JP2023018038A - Dosing regimes for treatment of synucleinopathies - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a method of treating or effecting prophylaxis of a subject having or at risk of a synucleinopathy.

SOLUTION: A method comprises intravenously administering to a subject a dose of 3000-5000 mg of an antibody against alpha-synuclein at intervals of 3-5 weeks, or administering another regime that delivers the antibody to the subject with substantially the same area under the curve.

SELECTED DRAWING: None

COPYRIGHT: (C)2023,JPO&INPIT

Description

SEQUENCE LISTING A Sequence Listing containing SEQ ID NOs: 1-151 is attached and is incorporated by reference in its entirety. 503725 SEQ LIST. txt was created on September 27, 2017 in ASCII format and is 127,151 bytes in size.

Lewy body disease (LBD) is characterized by degeneration of the dopaminergic system, motor changes, cognitive impairment, and the formation of Lewy bodies (LBs) and/or Lewy neurites. [McKeith et al., Neurology (1996) 47:1113-24]. Lewy body disease includes Parkinson's disease (including idiopathic Parkinson's disease), and dementia with Lewy bodies (DLB). Lewy body disease is a common cause of motor impairment and cognitive decline in the aging population [Galasko et al., Arch. Neurol. (1994) 51:888-95]. Constipation is another common symptom in subjects with Lewy body disease [Ondo et al., Neurology 2012;78;1650-1654; Ashraf et al., Movement Disorders 12, 946-951 (1997)]. .

Alpha-synuclein is a protein normally associated with synapses and is thought to play a role in neuroplasticity, learning and memory. Alpha-synuclein has also been implicated in a central role in the pathogenesis of Lewy body disease. The protein can aggregate to form insoluble fibrils in pathological conditions. For example, alpha-synuclein accumulates in LB [Spillantini et al., Nature (1997) 388:839-40; Takeda et al., J. Pathol. (1998) 152:367-72; Wakabayashi et al., Neurosci. Lett. (1997) 239:45-8]. Mutations in the alpha-synuclein gene co-segregate with rare familial forms of Parkinsonism [Kruger et al., Nature Gen. (1998) 18:106-8; Polymeropoulos, et al., Science (1997) 276 :2045-7]. Overexpression of alpha-synuclein in transgenic mice [Masliah et al., Science (2000) 287:1265-9] and fruit flies [Feany et al., Nature (2000) 404:394-8] has been associated with Lewy body disease. mimics several pathological aspects of Soluble oligomers of alpha-synuclein can be neurotoxic [Conway et al., Proc Natl Acad Sci USA (2000) 97:571-576]. Accumulation of alpha-synuclein with similar morphological and neurological changes in diverse species and animal models such as humans, mice, and flies suggests that this molecule contributes to the development of Lewy body disease do.

Immunotherapy directed to alpha-synuclein has been reported to reduce alpha-synuclein deposition and behavioral symptoms in a mouse model of Lewy body disease [Masliah et al., PLoS ONE 6(4): e19338 doi:10.1371/journal.pone.0019338].

In one aspect, the invention provides a method of performing treatment or prophylaxis in a subject having or at risk for a synucleinopathic disease, wherein the subject is administered an antibody against alpha-synuclein at a dose of 3000-5000 mg. Methods are provided comprising administering intravenously at weekly intervals or administering another regimen that delivers the antibody in substantially the same area under the curve to the subject. In some such methods the dose is 3500-4500 mg.

In some such methods, an antibody is administered to multiple subjects having or at risk of having a synucleinopathic disease, and the subjects receive one or two fixed doses. In some such methods, subjects weighing less than 65 kg receive a dose of 3500 mg of antibody and subjects weighing 65 kg or greater receive a dose of 4500 mg.

In some such methods, the dose is 45-75 mg/kg. In some such methods, the dose is 50-70 mg/kg.

In some such methods the interval is 28 days. In some such methods, the subject receives at least 6 doses of the antibody at 3-5 week intervals. In some such methods, the subject receives at least 12 doses of the antibody at 3-5 week intervals. In some such methods, the subject receives at least 18 doses of the antibody at 3-5 week intervals. In some such methods the interval is 4 weeks. In some such methods, the subject receives the antibody every 4 weeks for at least 52 weeks.

Some such methods further comprise monitoring the subject for changes in movement, cognitive impairment, autonomic dysfunction, gastrointestinal dysfunction, visual hallucinations, or psychological symptoms.

In some such methods, monitoring
(a) a mobile device programmed to receive and transmit data obtained from sensors internal and/or external to the mobile device regarding movement disorders in a subject having or suspected of having a synucleinopathic disease; providing to a subject, after which the subject performs a series of exercises to reveal a movement disorder, if any, and sensors internal or external to the device acquire data on the movements;
(b) collecting data transmitted from the mobile device; and (c) comparing the data obtained from the subject to control data to assess the presence or extent of movement disorders in the subject.

In some such methods, the mobile device is programmed to receive and transmit data from at least two external sensors attached to the subject's upper and lower extremities. In some such methods, the mobile device acquires data from sensors on the subject's upper and lower extremities. In some such methods, a mobile device is carried by the subject and acquires data from internal sensors. In some such methods, the sequence of movements includes tapping the device, sitting and standing. In some such methods, monitoring indicates a reduction in motor impairment or a reduction in cognitive impairment in response to administering the antibody.

In some such methods, the antibody binds within residues 115-130 of alpha-synuclein.

In some such methods, the antibody binds within residues 118-126 of alpha-synuclein.

In some such methods, the antibody binds within residues 117-123 of alpha-synuclein. In some such methods, the antibody is NI-202.21D11. In some such methods, the antibody comprises three heavy chain CDRs designated as SEQ ID NOs:139-141, respectively, and three light chain CDRs designated as SEQ ID NOs:143-145, respectively. In some such methods, the antibody comprises a heavy chain variable region set forth in SEQ ID NO:138 and a light chain variable region set forth in SEQ ID NO:142.

In some such methods, the antibody comprises the three light chain Kabat CDRs of SEQ ID NO:5 and the three heavy chain Kabat CDRs of SEQ ID NO:10. In some such methods, the antibody comprises a heavy chain variable region of any of SEQ ID NOs:8-11 and a light chain variable region of any of SEQ ID NOs:3-5, preferably the heavy chain variable region is The heavy and light chain variable region of SEQ ID NO:10 is the light chain variable region of SEQ ID NO:5.

In some such methods, the antibody is of human IgG1 isotype. In some such methods, the antibody comprises a heavy chain constant region of SEQ ID NO:35 and a light chain constant region of SEQ ID NO:30, optionally absent the C-terminal lysine of SEQ ID NO:35. In some such methods, the antibody comprises a heavy chain constant region of SEQ ID NO:35 and a light chain constant region of SEQ ID NO:13, optionally absent the C-terminal lysine of SEQ ID NO:35.

In some such methods, the antibody is 9E4. In some such methods, the antibody comprises three heavy chain CDRs designated in SEQ ID NOs:146-148, respectively, and three light chain CDRs designated in SEQ ID NOs:149-151, respectively. In some such methods, the antibody comprises a heavy chain designated as SEQ ID NO:37 and a light chain designated as SEQ ID NO:32, and the C-terminal lysine of SEQ ID NO:37 may be absent.

In some such methods, the antibody binds within residues 1-20 of alpha-synuclein. In some such methods, the antibody binds within residues 4-15 of alpha-synuclein.

In some such methods, the antibody is NI-202.12F4. In some such methods, the antibody comprises three heavy chain CDRs designated as SEQ ID NOs: 131-133, respectively, and three light chain CDRs designated as SEQ ID NOs: 135-137, respectively. In some such methods, the antibody comprises a heavy chain variable region set forth in SEQ ID NO:130 and a light chain variable region set forth in SEQ ID NO:134.

In some such methods, the subject has a body weight of less than 65 kg and is administered a dose of 3500 mg of antibody every 4 weeks.

In some such methods, a loading dose of 2000 mg of antibody is administered prior to administration of antibody in the range of 3000-5000 mg, and 2000 mg or more but less than 3500 mg until a dose of 3500 mg is reached. Uptitration may be performed in one or more subsequent doses, all doses separated by intervals of 3-5 weeks.

In some such methods, the 3500 mg dose is administered at the next interval after the 2000 mg dose. In some such methods, the dose is gradually increased over multiple intervals prior to administration of the 3500 mg dose. In some such methods, the subject has a body weight greater than 65 kg and is administered a dose of 4500 mg of antibody every 4 weeks.

In some such methods, a loading dose of 2000 mg of antibody is administered prior to administration of 4500 mg of antibody, and one or more doses of 2000 mg or more but less than 4500 mg are administered until a dose of 4500 mg is reached. Escalating titrations with subsequent doses may be performed, all doses separated by intervals of 3-5 weeks.

In some such methods, the 4500 mg dose is administered at the next interval after the 2000 mg dose. In some such methods, the dose is gradually increased over multiple intervals prior to administration of the 4500 mg dose.

Some such methods further comprise monitoring the subject for changes in movement, cognitive impairment, autonomic dysfunction, gastrointestinal dysfunction, visual hallucinations, or psychological symptoms.

In some such methods, monitoring
(a) a mobile device programmed to receive and transmit data obtained from sensors internal and/or external to the mobile device regarding movement disorders in a subject having or suspected of having a synucleinopathic disease; providing to a subject, after which the subject performs a series of exercises to reveal a movement disorder, if any, and sensors internal or external to the device acquire data on the movements;
(b) collecting data transmitted from the mobile device; and (c) comparing the data obtained from the subject to control data to assess the presence or extent of movement disorders in the subject.

