidal fit. DETAILED DESCRIPTION OF THE INVENTION

[0013] Detailed Description of the Disclosure definition Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those commonly understood by those of ordinary skill in the art to which this disclosure pertains.Although any method and material similar or equivalent to those described herein can be used in carrying out the test of this disclosure, exemplary materials and methods are described herein.In describing and claiming this disclosure, the following terminology is used.It should also be understood that the terminology used herein is only for describing specific embodiments and is not intended to be limiting.

[0014] As used herein, each of the following terms has the meaning associated with it in this section.

[0015] Unless otherwise defined, all technical and scientific terms used herein generally have the same meaning as commonly understood by those of ordinary skill in the art to which this disclosure belongs. Generally, the nomenclature used herein and the laboratory procedures in cell culture, molecular genetics, analytical chemistry, immunology, nucleic acid chemistry and hybridization are well known and commonly used in the art. Standard techniques or modifications thereof are used for chemical synthesis and chemical analysis.

[0016] The articles "a" and "an" are used herein to refer to one or to more than one (i.e., to at least one) of the grammatical object of the article. By way of example, "an element" means one element or more than one element.

[0017] As used herein, the term "alpha-synuclein" or "alpha-Syn" or "alpha-syn" refers to a protein that is expressed primarily in brain tissue and is located primarily at the presynaptic terminals of neurons. In certain aspects, the present disclosure provides a protein having the sequence SEQ ID NO:245: Assumes human α-Syn with TIFF0007777071000001.tif26128.

[0018] As used herein, the term "α-Syn" refers to total α-Syn, α-Syn monomers, α-Syn fibrils, and / or α-Syn oligomers. As used herein, α-Syn oligomers refer to any multimeric assembly of α-Syn containing two or more α-Syn monomers.

[0019] As used herein, the term "about" is understood by those skilled in the art and varies to some extent depending on the context in which it is used.As used herein, when referring to a measurable value such as an amount, concentration, length of time, etc., the term "about" means to encompass a variation of ±20% or ±10%, more preferably ±5%, even more preferably ±1%, and even more preferably ±0.1% from the specified value, because such a variation is appropriate for carrying out the disclosed method.

[0020] As used herein, the term "affinity" of one molecule for another refers to the degree (or tightness) of binding between two molecules. Higher affinity means tighter binding between two molecules. Affinity is measured by the dissociation constant (or K d ) and has a lower magnitude (closer to zero) K d A higher value indicates a higher affinity.

[0021] As used herein, "amino acid" is meant to include both natural and synthetic amino acids, and both D and L amino acids. "Standard amino acid" refers to any of the 20 L-amino acids commonly found in naturally occurring peptides. "Non-standard amino acid residue" refers to any amino acid other than the standard amino acids, whether prepared synthetically or derived from a natural source. As used herein, "synthetic amino acid" also encompasses chemically modified amino acids, including, but not limited to, salts, amino acid derivatives (such as amides), and substitutions. Amino acids contained within peptides, particularly at the carboxy or amino termini, can be modified by methylation, amidation, acetylation, or substitution with other chemical groups that can alter the circulating half-life of the peptide without adversely affecting its activity. In addition, disulfide linkages may or may not be present in the peptide.

[0022] The term "antibody," as used herein, refers to an immunoglobulin molecule capable of specifically binding to a specific epitope on an antigen. Antibodies can be intact immunoglobulins derived from natural or recombinant sources, or can be immunoreactive portions of intact immunoglobulins. Antibodies are typically tetramers of immunoglobulin molecules. Antibodies in the present disclosure can exist in a variety of forms, including, for example, polyclonal antibodies, monoclonal antibodies, intracellular antibodies ("intrabodies"), Fv, Fab and F(ab)2, as well as single-chain antibodies (scFv), camelid antibodies and humanized antibodies (Harlow et al., 1999, Using Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory Press, NY; Harlow et al., 1989, Antibodies: A Laboratory Manual, Cold Spring Harbor, New York; Houston et al., 1988, Proc. Natl. Acad. Sci. USA 85:5879-5883; Bird et al., 1988, Science 242:423-426). As used herein, a "neutralizing antibody" is an immunoglobulin molecule that binds to an antigen and blocks its biological activity.

[0023] The term "antigen" or "Ag," as used herein, is defined as a molecule that elicits an immune response. This immune response may involve either antibody production or activation of specific immunocompetent cells, or both. Those skilled in the art will understand that virtually any macromolecule, including any protein or peptide, can function as an antigen. Furthermore, antigens can be derived from recombinant or genomic DNA. Therefore, those skilled in the art will understand that any DNA containing a nucleotide sequence or partial nucleotide sequence that encodes a protein that elicits an immune response encodes an "antigen," as that term is used herein. Furthermore, those skilled in the art will understand that an antigen need not be encoded by only one full-length nucleotide sequence of a gene. It will be readily apparent that the present disclosure includes, but is not limited to, the use of partial nucleotide sequences of multiple genes, and that these nucleotide sequences may be arranged in various combinations to elicit a desired immune response. Moreover, those skilled in the art will understand that an antigen need not be encoded by a "gene" at all. It will be readily apparent that an antigen may be generated or synthesized, or may be derived from a biological sample. Such biological samples may include, but are not limited to, tissue samples, tumor samples, cells, or biological fluids.

[0024] The "coding region" of a gene consists of nucleotide residues on the coding strand of the gene and nucleotides on the non-coding strand of the gene that are homologous or complementary, respectively, to the coding region of an mRNA molecule produced by transcription of the gene.

[0025] Additionally, the "coding region" of an mRNA molecule consists of the nucleotide residues of the mRNA molecule that coincide with the anticodon region of a transfer RNA molecule or encode a stop codon during translation of the mRNA molecule. Thus, the coding region may include nucleotide residues corresponding to amino acid residues not present in the mature protein encoded by the mRNA molecule (e.g., amino acid residues in a protein export signal sequence).

[0026] "Complementary," as used herein to refer to nucleic acids, refers to the broad concept of sequence complementarity between regions of two nucleic acid strands or between two regions of the same nucleic acid strand. It is known that an adenine residue in a first nucleic acid strand can form specific hydrogen bonds ("base pairing") with a residue in a second nucleic acid strand antiparallel to the first strand if that residue is thymine or uracil. Similarly, it is known that a cytosine residue in a first nucleic acid strand can base pair with a residue in a second nucleic acid strand antiparallel to the first strand if that residue is guanine. A first region of a nucleic acid is complementary to a second region of the same or different nucleic acid if at least one nucleotide residue in the first region can base pair with a residue in the second region when the two regions are arranged in an antiparallel fashion. Preferably, the first region comprises the first portion and the second region comprises the second portion such that when the first and second portions are arranged in an antiparallel manner, at least about 50%, preferably at least about 75%, at least about 90%, or at least about 95% of the nucleotide residues in the first portion are capable of base pairing with nucleotide residues in the second portion. More preferably, all nucleotide residues in the first portion are capable of base pairing with nucleotide residues in the second portion.

[0027] The term "delivery vehicle" is used herein as a generic term for any delivery vehicle capable of delivering a compound to a subject, including, but not limited to, dermal and transdermal delivery vehicles.

[0028] The term "DNA" as used herein is defined as deoxyribonucleic acid.

[0029] "Effective amount" or "therapeutically effective amount" are used interchangeably herein and refer to an amount of a compound, product, material, or composition as described herein that is effective to achieve a particular biological result. Such result may include, but is not limited to, the treatment of a disease or condition as determined by any means suitable in the art.

[0030] "Encode" refers to the inherent property of a particular nucleotide sequence in a polynucleotide, such as a gene, cDNA, or mRNA, to serve as a template for the synthesis of other polymers and macromolecules having either a given nucleotide sequence (i.e., rRNA, tRNA, and mRNA) or a given amino acid sequence in biological processes, and the biological properties that result therefrom. Thus, a gene encodes a protein if transcription and translation of its corresponding mRNA produces the protein in a cell or other biological system. Both the coding strand, whose nucleotide sequence is identical to the mRNA sequence and is usually provided in a sequence listing, and the non-coding strand, which is used as a template for transcription of the gene or cDNA, can be said to encode the protein or other product of the gene or cDNA.

[0031] As used herein, the term "fragment," as applied to a protein or peptide, refers to a subsequence of a larger protein or peptide. A "fragment" of a protein or peptide can be at least about 20 amino acids in length; e.g., at least about 50 amino acids in length; at least about 100 amino acids in length, at least about 200 amino acids in length, at least about 300 amino acids in length, and at least about 400 amino acids in length (and any integer value therebetween). As used herein, an antibody fragment refers to an active fragment thereof, i.e., a fragment having the same properties used to define an antibody according to the present disclosure, in certain embodiments, a high affinity for α-syn fibrils (composed of misfolded α-syn) and a low or high binding affinity for α-syn monomers. For convenience, when the term antibody is used, fragments thereof exhibiting the same properties are also contemplated.

[0032] As used herein, the term "fragment," as applied to nucleic acids, refers to a subsequence of a larger nucleic acid. A "fragment" of a nucleic acid can be at least about 15 nucleotides in length; e.g., at least about 50 nucleotides to about 100 nucleotides; at least about 100 to about 500 nucleotides, at least about 500 to about 1000 nucleotides, at least about 1000 nucleotides to about 1500 nucleotides; or about 1500 nucleotides to about 2500 nucleotides; or about 2500 nucleotides (and any integer value therebetween).

[0033] Conventional notation is used herein to describe polynucleotide sequences: the left-hand end of a single-stranded polynucleotide sequence is the 5' end; the left-hand direction of a double-stranded polynucleotide sequence is referred to as the 5' direction.

[0034] The direction of 5' to 3' addition of nucleotides to the nascent RNA transcript is referred to as the transcription direction. The DNA strand with the same sequence as the mRNA is referred to as the "coding strand;" the sequence on the DNA strand that is 5' of the reference point on the DNA is referred to as the "upstream sequence;" the sequence on the DNA strand that is 3' of the reference point on the DNA is referred to as the "downstream sequence."

[0035] An "individual," "patient," or "subject," as the term is used herein, includes members of any animal species, including, but not limited to, birds, humans and other primates, and other mammals, including commercially relevant mammals such as cattle, pigs, horses, sheep, cats, and dogs. Preferably, the subject is a human.

[0036] As used herein, "instructions" includes publications, records, diagrams, or any other medium of expression that can be used to communicate the usefulness of the disclosed compositions and compounds in the kit. The instructions for the kit may, for example, be attached to a container containing the disclosed compounds and / or compositions, or may be shipped together with the container containing the compounds and / or compositions. Alternatively, the instructions may be shipped separately from the container, with the intention that the instructions and the compounds will be used together by the recipient. Delivery of the instructions may be, for example, by physical delivery of a publication or other medium of expression that communicates the usefulness of the kit, or may be achieved by electronic transmission, for example, by computer, for example, by email, or by downloading from a website.

[0037] "Isolated" means altered or removed from the natural state. For example, a nucleic acid or peptide naturally occurring in a living animal is not "isolated," but the same nucleic acid or peptide partially or completely separated from the coexisting materials of its natural state is "isolated." An isolated nucleic acid or protein may exist in a substantially purified form, or it may exist in a non-native environment, such as, for example, in a host cell.

[0038] An "isolated nucleic acid" refers to a nucleic acid segment or fragment that has been separated from adjacent sequences as it naturally occurs, i.e., a DNA fragment that has been removed from sequences that normally flank the fragment, i.e., from sequences that flank the fragment in the genome in which it naturally occurs. The term also applies to nucleic acids that have been substantially purified from other components that naturally accompany the nucleic acid, i.e., RNA or DNA, or protein, with which it is naturally associated in a cell. Thus, the term includes recombinant DNA that is incorporated into, for example, a vector, an autonomously replicating plasmid, or virus, or into the genomic DNA of a prokaryote or eukaryote, or that exists as a separate molecule free from other sequences (i.e., cDNA, or genomic or cDNA fragments produced by PCR or restriction enzyme digestion). It also includes recombinant DNA that is part of a hybrid gene that encodes additional polypeptide sequences.

[0039] "Nucleic acid" refers to any nucleic acid, whether composed of deoxyribonucleosides or ribonucleosides, and whether composed of phosphodiester or modified linkages, such as phosphotriester, phosphoramidate, siloxane, carbonate, carboxymethyl ester, acetamidate, carbamate, thioether, bridged phosphoramidate, bridged methylene phosphonate, phosphorothioate, methylphosphonate, phosphorodithioate, bridged phosphorothioate, or sulfone linkages, and combinations of such linkages. The term nucleic acid also specifically includes nucleic acids composed of bases other than the five biologically occurring bases (adenine, guanine, thymine, cytosine, and uracil).

[0040] Unless otherwise specified, a "nucleotide sequence encoding an amino acid sequence" includes all nucleotide sequences that are degenerate versions of each other and encode the same amino acid sequence. The phrase "nucleotide sequence encoding a protein or RNA" can also include introns, to the extent that a nucleotide sequence encoding a protein may contain introns in some forms.

[0041] The term "oligonucleotide" typically refers to short polynucleotides, generally less than about 60 nucleotides. Where a nucleotide sequence is represented by a DNA sequence (i.e., A, T, G, C), it is understood to also include RNA sequences in which "U" replaces "T" (i.e., A, U, G, C).

[0042] As used herein, the term "pharmaceutical composition" refers to a mixture of at least one compound of the present disclosure with other chemical components, such as carriers, stabilizers, diluents, dispersants, suspending agents, thickeners, and / or excipients. Pharmaceutical compositions facilitate the administration of compounds to living organisms. Multiple techniques for administering compounds exist in the art, including, but not limited to, intravenous, oral, aerosol, parenteral, ocular, pulmonary, and topical administration.

[0043] "Pharmaceutically acceptable" refers to properties and / or substances that are acceptable to patients from a pharmacological / toxicological standpoint and to pharmacologists from a physical / chemical standpoint with respect to composition, formulation, stability, patient acceptability, and bioavailability. A "pharmaceutically acceptable carrier" refers to a medium that does not interfere with the effectiveness of the biological activity of the active ingredient and that is not toxic to the host to which it is administered.

[0044] The term "polynucleotide," as used herein, is defined as a chain of nucleotides. Furthermore, nucleic acids are polymers of nucleotides. Therefore, nucleic acids and polynucleotides are interchangeable when used herein. Those skilled in the art have the general knowledge that nucleic acids are polynucleotides, which can be hydrolyzed into monomeric "nucleotides." Monomeric nucleotides can be hydrolyzed into nucleosides. As used herein, polynucleotides include, but are not limited to, all nucleic acid sequences obtained by any means available in the art, including, but not limited to, recombinant means, i.e., cloning nucleic acid sequences from recombinant libraries or cell genomes using conventional cloning techniques and PCR™, and by synthetic means.

[0045] As used herein, the terms "protein," "peptide," and "polypeptide" are used interchangeably and refer to compounds containing amino acid residues covalently linked by peptide bonds. The term "peptide bond" refers to a covalent amide linkage formed by the loss of a water molecule between the carboxyl group of one amino acid and the amino group of a second amino acid. A protein or peptide must contain at least two amino acids, and there is no limit to the maximum number of amino acids that may comprise a protein or peptide sequence. A polypeptide includes any peptide or protein containing two or more amino acids connected to each other by peptide bonds. As used herein, the term refers to both short chains, commonly referred to in the art as peptides, oligopeptides, and oligomers, and longer chains, commonly referred to in the art as proteins, of which there are many types. "Protein" includes, for example, biologically active fragments, substantially homologous proteins, oligopeptides, homodimers, heterodimers, protein variants, modified proteins, derivatives, analogs, and fusion proteins, among others. Proteins include natural proteins, recombinant proteins, synthetic proteins, or combinations thereof.

[0046] The term "recombinant DNA," as used herein, is defined as DNA that is produced by joining pieces of DNA from different sources.

[0047] The term "recombinant polypeptide," as used herein, is defined as a polypeptide produced by using recombinant DNA methods.

[0048] The term "RNA" as used herein is defined as ribonucleic acid.

[0049] The term "therapeutic" as used herein means treatment and / or prophylaxis.

[0050] The term "treat," as used herein, means to reduce the frequency of symptoms experienced by a subject, or to administer an agent or compound to reduce the frequency and / or severity of symptoms. As used herein, "alleviate" is used interchangeably with the term "treat."