In some such methods, the mobile device is programmed to receive and transmit data from at least two external sensors attached to the subject's upper and lower extremities. In some such methods, the mobile device acquires data from sensors on the subject's upper and lower extremities. In some such methods, a mobile device is carried by the subject to acquire data from internal sensors. In some such methods, the sequence of movements includes tapping the device, sitting and standing. In some such methods, monitoring indicates a reduction in motor impairment or a reduction in cognitive impairment in response to administering the antibody.

In some such methods, the synucleinopathy is Parkinson's disease. In some such methods, the synucleinopathy is dementia with Lewy bodies. In some such methods, the synucleinopathy is a multiple system atrophy. In some such methods, the synucleinopathy is progressive supranuclear palsy. In some such methods, the synucleinopathy is REM sleep behavior disorder. In some such methods, the synucleinopathic disease is Alzheimer's disease with Lewy bodies in the amygdala. In some such methods, the subject has the disease.

In some such methods, the subject is not receiving symptomatic therapy for Parkinson's disease in combination with the antibody. In some such methods, the antibody is administered in combination with levodopa.

In another aspect, the invention provides a method of practicing treatment or prophylaxis in a subject having Lewy body disease, wherein the subject is administered an antibody to alpha-synuclein at a dose of 1300-1700 mg intravenously at intervals of 3-5 weeks. or administering another regimen that delivers the antibody in substantially the same area under the curve to the subject.

In some such methods the dose is 1500 mg. In some such methods, an antibody is administered to multiple subjects with Lewy body disease, and the subjects receive one or two fixed doses. In some such methods, the subject receives a dose of 18-25 mg/kg. In some such methods, the subject receives a dose of 20 mg/kg.

In some such methods the interval is 28 days. In some such methods, the subject receives at least 6 doses of the antibody at 3-5 week intervals. In some such methods, the subject receives at least 12 doses of the antibody at 3-5 week intervals. In some such methods, the subject receives at least 18 doses of the antibody at 3-5 week intervals. In some such methods the interval is 4 weeks. In some such methods, the subject receives the antibody every 4 weeks for at least 52 weeks.

Some such methods further comprise monitoring the subject for changes in movement, cognitive impairment, autonomic dysfunction, gastrointestinal dysfunction, visual hallucinations, or psychological symptoms.

In some such methods, monitoring
(a) programmed to receive and transmit data obtained from sensors internal and/or external to the device regarding movement disorders in a subject having or suspected of having Lewy body disease; providing a mobile device to a subject, after which the subject performs a series of exercises to reveal a motor impairment, if any, and sensors internal or external to the device acquire data regarding the exercise;
(b) collecting data transmitted from the mobile device; and (c) comparing the data obtained from the subject to control data to assess the presence or extent of movement disorders in the subject.

In some such methods, the mobile device is programmed to receive and transmit data from at least two external sensors attached to the subject's upper and lower extremities.

In some such methods, the mobile device acquires data from sensors on the subject's upper and lower extremities. In some such methods, a mobile device is carried by the subject to acquire data from internal sensors. In some such methods, the sequence of movements includes tapping the device, sitting and standing. In some such methods, monitoring indicates a reduction in motor impairment or a reduction in cognitive impairment in response to administering the antibody.

In some such methods, the antibody binds within residues 115-130 of alpha-synuclein.

In some such methods, the antibody binds within residues 118-126 of alpha-synuclein.

In some such methods, the antibody binds within residues 117-123 of alpha-synuclein.

In some such methods, the antibody is NI-202.21D11. In some such methods, the antibody comprises three heavy chain CDRs designated as SEQ ID NOs:139-141, respectively, and three light chain CDRs designated as SEQ ID NOs:143-145, respectively. In some such methods, the antibody comprises a heavy chain variable region set forth in SEQ ID NO:138 and a light chain variable region set forth in SEQ ID NO:142.

In some such methods, the antibody comprises the three light chain Kabat CDRs of SEQ ID NO:5 and the three heavy chain Kabat CDRs of SEQ ID NO:10. In some such methods, the antibody comprises a heavy chain variable region of any of SEQ ID NOs:8-11 and a light chain variable region of any of SEQ ID NOs:3-5, preferably the heavy chain variable region is The heavy chain variable region of SEQ ID NO:10 and the light chain variable region is the light chain variable region of SEQ ID NO:5.

In some such methods, the antibody is of human IgG1 isotype.

In some such methods, the antibody comprises a heavy chain constant region of SEQ ID NO:35, optionally absent a C-terminal lysine, and a light chain constant region of SEQ ID NO:30. In some such methods, the antibody comprises a heavy chain constant region of SEQ ID NO:35, optionally absent a C-terminal lysine, and a light chain constant region of SEQ ID NO:13.

In some such methods, the antibody comprises three heavy chain CDRs designated in SEQ ID NOs:146-148, respectively, and three light chain CDRs designated in SEQ ID NOs:149-151, respectively. In some such methods, the antibody comprises a heavy chain designated as SEQ ID NO:37 and a light chain designated as SEQ ID NO:32, and the C-terminal lysine of SEQ ID NO:37 may be absent.

In some such methods, the antibody binds within residues 1-20 of alpha-synuclein. In some such methods, the antibody binds within residues 4-15 of alpha-synuclein.

In some such methods, the antibody is NI-202.12F4. In some such methods, the antibody comprises three heavy chain CDRs designated as SEQ ID NOs: 131-133, respectively, and three light chain CDRs designated as SEQ ID NOs: 135-137, respectively. In some such methods, the antibody comprises a heavy chain variable region set forth in SEQ ID NO:130 and a light chain variable region set forth in SEQ ID NO:134.

In some such methods, the synucleinopathy is Parkinson's disease. In some such methods, the synucleinopathy is dementia with Lewy bodies. In some such methods, the synucleinopathy is a multiple system atrophy. In some such methods, the synucleinopathy is progressive supranuclear palsy. In some such methods, the synucleinopathy is REM sleep behavior disorder. In some such methods, the synucleinopathic disease is Alzheimer's disease with Lewy bodies in the amygdala. In some such methods, the subject has the disease.

In some such methods, the subject is not receiving symptomatic therapy for Parkinson's disease in combination with the antibody. In some such methods, the antibody is administered in combination with levodopa.

DEFINITIONS The term "antibody" includes intact antibodies and binding fragments thereof. Typically, fragments compete with the intact antibody from which they are derived for specific binding to a target. Fragments include separated heavy chains, separated light chains, Fab, Fab', F(ab')2, F(ab)c, Fv, single chain antibodies, and single domain antibodies. The term "antibody" also includes bispecific antibodies. A bispecific or bifunctional antibody is an artificial hybrid antibody having two different heavy/light chain pairs and two different binding sites [e.g. Songsivilai and Lachmann, Clin. Exp. Immunol., 79:315 -321 (1990); see Kostelny et al., J. Immunol., 148:1547-53 (1992)].

The basic antibody structural unit is a tetramer of subunits. Each tetramer contains two identical pairs of polypeptide chains, each pair having one "light" (about 25 kDa) and one "heavy" chain (about 50-70 kDa). The amino-terminal portion of each chain includes a variable region of about 100-110 or more amino acids primarily responsible for antigen recognition. When first expressed, this variable region is typically linked to a cleavable signal peptide. A variable region without a signal peptide is sometimes referred to as a mature variable region. Thus, for example, a light chain mature variable region refers to a light chain variable region without a light chain signal peptide. The carboxy-terminal portion of each chain defines a constant region primarily responsible for effector function. A constant region can include any or all of the CH1 region, hinge region, CH2 region, and CH3 region.

Light chains are classified as either kappa or lambda. Heavy chains are classified as gamma, mu, alpha, delta, or epsilon, and define the antibody's isotype as IgG, IgM, IgA, IgD and IgE, respectively. Within light and heavy chains, the variable and constant regions are joined by a "J" region of about 12 or more amino acids, and heavy chains also include a "D" region of about 10 or more amino acids. [See generally, Fundamental Immunology (Paul, W., ed., 2nd ed. Raven Press, N.Y., 1989), Ch. 7] (incorporated by reference in its entirety for all purposes).

The mature variable region of each light/heavy chain pair forms the antibody binding site. An intact antibody therefore has two binding sites. The two binding sites are the same, except for bifunctional or bispecific antibodies. The chains all exhibit the same general structure of relatively conserved framework regions (FR) joined by three hypervariable regions, also called complementarity determining regions (CDRs). The CDRs from the two chains of each pair are aligned by framework regions to allow binding to a specific epitope. From N-terminus to C-terminus, both light and heavy chains comprise regions FR1, CDR1, FR2, CDR2, FR3, CDR3 and FR4. Amino acid assignments to each region can be found in Kabat, Sequences of Proteins of Immunological Interest (National Institutes of Health, Bethesda, MD, 1987 and 1991), or Chothia & Lesk, J. Mol. Biol. 196:901-917 (1987). ); as defined by Chothia et al., Nature 342:878-883 (1989). Kabat also provides a widely used numbering convention (Kabat numbering) in which corresponding residues between different heavy chains or between different light chains are assigned the same number.

Percentage sequence identity is determined with antibody sequences maximally aligned by the Kabat numbering convention. After alignment, when an antibody region of interest (e.g., the entire mature variable region of a heavy or light chain) is compared to the same region of a reference antibody, the percentage sequence identity between the antibody region of interest and the reference antibody region is the The number of positions occupied by the same amino acid in both the antibody region and the reference antibody region divided by the total number of aligned positions in the two regions (not counting gaps) multiplied by 100 to convert to a percentage is.

For purposes of classifying amino acid substitutions as conservative or non-conservative, amino acids are grouped as follows: Group I (hydrophobic side chains): Norleucine, Met, Ala, Val, Leu, Ile; Group II. (Neutral hydrophilic side chains): Cys, Ser, Thr; Group III (acidic side chains): Asp, Glu; Group IV (basic side chains): Asn, Gln, His, Lys, Arg; residues affecting orientation): Gly, Pro; and group VI (aromatic side chains): Trp, Tyr, Phe. Conservative substitutions involve substitutions between amino acids in the same class. Non-conservative substitutions constitute exchanging a member of one of these classes for a member of another class.

Antibodies of the invention typically bind their designated targets with an affinity constant of at least 10 ⁶ , 10 ⁷ , 10 ⁸ , 10 ⁹ , or 10 ¹⁰ M ⁻¹ . Such binding is specific binding in that it is detectably stronger and distinguishable from non-specific binding occurring to at least one unrelated target. Specific binding can be the result of bond formation between specific functional groups or specific spatial fits (e.g. types of locks and keys), while non-specific binding is usually the result of van der Waals forces. is. However, specific binding does not necessarily imply that a monoclonal antibody binds to one and only one target.

The term "symptom" refers to subjective evidence of disease, such as a change in gait, as perceived by a subject. A "sign" refers to objective evidence of disease as observed by a physician.

An individual whose subject has at least one known risk factor (e.g., genetic, biochemical, family history, environmental exposure) and is at a statistically significantly higher risk of developing the disease than an individual without the risk factor. An individual is at increased risk of disease when placing Statistical significance means p<0.05.

Unless otherwise apparent from the context, the term "about" includes values falling within the greater of the standard deviation of the mean of the stated values or +/−5% of the stated value. .

The term "9E4 antibody" refers to any antibody in which each CDR is substantially that of 9E4, and thus includes murine, chimeric, veneered, and humanized 9E4. References to other antibodies such as 5C1 and 1H7 have corresponding meanings.

An antibody of the invention can be administered in combination with another therapy for the same indication as the antibody, meaning that the other therapy is administered at least once during the period in which the antibody is administered. and such period begins one month before the first dose of antibody and ends one month after the last dose. The other treatment can be administered at repeated intervals during this period, which may or may not be the same as the intervals at which the antibody is administered. Other treatments may be symptomatic treatments.

Treatment is symptomatic if it affects only the cause of the disease, ie, one or more symptoms thereof, rather than its etiology.

As used herein, the term "mobile device" refers to any portable device that includes sensors and data recording equipment suitable for obtaining datasets of activity measurements. Typically, mobile devices include sensors for measuring activity. This may also require a data processor and storage device, as well as a display to electronically simulate the activity test on the mobile device. Additionally, data from the subject's activity is recorded and compiled into a dataset, which is evaluated by the method of the present invention either on the mobile device itself or on a second device. Depending on the particular configuration envisioned, it may be necessary for the mobile device to include data transmission equipment to transmit the acquired data sets from the mobile device to one or more further devices. Particularly well suited as mobile devices according to the invention are smart phones, smart watches, wearable sensors, portable multimedia devices or tablet computers. Alternatively, a portable sensor with data recording equipment and, optionally, processing equipment may be used. Additionally, depending on the type of activity test being performed, the mobile device is configured to instruct the subject regarding the activities to be performed for the test.

Unless otherwise clear from the context, references to ranges include any integer within the range.

A therapeutic regimen refers to the combination of parameters that characterize the administration of an immunotherapeutic agent, including any or all of dose, frequency of administration, route of administration, and total duration of administration.

Brief Description of Sequences SEQ ID NO: 1 is the m9E4VL variable region.

SEQ ID NO:2 is 63102889Hu9E4VLFr variable region.

SEQ ID NO:3 is the Hu9E4VLv1 variable region.

SEQ ID NO: 4 is the Hu9E4VLv2 (no backmutation) variable region.

SEQ ID NO: 5 is the Hu9E4VLv3 variable region.

SEQ ID NO: 6 is the m9E4 VH variable region.

SEQ ID NO: 7 is the 1791009 Hu9E4 VHFr variable region.

SEQ ID NO:8 is the Hu9E4 VHv1 variable region.

SEQ ID NO:9 is the Hu9E4 VHv2 variable region.

SEQ ID NO: 10 is the Hu9E4 VHv3 variable region.

SEQ ID NO: 11 is the Hu9E4 VHv4 (no backmutation) variable region.

SEQ ID NO: 12 is the amino acid sequence of native human wild-type alpha-synuclein.

SEQ ID NO: 13 is the humanized 9E4 light chain constant region (with R) (common to v1, v2 and v3).

SEQ ID NO: 14 is the humanized 9E4 light chain constant region (with R) nucleotide sequence (common for v1, v2, v3).

SEQ ID NO: 15 is the amino acid sequence of an exemplary human IgG1 constant region.

SEQ ID NO: 16 is a nucleic acid sequence encoding an exemplary human IgG1 constant region.

SEQ ID NO: 17 is the Hu9E4VLv1 nucleotide sequence variable region.

SEQ ID NO: 18 is the Hu9E4VLv2 nucleotide sequence (no backmutations) variable region.

SEQ ID NO: 19 is the Hu9E4VLv3 nucleotide sequence variable region.

SEQ ID NO:20 is the Hu9E4 VHv1 nucleotide sequence variable region.

SEQ ID NO:21 is the Hu9E4 VHv2 nucleotide sequence variable region.

SEQ ID NO:22 is the Hu9E4 VHv3 nucleotide sequence variable region.

SEQ ID NO:23 is the Hu9E4 VHv4 nucleotide sequence (no backmutations) variable region.

SEQ ID NO:24 is the Hu9E4VL signal peptide (common for v1, v2, v3).

SEQ ID NO:25 is the Hu9E4VL signal peptide nucleotide sequence (common for v1, v2, v3).

SEQ ID NO:26 is the Hu9E4 VH signal peptide (common for v1, v2, v3).

SEQ ID NO:27 is the Hu9E4 VH signal peptide nucleotide sequence (common for v1, v2, v3, v4).

SEQ ID NO:28 is a Hu9E4VL surrogate.

SEQ ID NO:29 is a Hu9E4 VH surrogate.

SEQ ID NO:30 is the amino acid sequence of an exemplary human kappa light chain constant region.

SEQ ID NO:31 is the humanized 9E4 light chain constant region (no R) nucleotides.

Array (common for v1, v2, v3).

SEQ ID NO: 32 is the humanized 9E4 light chain type 3 (variable region + constant region, with arginine).

SEQ ID NO:33 is the humanized 9E4 light chain type 3 (variable region + constant region, no arginine).

SEQ ID NO:34 is the humanized 9E4 heavy chain type 3 (variable region + constant region).

SEQ ID NO:35 is the humanized 9E4 heavy chain constant region (G1m3 allotype; BIP type).

SEQ ID NO:36 is the humanized 9E4 heavy chain type 3 (variable region + constant region).

SEQ ID NO:37 is the humanized 9E4 heavy chain type 3 (variable region + alternative constant region G1m3 allotype).

SEQ ID NO:38 is the m5C1 antibody mature heavy chain variable region nucleotide sequence.

SEQ ID NO:39 is the m5C1 antibody heavy chain variable region amino acid sequence.

SEQ ID NO:40 is the m5C1 antibody heavy chain signal peptide nucleotide sequence.

SEQ ID NO:41 is the m5C1 antibody heavy chain signal peptide amino acid sequence.

SEQ ID NO:42 is the m5C1 antibody mature light chain variable region nucleotide sequence.

SEQ ID NO:43 is the m5C1 antibody light chain variable region amino acid sequence.

SEQ ID NO:44 is the m5C1 antibody light chain signal peptide nucleotide sequence.

SEQ ID NO:45 is the m5C1 antibody light chain signal peptide amino acid sequence.

SEQ ID NO:46 is the 5C1 immunogen.

SEQ ID NO:47 is the m5C1 heavy chain CDR1.

SEQ ID NO:48 is the m5C1 heavy chain CDR2.

SEQ ID NO:49 is the m5C1 heavy chain CDR3.

SEQ ID NO:50 is the m5C1 light chain CDR1.

SEQ ID NO:51 is the m5C1 light chain CDR2.

SEQ ID NO:52 is the m5C1 light chain CDR3.

SEQ ID NO:53 is the mouse 5C1 heavy chain variable region nucleotide sequence with the sequence encoding the signal peptide.

SEQ ID NO:54 is the mouse 5C1 heavy chain variable region sequence with signal peptide.

SEQ ID NO:55 is the mouse 5C1 light chain variable region nucleotide sequence with the sequence encoding the signal peptide.

SEQ ID NO:56 is the mouse 5C1 light chain variable region sequence with signal peptide.

SEQ ID NO:57 is the human VH acceptor FR (Accession number AAY42876.1).

SEQ ID NO:58 is humanized 5C1H1.

SEQ ID NO:59 is humanized 5C1H2.

SEQ ID NO:60 is humanized 5C1H3.