[0051] As used herein, "treating a disease, disorder, or condition" means reducing the frequency or severity of symptoms of the disease, disorder, or condition experienced by a subject. Treating a disease, disorder, or condition may or may not include complete eradication or elimination of symptoms.

[0052] The following abbreviations are used herein: BBB, blood-brain barrier; CDR, complementarity-determining region; DLB, dementia with Lewy bodies; FL, full length; LB, Lewy bodies; LN, Lewy neurites; MSA, multiple system atrophy; NAC, non-amyloid component; PD, Parkinson's disease; PDD, Parkinson's disease dementia; PFF, preformed fibrils; VH, heavy chain variable region; VL, light chain variable region.

[0053] Ranges: Throughout this disclosure, various aspects of the present disclosure may be presented in a range format. It should be understood that the description in range format is merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of the disclosure. Accordingly, the description of a range should be considered to have specifically disclosed all possible subranges as well as individual numerical values ​​within that range. For example, description of a range such as 1 to 6 should be considered to have specifically disclosed subranges such as 1 to 3, 1 to 4, 1 to 5, 2 to 4, 2 to 6, 3 to 6, etc., as well as individual numbers within that range, e.g., 1, 2, 2.7, 3, 4, 5, 5.3, and 6. This applies regardless of the breadth of the range.

[0054] explanation The present disclosure generally relates to certain monoclonal (mouse) antibodies, or fragments thereof, that recognize certain pathogenic forms of α-Syn, i.e., the protein misfolded in Parkinson's disease (PD) and related neurodegenerative disorders known as synucleinopathies. PD and dementia with Lewy bodies (DLB) are progressive neurodegenerative diseases for which no disease-modifying treatments exist. PD and DLB are characterized by the aggregation of a synaptic protein called α-synuclein, and compelling evidence suggests that the progression of these diseases is associated with the transcellular spread of pathogenic α-synuclein in the brains of affected individuals. Therefore, treatments targeting extracellular pathogenic α-synuclein have the potential to slow or halt disease progression.

[0055] At the start of this investigation, it was uncertain what desirable properties a therapeutic α-synuclein antibody would have. For example, targeting all α-synuclein species with a pan-α-synuclein antibody could have adverse effects in the central nervous system or in the blood, where α-synuclein is abundant in red blood cells. Furthermore, binding of a pan-α-synuclein antibody to a nonpathogenic form of the protein would reduce the antibody available to bind to misfolded species.

[0056] Previous studies have identified several antibodies that are effective in reducing α-synuclein aggregates in animal models of PD. These antibodies have different selectivity profiles (α-synuclein oligomers, C-terminally truncated α-synuclein, or fibrillar α-synuclein), and it is currently unclear whether antibodies directed against certain epitopes are more effective than others. Given that it is generally accepted that only approximately 0.1% of administered antibodies cross the BBB, it would likely be beneficial to utilize α-synuclein antibodies that preferentially bind to pathogenic species present in the brain rather than α-synuclein monomers. This would increase the effective concentration of antibodies that can target the α-synuclein that promotes disease progression without losing antibody to unproductive binding to monomers.

[0057] In a non-limiting aspect, this study aimed to identify antibodies highly selective for pathogenic misfolded α-synuclein. Mice were first immunized with misfolded α-synuclein to generate a library of B cell hybridomas. Antibodies produced by clonal hybridoma cultures were then subjected to a series of screening assays to select antibodies that exhibited high selectivity for misfolded α-synuclein and were able to inhibit the formation of LB-like structures in a primary neuron model of α-synucleinopathy. An exemplary candidate antibody, Syn9048, was tested for efficacy in a wild-type mouse model of pathological α-synuclein transmission. Six months of chronic administration of Syn9048 was well tolerated by mice, and Syn9048 was able to maintain striatal dopamine levels and reduce α-synuclein pathology, particularly in areas where α-synuclein inclusions likely arose from transcellular transmission of pathogenic α-synuclein. Across all measures, Syn9048 demonstrated improved efficacy over previously validated alpha-synuclein antibodies (Tran, et al., 2014, Cell Reports 7:2054-2065), indicating that Syn9048 has desirable properties for treating, ameliorating, and / or preventing the diseases contemplated herein. This study highlights the therapeutic potential of alpha-synuclein immunotherapy for the treatment of PD and DLB and provides a framework for screening alpha-synuclein antibodies to identify those with desirable properties.

[0058] In certain embodiments, the antibodies of the present disclosure are highly selective for pathogenic forms of alpha-synuclein and can be administered as therapeutic agents targeting such pathogenic protein forms. In certain embodiments, the antibodies of the present disclosure recognize a conformational epitope comprising amino acids 110-120 and / or 120-130 in combination with other regions of alpha-Syn.

[0059] In certain embodiments, antibodies of the present disclosure exhibit preferential binding to pathological forms of α-Syn (i.e., fibrils and / or oligomers) compared to native (i.e., monomeric) forms. In other embodiments, antibodies of the present disclosure reduce the formation of pathological α-Syn inclusions / fibrils that normally form in cultured neurons exposed to recombinant α-Syn fibrils. In yet other embodiments, antibodies of the present disclosure detect pathological α-Syn fibrils. In yet other embodiments, antibodies of the present disclosure are used as therapeutics to reduce the development / propagation of pathological α-Syn fibrils and / or oligomers in synucleinopathies. In yet other embodiments, antibodies of the present disclosure do not cross-react with Tau and / or β-amyloid proteins.

[0060] In certain embodiments, the antibodies of the present disclosure bind to α-Syn fibrils and / or oligomers at a concentration of about 10 -6 M, about 10 -7 M, about 10 -8 M, about 10 -9 M, about 10 -10 M, or about 10 -11 A dissociation constant K equal to or lower than M d In other embodiments, antibodies of the present disclosure bind to α-Syn monomers with an affinity that is at least about 10-fold, 30-fold, 100-fold, 300-fold, or 1000-fold lower than the affinity of the antibody for α-Syn fibrils. In yet other embodiments, antibodies of the present disclosure bind to α-Syn monomers with an affinity that is at least about 10-fold, 30-fold, 100-fold, 300-fold, or 1000-fold lower than the affinity of the antibody for α-Syn fibrils. -10 M, about 10 -9 M, about 10 -8 M, about 10 -7 M, about 10 -6 M, about 10 -5 M, about 10 -4 M, or about 10 -3 A dissociation constant K equal to or higher than M d In yet other embodiments, the antibodies of the present disclosure bind to α-Syn fibrils and monomers with approximately equal affinity. In yet other embodiments, the antibodies of the present disclosure bind to α-Syn fibrils, oligomers, and monomers with approximately equal affinity, at a binding affinity of about 10 -7 M, about 10 -8M, about 10 -9 M, about 10 -10 M, or about 10 -11 A dissociation constant K equal to or lower than M d The binding affinity of an antibody can be determined using a variety of art-recognized methods, including but not limited to those described elsewhere herein, such as isothermal titration calorimetry, surface plasmon resonance, immunoassays such as ELISA or RIA.

[0061] Compositions containing antibodies In one aspect, the present disclosure includes an isolated monoclonal antibody that selectively binds to α-Syn in fibrillar and / or oligomeric conformations and / or binds with high affinity to both soluble oligomeric and fibrillar α-Syn. In certain embodiments, the antibody comprises a heavy chain. In other embodiments, the heavy chain comprises three complementarity-determining regions (CDRs), i.e., CDR1, CDR2, and CDR3. In yet other embodiments, the light chain comprises three complementarity-determining regions (CDRs), i.e., CDR1, CDR2, and CDR3.

[0062] In one particular embodiment, the monoclonal antibody (designated herein as 9003) comprises light and heavy variable chains having the sequences shown below. TIFF0007777071000002.tif233159

[0063] In one particular embodiment, the monoclonal antibody (designated herein as 9004) comprises light and heavy variable chains having the sequences shown below. TIFF0007777071000003.tif234159

[0064] In one particular embodiment, the monoclonal antibody (designated herein as 9005) comprises light and heavy variable chains having the sequences shown below. TIFF0007777071000004.tif233159

[0065] In one particular embodiment, the monoclonal antibody (designated herein as 9009) comprises light and heavy variable chains having the sequences shown below. TIFF0007777071000005.tif233159

[0066] In one particular embodiment, the monoclonal antibody (designated herein as 9014) comprises light and heavy variable chains having the sequences shown below. TIFF0007777071000006.tif234159

[0067] In one particular embodiment, the monoclonal antibody (designated herein as 9018) comprises light and heavy variable chains having the sequences shown below. TIFF0007777071000007.tif234159

[0068] In one particular embodiment, the monoclonal antibody (designated herein as 9021) comprises light and heavy variable chains having the sequences shown below. TIFF0007777071000008.tif233159

[0069] In one particular embodiment, the monoclonal antibody (designated herein as 9023) comprises light and heavy variable chains with the sequences shown below. Individual positive clones with the correct VH and VL insert sizes were sequenced. One VH DNA sequence and two VL DNA sequences were obtained in the study. TIFF0007777071000009.tif227159TIFF0007777071000010.tif115157

[0070] In one particular embodiment, the monoclonal antibody (designated herein as 9060) comprises light and heavy variable chains having the sequences shown below. TIFF0007777071000011.tif235159

[0071] In one particular embodiment, the monoclonal antibody (designated herein as 9064) comprises light and heavy variable chains having the sequences shown below. TIFF0007777071000012.tif234159

[0072] In one particular embodiment, the monoclonal antibody (designated herein as 9071) comprises light and heavy variable chains having the sequences shown below. TIFF0007777071000013.tif235159

[0073] In one particular embodiment, the monoclonal antibody (designated herein as 9096) comprises light and heavy variable chains having the sequences shown below. TIFF0007777071000014.tif234157

[0074] In one particular embodiment, the monoclonal antibody (designated herein as 9110) comprises light and heavy variable chains having the sequences shown below. TIFF0007777071000015.tif234159

[0075] In one particular embodiment, the monoclonal antibody (designated herein as 9035) comprises light and heavy variable chains having the sequences shown below. TIFF0007777071000016.tif234157

[0076] In one particular embodiment, the monoclonal antibody (designated herein as 9047) comprises light and heavy variable chains having the sequences shown below. TIFF0007777071000017.tif235157

[0077] In one particular embodiment, the monoclonal antibody (designated herein as 9052) comprises light and heavy variable chains having the sequences shown below. TIFF0007777071000018.tif233157

[0078] In one particular embodiment, the monoclonal antibody (designated herein as 9061) comprises light and heavy variable chains with the sequences shown below. Individual positive clones with the correct VH and VL insert sizes were sequenced. One VH DNA sequence and two VL DNA sequences were obtained in the study. TIFF0007777071000019.tif233157TIFF0007777071000020.tif110157

[0079] In one particular embodiment, the monoclonal antibody (designated herein as 9092) comprises light and heavy variable chains having the sequences shown below. TIFF0007777071000021.tif235157

[0080] In one particular embodiment, the monoclonal antibody (designated herein as 9099) comprises light and heavy variable chains having the sequences shown below. TIFF0007777071000022.tif234157

[0081] In one particular embodiment, the monoclonal antibody (designated herein as 9100) comprises light and heavy variable chains having the sequences shown below. TIFF0007777071000023.tif234157

[0082] In certain embodiments, the isolated monoclonal antibody comprises a light chain variable region (VL) and a heavy chain variable region (VH). In certain embodiments, the VL comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO: 27, 46, 70, 114, 126, 185, 213, or 240. In certain embodiments, the VL comprises a CDR2 region comprising the amino acid sequence of SEQ ID NO: 29, 72, 116, 214, or 242. In certain embodiments, the VL comprises a CDR3 region comprising the amino acid sequence of SEQ ID NO: 31, 85, 118, 129, 152, 205, 216, or 244. In certain embodiments, the VH comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO: 11, 58, 92, 101, 144, 160, or 226. In certain embodiments, the VH comprises a CDR2 region comprising the amino acid sequence of SEQ ID NO: 13, 40, 60, 93, 102, 134, 145, 161, 200, or 228. In certain embodiments, the VH comprises a CDR3 region comprising the amino acid sequence of SEQ ID NO: 15, 62, 81, 147, 163, 194, 201, or 230.

[0083] In certain embodiments, the VL comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:27, 46, 70, 114, or 126.

[0084] In certain embodiments, the VL comprises a CDR2 region comprising the amino acid sequence of SEQ ID NO:29, 72, or 116.

[0085] In certain embodiments, the VL comprises a CDR3 region comprising the amino acid sequence of SEQ ID NO:31, 85, 118, 129, or 152.

[0086] In certain embodiments, the VL comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:70, 185, 213, or 240.

[0087] In certain embodiments, the VL comprises a CDR2 region comprising the amino acid sequence of SEQ ID NO:29, 72, 214, or 242.

[0088] In certain embodiments, the VL comprises a CDR3 region comprising the amino acid sequence of SEQ ID NO:152, 129, 205, 216, or 244.

[0089] In certain embodiments, the VL comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:27; a CDR2 region comprising the amino acid sequence of SEQ ID NO:29; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:31.

[0090] In certain embodiments, the VL comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:46; a CDR2 region comprising the amino acid sequence of SEQ ID NO:29; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:31.

[0091] In certain embodiments, the VL comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:70; a CDR2 region comprising the amino acid sequence of SEQ ID NO:29; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:152.

[0092] In certain embodiments, the VL comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:70; a CDR2 region comprising the amino acid sequence of SEQ ID NO:72; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:85.

[0093] In certain embodiments, the VL comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:70; a CDR2 region comprising the amino acid sequence of SEQ ID NO:72; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:205.

[0094] In certain embodiments, the VL comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:114; a CDR2 region comprising the amino acid sequence of SEQ ID NO:116; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:118.

[0095] In certain embodiments, the VL comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:126; a CDR2 region comprising the amino acid sequence of SEQ ID NO:72; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:129.

[0096] In certain embodiments, the VL comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:185; a CDR2 region comprising the amino acid sequence of SEQ ID NO:29; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:129.

[0097] In certain embodiments, the VL comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:213; a CDR2 region comprising the amino acid sequence of SEQ ID NO:214; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:216.

[0098] In certain embodiments, the VL comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:240; a CDR2 region comprising the amino acid sequence of SEQ ID NO:242; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:244.

[0099] In certain embodiments, the VH comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:11, 58, 92, 101, or 144.

[0100] In certain embodiments, the VH comprises a CDR2 region comprising the amino acid sequence of SEQ ID NO: 13, 40, 60, 93, 102, 134, or 145.

[0101] In certain embodiments, the VH comprises a CDR3 region comprising the amino acid sequence of SEQ ID NO: 15, 62, 81, or 147.

[0102] In certain embodiments, the VH comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:160, 58, or 226.

[0103] In certain embodiments, the VH comprises a CDR2 region comprising the amino acid sequence of SEQ ID NO:161, 145, 200, or 228.

[0104] In certain embodiments, the VH comprises a CDR3 region comprising the amino acid sequence of SEQ ID NO:163, 194, 201, or 230.

[0105] In certain embodiments, the VH comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:11; a CDR2 region comprising the amino acid sequence of SEQ ID NO:13; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:15.

[0106] In certain embodiments, the VH comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:11; a CDR2 region comprising the amino acid sequence of SEQ ID NO:40; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:15.

[0107] In certain embodiments, the VH comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:11; a CDR2 region comprising the amino acid sequence of SEQ ID NO:134; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:15.

[0108] In certain embodiments, the VH comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:58; a CDR2 region comprising the amino acid sequence of SEQ ID NO:60; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:62.

[0109] In certain embodiments, the VH comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:58; a CDR2 region comprising the amino acid sequence of SEQ ID NO:60; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:81.

[0110] In certain embodiments, the VH comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:58; a CDR2 region comprising the amino acid sequence of SEQ ID NO:200; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:201.

[0111] In certain embodiments, the VH comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:92; a CDR2 region comprising the amino acid sequence of SEQ ID NO:93; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:62.

[0112] In certain embodiments, the VH comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:101; a CDR2 region comprising the amino acid sequence of SEQ ID NO:102; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:62.

[0113] In certain embodiments, the VH comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:144; a CDR2 region comprising the amino acid sequence of SEQ ID NO:145; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:147.

[0114] In certain embodiments, the VH comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:160; a CDR2 region comprising the amino acid sequence of SEQ ID NO:145; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:163.

[0115] In certain embodiments, the VH comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:160; a CDR2 region comprising the amino acid sequence of SEQ ID NO:145; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:194.