SEQ ID NO:61 is humanized 5C1H4.

SEQ ID NO:62 is humanized 5C1H5.

SEQ ID NO:63 is the nucleic acid sequence encoding humanized 5C1 H1.

SEQ ID NO:64 is the nucleic acid sequence encoding humanized 5C1 H2.

SEQ ID NO:65 is the nucleic acid sequence encoding humanized 5C1H3.

SEQ ID NO:66 is the nucleic acid sequence encoding humanized 5C1 H4.

SEQ ID NO:67 is the nucleic acid sequence encoding humanized 5C1H5.

SEQ ID NO: 68 is the human VL acceptor FR (accession number CAB51293.1).

SEQ ID NO: 69 is humanized 5C1L1.

SEQ ID NO:70 is humanized 5C1L2.

SEQ ID NO:71 is humanized 5C1L3.

SEQ ID NO:72 is humanized 5C1L4.

SEQ ID NO:73 is the nucleic acid sequence encoding humanized 5C1 L1.

SEQ ID NO:74 is the nucleic acid sequence encoding humanized 5C1 L2.

SEQ ID NO:75 is the nucleic acid sequence encoding humanized 5C1L3.

SEQ ID NO:76 is the nucleic acid sequence encoding humanized 5C1 L4.

SEQ ID NO:77 is a nucleic acid sequence encoding an exemplary human kappa light chain constant region.

SEQ ID NO: 78 is according to Jensen et al. The non-amyloid component (NAC) domain of alpha-synuclein as reported by.

SEQ ID NO:79 is according to Ueda et al. The non-amyloid component (NAC) domain of alpha-synuclein as reported by.

SEQ ID NO:80 is the m1H7 antibody heavy chain variable nucleotide sequence.

SEQ ID NO:81 is the m1H7 antibody heavy chain variable amino acid sequence.

SEQ ID NO:82 is the m1H7 antibody light chain variable nucleotide sequence.

SEQ ID NO:83 is the m1H7 antibody light chain variable amino acid sequence.

SEQ ID NO:84 is the mature m1H7 antibody heavy chain variable nucleotide sequence.

SEQ ID NO:85 is the mature m1H7 antibody heavy chain variable amino acid sequence.

SEQ ID NO:86 is the mature m1H7 antibody light chain variable nucleotide sequence.

SEQ ID NO:87 is the mature m1H7 antibody light chain variable amino acid sequence.

SEQ ID NO:88 is the m1H7 antibody heavy chain CDR1 (Kabat definition).

SEQ ID NO:89 is the m1H7 antibody heavy chain CDR2 (Kabat definition).

SEQ ID NO:90 is the m1H7 antibody heavy chain CDR3 (Kabat definition).

SEQ ID NO:91 is the m1H7 antibody light chain CDR1 (Kabat definition).

SEQ ID NO:92 is the m1H7 antibody light chain CDR2 (Kabat definition).

SEQ ID NO:93 is the m1H7 antibody light chain CDR3 (Kabat definition).

SEQ ID NO:94 is the Hu1H7VHv1 nucleotide sequence.

SEQ ID NO:95 is the Hu1H7VHv1 amino acid sequence.

SEQ ID NO:96 is the Hu1H7VHv2 nucleotide sequence.

SEQ ID NO:97 is the Hu1H7VHv2 amino acid sequence.

SEQ ID NO:98 is the Hu1H7VHv3 nucleotide sequence.

SEQ ID NO:99 is the Hu1H7VHv3 amino acid sequence.

SEQ ID NO: 100 is the Hu1H7VHv4 nucleotide sequence.

SEQ ID NO: 101 is the Hu1H7VHv4 amino acid sequence.

SEQ ID NO: 102 is the Hu1H7VHv5 nucleotide sequence.

SEQ ID NO: 103 is the Hu1H7VHv5 amino acid sequence.

SEQ ID NO: 104 is the Hu1H7VH signal peptide nucleotide sequence.

SEQ ID NO: 105 is the Hu1H7 VH signal peptide amino acid sequence.

SEQ ID NO: 106 is the Hu1H7VH signal peptide nucleotide sequence.

SEQ ID NO: 107 is the Hu1H7 VH signal peptide amino acid sequence.

SEQ ID NO: 108 is the Hu1H7VLv1 nucleotide sequence.

SEQ ID NO: 109 is the Hu1H7VLv1 amino acid sequence.

SEQ ID NO: 110 is the Hu1H7VLv2 nucleotide sequence.

SEQ ID NO: 111 is the Hu1H7VLv2 amino acid sequence.

SEQ ID NO: 112 is the Hu1H7VLv3 nucleotide sequence.

SEQ ID NO: 113 is the Hu1H7VLv3 amino acid sequence.

SEQ ID NO: 114 is the Hu1H7VLv4 nucleotide sequence.

SEQ ID NO: 115 is the Hu1H7VLv4 amino acid sequence.

SEQ ID NO: 116 is the Hu1H7VL signal peptide nucleotide sequence.

SEQ ID NO: 117 is the amino acid sequence of BAC02037 (GI-21670055) human acceptor used for the heavy chain framework.

SEQ ID NO: 118 is the amino acid sequence of AAY33358 GI-63102905) human acceptor used for the light chain framework.

SEQ ID NO: 119 is Hu1H7VH with no backmutations or CDR mutations.

SEQ ID NO: 120 is Hu1H7VL with no backmutations or CDR mutations.

SEQ ID NO: 121 is a Hu1H7 VH surrogate.

SEQ ID NO: 122 is a Hu1H7VL surrogate.

SEQ ID NO: 123 is a Hu1H7VH CDR3 surrogate.

SEQ ID NO: 124 is humanized 1H7 light chain type 3 (variable region + constant region, with arginine).

SEQ ID NO: 125 is humanized 1H7 light chain type 3 (variable region + constant region, no arginine).

SEQ ID NO: 126 is the humanized 1H7 heavy chain type 3 (variable region + constant region).

SEQ ID NO: 127 is the humanized 1H7 heavy chain type 3 (variable region + constant region G1m3 allotype).

SEQ ID NO: 128 is the humanized 1H7 heavy chain constant region (IgG2).

SEQ ID NO: 129 is the humanized 1H7 heavy chain constant region (G1m1 allotype).

SEQ ID NO: 130 is the NI-202.12F4-VHA1b variable heavy chain (VH) sequence.

SEQ ID NO: 131 is the NI-202.12F4-VHA1b CDR1 sequence.

SEQ ID NO: 132 is the NI-202.12F4-VHA1b CDR2 sequence.

SEQ ID NO: 133 is the NI-202.12F4-VHA1b CDR3 sequence.

SEQ ID NO: 134 is the NI-202.12F4-VLa1 variable light chain (VL) sequence.

SEQ ID NO: 135 is the NI-202.12F4-VLa1 CDR1 sequence.

SEQ ID NO: 136 is the NI-202.12F4-VLa1 CDR2 sequence.

SEQ ID NO: 137 is the NI-202.12F4-VLa1 CDR3 sequence.

SEQ ID NO: 138 is the NI-202.21D11-VH sequence.

SEQ ID NO: 139 is the NI-202.21D11-VH CDR1 sequence.

SEQ ID NO: 140 is the NI-202.21D11-VH CDR2 sequence.

SEQ ID NO: 141 is the NI-202.21D11-VH CDR3 sequence.

SEQ ID NO: 142 is the NI-202.21D11-VK sequence.

SEQ ID NO: 143 is the NI-202.21D11 VK CDR1 sequence.

SEQ ID NO: 144 is the NI-202.21D11 VK CDR2 sequence.

SEQ ID NO: 145 is the NI-202.21D11 VK CDR3 sequence.

SEQ ID NO: 146 is the 9E4 VH CDR1 sequence.

SEQ ID NO: 147 is the 9E4 VH CDR2 sequence.

SEQ ID NO: 148 is the 9E4 VH CDR3 sequence.

SEQ ID NO: 149 is the 9E4 VL CDR1 sequence.

SEQ ID NO: 150 is the 9E4 VL CDR2 sequence.

SEQ ID NO: 151 is the 9E4 VL CDR3 sequence.

I. Overview The present invention provides dosing regimens for immunotherapy of synucleinopathic diseases. Subjects receive antibodies against alpha-synuclein intravenously at doses of 3000-5000 mg or 1300-1700 mg every 3-5 weeks. Such doses may be preceded by one or more lower loading doses.

II. Target Molecule Native human wild-type alpha-synuclein has the following amino acid sequence:
MDVFMKGLSK AKEGVVAAAE KTKQGVAEAA GKTKEGVLYV GSKTKEGVVH GVATVAEKTK EQVTNVGGAV VTGVTAVAQK TVEGAGSIAA ATGFVKKDQL GKNEEGAPQE GILEDMPVDP DNEAYEMPSE EGYQDYEPEA (SEQ ID NO: 12)
[Ueda et al., Proc. Natl. Acad. Sci. USA (1993) 90:11282-6]; GenBank accession number: P37840. The protein has three recognized domains: an N-terminal KTKE repeat domain covering amino acids 1-61; a NAC (non-amyloid component) domain spanning approximately amino acids 60-95; and a C-terminal acidic domain spanning approximately amino acids 98-140. have.