[0116] In certain embodiments, the VH comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:160; a CDR2 region comprising the amino acid sequence of SEQ ID NO:161; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:163.

[0117] In certain embodiments, the VH comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO:226; a CDR2 region comprising the amino acid sequence of SEQ ID NO:228; and a CDR3 region comprising the amino acid sequence of SEQ ID NO:230.

[0118] In certain embodiments, the VL is selected from the group consisting of SEQ ID NO:27-SEQ ID NO:28-SEQ ID NO:29-SEQ ID NO:30-SEQ ID NO:31, SEQ ID NO:46-SEQ ID NO:28-SEQ ID NO:29-SEQ ID NO:30-SEQ ID NO:31, SEQ ID NO:70-SEQ ID NO:71-SEQ ID NO:72-SEQ ID NO:73-SEQ ID NO:85, SEQ ID NO:70-SEQ ID NO:127-SEQ ID NO:29-SEQ ID NO:151-SEQ ID NO:152, SEQ ID NO:70-SEQ ID NO:127-SEQ ID NO:29-SEQ ID NO:172-SEQ ID NO:152, SEQ ID NO:70-SEQ ID NO:127-SEQ ID NO:72-SEQ ID NO: SEQ ID NO:204-SEQ ID NO:205, SEQ ID NO:114-SEQ ID NO:115-SEQ ID NO:116-SEQ ID NO:117-SEQ ID NO:118, SEQ ID NO:126-SEQ ID NO:127-SEQ ID NO:72-SEQ ID NO:128-SEQ ID NO:129, SEQ ID NO:185-SEQ ID NO:186-SEQ ID NO:29-SEQ ID NO:187-SEQ ID NO:129, SEQ ID NO:213-SEQ ID NO:115-SEQ ID NO:214-SEQ ID NO:215-SEQ ID NO:216, or SEQ ID NO:240-SEQ ID NO:241-SEQ ID NO:242-SEQ ID NO:243-SEQ ID NO:244.

[0119] In certain aspects, the VH is selected from the group consisting of SEQ ID NO:11-SEQ ID NO:12-SEQ ID NO:13-SEQ ID NO:14-SEQ ID NO:15, SEQ ID NO:11-SEQ ID NO:12-SEQ ID NO:40-SEQ ID NO:41-SEQ ID NO:15, SEQ ID NO:11-SEQ ID NO:12-SEQ ID NO:134-SEQ ID NO:135-SEQ ID NO:15, SEQ ID NO:58-SEQ ID NO:59-SEQ ID NO:60-SEQ ID NO:61-SEQ ID NO:62, SEQ ID NO:58-SEQ ID NO:59-SEQ ID NO:60-SEQ ID NO:80-SEQ ID NO:81, SEQ ID NO:58-SEQ ID NO:59-SEQ ID NO:200-SEQ ID NO:94-SEQ ID NO:201, SEQ ID NO:92-SEQ ID NO:59-SEQ ID NO:93-SEQ ID NO:94-SEQ ID NO:62, SEQ ID NO:101-SEQ ID NO:59-SEQ ID NO:102-SEQ ID NO:103-SEQ ID NO:62, SEQ ID NO:144-SEQ ID NO:59-SEQ ID NO:145-SEQ ID NO:146-SEQ ID NO:147, SEQ ID NO:160-SEQ ID NO:59-SEQ ID NO:145-SEQ ID NO:177-SEQ ID NO:163, SEQ ID NO:160-SEQ ID NO:59-SEQ ID NO:145-SEQ ID NO:193-SEQ ID NO:194, SEQ ID NO:160-SEQ ID NO:59-SEQ ID It includes the amino acid sequence of SEQ ID NO:161-SEQ ID NO:162-SEQ ID NO:163, or SEQ ID NO:226-SEQ ID NO:227-SEQ ID NO:228-SEQ ID NO:229-SEQ ID NO:230.

[0120] In certain embodiments, the VL is selected from the group consisting of SEQ ID NO:27-SEQ ID NO:28-SEQ ID NO:29-SEQ ID NO:30-SEQ ID NO:31, SEQ ID NO:46-SEQ ID NO:28-SEQ ID NO:29-SEQ ID NO:30-SEQ ID NO:31, SEQ ID NO:70-SEQ ID NO:71-SEQ ID NO:72-SEQ ID NO:73-SEQ ID NO:85, SEQ ID NO:70-SEQ ID NO:127-SEQ ID NO:29-SEQ ID NO:151-SEQ ID NO:152, SEQ ID NO:114-SEQ ID NO:115-SEQ ID NO:116-SEQ ID NO:117-SEQ ID NO:118, or SEQ ID NO:126-SEQ ID NO:127-SEQ ID NO:72-SEQ Contains the amino acid sequences of SEQ ID NO:128-SEQ ID NO:129.

[0121] In certain aspects, the VH is selected from the group consisting of SEQ ID NO:11-SEQ ID NO:12-SEQ ID NO:13-SEQ ID NO:14-SEQ ID NO:15, SEQ ID NO:11-SEQ ID NO:12-SEQ ID NO:40-SEQ ID NO:41-SEQ ID NO:15, SEQ ID NO:11-SEQ ID NO:12-SEQ ID NO:134-SEQ ID NO:135-SEQ ID NO:15, SEQ ID NO:58-SEQ ID NO:59-SEQ ID NO:60-SEQ ID NO:61-SEQ ID NO:62, SEQ ID NO:58-SEQ ID NO:59-SEQ ID NO:60-SEQ ID NO:80-SEQ ID NO:81, SEQ ID NO:92-SEQ ID NO:59-SEQ ID NO:93-SEQ ID NO:94-SEQ ID NO: NO:62, SEQ ID NO:101-SEQ ID NO:59-SEQ ID NO:102-SEQ ID NO:103-SEQ ID NO:62, or SEQ ID NO:144-SEQ ID NO:59-SEQ ID NO:145-SEQ ID NO:146-SEQ ID NO:147.

[0122] In certain embodiments, the VL comprises the amino acid sequence of SEQ ID NO:70-SEQ ID NO:127-SEQ ID NO:29-SEQ ID NO:172-SEQ ID NO:152, SEQ ID NO:70-SEQ ID NO:127-SEQ ID NO:72-SEQ ID NO:204-SEQ ID NO:205, SEQ ID NO:185-SEQ ID NO:186-SEQ ID NO:29-SEQ ID NO:187-SEQ ID NO:129, SEQ ID NO:213-SEQ ID NO:115-SEQ ID NO:214-SEQ ID NO:215-SEQ ID NO:216, or SEQ ID NO:240-SEQ ID NO:241-SEQ ID NO:242-SEQ ID NO:243-SEQ ID NO:244.

[0123] In certain embodiments, the VH comprises the amino acid sequence of SEQ ID NO:58-SEQ ID NO:59-SEQ ID NO:200-SEQ ID NO:94-SEQ ID NO:201, SEQ ID NO:160-SEQ ID NO:59-SEQ ID NO:145-SEQ ID NO:177-SEQ ID NO:163, SEQ ID NO:160-SEQ ID NO:59-SEQ ID NO:145-SEQ ID NO:193-SEQ ID NO:194, SEQ ID NO:160-SEQ ID NO:59-SEQ ID NO:161-SEQ ID NO:162-SEQ ID NO:163, or SEQ ID NO:226-SEQ ID NO:227-SEQ ID NO:228-SEQ ID NO:229-SEQ ID NO:230.

[0124] In certain embodiments, the monoclonal antibody is humanized.

[0125] In certain embodiments, the monoclonal antibody is labeled.

[0126] In certain embodiments, the antibodies of the present disclosure are capable of crossing the blood-brain barrier (BBB). In certain embodiments, the antibodies of the present disclosure are bispecific and IgG-like or non-IgG-like. The antibodies can bind to alpha-synuclein using any of the CDR sequences listed herein, as described elsewhere herein, and can also bind to a BBB target receptor that allows the antibody to be transported through the BBB (in a non-limiting example, via receptor-mediated transcytosis). A non-limiting example of such a BBB target receptor is the transferrin receptor, which normally activates a molecular channel that transports iron to the brain. Anti-human transferrin receptor antibodies contemplated within the present invention include those listed in US20160369001, which is incorporated herein by reference in its entirety. Other non-limiting examples of such BBB target receptors include the low-density lipoprotein (LDL) receptor and the insulin receptor. In certain embodiments, the antibodies of the present disclosure include single-chain anti-BBB target receptor antibodies. In certain embodiments, the antibodies of the present disclosure have an Fc fragment that is engineered to be capable of binding to a BBB target receptor. The approach contemplated in this disclosure is described in Pulgar, Front. Neurosci., January 2019, Vol. 12, Article 2019, and Kariolis, et al., Science Translational Medicine 12(545), eaay1359, which are incorporated herein by reference in their entirety.

[0127] The present disclosure further provides isolated polynucleotides (including RNA and / or DNA) encoding antibodies or antigen-binding fragments thereof, such as nucleic acids encoding one or more CDRs, or variable heavy or light chain regions, of the alpha-Syn antibodies of the present disclosure. Nucleic acids include DNA and RNA.

[0128] In certain aspects, the disclosure provides isolated polynucleotides comprising the nucleic acid sequences of SEQ ID NOs: 3, 5, 7, 19, 21, 23, 35, 43, 44, 50, 52, 54, 66, 68, 75, 77, 88, 89, 96, 97, 99, 106, 108, 110, 121, 132, 138, 139, 141, 148, 150, 155, 157, 167, 168, 170, 175, 180, 191, 195, 197, 198, 203, 207, 209, 211, 219, 221, 223, 233, 235, and 237.

[0129] In certain aspects, the disclosure provides isolated polynucleotides comprising at least one nucleic acid sequence selected from the group consisting of SEQ ID NOs: 3, 5, 7, 35, 50, 52, 54, 75, 77, 88, 89, 97, 99, 132, 138, 139, 141, 155, 157, 175, 191, 197, 198, 219, 221, and 223.

[0130] In certain aspects, the disclosure provides isolated polynucleotides comprising at least one nucleic acid sequence selected from the group consisting of SEQ ID NOs: 19, 21, 23, 43, 44, 66, 68, 96, 106, 108, 110, 121, 148, 150, 167, 168, 170, 180, 195, 203, 207, 209, 211, 233, 235, and 237.

[0131] In certain embodiments, the antibody has a VL encoded by a group of nucleic acid sequences comprising the set of nucleic acid sequences selected from the group consisting of SEQ ID NOs: 19, 21, 23; SEQ ID NOs: 19, 43, 44; SEQ ID NOs: 19, 66, 68; SEQ ID NOs: 19, 66, 96; SEQ ID NOs: 19, 66, 203; SEQ ID NOs: 19, 148, 150; SEQ ID NOs: 106, 108, 110; SEQ ID NOs: 121, 66, 68; SEQ ID NOs: 167, 168, 170; SEQ ID NOs: 167, 168, 195; SEQ ID NOs: 180, 168, 68; SEQ ID NOs: 213, 214, 216; and SEQ ID NOs: 233, 235, 237.

[0132] In certain embodiments, the antibody has a VH encoded by a group of nucleic acid sequences comprising the set of nucleic acid sequences selected from the group consisting of SEQ ID NO:3,5,7; SEQ ID NO:3,35,7; SEQ ID NO:3,132,7; SEQ ID NO:50,52,54; SEQ ID NO:50,75,77; SEQ ID NO:50,139,175; SEQ ID NO:50,139,191; SEQ ID NO:50,155,157; ​​SEQ ID NO:50,197,198; SEQ ID NO:88,89,54; SEQ ID NO:97,99,54; SEQ ID NO:138,139,141; and SEQ ID NO:219,221,223.

[0133] In certain aspects, the present disclosure provides an autonomously replicating or integrating mammalian cell vector comprising a recombinant nucleic acid of the present disclosure. In other aspects, the present disclosure provides a vector comprising a recombinant nucleic acid of the present disclosure. In still other aspects, the recombinant nucleic acid of the present disclosure encodes an antibody comprising a light chain variable region (VL) and a heavy chain variable region (VH). In certain aspects, the VL comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO: 27, 46, 70, 114, 126, 185, 213, or 240. In certain aspects, the VL comprises a CDR2 region comprising the amino acid sequence of SEQ ID NO: 29, 72, 116, 214, or 242. In certain aspects, the VL comprises a CDR3 region comprising the amino acid sequence of SEQ ID NO: 31, 85, 118, 129, 152, 205, 216, or 244. In certain embodiments, the VH comprises a CDR1 region comprising the amino acid sequence of SEQ ID NO: 11, 58, 92, 101, 144, 160, or 226. In certain embodiments, the VH comprises a CDR2 region comprising the amino acid sequence of SEQ ID NO: 13, 40, 60, 93, 102, 134, 145, 161, 200, or 228. In certain embodiments, the VH comprises a CDR3 region comprising the amino acid sequence of SEQ ID NO: 15, 62, 81, 62, 147, 163, 194, 201, or 230. In yet other embodiments, the vector comprises a plasmid or virus. In yet other embodiments, the vector comprises a mammalian cell expression vector. The expression vector can comprise a nucleic acid sequence that directs and / or controls expression of the inserted polynucleotide. Such nucleic acid sequences can include regulatory sequences, including promoter sequences, terminator sequences, polyadenylation sequences, and enhancer sequences. Systems for cloning and expression of a polypeptide in a variety of cells are well known in the art.

[0134] The present disclosure further provides a host cell comprising an expression vector of the present disclosure. In certain aspects, the host cell is isolated. In other aspects, the host cell is a non-human cell. In yet other aspects, the host cell is mammalian.

[0135] The antibodies of the present disclosure can be mammalian antibodies, such as primate, human, rodent, rabbit, ovine, porcine, or equine antibodies. The antibodies can be of any class or isotype, such as IgM or IgG. In certain embodiments, the antibodies are IgG.

[0136] The present disclosure further provides a kit comprising an antibody of the present disclosure. The antibody can be an intact immunoglobulin molecule or a fragment thereof, such as a Fab, F(ab)2, or Fv fragment. The antibody can be labeled as described elsewhere herein. The kit can be for use in a method for determining whether a subject has a neurodegenerative disease and / or for treating, ameliorating, and / or preventing a subject suffering from or suspected of suffering from a neurodegenerative disease. The kit can further include any other reagents or equipment, such as buffers, applicators, etc., required to perform the method of the present disclosure.

[0137] The following table shows the relationship between the SEQ ID NOs used in the priority document (US Provisional Patent Application No. 62 / 937,636) and the SEQ ID NOs used in this disclosure.

[0138] TIFF0007777071000024.tif207150TIFF0007777071000025.tif243150TIFF0007777071000026.tif24315 0TIFF0007777071000027.tif243150TIFF0007777071000028.tif243150TIFF0007777071000029.tif82150

[0139] Non-limiting examples of the production of these antibodies are illustrated in the examples and figures provided herein. In certain aspects, the present disclosure includes pharmaceutical compositions comprising each of these antibodies in combination with one or more pharmaceutically acceptable excipients. In some aspects, the pharmaceutical compositions are formulated for parenteral delivery. In other aspects, the antibodies are humanized.

[0140] Methods of Treating, Ameliorating, and / or Preventing Synucleinopathies In one aspect, the present disclosure provides a method for treating, ameliorating, and / or preventing a synucleinopathy, comprising administering to a patient a therapeutically effective amount of an isolated monoclonal antibody of the present disclosure. In another aspect, the present disclosure provides an isolated monoclonal antibody of the present disclosure for use as a medicament for treating, ameliorating, and / or preventing a synucleinopathy. In yet another aspect, the present disclosure provides use of an isolated monoclonal antibody of the present disclosure in the manufacture of a medicament for treating, ameliorating, and / or preventing a synucleinopathy. In yet another aspect, the present disclosure provides an isolated monoclonal antibody of the present disclosure for use in a method for treating, ameliorating, and / or preventing a synucleinopathy.

[0141] In certain embodiments, the antibody is humanized. In other embodiments, the antibody is administered as a pharmaceutical composition.

[0142] The monoclonal antibodies described above can be used to treat, ameliorate, and / or prevent synucleinopathies by reducing α-Syn pathology in neurons induced by α-Syn fibrils and / or oligomers. In certain embodiments, α-Syn-associated neurodegenerative disorders include, but are not limited to, Parkinson's disease, dementia (such as Parkinson's disease with dementia and / or dementia with Lewy bodies), Alzheimer's disease, Down's syndrome, multiple system atrophy, prion diseases, and other α-Syn-associated neurodegenerative disorders. The antibodies can be administered systemically or directly to the site where α-Syn fibrils, e.g., Lewy bodies, are observed or believed to be present. In a non-limiting example, the antibodies can be administered by injection into a blood vessel supplying the brain or into the brain itself. The subject can be a mammal, e.g., a human or non-human mammal.

[0143] Methods for detecting synucleinopathies In yet another aspect, the present disclosure provides a method for detecting synucleinopathies in patients. In other embodiments, the antibodies of the present disclosure can be used as a diagnostic tool for α-Syn-related neurodegenerative disorders, including but not limited to Parkinson's disease, dementia (such as Parkinson's disease with dementia and / or dementia with Lewy bodies), Alzheimer's disease, Down's syndrome, multiple system atrophy, prion disease, and other α-Syn-related neurodegenerative disorders. In certain embodiments, the method is performed in vitro. In certain embodiments, the method is performed ex vivo.

[0144] In certain embodiments, a method for detecting a synucleinopathy in a subject comprises administering a labeled isolated monoclonal antibody of the present disclosure to the subject and detecting the presence or absence of any α-Syn fibril and / or oligomer-antibody complexes in the subject. The presence of the complex indicates the presence of α-Syn fibrils and / or oligomers in the subject. In certain embodiments, if α-Syn fibrils and / or oligomers are present in the subject, the subject has a neurodegenerative disease. In other embodiments, if α-Syn fibrils or oligomers are not present in the subject, the subject does not have a neurodegenerative disease. In still other embodiments, if the subject has a neurodegenerative disease, the individual is recommended to receive a therapy and / or pharmacological treatment for the neurodegenerative disease. In still other embodiments, if the subject has a neurodegenerative disease, the individual is provided with a therapy and / or pharmacological treatment for the neurodegenerative disease.

[0145] In certain embodiments, the method further comprises comparing the level of antibody / α-Syn fibril and / or oligomer complexes formed in the subject with the level of antibody / α-Syn fibril and / or oligomer complexes formed in a reference subject. The reference subject can be a subject known to have no α-Syn fibrils and / or oligomers, a subject known to have detectable α-Syn fibrils and / or oligomers, and / or a subject known to have a particular level of α-Syn fibrils and / or oligomers. The reference subject can also be the same subject being treated or evaluated, but corresponding to a previous α-Syn fibril and / or oligomer detection experiment, as a means of assessing disease progression and / or treatment efficacy in the subject.

[0146] In yet another aspect, the present disclosure provides methods for detecting α-Syn fibrils in a sample. In certain embodiments, the antibodies of the present disclosure can be used as diagnostic tools to detect the presence of α-Syn fibrils, oligomers, or other misfolded α-Syn species in a sample.

[0147] In certain embodiments, a method for detecting α-Syn fibrils, oligomers, or other misfolded α-Syn species in a sample (e.g., from a subject) comprises contacting the sample with a labeled isolated monoclonal antibody of the present disclosure and detecting the presence or absence of any complexes between the antibody and α-Syn fibrils, oligomers, or other misfolded α-Syn species in the sample. Detection of complexes indicates the presence of α-Syn fibrils, oligomers, or other misfolded α-Syn species in the sample. The sample can be, by non-limiting example, cerebrospinal fluid (CSF), blood, urine, saliva, or tissue from the brain, gastrointestinal tract, colon, skin, or salivary glands. In certain embodiments, the sample is a CSF sample and / or a brain tissue sample. In other embodiments, the sample is used directly after removal from the subject. In other embodiments, the sample is pre-treated during use in the method.

[0148] In certain embodiments, the method further comprises comparing the level of antibody-α-syn fibrils / oligomers or other antibody-misfolded α-syn complexes formed in the sample with the level of antibody-α-syn fibrils / oligomers or other antibody-misfolded α-syn complexes formed in a reference sample. The reference sample can be from a subject known to have no α-syn fibrils / oligomers or other misfolded α-syn species, a subject known to have detectable α-syn fibrils / oligomers or other misfolded α-syn species, and / or a subject known to have a particular level of α-syn fibrils / oligomers or other misfolded α-syn species. The reference sample can also be from the same subject being treated or evaluated, but corresponding to a previous detection of α-syn fibrils / oligomers or other misfolded α-syn species, as a means of assessing disease progression and / or treatment efficacy in the subject.

[0149] In certain embodiments, the level of detected α-Syn fibrils / oligomers or other misfolded α-Syn species in a subject or a sample derived from a subject correlates with the severity or progression of a neurodegenerative disease in the subject. In other embodiments, the disclosed methods can be used to monitor the severity or progression of a neurodegenerative disease in a subject. In yet other embodiments, the disclosed methods can be used to monitor the effectiveness of therapy and / or pharmacological intervention in a subject suffering from or suspected of suffering from a neurodegenerative disease.

[0150] In certain aspects, the sample comprises or is an in vitro sample, hi certain aspects, the sample comprises or is an ex vivo sample.

[0151] Methods for detecting the formation of complexes between antibodies and α-syn fibrils / oligomers or other misfolded α-syn species include, but are not limited to, radioimmunoassays, enzyme-linked immunosorbent assays (ELISAs), sandwich immunoassays, fluorescent immunoassays, precipitation reactions, gel immunodiffusion assays, agglutination assays, protein A immunoassays, immunoelectrophoretic assays, electrophoresis, Western blotting, or any other techniques known in the art.

[0152] In certain embodiments, the antibodies of the present disclosure can be combined with a label to detect excess α-Syn levels in a patient or sample. In other embodiments, the antibodies of the present disclosure can be combined with a label to detect α-Syn fibrils / oligomers or other misfolded α-Syn species in a patient or sample. Methods for labeling antibodies are known in the art, and a wide variety of approaches can be used. In certain embodiments, the label can be, but is not limited to, F 18 , I 123 , In 111 , I 131 , C 14 , H 3 , Tc 99m , P 32 , I 125 , Ga 68In other embodiments, the label is a radioactive label such as, but not limited to, fluorescein or rhodamine. In still other embodiments, the label is an imaging agent such as, but not limited to, gadolinium (Gd), dysprosium, and iron, a magnetic agent, or the like. Other labels include nuclear magnetic resonance active labels, positron-emitting isotopes detectable by PET scanners, chemiluminescent, and enzyme markers. Non-limiting imaging techniques include electron microscopy, confocal microscopy, light microscopy, positron emission tomography (PET), gamma scintigraphy, magnetic resonance imaging (MRI), functional magnetic resonance imaging (FMRI), magnetoencephalography (MEG), and single-photon emission computed tomography (SPECT). In still other embodiments, the label is on a secondary antibody that binds to the primary antibody comprising the sequence described above.

[0153] Administration / Dosage / Formulation The compounds and / or compositions of the present disclosure can be administered to patients, preferably mammals, more preferably humans, using known procedures at dosages and for periods effective to carry out the therapeutic and / or imaging methods envisioned in the present disclosure. The effective amount of compound required for adequate therapeutic treatment and / or imaging signal can vary depending on factors such as the state of the disease or disorder in the patient; the age, sex, and weight of the patient; and / or the device used to detect the compounds of the present disclosure. Those skilled in the art will be able to study the relevant factors and make the determination regarding the effective amount of therapeutic compound and / or imaging compound without undue experimentation.

[0154] Actual dosage levels of the active ingredients in the pharmaceutical compositions of the present disclosure may be varied to provide an amount of the active ingredient that is effective to achieve successful treatment and / or imaging for a particular patient, composition, and mode of administration without being toxic to the patient.

[0155] In certain embodiments, the compositions of the present disclosure are formulated using one or more pharmaceutically acceptable excipients or carriers.In certain embodiments, the pharmaceutical compositions of the present disclosure comprise an effective amount of the compound of the present disclosure and a pharmaceutically acceptable carrier.

[0156] The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (e.g., glycerol, propylene glycol, and liquid polyethylene glycol, etc.), suitable mixtures thereof, and vegetable oils. Proper fluidity can be maintained, for example, by the use of a coating such as lecithin, by maintaining the required particle size in the case of dispersions, and by the use of surfactants. Prevention of microbial action can be achieved by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, etc. In many cases, it will be preferable to include isotonic agents, for example, sugars, sodium chloride, or polyalcohols such as mannitol and sorbitol, in the composition. Prolonged absorption of injectable compositions can be achieved by including agents that delay absorption, for example, aluminum monostearate and gelatin in the composition.

[0157] The formulations may be used in admixture with conventional excipients, i.e., pharmaceutically acceptable organic or inorganic carrier substances suitable for oral, parenteral, nasal, intravenous, subcutaneous, enteral, or any other suitable mode of administration known in the art. Pharmaceutical preparations may be sterilized and, if desired, may be mixed with auxiliary substances, such as lubricants, preservatives, stabilizers, wetting agents, emulsifiers, salts for influencing osmotic pressure, buffers, colorants, flavorings, and / or aromatic substances.

[0158] The administration route of any of the compositions of the present disclosure includes oral, nasal, rectal, intravaginal, parenteral, buccal, sublingual or topical.The compound for use in the present disclosure can be formulated for administration by any suitable route, for example, oral or parenteral, for example, transdermal, transmucosal (for example, sublingual, lingual, (trans)buccal, (trans)urethral, ​​vaginal (for example, transvaginal and perivaginal), (intra)nasal and (trans)rectal), intravesical, intrapulmonary, intraduodenal, intragastric, intraspinal, subcutaneous, intramuscular, intradermal, intraarterial, intravenous, intrabronchial, inhalation and topical administration.

[0159] Suitable compositions and dosage forms include, for example, tablets, capsules, caplets, pills, gelcaps, troches, dispersions, suspensions, liquids, syrups, granules, beads, transdermal patches, gels, powders, pellets, magmas, lozenges, creams, pastes, plasters, lotions, discs, suppositories, liquid sprays for nasal or oral administration, dry powder or aerosolized formulations for inhalation, compositions and formulations for intravesical administration, etc. It should be understood that the formulations and compositions that would be useful in the present disclosure are not limited to the specific formulations and compositions described herein.

[0160] Parenteral administration As used herein, "parenteral administration" of a pharmaceutical composition includes any administration route characterized by physical breaching of the target tissue, and administration of the pharmaceutical composition through an incision in the tissue. Thus, parenteral administration includes, but is not limited to, administration of the pharmaceutical composition by injection of the composition, application of the composition through a surgical incision, application of the composition through a non-surgical wound that penetrates the tissue, etc. In particular, parenteral administration is intended to include, but is not limited to, subcutaneous, intravenous, intraperitoneal, intramuscular, intrasternal injection, and kidney dialysis infusion techniques.

[0161] Pharmaceutical compositions suitable for parenteral administration include an active ingredient in combination with a pharmaceutically acceptable carrier, such as sterile water or sterile isotonic saline. Such formulations may be prepared, packaged, or sold in a form suitable for bolus administration or continuous administration. Injectable formulations may be prepared, packaged, or sold in unit dosage form, for example, in ampoules or in multi-dose containers containing a preservative. Formulations for parenteral administration include, but are not limited to, suspensions, solutions, emulsions in oily or aqueous vehicles, pastes, and implantable sustained-release or biodegradable formulations. Such formulations may further comprise one or more additional ingredients, including, but not limited to, suspending agents, stabilizers, or dispersing agents. In certain embodiments of formulations for parenteral administration, the active ingredient is provided in a dry (i.e., powdered or granular) form for reconstitution with a suitable vehicle (e.g., sterile, pyrogen-free water) prior to parenteral administration of the reconstituted composition.

[0162] Pharmaceutical compositions may be prepared, packaged, or sold in the form of a sterile injectable aqueous or oily suspension or solution. These suspensions or solutions may be formulated according to known techniques and may contain, in addition to the active ingredient, additional ingredients such as dispersing agents, wetting agents, or suspending agents described herein. Such sterile injectable formulations may be prepared using a non-toxic parenterally acceptable diluent or solvent, such as water or 1,3-butanediol. Other acceptable diluents and solvents include, but are not limited to, Ringer's solution, isotonic sodium chloride solution, and fixed oils such as synthetic mono- or diglycerides. Other useful parenterally administrable formulations include those comprising the active ingredient in microcrystalline form, in a liposomal preparation, or as a component of a biodegradable polymer system. Sustained-release or implantable compositions may include pharmaceutically acceptable polymeric or hydrophobic materials, such as emulsions, ion exchange resins, poorly soluble polymers, or poorly soluble salts.

[0163] Additional Dosage Forms Additional dosage forms of the present disclosure include those described in U.S. Patent Nos. 6,340,475; 6,488,962; 6,451,808; 5,972,389; 5,582,837; and 5,007,790. Additional dosage forms of the present disclosure also include those described in U.S. Patent Application Nos. 2003 / 0147952; 2003 / 0104062; 2003 / 0104053; 2003 / 0044466; 2003 / 0039688; and 2002 / 0051820. Additional dosage forms of the present disclosure also include those described in PCT Application Nos. WO 03 / 35041; WO 03 / 35040; WO 03 / 35029; WO 03 / 35177; WO 03 / 35039; WO 02 / 96404; WO 02 / 32416; WO 01 / 97783; WO 01 / 56544; WO 01 / 32217; WO 98 / 55107; WO 98 / 11879; WO 97 / 47285; WO 93 / 18755; and WO 90 / 11757. [Example]

[0164] Experimental Example The present disclosure will be further described in detail by reference to the following experimental examples. Unless otherwise specified, these examples are provided for illustrative purposes only and are not intended to be limiting. Therefore, the present disclosure should in no way be construed as being limited to the following examples, but rather as encompassing any and all variations that are evident as a result of the teachings provided herein.

[0165] Without further description, it is believed that one of ordinary skill in the art can, using the preceding description and the following illustrative examples, practice the methods of the present disclosure as claimed. The following examples, therefore, specifically point out preferred embodiments of the present disclosure, and are not to be construed as limiting in any way the remainder of the disclosure.

[0166] animal Mice used for antibody production were Balb / c; mice used for primary neuronal cultures were CD-1 (Charles River, Cat# CRL:22, RRID:IMSR_CRL:22), and mice used for in vivo studies were B6C3F1 (Charles River, Cat# CRL:31, RRID:IMSR_CRL:31).

[0167] Primary hippocampal neuron cultures Primary hippocampal neuron cultures were prepared from embryonic day (E) 16–18 CD1 embryos as previously described (Henderson, et al., 2017, J Neurosci.; Henderson, et al., 2018, Acta Neuropathologica Comm. 6:45). Dissociated hippocampal neurons were plated at 17,500 cells / well (96-well plates) in Neurobasal medium (ThermoFisher 21103049) supplemented with B27 (ThermoFisher 17504044), 2 mM GlutaMax (ThermoFisher 35050061), and 100 U / mL penicillin / streptomycin (ThermoFisher 15140122).

[0168] α-synuclein PFF Purification of recombinant α-synuclein and production of α-synuclein PFFs were performed as described elsewhere (Luk, et al., 2009, Proc Natl Acad Sci USA 106:20051-20056; Volpicelli-Daley, et al., 2014, Nature Protocols 9:2135-2146). The pRK172 plasmid containing the gene of interest was transformed into BL21(DE3)RIL-competent E. coli (Agilent Technologies Cat#230245). A single colony from this transformation was expanded in Terrific Broth (12 g / L Bacto-tryptone, 24 g / L yeast extract, 4% (vol / vol) glycerol, 17 mM KH2PO4, and 72 mM K2HPO4) containing ampicillin. The bacterial pellet from the growth was sonicated, and the sample was boiled to precipitate unwanted proteins. The supernatant was dialyzed overnight against 10 mM Tris (pH 7.6), 50 mM NaCl, and 1 mM EDTA. The protein was filtered through a 0.22 μm filter and concentrated using an Amicon Ultra-15 centrifugal filter unit (Millipore Sigma Cat# UFC901008). The protein was then loaded onto a Superdex 200 column, and 1 mL fractions were collected. The fractions were run on SDS-PAGE and stained with Coomassie blue to select fractions highly enriched in α-synuclein. These fractions were combined and dialyzed overnight in 10 mM Tris (pH 7.6), 50 mM NaCl, and 1 mM EDTA. The dialyzed fraction was applied to a MonoQ column (GE Health, HiTrap Q HP 645932) and run using a linear gradient from 25 mM NaCl to 1 M NaCl. Collected fractions were run on SDS-PAGE and stained with Coomassie blue. Fractions highly enriched in α-synuclein were collected and dialyzed into DPBS. The protein was filtered through a 0.22 μm filter and concentrated to 5 mg / mL using an Amicon Ultra-15 centrifugal filter unit (Millipore Sigma Cat# UFC901008). The monomer was aliquoted and frozen at -80°C.To prepare α-synuclein PFFs, α-synuclein monomers were shaken at 1,000 rpm for 7 days. Conversion to PFFs was verified by sedimentation at 100,000 × g for 60 min and thioflavin T fluorescence.