Unless otherwise clear from the context, references to alpha-synuclein or fragments thereof include the native human wild-type amino acid sequence indicated above and human allelic variants thereof, particularly human allelic variants associated with Lewy body disease. variants (e.g. variants E46K, A30P and A53T; the first letter indicates the amino acid in SEQ ID NO: 12, the number indicates the codon position in SEQ ID NO: 12, the second letter indicates the amino acid in the allelic variant); pointing). Such variants may be present individually or in any combination in any of the aspects of the invention described below, as appropriate.

III. Synucleinopathies Synucleinopathies are characterized by excessive accumulation of alpha-synuclein, which may or may not be accompanied by the formation of Lewy bodies (LBs). For example, Lewy body disease (LBD) is characterized by degeneration of the dopaminergic system, motor changes, cognitive impairment, and formation of Lewy bodies. [McKeith et al., Neurology (1996) 47:1113-24]. Lewy bodies are globular protein deposits found in nerve cells. Their presence in the brain disrupts the normal functioning of the brain and interferes with the action of chemical messengers including acetylcholine and dopamine. Synucleinopathies include Parkinson's disease (including idiopathic Parkinson's disease), dementia with Lewy bodies (DLB) [also known as diffuse Lewy body disease (DLBD)], Lewy body variant of Alzheimer's disease ( LBV), Combined Alzheimer's and Parkinson disease, progressive supranuclear palsy (PSP), and multiple system atrophy (MSA; e.g., olivopontocerebellar atrophy, striatonigral degeneration , and Shy-Drager syndrome). DLB shares symptoms of both Alzheimer's disease and Parkinson's disease. DLB differs from Parkinson's disease primarily in the location of Lewy bodies. In DLB, Lewy bodies are formed primarily in the cortex. In Parkinson's disease they are mainly formed in the substantia nigra. Other synucleinopathies include pure autonomic failure, Lewy body dysphagia, episodic LBD, and hereditary LBD (eg, mutations in the alpha-synuclein genes, PARK3 and PARK4).

IV. Immunotherapy Agents Passive immunotherapy involves treatment with antibodies that specifically bind to alpha-synuclein.

1. Antibody A. Binding Specificity and Functional Properties The methods of the invention are directed to epitopes within residues 1-15, 4-15, 1-20, 91-99, 115-130, 117-123 or 118-126 of human alpha-synuclein. can include an antibody that specifically binds to Antibodies can be monoclonal or polyclonal antibodies. A polyclonal serum can be specific for an epitope or amino acid range within alpha-synuclein in that it lacks antibody binding outside of that epitope or range. Antibodies can be non-human, chimeric, veneered, humanized, or human, among other possibilities.

Some antibodies are bispecific, with one arm directed against alpha-synuclein and the other arm directed against insulin receptor, insulin-like growth factor (IGF) receptor, leptin an antibody that binds to a receptor expressed in the blood-brain barrier, such as a receptor, or a lipoprotein receptor, or preferably a transferrin receptor (Friden et al., PNAS 88:4771-4775, 1991; Friden et al. ., Science 259:373-377, 1993). Such bispecific antibodies can translocate across the blood-brain barrier by receptor-mediated transcytosis. Brain uptake of bispecific antibodies can be further enhanced by engineering the bispecific antibody to reduce its affinity for blood-brain barrier receptors. Reduced affinity for the receptor resulted in a broader distribution in the brain (eg Atwal. et al. Sci. Trans. Med. 3, 84ra43, 2011; Yu et al. Sci. Trans (see Med. 3, 84ra44, 2011).

Exemplary bispecific antibodies can also be: (1) a dual variable domain antibody (DVD-Ig) in which each light and heavy chain contains two variable domains in tandem through a short peptide linkage [ in Wu et al., Generation and Characterization of a Dual Variable Domain Immunoglobulin (DVD-Ig ^™ ) Molecule, Antibody Engineering, Springer Berlin Heidelberg (2010)]; Tandab, a fusion of two single-chain diabodies resulting in a bispecific antibody; (3) a flexibody, a combination of scFv with diabodies resulting in a multivalent molecule; (4) Based on the "dimerization and docking domain" in protein kinase A, a so-called "dock and lock"molecules; (5) so-called Scorpion molecules, eg, comprising two scFvs fused to both ends of a human Fc region. Examples of platforms useful for preparing bispecific antibodies include BiTE (Micromet), DART (MacroGenics), Fcab and Mab2 (F-star), Fc-engineered IgGl (Xencor) or DuoBody (Fab arm exchange based on Genmab).

Another exemplary antibody is mAb 9E4, which binds to an epitope within residues 118-126 of human alpha-synuclein. The cell line designated JH17.9E4.3.37.1.14.2, which produces antibody 9E4, has ATCC Accession No. PTA-8221 and was approved by American on February 26, 2007 under the provisions of the Budapest Treaty. Deposited in the Type Culture Collection (ATCC, PO Box 1549, Manassas, Va. 20108). Mouse light and heavy chain variable sequences are SEQ ID NO: 1 and SEQ ID NO: 6, respectively. Mouse mAb 9E4 has VH-CDR1 of SEQ ID NO: 146, VH-CDR2 of SEQ ID NO: 147, VH-CDR3 of SEQ ID NO: 148, VL-CDR1 of SEQ ID NO: 149, VL-CDR2 of SEQ ID NO: 150, and Contains VL-CDR3.

Another exemplary antibody is mAb 5C1. Mouse light and heavy chain variable sequences are SEQ ID NO:43 and SEQ ID NO:39, respectively.

Another exemplary antibody is mAb 1H7. The cell line designated JH17.1H7.4.24.34, which produces antibody 1H7, has ATCC Accession No. PTA-8220 and was added to the American Type Culture Collection on February 26, 2007 under the provisions of the Budapest Treaty. ATCC, PO Box 1549, Manassas, Va. 20108). Mouse light and heavy chain variable sequences are SEQ ID NO:87 and SEQ ID NO:85, respectively.

Such antibodies are described in WO06/020581 and US Application Nos. 61/591,835, 61/711,207, 13/750,983, 61/711,204, 61 /719,281, 61/840,432, 61/872,366, 14/049,169, 61/553,131, 61/711,204 , 61/843,011, and 13/662,261, all of which are incorporated herein by reference for all purposes.

Another exemplary antibody is NI-202.12F4, which binds to an epitope within residues 4-15 of alpha-synuclein, with the major contributor to the epitope being residue 10. (US 8,940,276; see WO2012177972, US 9,580,493). NI-202.12F4 contains the variable heavy chain (VHA1b) sequence of SEQ ID NO:130 and the variable light chain (VLa1) sequence of SEQ ID NO:134. NI-202.12F4 has VHA1b-CDR1 of SEQ ID NO: 131, VHA1b-CDR2 of SEQ ID NO: 132, VHA1b-CDR3 sequence of SEQ ID NO: 133 (Ala-His), VLa1-CDR1 sequence of SEQ ID NO: 135, VLa1-CDR2 sequence, and VLa1-CDR3 sequence of SEQ ID NO:137. Another exemplary antibody is NI-202.21D11, which binds to an epitope within residues 117-123 of alpha-synuclein, with residues 121 and 122 contributing most to the epitope. (see US 9,580,493). NI-202.21D11 contains the variable heavy chain (VH) sequence of SEQ ID NO:138 and the variable light chain (VK) sequence of SEQ ID NO:142. NI-202.21D11 has the VH-CDR1 of SEQ ID NO: 139, the VH-CDR2 of SEQ ID NO: 140, the VH-CDR3 sequence of SEQ ID NO: 141, the VK-CDR1 sequence of SEQ ID NO: 143, the VK-CDR2 sequence of SEQ ID NO: 144, and the VK-CDR3 sequence of SEQ ID NO:145.

B. Humanized Antibodies Humanized antibodies are genetically engineered antibodies in which the CDRs from a non-human “donor” antibody have been grafted onto human “acceptor” antibody sequences (see, eg, Queen et al., US Pat. No. 5,530,101 and ibid.). 5,585,089; Winter et al., US 5,225,539; Carter, US 6,407,213; Adair, US 5,859,205; . The acceptor antibody sequence can be, for example, a mature human antibody variable region sequence, a composite of such sequences, a consensus sequence of human antibody sequences (e.g., Kabat, 1991, supra light and heavy chain variable region consensus sequences), or It can be a germline variable region sequence. Thus, a humanized antibody of the invention may have wholly or substantially three light chain CDRs and three heavy chain CDRs as defined by Kabat from a murine antibody (donor antibody) such as 9E4, 5C1 or 1H7, and antibodies having mature variable region framework sequences and constant regions, if present, that are wholly or substantially from human antibody sequences. Similarly, a humanized heavy chain has the three heavy chain CDRs as defined by Kabat from a donor antibody heavy chain and, if present, a mature heavy chain from wholly or substantially human antibody heavy chain sequence. It includes a heavy chain having a chain variable sequence and a heavy chain constant region sequence. Similarly, a humanized light chain includes the three light chain CDRs as defined by Kabat from a donor antibody light chain and, if present, a mature CDR from a wholly or substantially human antibody light chain sequence. It includes a light chain having a light chain variable sequence and a light chain constant region sequence. The mature variable region framework sequence of the antibody chain or the constant region sequence of the antibody chain has at least 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity is substantially from a human mature variable region framework sequence or a human constant region sequence, respectively. A CDR is substantially identified from a corresponding donor CDR if it exhibits at least 85% identity to the corresponding donor CDR, except for CDRH2 where at least 65% identity is required. belongs to.