[0169] α-synuclein PFF treatment primary neurons For neuronal treatment, mouse α-synuclein PFFs prepared at a concentration of 5 mg / mL were vortexed and diluted to 100 μg / mL with Dulbecco's phosphate-buffered saline (DPBS, Corning Cat#21-031-CV). They were then sonicated (Diagenode Biorupter UCD-300 batch sonicator) for 10 cycles of 30 seconds on and 30 seconds off at the high setting. The α-synuclein PFFs were then diluted to 5 μg / mL in neuronal medium and added to neuronal cultures at the indicated concentrations.

[0170] mouse Mouse α-synuclein PFFs prepared at a concentration of 5 mg / mL were vortexed and diluted to 2 mg / mL with DPBS. They were then sonicated (Diagenode Biorupter UCD-300 batch sonicator) for 10 cycles of 30 seconds on and 30 seconds off at the high setting. Mice were injected at 3 months of age. Mice were injected unilaterally with 5 μg of α-synuclein PFFs (2.5 μL) via a single needle insertion into the right forebrain (coordinates: +0.2 mm relative to the bregma, +2.0 mm from the midline) targeting the dorsal striatum (2.6 mm below the dura). Injections were performed at a rate of 0.4 μL / min using a 10 μL syringe (Hamilton, NV). After 6 months, mice were perfused transcardially with PBS, and brains were removed and fixed overnight in 70% ethanol, 150 mM NaCl (pH 7.4).

[0171] antibody production Mouse monoclonal antibodies were raised by administering sonicated human α-synuclein PFFs (0.05 mg α-synuclein / mouse) emulsified in complete Freund's adjuvant, followed by two successive boosts of 0.025 mg α-synuclein emulsified in incomplete Freund's adjuvant 3 and 6 weeks after the initial injection, as previously described (Gibbons, et al., 2018, Neuropathol Exp Neurol 77:216-228). Nine weeks after the initial antigen injection, mice received an intravenous boost of 0.025 mg / mouse. Four days after the intravenous injection, spleens were dissociated into single-cell suspensions and fused with SP2 cells by treatment with a mixture of 50% polyethylene glycol and 5% DMSO for 1 minute. Hybridoma cells were selected for 7 days in medium containing 5.7 μM azaserine-10; 100 μM hypoxanthine (Sigma Cat# A9666) and cultured in Kennett's HY (90% DMEM, 10% NCTC135, 4.15 g / L glucose, 3.55 g / L NaHCO) supplemented with 20% fetal bovine serum (FBS; Atlanta Biological Cat# E0118), 100 U / mL penicillin / 100 μg / mL streptomycin (Gibco Cat# 15140-122), 2 mM L-glutamine (Corning Cat# 25-005-Cl), and OPI medium supplement (1 mM oxaloacetate, 0.45 mM pyruvate, 0.2 U / mL insulin; Sigma Cat# 05003). Monoclonal populations were isolated by limiting dilution to 0.3 cells / well in 96-well plates. Antibodies were further characterized as described below. Hybridomas selective for pathological misfolded α-syn were expanded and subcloned at least twice.

[0172] Epitope mapping Recombinant α-synuclein constructs were produced in E. coli as previously described (Volpicelli-Daley, et al., 2014, Nature Protocols 9:2135-2146). Total protein concentration in each sample was determined by bicinchoninic acid colorimetric assay (Fisher Cat#23223 and 23224) using bovine serum albumin (Thermo Fisher Cat#23210) as a standard. Proteins were separated on a 5-20% gradient polyacrylamide gel with equal protein loading (α-synuclein 250 ng / well). Proteins were transferred to a 0.2 μm nitrocellulose membrane and detected with primary antibodies (1:1000). Primary antibodies were detected using IRDye 800 (LI-COR 925-32210) or IRDye 680 (LI-COR 925-68071) secondary antibodies, scanned on a LI-COR Odyssey Imaging System, and analyzed using Image Studio software.

[0173] Primary neuron immunotherapy assay Neurons were treated using a modified version of a previously described procedure (Tran, et al., 2014, Cell Reports 7:2054-2065). Neurons were fed every 3 days after plating until 10 DIV. At that time, 125 μL of medium was removed from each well, and sterile α-synuclein antibody was added in 20 μL of fresh neuronal medium and incubated at 37°C for 30 minutes. 125 μg of freshly sonicated PFFs was added in an additional 20 μL of neuronal medium. Neurons were fed at 1 and 4 DPT and fixed and stained 7 days post-transduction as previously described (Tran, et al., 2014, Cell Reports 7:2054-2065). Data are reported as normalized pS129 α-synuclein area divided by NeuN count. For initial clone selection, antibodies were included at a 1:1 molar ratio to α-synuclein PFF. For further characterization of high-priority clones, antibodies were included at molar ratios ranging from 2:1 to 0.0003:1 (antibody:PFF).

[0174] immunocytochemistry Primary neuronal or cell line cultures were fixed with 4% paraformaldehyde and 4% sucrose in phosphate-buffered saline and washed five times in PBS. Immunostaining of neuronal cultures was performed as previously described (Henderson, et al., 2018, Acta Neuropathologica Comm. 6:45). Cells were permeabilized in 3% BSA + 0.3% TX-100 in PBS for 15 minutes at room temperature. After PBS washing, cells were blocked with 3% BSA in PBS for 50 minutes and then incubated with primary antibodies for 2 hours at room temperature. Primary antibodies used were those targeting pS129 α-synuclein (81A, CNDR, 1:5,000) and NeuN (Millipore Cat#MAB377, 1:1,500). Cells were washed five times with PBS and incubated with secondary antibodies for 1 hour at room temperature. After washing five times with PBS, cells were incubated in DAPI (ThermoFisher Cat# D21490, 1:10,000) in PBS. The 96-well plates were imaged using an In Cell Analyzer 2200 (GE Healthcare) and analyzed using the accompanying software. A standard intensity-based threshold was applied equally across the plate to the pS129 α-synuclein channel to quantify the positive area. For NeuN quantification, an object-based analysis was applied to identify objects of a specified size and intensity. All quantifications were optimized and applied equally across all conditions.

[0175] Sandwich ELISA 384-well Maxisorp clear plates (Thermo Fisher Scientific, Cat# 12565347) were coated with 30 μL (50 ng) of antibody solution per well in Takeda coating buffer. Plates were then spun at 1000 × g for 1 minute and incubated overnight at 4°C. Plates were washed four times with PBST (PBS containing 0.05% Tween) and blocked overnight at 4°C using Block Ace blocking solution (95 μL per well) (AbD Serotec). Serial doubling dilutions of human wild-type α-synuclein fibrils and monomers were made in buffer C (0.02 M sodium phosphate buffer, 2 mM EDTA, 0.4 M NaCl, 1% BSA, 0.005% thimerosal), starting at 256 μg / mL of monomer and 25.6 μg / mL of fibrils. The fibrils were then sonicated (Diagenode Biorupter UCD-300 batch sonicator) at the high setting for 10 cycles of 30 seconds on and 30 seconds off before dilution. The dilutions were added to each well (30 μL per well) and incubated overnight at 4°C. The plate was washed four times with PBST. 30 μL of MJF-R1 (1:3,000, Abcam Cat# ab138501) in buffer C was added to each well, and the plate was incubated at 37°C for 4 hours. After washing four times with PBST, a 1:10k diluted goat-anti-rabbit IgG-HRP conjugate (Cell Signaling Technology) was added to the plate, and the plate was incubated at 37°C for 1 hour. After washing four times with PBST, the plates were developed with 30 μL per well of room temperature 1-Step Ultra TMB-ELISA Substrate Solution (Thermo Fisher Scientific, Cat# 34029) for 10–15 min, and the reaction was quenched with 10% phosphoric acid (30 μL per well). Plates were read at 384–450 nm on a SpectraMax M5 plate reader (Molecular Devices).

[0176] Antibody purification (small amount preparation) Small amounts of antibody for in vitro experiments were purified using the DynaBead magnetic system (Invitrogen, Cat#10003D) using 1 mL of Protein A beads and 1 mL of Protein G beads per 50 mL of supernatant. The beads were first washed three times with PBS on a rotator with a 10-minute wash time. The supernatant was added and incubated with the beads for 4 hours at room temperature or overnight at 4°C. The supernatant was removed, and the beads were washed three times with PBS. Elution buffer (100 mM glycine, pH 2.8) was added to the beads for 30 seconds and immediately neutralized with 1.5 M Tris base. The buffer solution was removed, placed in an Eppendorf tube with a 50 kD filter, and spun three times according to the manufacturer's protocol to complete the buffer exchange. The antibody was stored at 1 mg / mL.

[0177] Antibody purification (large scale preparation) For large-scale preparation of hybridoma supernatants, antibodies were purified using a HiTrap MabSelect SuRe column (GE Healthcare Life Sciences, Cat# 11003494) on an AKTA Pure FPLC system (GE Healthcare Life Sciences). The supernatant was sterile filtered through a 0.2 μm filter and loaded onto the column. After washing, the antibody was eluted with 100 mM glycine, 150 mM NaCl (pH 3.0). The eluate was immediately neutralized with 1 M Tris base (pH 9.0). Antibody-containing fractions were selected and pooled using a UV trace. The antibody was concentrated using an Amicon Ultra-15 50K centrifugal filter unit (Millipore Sigma, Cat# UFC905024) and dialyzed into phosphate-buffered saline (pH 7.2). The antibody was then sterile filtered through a 0.2 μm filter, and the protein concentration in each sample was determined by bicinchoninic acid colorimetric assay (Fisher Cat# 23223 and 23224) using bovine serum albumin (Thermo Fisher Cat# 23210) as a standard. Samples were run on a 15% SDS-PAGE gel and Coomassie stained to confirm the presence of heavy and light chains and protein purity. After purification from hybridoma supernatants, the antibody was frozen in 1 mL aliquots and stored at -20°C.

[0178] In vivo antibody administration Immediately prior to use, the antibody was thawed and kept on ice until administration. Mice were weighed and administered intraperitoneally with antibody to a final concentration of 30 mg / kg body weight. Injection sites were rotated between injections, and mice were monitored for injection-related adverse events.

[0179] Striatal dopamine / DOPAC detection After transcardial perfusion, 1 mm coronal sections were cut from the rostral brain approximately from anterior to posterior +1 mm. The dorsal striatum was manually dissected from both the right (ipsilateral) and left (contralateral) sides of the brain and flash-frozen on dry ice for liquid chromatography-mass spectrometry (LC-MS) analysis of dopamine (DA) and dihydroxyphenylacetic acid (DOPAC). The frozen tissue was suspended in 10 μL / mg tissue of Milli-Q water and sonicated at a power level of 1.5 using a QSONICA MICROSON™ XL-2000 sonicator with 15–20 short pulses until the solution was homogenous. The lysate was briefly spun down, and 30 μL was transferred to a new tube containing 30 μL of 0.4 M perchloric acid. The remaining lysate was suspended in 2x RIPA buffer containing protease inhibitors for protein level assays. The perchloric acid samples were spun at 3,000 x g and 4°C for 15 minutes. Two volumes of 0.4 M sodium acetate were added to the supernatant and spin-filtered through a 0.65 μm filter. DA and DOPAC were then quantified using a Waters Acquity UPLC-TQD LC-MS system. Ten microliters of each sample was injected onto a Waters Acquity HSS T3, C18, 1.8 μm, 2.1 × 100 mm column at 35 °C and 0.4 mL / min. The mobile phase B was held at 0% B for 1 min after injection, followed by a gradient separation from 0 to 25% B over 2–3 min, followed by wash and equilibration steps (A: 0.1% (v / v) formic acid in water; B: acetonitrile containing 0.1% (v / v) formic acid). Multiple reaction monitoring of specific collision-induced ion transitions was used to detect the compounds (dopamine, ES+ 154 > 137; 3,4-dihydrophenylacetic acid, ES- 167 > 123). Peak areas were quantified against standard curves analyzed simultaneously using Waters QuaLynx software (dopamine hydrochloride (DA), Sigma Cat# H8502-5G; 3,4-dihydrophenylacetic acid (DOPAC), Sigma Cat# 850217-1G) and normalized to total protein levels.

[0180] immunohistochemistry After perfusion and fixation, brains were embedded in paraffin blocks, cut into 6 μm sections, and mounted on glass slides. Slides were then stained using standard immunohistochemistry as described elsewhere herein. Slides were deparaffinized with two consecutive 5-minute washes in xylene, followed by 1-minute washes in a descending series of ethanol: 100%, 100%, 95%, 80%, and 70%. Slides were then incubated in deionized water for 1 minute before antigen retrieval as described. After antigen retrieval, slides were incubated in 5% hydrogen peroxide in methanol to quench endogenous peroxidase activity. Slides were washed in tap water for 10 minutes and 0.1 M Tris for 5 minutes, then blocked in 0.1 M Tris / 2% fetal bovine serum (FBS). Slides were incubated overnight in primary antibodies. The following primary antibodies were used: For misfolded α-synuclein, microwave antigen retrieval was performed using Syn506 at a final concentration of 0.4 μg / mL (citric acid-based antigen unmasking solution (Vector H-3300) at 95°C for 15 minutes). For staining midbrain dopaminergic neurons, formic acid antigen retrieval was performed using tyrosine hydroxylase (TH-16; Sigma-Aldrich T2928, RRID:AB_477569) at 1:5,000.

[0181] The primary antibody was washed off with 0.1M Tris for 5 minutes, followed by incubation with goat anti-rabbit (Vector BA1000, RRID:AB_2313606) or horse anti-mouse (Vector BA2000, RRID:AB_2313581) biotinylated IgG at 1:1000 in 0.1M Tris / 2% FBS for 1 hour. The biotinylated antibody was washed off with 0.1M Tris for 5 minutes, followed by incubation with avidin-biotin solution (Vector PK-6100, RRID:AB_2336819) for 1 hour. Slides were then rinsed in 0.1 M Tris for 5 minutes, developed with ImmPACT DAB peroxidase substrate (Vector SK-4105, RRID:AB_2336520), and briefly counterstained with Harris hematoxylin (Fisher 67-650-01). Slides were washed in tap water for 5 minutes, dehydrated in ascending ethanol streams: 70%, 80%, 95%, 100%, and 100% for 1 minute each, then washed twice in xylene for 5 minutes and coverslipped in Cytoseal Mounting Media (Fisher 23-244-256). Digitized slides were then used for quantitative pathology.

[0182] Quantitative histology To assay for antibody binding to Lewy body α-synuclein, undiluted hybridoma supernatant or supernatant diluted 1:3 in PBS was added directly to sections of human amygdala tissue containing abundant Lewy bodies. Tissues were processed in parallel and developed with DAB reagent. A similar 1 mm section from each tissue section was then 2 Sections were used to assay the ability of the antibody to preferentially bind to Lewy bodies. Staining intensity was manually thresholded to highlight only Lewy bodies that were unaffected by antibody treatment and analyzed for mean optical density. A standardized cutoff was then used to assay the mean optical density for all sections. The mean optical density of Lewy body staining divided by the mean optical density of the tissue section was calculated and reported as the Lewy body discrimination index.

[0183] All section selection, annotation, and quantification of mice was performed blinded to treatment group. All quantification was performed using HALO quantitative pathology software (Indica Labs). Every 10th slide through the midbrain was stained for tyrosine hydroxylase (TH). TH-stained sections were used to annotate SNs, and blinded cell counts were performed manually on all sections. The sum of all sections was multiplied by 10 to estimate the total number obtained by counting all sections. SN annotations drawn on TH-stained sections were then transferred to serial sections stained for misfolded α-synuclein (Syn506). Amygdala regions were also annotated every 10th section along the length of the amygdala. A single analysis algorithm was then applied equally to all stained sections to quantify the percentage of area occupied by Syn506 staining. Specifically, the analysis included all DAB signals above an optical density threshold of 0.157, empirically determined to be free of any background signal. This signal was then normalized to the total tissue area, using a minimum tissue optical density of 0.02 to exclude any areas where the tissue was split.

[0184] Antibody Sequencing Total RNA was isolated from hybridoma cells according to the TRIZOL® Reagent technical manual. Total RNA was then reverse transcribed into cDNA using either an isotype-specific antisense primer or a universal primer according to the PRIMESCRIPT™ 1st Strand cDNA Synthesis Kit technical manual. Heavy and light chain antibody fragments were amplified according to the GenScript Rapid Amplification of cDNA Ends (RACE) standard operating procedure (SOP). The amplified antibody fragments were separately cloned into standard cloning vectors. Colony PCR was performed to screen for clones with the correct insert size. Consensus sequences are provided elsewhere herein.

[0185] statistical analysis All statistical analyses were performed in GraphPad Prism 7. The analysis used for each dataset is described in the figure legend. The number of samples (n) is described in the respective figure legend. For primary neuron experiments, n represents the number of independent wells assayed. For in vivo experiments, n represents the number of mice.

[0186] Example 1: There are concerns that pan-α-synuclein antibodies may have drawbacks as a therapeutic approach for PD. For example, there is conflicting data regarding whether reducing overall α-synuclein levels in the brain is harmful to neuronal function. Because blood concentrations of antibodies are roughly 1,000-fold higher than in the brain, the abundance of α-synuclein in red blood cells also increases the likelihood that pan-α-synuclein antibodies will cause on-target side effects in the blood. Additionally, serum α-synuclein could act as a receptor for pan-α-synuclein antibodies, thereby reducing their association with their intended target in the brain.

[0187] Therefore, this study was directed, in part, at identifying antibodies highly selective for pathogenic misfolded α-synuclein. To develop these antibodies, we first immunized mice with misfolded α-synuclein preformed fibrils (PFFs) formed from recombinant human α-synuclein (Figure 1A). The mice mounted an immune response against the α-synuclein PFF immunogen, and antibody-producing B cells were harvested and fused with myeloma cells to generate antibody-producing hybridoma cells (Figure 1B). Monoclonal hybridomas were isolated by limiting dilution, and antibody-containing supernatants from these clones were tested in several assays to determine which hybridoma clones produced antibodies with desirable properties (Figure 1C). Candidate antibodies were further subcloned to ensure monoclonality (Figure 1D), and the properties of the antibodies were confirmed through screening to ensure retention of properties (Figure 1E). After validation of the results of the different screening assays, non-exhaustive candidate antibodies were selected for efficacy testing in an in vivo model of PD (Figure 1F), and the candidate antibodies were compared with previously characterized immunotherapeutic antibodies.

[0188] Example 2: Immunohistochemistry reveals antibodies that preferentially bind to LB In a non-limiting aspect, certain antibodies of interest have a higher binding preference for misfolded LB α-synuclein over monomeric α-synuclein. To rapidly screen antibody selectivity for pathological α-synuclein, amygdala sections from PD patients with abundant LB pathology were immunostained. Hybridoma supernatants were used undiluted or at a 1:3 dilution on parallel-processed screening slides to allow direct comparison of immunostaining results. Most antibodies tested showed a binding preference for LB α-synuclein over normal synaptic α-synuclein in the neuropil (Figure 2A). To better compare the relative binding of antibodies to LB versus neuropil pools, an LB discrimination index (optical density of LB / optical density of all tissue) was calculated (Figure 2B). A value of 1 indicates indistinguishability of LB from normal synaptic neuropil staining. Although most of the antibodies had a preference for LB, a cutoff of 1.5 was established to eliminate antibodies that showed relatively non-selective staining of LB from further consideration.

[0189] Example 3: Epitope mapping confirms that preferred antibodies recognize human and mouse alpha-synuclein While the selected immunotherapeutic antibodies could, in principle, be used in humans, this study identified antibodies capable of recognizing both mouse and human α-synuclein, allowing for further characterization in mouse primary neurons and wild-type mouse models of PD. The antibodies were assayed by Western blot to determine whether they bound to human and mouse α-synuclein (Figures 3A-3C). Multiple truncated human α-synuclein forms (Figure 3A) were also analyzed in parallel to determine the epitope within α-synuclein recognized by each antibody. All antibodies recognized portions of the immunogenic C-terminus of α-synuclein, with the majority recognizing the distant C-terminal amino acids (120-140; Figure 3B). Another large group of antibodies recognized slightly more internal epitopes (110–120), and only a few antibodies recognized regions closer to the aggregation-prone NAC (the non-Aβ component of the Alzheimer's disease amyloid domain of α-synuclein (residues 61–95)) (Fig. 3C). Most antibodies recognized both human and mouse α-synuclein, although there were examples of antibodies in each epitope class that showed preferential binding to human α-synuclein. Only two antibodies showed evidence of cross-reactivity with β-synuclein (Fig. 10).

[0190] Example 4: Sandwich ELISA identifies antibodies with a preference for misfolded α-synuclein Antibodies that demonstrated selectivity for LBs in human tissues by immunohistochemistry and recognized both human and mouse α-synuclein were further tested for selectivity by assessing their ability to bind to monomeric human α-synuclein and α-synuclein PFFs in a sandwich ELISA format. This assay preserves the conformation of α-synuclein by allowing solution-state α-synuclein to bind to immobilized antibodies, allowing broad affinity detection. The antibody of interest was coated onto an ELISA plate, and the antibody was incubated with either α-synuclein monomer or PFFs at increasing concentrations of α-synuclein to determine the antibody's relative affinity for each form of α-synuclein. The previously characterized Syn211 antibody (Giasson, et al., 2000, J. Neurosci. Res. 59:528-533) was also coated onto each plate as a nonselective antibody control. Bound α-synuclein was detected with a monoclonal antibody (MJF-R1, Abcam, Cat#138501) and a goat-anti-rabbit IgG-HRP conjugate.

[0191] The capture antibodies could be roughly evenly divided into three categories: 17 non-binding (Figures 4A & 11), 18 non-selective (Figures 4B & 12), and 19 PFF-selective (Figures 4C & 13). Without wishing to be bound by any theory, because nearly all of the non-binding antibodies bound to LB pathology and detected α-synuclein by immunoblotting, we can suggest that the lack of activity in the sandwich ELISA affected the ability of immobilization on the ELISA plate wells to capture α-synuclein.

[0192] EC values ​​were calculated for each antibody by fitting the absorbance values ​​to a sigmoidal dose curve. 50 The values ​​were then calculated using Syn211 as a non-selective control using the following equation: PFF priority values ​​were calculated for each antibody using TIFF0007777071000030.tif14143.

[0193] The values ​​are summarized in Table 1. To ensure that the apparent conformational selectivity was not due to a shared epitope between the capture and detection antibodies that reduced detection of α-synuclein monomers, four of the most selective antibodies were evaluated by sandwich ELISA using the alternative polyclonal detection antibody SNL4 (Figure 14). This assay confirmed the conformational selectivity of the α-synuclein antibodies.

[0194] Example 5: Primary neuron immunotherapy assay reveals differential efficacy of α-synuclein antibodies to block LB-like pathology While the intrinsic properties of an antibody are important, we also investigated its ability to prevent the induction of α-synuclein pathology, a property critical for therapeutic success. To achieve this, we developed a neuronal immunotherapy assay that allows for co-treatment of antibodies with α-synuclein PFFs in a high-content format. After 10 days in culture, neurons were treated with purified antibodies of interest. Thirty minutes later, neurons were treated with human α-synuclein PFFs, which seed the recruitment of endogenous mouse α-synuclein to LB- and LN-like inclusions. Seven days later, neurons were fixed and assayed for pathological pS129 α-synuclein and neuron number (NeuN; Figure 5A). Antibodies demonstrated broad efficacy in this assay, with 22 of 41 antibodies causing a greater than 75% reduction in pathology (Figure 5B). The most potent antibody, Syn9048, reduced pathology by a remarkable 97%.

[0195] Thorough characterization of α-synuclein antibodies in multiple assays allowed us to quantitatively compare antibodies and select preferred antibodies for subcloning and further screening (Figures 6A-6B). In certain embodiments, certain antibodies of interest recognize both mouse and human α-synuclein, preferentially bind to LBs in human tissues, exhibit greater affinity for misfolded α-synuclein than monomeric α-synuclein, and reduce PFF-seeded α-synuclein pathology in primary neurons. Based on these criteria, two antibodies (Syn9063 and Syn9048) were selected for subcloning, large-scale production, and in vivo testing.

[0196] Example 6: Subclones retain the properties of the parent clone One of the key features of the selected antibodies was their preference for misfolded α-synuclein. Therefore, supernatants from clones derived during additional subcloning of the Syn9063 and Syn9048 hybridomas were analyzed using a sandwich ELISA platform to identify clones producing the desired antibodies. All but one of the Syn9063 subclones showed a preference for α-synuclein PFFs similar to that of the parent clone (Figure 7A). However, upon clonally expanding, the Syn9063 subclones had very low antibody productivity, which precludes in vivo use, where larger amounts of antibody would be required. All Syn9048 clones showed high selectivity for α-synuclein PFFs (Figure 7B), and clone #3 was used for further studies due to the high antibody productivity obtained from rotary cultures.

[0197] To gain further understanding of the antibody concentration required to effectively inhibit the induction of α-synuclein pathology, purified Syn9048 was diluted across several log concentrations and evaluated as described above in primary hippocampal neuron assays seeded with human α-synuclein PFFs. Syn9048 was measured at an antibody:α-synuclein molar ratio of IC 50At a molar ratio of 0.006 (1:166), Syn9048 dose-dependently reduced neuronal α-synuclein pathology (Figures 7C-7D). Syn9048 showed near-complete inhibition of α-synuclein pathology at molar ratios of 0.03 or greater, suggesting that only one antibody per 33 α-synuclein monomer units is sufficient to completely inhibit α-synuclein seeding in neurons. Syn9048 also inhibited α-synuclein pathology induced by mouse α-synuclein PFFs with a similar IC. 50 (Figure 15), suggesting that Syn9048 is suitable for testing in non-transgenic mouse models of PD.

[0198] Example 7: In vivo immunotherapy is well tolerated and improves dopaminergic tone Several α-synuclein antibodies have been tested in animal models for their ability to prevent PD-like pathology. However, in the absence of prodromal disease biomarkers, it is unlikely that patients with neurodegenerative disorders can receive immunotherapy treatment before the symptomatic stage of the disease, even when brain pathology is already established.

[0199] Therefore, we tested the efficacy of Syn9048 in reducing pathology and rescuing neuronal function after pathology has developed. Using a previously established model of α-synuclein pathology induction and transmission (Luk, et al., 2012, Science 338:949-953), non-transgenic mice were injected with 5 μg of α-synuclein PFFs into the dorsal striatum at 2–3 months of age. Prior to antibody treatment, mice were given a week to allow for BBB recovery and for pathogenic α-synuclein to be internalized by neurons and induce pathology. Mice then received weekly Syn9048 or isotype control antibody treatment (30 mg / kg intraperitoneally) for the following 6 months (Figure 8A, 16 mice / group). An additional group of mice was treated with a comparative antibody, Syn303, previously validated in an in vivo immunotherapy model (Tran, et al., 2014, Cell Reports 7:2054-2065). To minimize inhibition of early PFF seeding in neurons, antibody administration was delayed by one week after intrastriatal PFF injection. In certain embodiments, the site of secondary α-synuclein pathology resulting from pathology transmission is most affected by antibody treatment. All three groups of mice steadily gained weight over the course of the study, suggesting that passive immunotherapy was well tolerated (Figure 8B).

[0200] Dopaminergic neuron loss in the substantia nigra (SN) is a major feature of PD and is recapitulated in PFF-injected mouse models (Henderson, et al., 2019, Nature Neuroscience 22:1248-1257; Luk, et al., 2012, Science 338:949-953) (Figure 8C). IgG1-treated mice had dramatic tyrosine hydroxylase (TH)-positive neuron loss ipsilateral to the injection site, and this neuron loss was not abolished by treatment with either Syn303 or Syn9048 (Figures 8D-8E). Because dopaminergic neuron loss is consistent with these cells being directly associated with the injection site, antibody treatment 1 week after PFF injection would not be expected to block the development of pathology in these neurons. Dopaminergic tone is also reduced in the dorsal striatum in response to PFF injection (Figure 8F). Somewhat surprisingly, Syn9048, but not Syn303, was able to rescue the loss of striatal dopamine and DOPAC (Figures 8F-8G). Thus, while neurons are still lost, the function of remaining neurons can be improved by Syn9048 treatment, likely due to reduced α-synuclein pathology.

[0201] Example 8: Syn9048 reduces α-synuclein pathology in the SN and amygdala To better understand how α-synuclein pathology changes in mice after passive immunotherapy, we performed quantitative pathology analysis of misfolded α-synuclein (Syn506) in the SN (Figures 9A-9C) and amygdala (Figures 9D-9F). Syn303 and Syn9048 reduced mean pathology in the SN contralateral (Figure 9B) and ipsilateral (Figure 9C) to the injection site, but only the reduction by Syn9048 in the ipsilateral SN was statistically significant. Pathology reduction was also more evident in the contralateral amygdala (Figure 9E) than in the ipsilateral amygdala (Figure 9F), and more pronounced in Syn9048-treated animals than in Syn303-treated animals. Only the reduction by Syn9048 in the contralateral amygdala was statistically significant. The reduction of ipsilateral SN pathology by Syn9048, although modest, may account for the improvement in striatal dopamine and DOPAC levels. Moreover, the greater effect of antibody treatment on contralateral α-synuclein pathology suggests that efficacy is greater at sites where extracellular α-synuclein propagation is involved in pathology induction.

[0202] (Table 1) Measurement summary TIFF0007777071000031.tif99169TIFF0007777071000032.tif241169TIFF0007777071000033.tif66169NB=Unbound