Certain amino acids from the human mature variable region framework residues may be selected for substitution based on their possible influence on CDR conformation and/or binding to antigen. Investigation of such possible effects is by modeling, examination of amino acid characteristics at particular positions, or empirical observation of the effects of particular amino acid substitutions or mutagenesis.

For example, when an amino acid differs between a mouse mature variable region framework residue and a selected human mature variable region framework residue, the amino acid is
(1) directly non-covalently binds to the antigen;
(2) flanking the CDR regions;
(3) otherwise interact with the CDR region (e.g., be within about 6 Å of the CDR region);
(4) Human framework amino acids can be replaced by equivalent framework amino acids from the murine antibody when reasonably expected to mediate interactions between heavy and light chains.

An exemplary humanized form of the murine 9E4 antibody comprises three exemplary humanized light chain mature variable regions (Hu9E4VLv1-v3; SEQ ID NOS:3-5) and four exemplary humanized heavy chain mature variable regions (Hu9E4VHv1 ~v4; SEQ ID NOS:8-11).

Exemplary light and heavy chain mature variable regions may be paired in any combination. An exemplary combination is Hu9E4VLv3 (SEQ ID NO:5) and Hu9E4VHv3 (SEQ ID NO:10). An exemplary heavy chain constant region is SEQ ID NO:35 with or without a C-terminal lysine. This constant region may be linked to any of the heavy chain variable regions. An exemplary light chain constant region is SEQ ID NO:30. This constant region may be linked to any of the light chain variable regions. An exemplary full length heavy chain has the amino acid sequence of SEQ ID NO:37 with or without a C-terminal lysine. An exemplary full length light chain has the amino acid sequence of SEQ ID NO:32. An exemplary combination is an antibody comprising SEQ ID NO:37 (with or without C-terminal lysine) and SEQ ID NO:32. Nucleic acids encoding the various heavy and light chains described immediately above are provided in SEQ ID NOS: 17-23. Exemplary humanized antibodies have heavy and light chains characterized by SEQ ID NOS: 37 (with or without C-terminal lysine) and 32.

Exemplary humanized murine 5C1 antibodies have four exemplary humanized light chain mature variable regions (5C1L1-L4; SEQ ID NOS: 69-72) and five exemplary humanized heavy chain mature variable regions ( 5C1H1-H4; SEQ ID NOs:58-62). Exemplary light and heavy chain mature variable regions may be paired in any combination. An exemplary heavy chain constant region is SEQ ID NO:35 with or without a C-terminal lysine. This constant region may be linked to any of the heavy chain variable regions. An exemplary light chain constant region is SEQ ID NO:30. This constant region may be linked to any of the light chain variable regions. Nucleic acids encoding the various heavy and light chains described immediately above are provided in SEQ ID NOs:73-76 and 63-67.

An exemplary humanized murine 1H7 antibody has four exemplary humanized light chain mature variable regions (Hu1H7VLv1-v4; SEQ ID NOs: 109, 111, 113, and 115, respectively), and five exemplary humanized Includes heavy chain mature variable regions (Hu1H7 VHv1-v5; SEQ ID NOs: 95, 97, 99, 101, and 103, respectively). Exemplary light and heavy chain mature variable regions may be paired in any combination. An exemplary combination is Hu1H7VHv3 (SEQ ID NO:99) and Hu1H7VLv3 (SEQ ID NO:113). An exemplary heavy chain constant region is SEQ ID NO:35 with or without a C-terminal lysine. This constant region may be linked to any of the heavy chain variable regions. An exemplary light chain constant region is SEQ ID NO:30. This constant region may be linked to any of the light chain variable regions. An exemplary full-length heavy chain has the amino acid sequence of SEQ ID NO: 127 (with G1m3). An exemplary full length light chain has the amino acid sequence of SEQ ID NO: 125 (without arginine). An exemplary combination is an antibody comprising SEQ ID NO:127 and SEQ ID NO:125. Nucleic acids encoding the various heavy and light chains described immediately above are provided in SEQ ID NOs: 108, 110, 112, 114, 94, 96, 98, 100, 102.

Such humanized antibodies are described in WO 06/020581 and US 13/750,983, 14/049,169, 13/662,261, and 61/843,011.

In these or other antibodies described herein, one or more terminal residues may be cleaved during post-translational processing, particularly for the terminal lysine of the heavy chain constant region.

C. Chimeric and Veneered Antibodies The present invention further provides chimeric and veneered non-human antibodies, particularly 9E4, 5C1 or 1H7.

Chimeric antibodies are antibodies in which mature variable regions of light and heavy chains of a non-human antibody (eg, murine) are combined with light and heavy chain constant regions from an antibody of a different species. Typically, the light and heavy chain constant regions are of human origin, although the constant regions may optionally be derived from rat (e.g., to facilitate testing of non-human antibodies in appropriate animal models). can originate from different non-human species such as Such antibodies retain substantially or entirely the binding specificity of the non-human (e.g., murine) antibody that supplies the variable region and are about two-thirds human (or a different non-human species) sequence. .

A veneered antibody retains some and usually all of the CDRs of the non-human antibody and some of the non-human variable region framework residues, but other variable region framework residues that may contribute to B or T cell epitopes. A class of humanized antibodies in which groups, such as exposed residues (Padlan, Mol. Immunol. 28:489, 1991), are replaced with residues from the corresponding positions in the human antibody sequence. The result is an antibody whose CDRs are wholly or substantially from a non-human antibody and in which the variable region framework of the non-human antibody has been made more human-like by substitutions. A veneered version of 9E4 is included in the present invention.

D. Selection of Constant Regions The heavy and light chain variable regions of a chimeric, veneered or humanized antibody may be joined to at least a portion of a human constant region. The choice of constant region depends in part on whether antibody-dependent cell-mediated cytotoxicity, antibody-dependent cell phagocytosis and/or complement-dependent cytotoxicity is desired. For example, human isotopes IgG1 and IgG3 have complement dependent cytotoxicity, while human isotypes IgG2 and IgG4 do not. Human IgG1 and IgG3 also induce stronger cell-mediated effector functions than human IgG2 and IgG4. Light chain constant regions can be lambda or kappa. An exemplary human light chain kappa constant region has the amino acid sequence of SEQ ID NO:13. Some such light chain kappa constant regions may be encoded by nucleic acid sequences. The N-terminal arginine of SEQ ID NO:13 may be omitted, in which case the light chain kappa constant region has the amino acid sequence of SEQ ID NO:30. Some such light chain kappa constant regions may be encoded by nucleic acid sequences. An exemplary human IgG1 heavy chain constant region has the amino acid sequence of the heavy chain constant region component of SEQ ID NO: 15 (with or without C-terminal lysine) or SEQ ID NO: 37 (with or without C-terminal lysine). Some such heavy chain constant regions may be encoded by nucleic acid sequences. Antibodies can be produced as tetramers containing two light and two heavy chains, as separate heavy chains, light chains, as Fab, Fab', F(ab')2, and Fv, or as heavy and light chains. Chain mature variable domains can be expressed as single chain antibodies linked through spacers.

Human constant regions exhibit allotypic and isoallotypic variation between different individuals, ie, the constant regions may differ at one or more polymorphic positions in different individuals. Isoallotypes differ from allotypes in that sera that recognize the isoallotype bind to non-polymorphic regions of one or more other isotypes. Thus, for example, another heavy chain constant region is an IgG1 G1m3 allotype heavy chain constant region, having the amino acid sequence encoding the constant region of SEQ ID NO:37. Yet another heavy chain constant region has the amino acid sequence encoding the constant region of SEQ ID NO:37, except lacking the C-terminal lysine.

One or more amino acids at the amino or carboxy terminus of the light and/or heavy chain may be deleted or derivatized in part or all of the molecule, such as the C-terminal lysine of the heavy chain. Substitutions are made in the constant region to reduce or increase effector functions such as complement-mediated cytotoxicity or ADCC (e.g. Winter et al., US 5,624,821; Tso et al., US 5,834,597; and Lazar et al., Proc. Natl. Acad. Sci. 2004). Exemplary substitutions include Gln at position 250 and/or Leu at position 428 to increase the half-life of the antibody (EU numbering is used for the constant regions in this paragraph). Substitutions at any or all of positions 234, 235, 236 and/or 237 reduce affinity for Fcγ receptors, particularly FcγRI receptors (see, eg, US Pat. No. 6,624,821). Some antibodies have alanine substitutions at positions 234, 235 and 237 of human IgG1 to reduce effector function. Positions 234, 236 and/or 237 in human IgG2 may be substituted by alanine and position 235 by glutamine (see, eg, US Pat. No. 5,624,821).

E. Expression of Recombinant Antibodies Antibodies can be produced by recombinant expression. Antibody-encoding nucleic acids can be codon-optimized for expression in the desired cell type (eg, CHO or Sp2/0). Recombinant nucleic acid constructs typically include an expression control sequence operably linked to the coding sequences of antibody chains, including naturally associated or heterologous promoter regions. The expression control sequences can be eukaryotic promoter systems in vectors capable of transforming or transfecting eukaryotic host cells. Once the vector has been incorporated into a suitable host, the host is maintained under conditions suitable for high level expression of the nucleotide sequences and recovery and purification of the cross-reacting antibody. The vector or vectors encoding antibody chains may also contain selectable genes such as dihydrofolate reductase to allow amplification of the copy number of the nucleic acid encoding the antibody chains.