[0203] List of modes The following exemplary aspects are provided, the numbering of which should not be construed as indicating any level of importance. Aspect 1 provides the following: An isolated monoclonal antibody comprising a light chain variable region (VL) and a heavy chain variable region (VH), The VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO: 27, 46, 70, 114, 126, 185, 213, or 240; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29, 72, 116, 214, or 242; and a CDR3 region comprising the amino acid sequence of SEQ ID NO: 31, 85, 118, 129, 152, 205, 216, or 244 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO: 11, 58, 92, 101, 144, 160, or 226; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 13, 40, 60, 93, 102, 134, 145, 161, 200, or 228; and a CDR3 region comprising the amino acid sequence of SEQ ID NO: 15, 62, 81, 147, 163, 194, 201, or 230 Including, The isolated monoclonal antibody. Aspect 2 provides the following: The VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO: 27, 46, 70, 114, or 126; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29, 72, or 116; and a CDR3 region comprising the amino acid sequence of SEQ ID NO: 31, 85, 118, 129, or 152 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO: 11, 58, 92, 101, or 144; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 13, 40, 60, 93, 102, 134, or 145; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 15, 62, 81, or 147 Including, The monoclonal antibody of embodiment 1. Aspect 3 provides the following: The VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:70, 185, 213, or 240; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29, 72, 214, or 242; and a CDR3 region comprising the amino acid sequence of SEQ ID NO: 152, 129, 205, 216, or 244 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO: 160, 58, or 226; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 161, 145, 200, or 228; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 163, 194, 201, or 230 Including, The monoclonal antibody of embodiment 1. Aspect 4 provides the following: The monoclonal antibody of any of embodiments 1 to 3, wherein at least one of the following applies: (a) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:27; A CDR2 region comprising the amino acid sequence of SEQ ID NO:29; and CDR3 region comprising the amino acid sequence of SEQ ID NO:31 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:11; A CDR2 region comprising the amino acid sequence of SEQ ID NO:13; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 15 including; (b) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:46; A CDR2 region comprising the amino acid sequence of SEQ ID NO:29; and CDR3 region comprising the amino acid sequence of SEQ ID NO:31 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:11; A CDR2 region comprising the amino acid sequence of SEQ ID NO:40; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 15 including; (c) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:70; A CDR2 region comprising the amino acid sequence of SEQ ID NO:72; and CDR3 region comprising the amino acid sequence of SEQ ID NO:85 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:58; A CDR2 region comprising the amino acid sequence of SEQ ID NO:60; and CDR3 region comprising the amino acid sequence of SEQ ID NO:62 including; (d) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:70; A CDR2 region comprising the amino acid sequence of SEQ ID NO:72; and CDR3 region comprising the amino acid sequence of SEQ ID NO:85 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:58; A CDR2 region comprising the amino acid sequence of SEQ ID NO:60; and CDR3 region comprising the amino acid sequence of SEQ ID NO:81 including; (e) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:70; A CDR2 region comprising the amino acid sequence of SEQ ID NO:72; and CDR3 region comprising the amino acid sequence of SEQ ID NO:85 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:92; a CDR2 region comprising the amino acid sequence of SEQ ID NO:93; and CDR3 region comprising the amino acid sequence of SEQ ID NO:62 including; (f) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:114; A CDR2 region comprising the amino acid sequence of SEQ ID NO:116; and CDR3 region comprising the amino acid sequence of SEQ ID NO:118 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:101; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 102; and CDR3 region comprising the amino acid sequence of SEQ ID NO:62 including; (g) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:126; A CDR2 region comprising the amino acid sequence of SEQ ID NO:72; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 129 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:101; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 102; and CDR3 region comprising the amino acid sequence of SEQ ID NO:62 including; (h) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:46; A CDR2 region comprising the amino acid sequence of SEQ ID NO:29; and CDR3 region comprising the amino acid sequence of SEQ ID NO:31 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:11; A CDR2 region comprising the amino acid sequence of SEQ ID NO:134; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 15 including; (i) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:70; A CDR2 region comprising the amino acid sequence of SEQ ID NO:29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 152 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:144; A CDR2 region comprising the amino acid sequence of SEQ ID NO:145; and CDR3 region comprising the amino acid sequence of SEQ ID NO:147 including; (j) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:70; A CDR2 region comprising the amino acid sequence of SEQ ID NO:29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 152 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:160; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 161; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 163 including; (k) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:185; A CDR2 region comprising the amino acid sequence of SEQ ID NO:29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 129 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:160; A CDR2 region comprising the amino acid sequence of SEQ ID NO:145; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 163 including; (l) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:70; A CDR2 region comprising the amino acid sequence of SEQ ID NO:29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 152 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:160; A CDR2 region comprising the amino acid sequence of SEQ ID NO:145; and CDR3 region comprising the amino acid sequence of SEQ ID NO:194 including; (m) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:70; A CDR2 region comprising the amino acid sequence of SEQ ID NO:72; and CDR3 region comprising the amino acid sequence of SEQ ID NO:205 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:58; A CDR2 region comprising the amino acid sequence of SEQ ID NO:200; and CDR3 region comprising the amino acid sequence of SEQ ID NO:201 including; (n) The VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:213; A CDR2 region comprising the amino acid sequence of SEQ ID NO:214; and CDR3 region comprising the amino acid sequence of SEQ ID NO:216 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:58; A CDR2 region comprising the amino acid sequence of SEQ ID NO:200; and CDR3 region comprising the amino acid sequence of SEQ ID NO:201 including; (o) the VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO:240; A CDR2 region comprising the amino acid sequence of SEQ ID NO:242; and CDR3 region comprising the amino acid sequence of SEQ ID NO:244 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO:226; A CDR2 region comprising the amino acid sequence of SEQ ID NO:228; and CDR3 region comprising the amino acid sequence of SEQ ID NO:230 Including. Aspect 5 provides the following: The VL is SEQ ID NO:27-SEQ ID NO:28-SEQ ID NO:29-SEQ ID NO:30-SEQ ID NO:31, SEQ ID NO:46-SEQ ID NO:28-SEQ ID NO:29-SEQ ID NO:30-SEQ ID NO:31, SEQ ID NO:70-SEQ ID NO:71-SEQ ID NO:72-SEQ ID NO:73-SEQ ID NO:85, SEQ ID NO:70-SEQ ID NO:127-SEQ ID NO:29-SEQ ID NO:151-SEQ ID NO:152, SEQ ID NO:70-SEQ ID NO:127-SEQ ID NO:29-SEQ ID NO:172-SEQ ID NO:152, SEQ ID NO:70-SEQ ID NO:127-SEQ ID NO:72-SEQ ID NO:204-SEQ ID NO:205, SEQ ID NO:114-SEQ ID NO:115-SEQ ID NO:116-SEQ ID NO:117-SEQ ID NO:118, SEQ ID NO:126-SEQ ID NO:127-SEQ ID NO:72-SEQ ID NO:128-SEQ ID NO:129, SEQ ID NO:185-SEQ ID NO:186-SEQ ID NO:29-SEQ ID NO:187-SEQ ID NO:129, SEQ ID NO:213-SEQ ID NO:115-SEQ ID NO:214-SEQ ID NO:215-SEQ ID NO:216, or SEQ ID NO:240-SEQ ID NO:241-SEQ ID NO:242-SEQ ID NO:243-SEQ ID NO:244 comprising the amino acid sequence and the VH is SEQ ID NO:11-SEQ ID NO:12-SEQ ID NO:13-SEQ ID NO:14-SEQ ID NO:15, SEQ ID NO:11-SEQ ID NO:12-SEQ ID NO:40-SEQ ID NO:41-SEQ ID NO:15, SEQ ID NO:11-SEQ ID NO:12-SEQ ID NO:134-SEQ ID NO:135-SEQ ID NO:15, SEQ ID NO:58-SEQ ID NO:59-SEQ ID NO:60-SEQ ID NO:61-SEQ ID NO:62, SEQ ID NO:58-SEQ ID NO:59-SEQ ID NO:60-SEQ ID NO:80-SEQ ID NO:81, SEQ ID NO:58-SEQ ID NO:59-SEQ ID NO:200-SEQ ID NO:94-SEQ ID NO:201, SEQ ID NO:92-SEQ ID NO:59-SEQ ID NO:93-SEQ ID NO:94-SEQ ID NO:62, SEQ ID NO:101-SEQ ID NO:59-SEQ ID NO:102-SEQ ID NO:103-SEQ ID NO:62, SEQ ID NO:144-SEQ ID NO:59-SEQ ID NO:145-SEQ ID NO:146-SEQ ID NO:147, SEQ ID NO:160-SEQ ID NO:59-SEQ ID NO:145-SEQ ID NO:177-SEQ ID NO:163, SEQ ID NO:160-SEQ ID NO:59-SEQ ID NO:145-SEQ ID NO:193-SEQ ID NO:194, SEQ ID NO:160-SEQ ID NO:59-SEQ ID NO:161-SEQ ID NO:162-SEQ ID NO:163, or SEQ ID NO:226-SEQ ID NO:227-SEQ ID NO:228-SEQ ID NO:229-SEQ ID NO:230 comprising the amino acid sequence The monoclonal antibody according to any one of aspects 1 to 4. Aspect 6 provides the following: The VL is SEQ ID NO:27-SEQ ID NO:28-SEQ ID NO:29-SEQ ID NO:30-SEQ ID NO:31, SEQ ID NO:46-SEQ ID NO:28-SEQ ID NO:29-SEQ ID NO:30-SEQ ID NO:31, SEQ ID NO:70-SEQ ID NO:71-SEQ ID NO:72-SEQ ID NO:73-SEQ ID NO:85, SEQ ID NO:70-SEQ ID NO:127-SEQ ID NO:29-SEQ ID NO:151-SEQ ID NO:152, SEQ ID NO:114-SEQ ID NO:115-SEQ ID NO:116-SEQ ID NO:117-SEQ ID NO:118, or SEQ ID NO:126-SEQ ID NO:127-SEQ ID NO:72-SEQ ID NO:128-SEQ ID NO:129 comprising the amino acid sequence and the VH is SEQ ID NO:11-SEQ ID NO:12-SEQ ID NO:13-SEQ ID NO:14-SEQ ID NO:15, SEQ ID NO:11-SEQ ID NO:12-SEQ ID NO:40-SEQ ID NO:41-SEQ ID NO:15, SEQ ID NO:11-SEQ ID NO:12-SEQ ID NO:134-SEQ ID NO:135-SEQ ID NO:15, SEQ ID NO:58-SEQ ID NO:59-SEQ ID NO:60-SEQ ID NO:61-SEQ ID NO:62, SEQ ID NO:58-SEQ ID NO:59-SEQ ID NO:60-SEQ ID NO:80-SEQ ID NO:81, SEQ ID NO:92-SEQ ID NO:59-SEQ ID NO:93-SEQ ID NO:94-SEQ ID NO:62, SEQ ID NO:101-SEQ ID NO:59-SEQ ID NO:102-SEQ ID NO:103-SEQ ID NO:62, or SEQ ID NO:144-SEQ ID NO:59-SEQ ID NO:145-SEQ ID NO:146-SEQ ID NO:147 comprising the amino acid sequence The monoclonal antibody according to any one of embodiments 1 to 5. Aspect 7 provides the following: The VL is SEQ ID NO:70-SEQ ID NO:127-SEQ ID NO:29-SEQ ID NO:172-SEQ ID NO:152, SEQ ID NO:70-SEQ ID NO:127-SEQ ID NO:72-SEQ ID NO:204-SEQ ID NO:205, SEQ ID NO:185-SEQ ID NO:186-SEQ ID NO:29-SEQ ID NO:187-SEQ ID NO:129, SEQ ID NO:213-SEQ ID NO:115-SEQ ID NO:214-SEQ ID NO:215-SEQ ID NO:216, or SEQ ID NO:240-SEQ ID NO:241-SEQ ID NO:242-SEQ ID NO:243-SEQ ID NO:244 comprising the amino acid sequence and the VH is SEQ ID NO:58-SEQ ID NO:59-SEQ ID NO:200-SEQ ID NO:94-SEQ ID NO:201, SEQ ID NO:160-SEQ ID NO:59-SEQ ID NO:145-SEQ ID NO:177-SEQ ID NO:163, SEQ ID NO:160-SEQ ID NO:59-SEQ ID NO:145-SEQ ID NO:193-SEQ ID NO:194, SEQ ID NO:160-SEQ ID NO:59-SEQ ID NO:161-SEQ ID NO:162-SEQ ID NO:163, or SEQ ID NO:226-SEQ ID NO:227-SEQ ID NO:228-SEQ ID NO:229-SEQ ID NO:230 comprising the amino acid sequence The monoclonal antibody according to any one of embodiments 1 to 5. Aspect 8 provides the following: The monoclonal antibody of any of embodiments 1 to 7, which is humanized. Aspect 9 provides the following: The monoclonal antibody of any one of embodiments 1 to 8, which is labeled. Aspect 10 provides the following: A pharmaceutical composition comprising the monoclonal antibody of any of embodiments 1 to 9 and at least one pharmaceutical excipient. Aspect 11 provides the following: 1. An isolated polynucleotide comprising at least one of the nucleic acid sequences of SEQ ID NOs:3, 5, 7, 19, 21, 23, 35, 43, 44, 50, 52, 54, 66, 68, 75, 77, 88, 89, 96, 97, 99, 106, 108, 110, 121, 132, 138, 139, 141, 148, 150, 155, 157, 167, 168, 170, 175, 180, 191, 195, 197, 198, 203, 207, 209, 211, 219, 221, 223, 233, 235, and 237. Aspect 12 provides the following: At least one nucleic acid sequence selected from the group consisting of SEQ ID NOs: 3, 5, 7, 35, 50, 52, 54, 75, 77, 88, 89, 97, 99, 132, 138, 139, 141, 155, 157, 175, 191, 197, 198, 219, 221, and 223; and At least one nucleic acid sequence selected from the group consisting of SEQ ID NOs: 19, 21, 23, 43, 44, 66, 68, 96, 106, 108, 110, 121, 148, 150, 167, 168, 170, 180, 195, 203, 207, 209, 211, 233, 235, and 237. 12. The isolated polynucleotide of embodiment 11, comprising: Aspect 13 provides the following: At least one nucleic acid sequence selected from the group consisting of SEQ ID NOs:3, 5, 7; SEQ ID NOs:3, 35, 7; SEQ ID NOs:3, 132, 7; SEQ ID NOs:50, 52, 54; SEQ ID NOs:50, 75, 77; SEQ ID NOs:50, 139, 175; SEQ ID NOs:50, 139, 191; SEQ ID NOs:50, 155, 157; SEQ ID NOs:50, 197, 198; SEQ ID NOs:88, 89, 54; SEQ ID NOs:97, 99, 54; SEQ ID NOs:138, 139, 141; and SEQ ID NOs:219, 221, 223; and At least one nucleic acid sequence selected from the group consisting of SEQ ID NOs: 19, 21, 23; SEQ ID NOs: 19, 43, 44; SEQ ID NOs: 19, 66, 68; SEQ ID NOs: 19, 66, 96; SEQ ID NOs: 19, 66, 203; SEQ ID NOs: 19, 148, 150; SEQ ID NOs: 106, 108, 110; SEQ ID NOs: 121, 66, 68; SEQ ID NOs: 167, 168, 170; SEQ ID NOs: 167, 168, 195; SEQ ID NOs: 180, 168, 68; SEQ ID NOs: 213, 214, 216; and SEQ ID NOs: 233, 235, 237. 13. The isolated polynucleotide of any of embodiments 11-12, comprising: Aspect 14 provides the following: 1. A method of treating, ameliorating, and / or preventing a synucleinopathy in a subject in need thereof, comprising: The method, comprising the step of administering to the subject a therapeutically effective amount of at least one isolated monoclonal antibody according to any one of aspects 1 to 9. Aspect 15 provides the following: 15. The method of embodiment 14, wherein said synucleinopathy is at least one from the group consisting of Parkinson's disease, Parkinson's disease with dementia, dementia with Lewy bodies, Alzheimer's disease, Down syndrome, multiple system atrophy, prion diseases, and other α-Syn-related neurodegenerative disorders. Aspect 16 provides the following: The method of any of aspects 14 to 15, wherein the antibody is provided to the subject as a pharmaceutical composition. Aspect 17 provides the following: The method of any of aspects 14 to 16, wherein the antibody is administered to the subject parenterally. Aspect 18 provides the following: 1. A method for detecting a synucleinopathy in a subject, comprising: administering to the subject at least one labeled isolated monoclonal antibody of any of Aspects 1-9, and detecting the presence or absence of complexes between the labeled isolated monoclonal antibody and any α-syn fibrils, oligomers, and / or other misfolded α-syn species present in the subject; If the complex is detected, the subject has a synucleinopathy. The method. Aspect 19 provides the following: 1. A method for detecting total α-Syn, α-Syn fibril, and / or α-Syn oligomeric species in a sample, comprising: contacting the sample with at least one labeled isolated monoclonal antibody of any of Aspects 1-9; and detecting the presence or absence of complexes between the labeled isolated monoclonal antibody and total α-Syn, α-Syn monomers, α-Syn fibrils, and / or α-Syn oligomeric species present in the sample; If the complex is detected, total α-Syn, α-Syn monomer, α-Syn fibril, and / or α-Syn oligomer species are present in the sample. The method. Aspect 20 provides the following: 20. The method of embodiment 19, wherein said sample comprises an in vitro and / or ex vivo sample. Aspect 21 provides the following: An autonomously replicating or integrating mammalian cell vector comprising a recombinant nucleic acid encoding an antibody comprising a light chain variable region (VL) and a heavy chain variable region (VH), The VL is a CDR1 region comprising the amino acid sequence of SEQ ID NO: 27, 46, 70, 114, 126, 185, 213, or 240; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29, 72, 116, 214, or 242; and a CDR3 region comprising the amino acid sequence of SEQ ID NO: 31, 85, 118, 129, 152, 205, 216, or 244 Includes; and the VH is a CDR1 region comprising the amino acid sequence of SEQ ID NO: 11, 58, 92, 101, 144, 160, or 226; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 13, 40, 60, 93, 102, 134, 145, 161, 200, or 228; and a CDR3 region comprising the amino acid sequence of SEQ ID NO: 15, 62, 81, 147, 163, 194, 201, or 230 Including, The vector. Aspect 22 provides the following: 22. The cellular vector of embodiment 21, comprising a plasmid or a virus. Aspect 23 provides the following: 23. The cell vector of any of embodiments 21 to 22, comprising a mammalian cell expression vector. Aspect 24 provides the following: 24. The cellular vector of any of embodiments 21 to 23, further comprising at least one nucleic acid sequence that directs and / or controls the expression of said antibody. Embodiment 25 provides the following: 25. An isolated host cell comprising at least one vector of any of aspects 21 to 24. Embodiment 26 provides the following: 26. The host cell of embodiment 25, which is a non-human cell. Aspect 27 provides the following: 27. The cell vector of any one of embodiments 25 to 26, which is a mammalian cell.

[0204] The disclosures of each and every patent, patent application and publication cited herein are hereby incorporated by reference in their entirety.

[0205] While the present disclosure has been disclosed with reference to specific embodiments, it will be apparent that other embodiments and variations of the present disclosure may be devised by those skilled in the art without departing from the true spirit and scope of the present disclosure, and it is intended that the appended claims be construed to include all such embodiments and equivalent variations.

Claims

1. A monoclonal antibody against alpha-synuclein, comprising a light chain variable region (VL) and a heavy chain variable region (VH), to which at least one of the following applies: (a) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 27; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 31 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 11; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 13; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 15 containing; (b) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 46; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 31 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 11; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 40; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 15 containing; (c) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 72; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 85 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 58; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 60; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 62 containing; (d) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 72; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 85 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 58; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 60; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 81 containing; (e) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 72; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 85 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 92; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 93; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 62 containing; (f) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 114; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 116; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 118 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 101; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 102; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 62 containing; (g) The VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 126; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 72; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 129 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 101; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 102; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 62 containing; (h) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 46; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 31 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 11; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 134; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 15 containing; (i) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 152 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 144; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 145; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 147 containing; (j) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 152 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 160; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 161; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 163 containing; (k) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 185; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 129 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 160; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 145; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 163 containing; (l) The VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 152 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 160; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 145; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 194 containing; (m) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 72; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 205 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 58; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 200; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 201 containing; (n) The VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 213; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 214; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 216 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 58; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 200; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 201 Including.

2. The VL is SEQ ID NO: 27-SEQ ID NO: 28-SEQ ID NO: 29-SEQ ID NO: 30-SEQ ID NO: 31, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 46-SEQ ID NO: 28-SEQ ID NO: 29-SEQ ID NO: 30-SEQ ID NO: 31, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 70-SEQ ID NO: 71-SEQ ID NO: 72-SEQ ID NO: 73-SEQ ID NO: 85, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 70-SEQ ID NO: 127-SEQ ID NO: 29-SEQ ID NO: 151-SEQ ID NO: 152, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 70-SEQ ID NO: 127-SEQ ID NO: 29-SEQ ID NO: 172-SEQ ID NO: 152, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 70-SEQ ID NO: 127-SEQ ID NO: 72-SEQ ID NO: 204-SEQ ID NO: 205, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 114-SEQ ID NO: 115-SEQ ID NO: 116-SEQ ID NO: 117-SEQ ID NO: 118, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 126-SEQ ID NO: 127-SEQ ID NO: 72-SEQ ID NO: 128-SEQ ID NO: 129, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO:185-SEQ ID NO:186-SEQ ID NO:29-SEQ ID NO:187-SEQ ID NO:129, wherein each sequence is arranged consecutively in the order listed; or SEQ ID NO: 213-SEQ ID NO: 115-SEQ ID NO: 214-SEQ ID NO: 215-SEQ ID NO: 216, where each sequence is arranged consecutively in the order listed. comprising the amino acid sequence and the VH is SEQ ID NO: 11-SEQ ID NO: 12-SEQ ID NO: 13-SEQ ID NO: 14-SEQ ID NO: 15, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 11-SEQ ID NO: 12-SEQ ID NO: 40-SEQ ID NO: 41-SEQ ID NO: 15, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 11-SEQ ID NO: 12-SEQ ID NO: 134-SEQ ID NO: 135-SEQ ID NO: 15, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 58-SEQ ID NO: 59-SEQ ID NO: 60-SEQ ID NO: 61-SEQ ID NO: 62, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 58-SEQ ID NO: 59-SEQ ID NO: 60-SEQ ID NO: 80-SEQ ID NO: 81, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 58-SEQ ID NO: 59-SEQ ID NO: 200-SEQ ID NO: 94-SEQ ID NO: 201, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 92-SEQ ID NO: 59-SEQ ID NO: 93-SEQ ID NO: 94-SEQ ID NO: 62, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 101-SEQ ID NO: 59-SEQ ID NO: 102-SEQ ID NO: 103-SEQ ID NO: 62, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 144-SEQ ID NO: 59-SEQ ID NO: 145-SEQ ID NO: 146-SEQ ID NO: 147, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 160-SEQ ID NO: 59-SEQ ID NO: 145-SEQ ID NO: 177-SEQ ID NO: 163, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 160-SEQ ID NO: 59-SEQ ID NO: 145-SEQ ID NO: 193-SEQ ID NO: 194, wherein each sequence is arranged consecutively in the order listed; or SEQ ID NO: 160-SEQ ID NO: 59-SEQ ID NO: 161-SEQ ID NO: 162-SEQ ID NO: 163, where each sequence is arranged consecutively in the order listed. comprising the amino acid sequence The monoclonal antibody of claim 1.

3. The VL is SEQ ID NO: 27-SEQ ID NO: 28-SEQ ID NO: 29-SEQ ID NO: 30-SEQ ID NO: 31, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 46-SEQ ID NO: 28-SEQ ID NO: 29-SEQ ID NO: 30-SEQ ID NO: 31, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 70-SEQ ID NO: 71-SEQ ID NO: 72-SEQ ID NO: 73-SEQ ID NO: 85, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 70-SEQ ID NO: 127-SEQ ID NO: 29-SEQ ID NO: 151-SEQ ID NO: 152, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 114-SEQ ID NO: 115-SEQ ID NO: 116-SEQ ID NO: 117-SEQ ID NO: 118, wherein each sequence is arranged consecutively in the order listed; or SEQ ID NO: 126-SEQ ID NO: 127-SEQ ID NO: 72-SEQ ID NO: 128-SEQ ID NO: 129, wherein each sequence is arranged consecutively in the order listed; comprising the amino acid sequence and the VH is SEQ ID NO: 11-SEQ ID NO: 12-SEQ ID NO: 13-SEQ ID NO: 14-SEQ ID NO: 15, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 11-SEQ ID NO: 12-SEQ ID NO: 40-SEQ ID NO: 41-SEQ ID NO: 15, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 11-SEQ ID NO: 12-SEQ ID NO: 134-SEQ ID NO: 135-SEQ ID NO: 15, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 58-SEQ ID NO: 59-SEQ ID NO: 60-SEQ ID NO: 61-SEQ ID NO: 62, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 58-SEQ ID NO: 59-SEQ ID NO: 60-SEQ ID NO: 80-SEQ ID NO: 81, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 92-SEQ ID NO: 59-SEQ ID NO: 93-SEQ ID NO: 94-SEQ ID NO: 62, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO:101-SEQ ID NO:59-SEQ ID NO:102-SEQ ID NO:103-SEQ ID NO:62, wherein each sequence is arranged consecutively in the order listed; or SEQ ID NO: 144-SEQ ID NO: 59-SEQ ID NO: 145-SEQ ID NO: 146-SEQ ID NO: 147, wherein each sequence is arranged consecutively in the order listed; comprising the amino acid sequence The monoclonal antibody of claim 1.

4. The VL is SEQ ID NO: 70-SEQ ID NO: 71-SEQ ID NO: 72-SEQ ID NO: 73-SEQ ID NO: 85, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 70-SEQ ID NO: 127-SEQ ID NO: 29-SEQ ID NO: 172-SEQ ID NO: 152, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 70-SEQ ID NO: 127-SEQ ID NO: 72-SEQ ID NO: 204-SEQ ID NO: 205, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO:185-SEQ ID NO:186-SEQ ID NO:29-SEQ ID NO:187-SEQ ID NO:129, wherein each sequence is arranged consecutively in the order listed; or SEQ ID NO: 213-SEQ ID NO: 115-SEQ ID NO: 214-SEQ ID NO: 215-SEQ ID NO: 216, where each sequence is arranged consecutively in the order listed. comprising the amino acid sequence and the VH is SEQ ID NO: 58-SEQ ID NO: 59-SEQ ID NO: 60-SEQ ID NO: 80-SEQ ID NO: 81, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 58-SEQ ID NO: 59-SEQ ID NO: 200-SEQ ID NO: 94-SEQ ID NO: 201, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 160-SEQ ID NO: 59-SEQ ID NO: 145-SEQ ID NO: 177-SEQ ID NO: 163, wherein each sequence is arranged consecutively in the order listed; SEQ ID NO: 160-SEQ ID NO: 59-SEQ ID NO: 145-SEQ ID NO: 193-SEQ ID NO: 194, wherein each sequence is arranged consecutively in the order listed; or SEQ ID NO: 160-SEQ ID NO: 59-SEQ ID NO: 161-SEQ ID NO: 162-SEQ ID NO: 163, where each sequence is arranged consecutively in the order listed. comprising the amino acid sequence The monoclonal antibody of claim 1.

5. The monoclonal antibody of any one of claims 1 to 4, which is humanized.

6. The monoclonal antibody of any one of claims 1 to 5, which is labeled.

7. A pharmaceutical composition comprising the monoclonal antibody of any one of claims 1 to 6 and at least one pharmaceutical excipient.

8. 1. An isolated polynucleotide encoding a heavy chain variable region (VH) and a light chain variable region (VL) of a monoclonal antibody against alpha-synuclein, comprising: (a) a CDR1 region comprising the amino acid sequence of SEQ ID NO: 27; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 31 a first isolated polynucleotide encoding a VL comprising: CDR1 region comprising the amino acid sequence of SEQ ID NO: 11; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 13; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 15 a second isolated polynucleotide encoding a VH comprising: (b) a CDR1 region comprising the amino acid sequence of SEQ ID NO: 46; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 31 a first isolated polynucleotide encoding a VL comprising: CDR1 region comprising the amino acid sequence of SEQ ID NO: 11; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 40; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 15 a second isolated polynucleotide encoding a VH comprising: (c) a CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 72; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 85 a first isolated polynucleotide encoding a VL comprising: CDR1 region comprising the amino acid sequence of SEQ ID NO: 58; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 60; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 62 a second isolated polynucleotide encoding a VH comprising: (d) a CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 72; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 85 a first isolated polynucleotide encoding a VL comprising: CDR1 region comprising the amino acid sequence of SEQ ID NO: 58; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 60; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 81 a second isolated polynucleotide encoding a VH comprising: (e) a CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 72; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 85 a first isolated polynucleotide encoding a VL comprising: CDR1 region comprising the amino acid sequence of SEQ ID NO: 92; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 93; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 62 a second isolated polynucleotide encoding a VH comprising: (f) a CDR1 region comprising the amino acid sequence of SEQ ID NO: 114; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 116; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 118 a first isolated polynucleotide encoding a VL comprising: CDR1 region comprising the amino acid sequence of SEQ ID NO: 101; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 102; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 62 a second isolated polynucleotide encoding a VH comprising: (g) a CDR1 region comprising the amino acid sequence of SEQ ID NO: 126; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 72; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 129 a first isolated polynucleotide encoding a VL comprising: CDR1 region comprising the amino acid sequence of SEQ ID NO: 101; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 102; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 62 a second isolated polynucleotide encoding a VH comprising: (h) a CDR1 region comprising the amino acid sequence of SEQ ID NO: 46; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 31 a first isolated polynucleotide encoding a VL comprising: CDR1 region comprising the amino acid sequence of SEQ ID NO: 11; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 134; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 15 a second isolated polynucleotide encoding a VH comprising: (i) a CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 152 a first isolated polynucleotide encoding a VL comprising: CDR1 region comprising the amino acid sequence of SEQ ID NO: 144; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 145; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 147 a second isolated polynucleotide encoding a VH comprising: (j) a CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 152 a first isolated polynucleotide encoding a VL comprising: CDR1 region comprising the amino acid sequence of SEQ ID NO: 160; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 161; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 163 a second isolated polynucleotide encoding a VH comprising: (k) a CDR1 region comprising the amino acid sequence of SEQ ID NO: 185; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 129 a first isolated polynucleotide encoding a VL comprising: CDR1 region comprising the amino acid sequence of SEQ ID NO: 160; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 145; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 163 a second isolated polynucleotide encoding a VH comprising: (l) a CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 152 a first isolated polynucleotide encoding a VL comprising: CDR1 region comprising the amino acid sequence of SEQ ID NO: 160; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 145; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 194 a second isolated polynucleotide encoding a VH comprising: (m) a CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 72; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 205 a first isolated polynucleotide encoding a VL comprising: CDR1 region comprising the amino acid sequence of SEQ ID NO: 58; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 200; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 201 a second isolated polynucleotide encoding a VH comprising: (n) a CDR1 region comprising the amino acid sequence of SEQ ID NO: 213; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 214; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 216 a first isolated polynucleotide encoding a VL comprising: CDR1 region comprising the amino acid sequence of SEQ ID NO: 58; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 200; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 201 a second isolated polynucleotide encoding a VH comprising An isolated polynucleotide comprising:

9. A pharmaceutical composition comprising the isolated monoclonal antibody of any one of claims 1 to 6 for the treatment, amelioration and / or prevention of a synucleopathic disease.

10. 10. The pharmaceutical composition of claim 9, wherein the synucleinopathy is at least one from the group consisting of Parkinson's disease, Parkinson's disease with dementia, dementia with Lewy bodies, Alzheimer's disease, Down's syndrome, multiple system atrophy, prion diseases, and other alpha-Syn-related neurodegenerative disorders.

11. 11. The pharmaceutical composition of claim 9 or 10, formulated for parenteral administration.

12. 10. A pharmaceutical composition comprising the labeled isolated monoclonal antibody of claim 6 for use in a method for detecting a synucleinopathy in a subject, the method comprising: administering to the subject at least one labeled isolated monoclonal antibody of claim 6; and detecting the presence or absence of complexes between the labeled isolated monoclonal antibody and any α-syn fibrils, oligomers, and / or other misfolded α-syn species present in the subject; If the complex is detected, the subject has a synucleinopathy. The pharmaceutical composition.

13. 1. A method for detecting total α-Syn, α-Syn fibril, and / or α-Syn oligomeric species in a sample, comprising: contacting the sample with at least one labeled isolated monoclonal antibody of claim 6; and detecting the presence or absence of complexes between the labeled isolated monoclonal antibody and total α-Syn, α-Syn monomers, α-Syn fibrils, and / or α-Syn oligomeric species present in the sample; If the complex is detected, total α-Syn, α-Syn monomer, α-Syn fibril, and / or α-Syn oligomer species are present in the sample. The method.

14. 14. The method of claim 13, wherein the sample comprises an in vitro and / or ex vivo sample.

15. An autonomously replicating or integrating mammalian cell vector comprising a recombinant nucleic acid encoding an antibody against alpha-synuclein comprising a light chain variable region (VL) and a heavy chain variable region (VH), wherein at least one of the following applies: (a) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 27; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 31 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 11; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 13; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 15 containing; (b) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 46; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 31 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 11; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 40; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 15 containing; (c) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 72; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 85 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 58; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 60; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 62 containing; (d) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 72; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 85 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 58; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 60; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 81 containing; (e) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 72; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 85 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 92; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 93; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 62 containing; (f) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 114; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 116; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 118 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 101; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 102; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 62 containing; (g) The VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 126; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 72; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 129 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 101; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 102; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 62 containing; (h) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 46; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 31 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 11; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 134; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 15 containing; (i) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 152 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 144; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 145; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 147 containing; (j) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 152 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 160; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 161; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 163 containing; (k) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 185; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 129 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 160; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 145; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 163 containing; (l) The VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 29; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 152 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 160; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 145; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 194 containing; (m) the VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 70; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 72; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 205 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 58; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 200; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 201 containing; (n) The VL is CDR1 region comprising the amino acid sequence of SEQ ID NO: 213; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 214; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 216 Including; and the VH is CDR1 region comprising the amino acid sequence of SEQ ID NO: 58; A CDR2 region comprising the amino acid sequence of SEQ ID NO: 200; and CDR3 region comprising the amino acid sequence of SEQ ID NO: 201 Including.

16. 16. The vector of claim 15, comprising a plasmid or a virus.

17. 16. The vector of claim 15, comprising a mammalian cell expression vector.

18. 16. The vector of claim 15, further comprising at least one nucleic acid sequence that directs and / or controls the expression of the antibody.

19. An isolated host cell comprising at least one vector according to any one of claims 15 to 18.

20. 20. The cell of claim 19, which is a non-human cell.

21. 20. The cell of claim 19, which is a mammalian cell.

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