E. coli is a prokaryotic host that is particularly useful for the expression of antibodies, especially antibody fragments. Microorganisms such as yeast are also useful for expression. Saccharomyces is an exemplary yeast host, and suitable vectors have expression control sequences, origins of replication, termination sequences and the like as desired. Typical promoters include 3-phosphoglycerate kinase and other glycolytic enzymes. Inducible yeast promoters include promoters from alcohol dehydrogenase, isocytochrome C, and enzymes involved in maltose and galactose utilization, among others.

Mammalian cells can be used to express nucleotide segments encoding immunoglobulins or fragments thereof. See Winnacker, From Genes to Clones, (VCH Publishers, NY, 1987). Several suitable host cell lines capable of secreting intact heterologous proteins have been developed in the art, including CHO cell lines, various COS cell lines, HeLa cells, HEK293 cells. , L-cells, and non-antibody-producing myeloma, including Sp2/0 and NS0. It may be advantageous to use non-human cells. Expression vectors for these cells contain expression control sequences such as origins of replication, promoters, enhancers [Queen et al., Immunol. Rev. 89:49 (1986)], as well as ribosome binding sites, RNA splice sites, polyadenylation sites. sites and necessary processing information sites such as transcription terminator sequences. Preferred expression control sequences are promoters derived from endogenous genes, cytomegalovirus, SV40, adenovirus, bovine papilloma virus, and the like. See Co et al., J. Immunol. 148:1149 (1992).

Once vectors encoding antibody heavy and light chains have been introduced into cell culture, cell pools can be screened for growth productivity and product quality in serum-free medium. Top producer cell pools can then be subjected to FACS-based single cell cloning to generate monoclonal lines. A specific productivity of greater than 50 or 100 pg per cell per day may be advantageous, corresponding to a product titer of greater than 7.5 g per L of culture. Antibodies produced by single cell clones can also be tested for turbidity, filtration properties, PAGE, IEF, UV scan, HP-SEC, carbohydrate-oligosaccharide mapping, mass spectrometry, and binding assays such as ELISA or Biacore. . Selected clones can then be pooled in multiple vials and stored frozen for later use.

Once the antibodies are expressed, they can be purified according to standard procedures in the art, including protein A capture, column chromatography (e.g., hydrophobic interaction or ion exchange), viral isolation. low pH for activation, etc. [see generally Scopes, Protein Purification (Springer-Verlag, NY, 1982)].

Methodologies for commercial production of antibodies include codon optimization, selection of promoters, transcriptional elements, and terminators, serum-free single cell cloning, cell banking, use of selectable markers, CHO terminators, serum free single cell cloning, improved protein titers (e.g. US 5,786,464, US 6,114,148, US 6,063,598 , US 7,569,339, WO 2004/050884, WO 2008/012142, WO 2008/012142, WO 2005/019442, WO 2008/107388, and WO 2009/027471, and US 5,888,809).

V. Formulations The formulations (also known as pharmaceutical compositions) of the present invention comprise an antibody (eg, a chimeric, veneered or humanized form of murine 9E4 (ATCC Accession No. PTA-8221)) or an antigen-binding fragment thereof, a buffer agent, one or more sugars and/or polyols and a surfactant, and has a pH within the range of about 5 to about 7.5. Formulations may be prepared for storage in liquid or lyophilized form. When stored in lyophilized form, the formulation can be reconstituted with liquid (eg, sterile water) to the concentrations and properties described herein. When a lyophilized composition is described as reconstitutable by the addition of water to produce a formulation of a specified ingredient concentration and pH, the lyophilized formulation can be reconstituted simply by the addition of water (i.e., additional It means that it can be so reconstituted by supplying amounts of ingredients or adding acids or bases to change the pH). The concentration and properties of the pre-lyophilized liquid formulation may also follow those described below when the lyophilized formulation is reconstituted to the same volume as the formulation pre-lyophilized. If the volumes are different, the concentrations of the formulations should be adjusted proportionally. For example, if the reconstituted volume is half the pre-lyophilized volume, the concentration of the component in the pre-lyophilized formulation should be half of the concentration in the reconstituted formulation.

Some formulations include fillers, which may or may not be the same as the sugar/polyol component. Typically, the formulations are sterile, which is accomplished by sterile filtration, for example using a 0.2 μm or 0.22 μm filter. Some formulations have a bioburden of ≦about 3 CFU/30 mL. Some formulations contain < about 0.1 EU/mg bacterial endotoxin. The formulations of the present invention are also generally stable upon freezing and thawing with low to undetectable levels of fragmentation and/or aggregation as further defined below. Still other formulations are stable at 40° C. for at least 3 months after reconstitution of the lyophilized cake. In some formulations, less than about 10% of the antibody is present as aggregates in the formulation. In some formulations, about 5% or less of the antibody is present as aggregates in the formulation.

In some formulations, the antibody is present at a concentration within the range of about 5 mg/mL to about 100 mg/mL. In some formulations, the antibody is present at a concentration within the range of about 5 mg/mL to about 50 mg/mL. In some formulations, the antibody is present at a concentration within the range of about 25 mg/mL to about 50 mg/mL. For example, the antibody may be present at a concentration of about 35-45 mg/ml or about 40 mg/ml. The antibody may be present in sterile liquid dosage forms from about 50 mg/vial to about 500 mg/vial, or more. The antibody may be present in a lyophilized dosage form from about 40 mg/vial to about 500 mg/vial. For example, the antibody may be present in a sterile liquid or lyophilized dosage form of about 250-350 mg/vial or about 200 mg/vial.

Antibodies used in the disclosed formulations can be cytotoxic agents, radiotherapeutic agents, immunomodulatory agents, second antibodies (e.g., to form antibody heteroconjugates), or enhance the activity of the formulated antibody. Or it can be coupled with a therapeutic moiety such as any other enhancing bioactive agent. However, antibodies are usually used in naked form (ie, not conjugated to any of these moieties).

The formulated antibody can be any of the antibodies described above, including any of the chimeric, veneered or humanized forms of antibody 9E4 described above.

Buffers are used in the disclosed formulations to achieve a suitable pH for the antibody, such as histidine, succinate, and citrate buffers. Some formulations have a pH within the range of about 5.5 to about 7, such as 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, Has a pH of 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, or 7.0. Some formulations have a pH between about 5.5 and about 6.5. Some formulations have a pH of about 6.0 and others have a pH of about 6.5. In some formulations, the citrate or succinate buffer is present at a concentration within the range of about 10 mM to about 30 mM, eg, about 15-25 mM or about 20 mM. Some citrate buffers contain sodium citrate dehydrate and citric acid monohydrate at concentrations within the ranges of about 15 mM to about 20 mM and about 2 mM to about 6 mM, respectively.

Suitable sugars and/or polyols for formulation include trehalose, sucrose, mannitol, or combinations thereof. Sugars/polyols act as bulking agents, cryoprotectants, and/or tonicity modifiers. For example, some formulations have trehalose present in a concentration within the range of about 220 mM to about 260 mM, sucrose present in a concentration within the range of about 220 mM to about 260 mM, or sucrose present in a concentration within the range of about 20 mM to about 40 mM. and mannitol present at a concentration within the range of about 200 mM to about 220 mM. Some formulations contain trehalose present at a concentration of about 230 mM or 240 mM. Other formulations include sucrose present at a concentration of about 230 mM or 240 mM. Other formulations include a mixture of sucrose present at a concentration of about 50 mM and mannitol present at a concentration of about 200 mM. Another formulation includes a mixture of sucrose present at a concentration of about 28 mM and mannitol present at a concentration of about 212 mM. Some such formulations are characterized by an osmolality within the range of about 250-400, 300-400, or 300-350 mOsm/kg, such as 335 mOsm/kg.

The formulation may contain a surfactant to reduce antibody aggregation and absorption to surfaces. Suitable surfactants include polysorbate 20 (PS20) present at a concentration within the range of about 0.005% to about 0.05% by weight. PS20 protects against aggregation or significant increase in turbidity that would otherwise occur in formulations of 9E4 antibodies. Polysorbate 20 may be present at a concentration within the range of about 0.01% to about 0.05%. For example, the concentrations are 0.005%, 0.01%, 0.015%, 0.02%, 0.025%, 0.03%, 0.035%, 0.04%, 0.045%, or 0.05%. Alternatively, in some formulations polysorbate 20 is approximately 0.05 g/L, 0.1 g/L, 0.15 g/L, 0.2 g/L, 0.25 g/L, 0.3 g/L It is present at concentrations within the range of L, 0.35 g/L, 0.4 g/L, 0.45 g/L, or 0.5 g/L. Some formulations contain polysorbate 20 at a concentration of 0.2 g/L (ie, 0.163 mmol/L).

Exemplary formulations (liquid, pre-lyophilized or reconstituted after lyophilization) are characterized by a pH within the range of about 5.5 to about 7, and (a) about 10 mg/ml to about 50 mg/ml a chimeric, veneered, or humanized form of antibody 9E4, or a fragment thereof that specifically competes with 9E4 for binding to antigen; citrate or succinate buffer present; (c) trehalose present at a concentration within the range of about 220 mM to about 260 mM, sucrose present at a concentration within the range of about 220 mM to about 260 mM, and from about 20 mM to about and ( d) Polysorbate 20 present at a concentration within the range of about 0.005% to about 0.05% by weight. For example, a formulation comprises (a) a light chain having an amino acid sequence set forth as SEQ ID NO:32 and a heavy chain comprising an amino acid sequence set forth as SEQ ID NO:37 with or without a C-terminal lysine at a concentration of about 40 mg/mL (b) citrate buffer at a concentration of about 20 mM; (c) trehalose at a concentration of about 230 mM; (d) polysorbate 20 at a concentration of about 0.02%; can contain.

Some lyophilized formulations comprise (a) a humanized form of antibody 9E4 or an antigen-binding fragment thereof; (b) citrate; (c) trehalose; A lyophilized formulation may contain about 200 mg of antibody. Some lyophilized formulations can be reconstituted with sterile water. Some lyophilized formulations contain 100-300 or 150-250 mg 9E4 antibody, 15-35 or 20-25 mg sodium citrate dehydrate, 1.65-2.75 or 2-2.3 mg citric acid Monohydrate, 360-500 or 400-470 mg trehalose dehydrate, and 0.5-1.5 mg or 0.75-1.25 mg polysorbate 20. An exemplary lyophilized formulation is 200 mg 9E4 antibody (e.g., humanized 9E4 antibody), 25 mg sodium citrate dehydrate, 2.15 mg citric acid monohydrate, 435 mg trehalose dehydrate, and 1 mg Contains polysorbate 20. Another exemplary lyophilized formulation is 200 mg 9E4 antibody (e.g., humanized 9E4 antibody), 25 mg sodium citrate dehydrate, 3.15 mg citric acid monohydrate, 435 mg trehalose dehydrate, and Contains 1 mg of polysorbate 20. Such formulations can be reconstituted to a volume of about 5 ml. Other lyophilized formulations contain the same proportions but different amounts as any disclosed in this paragraph (e.g., 400 mg antibody, 50 mg sodium citrate, 4.3 mg citric acid monohydrate, 870 mg trehalose Dehydrated and 2 mg polysorbate 20) of the same ingredients.

(b) a citrate buffer present at a concentration of about 10-30 or 15-25 mM, preferably about 20 mM; (c) trehalose present at a concentration of about 160-330 or 200-260 mM, such as about 230 mM; (d) about 0.1-0.3 or 0.15-0.25 g/L, such as about 0.25 g/L; Polysorbate 20 present at a concentration of 2 g/L; and (e) reconstituted to a pH of about 5.5-6.5, eg about 6.0.

The liquid or reconstituted lyophilized formulation may be substantially isotonic, implied by an osmolality of about 250-350 mOsm/kg water. Some formulations have an osmolality of about 335 mOsm/kg. Some formulations have an osmolality of 270-300 mOsm/kg. Liquid or reconstituted lyophilized formulations can also be hypertonic >350 mOsm/kg water or hypotonic (<250 mOsm/kg water).

Any of the formulations described may be made without pharmaceutical excipients or carriers and the like other than those listed as ingredients herein. Such formulations may be described as consisting of or consisting essentially of the recited ingredients when minor amounts of other ingredients are present that do not affect the properties of the formulation. The formulations are approved by the FDA for the preparation of drugs for human administration or may be made under acceptable good manufacturing practices (GMP).

VI. Diagnostic criteria for synucleinopathic disease The methods of the present invention include those diagnosed as having Lewy body disease by a qualified medical practitioner, or as evidenced by genetic or biochemical markers, family history or prodromal symptoms of the disease. It is commonly performed on subjects at that elevated risk compared to the general population. Such individuals include anyone who has been given a previous prescription for treatment or prevention of Lewy body disease. Diagnosis of synucleinopathy is recognized in the art for probable or probable Lewy body disease such as DSM-V or DSM IV-TR, the Lewy Body dementia association, and the Parkinson's disease society Can be based on criteria. However, diagnosis can also be based on the presence of any sign or symptom of Lewy body disease that leads the treating physician to conclude that the subject is likely to have Lewy body disease. Exemplary criteria for diagnosing probable or probable PD are provided below.
Group A: Occurrence of resting tremor, bradykinesia, rigidity and asymmetry Group B characteristics: Suggestion of selective diagnosis Marked postural instability in the first 3 years after symptom onset Freezing phenomenon in the first 3 years Hallucinations unrelated to medication in the first 3 years Dementia preceding motor symptoms or in the first year Supranuclear gaze palsy (other than limitation of upward gaze) or slowing of vertical saccades Severe symptoms unrelated to medication dysautonomia

Recording of conditions known to cause parkinsonism and plausibly associated with the subject's symptoms (eg, a well-located focal brain injury or use of neuroleptics within the past 6 months).

Criteria for possible diagnosis of Parkinson's disease:
At least 2 of the 4 features in Group A are present; at least 1 of these is tremor or bradykinesia and none of the features in Group B are present or symptoms have been present for less than 3 years. , none of the features in Group B are present to date; and either a substantial and sustained response to levodopa or a dopamine agonist has been documented, or the subject has had adequate testing for both levodopa and a dopamine agonist. Not done.

Diagnostic criteria for high likelihood of Parkinson's disease:
At least 3 or 4 features in group A are present, none of the features in group B are present, and substantial and sustained responses to levodopa or dopamine agonists are documented.

Exemplary criteria for the diagnosis of dementia with Lewy bodies are:
Presence of dementia At least two of the three core characteristics:
attention and concentration sway,
recurrent, well-formed visual hallucinations and spontaneous Parkinsonian-like motor signs.
Suggestive clinical features include the following:
Rapid eye movement (REM) sleep behavior disorder Severe neuroleptic hypersensitivity Low dopamine transporter uptake in the basal ganglia demonstrated by SPECT or PET imaging

In the absence of two core features, a probable diagnosis of DLB can also be made when at least one suggestive feature is present with one core feature in addition to dementia.

The possibility of DLB can be diagnosed with the presence of one core or suggestive feature in addition to dementia.

Early warning signs of Lewy body disease include, for example, EEG slowing, neuropsychiatric manifestations (depression, dementia, hallucinations, anxiety, apathy, anhedonia), autonomic changes ( orthostatic hypotension, bladder disorders, constipation, fecal incontinence, drooling, dysphagia, sexual dysfunction, changes in cerebral blood flow), sensory changes (abnormal sensations of smell, pain, and color discrimination), sleep disorders [REM sleep behavior disorder (RBD), restless leg syndrome/regular extreme exercise, hypersomnia, insomnia] and miscellaneous other signs and symptoms (fatigue, diplopia, blurred vision, seborrhea, weight loss/gain ). Genetic markers of risk for PD include mutations in the alpha-synuclein or Parkin, UCHLI, and CYP2D6 genes, particularly mutations at positions 30 and 53 of the alpha-synuclein gene. Although none of these genetic markers or early warning signs are diagnostic of Lewy body disease by themselves, they may contribute to the diagnosis of Lewy body disease individually or in combination.

VII. Treatment Regimens In therapeutic applications, regimens (dose, frequency and route of administration) known or suspected to be effective for ameliorating at least one sign or symptom or at least inhibiting further exacerbation of disease The antibody is administered to a subject diagnosed as having a synucleinopathic disease in. In prophylactic applications, there is still an increased risk of synucleinopathic disease in regimens known or suspected to be effective in inhibiting or delaying the onset of at least one sign or symptom of the disease. An antibody is administered to a subject who does not have sufficient symptoms to be diagnosed with the disease.

p<0.2, 0.1, 0.05 or 0.01 or even 0.001 in controlled animal models or clinical trials (e.g., Phase II, Phase II/III or Phase III trials) A regimen is considered therapeutically or prophylactically effective if it demonstrates a more favorable outcome in treated subjects compared to control subjects at the level of . However, due to variability in genetics, subject characteristics and circumstances, and disease subtypes among individual subjects, regimens that are effective in one subject may not be effective in another, or to varying degrees. can be effective.

An exemplary dosage range for the antibody is 3000-5000 mg of antibody to alpha-synuclein administered intravenously at intervals of 3-5 weeks, such as every 4 weeks. In some subjects, the dosage is 3500-4500 mg every 3-5 weeks, such as every 4 weeks. Subjects can receive the same or different dosages from each other (eg, depending on the weight of the subject). In some methods, subjects receive one of two fixed doses. For example, subjects weighing less than 65 kg may receive 3500 mg, and subjects weighing 65 kg or greater may receive 4500 mg. In some methods, the dosage range for at least some subjects is within the range of 45-75 mg/kg, such as 50-70 mg/kg, 45 mg/kg, 60 mg/kg or 65 mg/kg. Dosages are usually administered at intervals of 3 to 5 weeks, eg, every 28 days or every 4 weeks, or on multiple occasions every calendar month. A subject may receive at least 6, 9, 12 or 18 doses at such intervals, or may be administered while symptoms of the condition persist or for the remainder of the subject's life. In some regimens, an initial loading dose of 2000 mg is administered, followed by doses in the range of 2000 mg or more but less than the intended target dose until the intended target dose is reached. For example, subjects may receive an initial loading dose of 2000 mg followed by incremental titration to a dose of 3500 mg or a dose of 4500 mg. Incremental titration can be performed in a single subsequent dose or in gradual increments over multiple doses until a target dose or dose within a target range is reached. For example, a subject may receive a starting dose of 2000 mg followed by subsequent doses of 3500 mg. Alternatively, the subject may receive a starting dose of 2000 mg, followed by one or more subsequent doses of 2000 mg or more but less than 3500 mg, and subsequent doses of 3500 mg. Similarly, subjects may receive a starting dose of 2000 mg followed by subsequent doses of 4500 mg. Alternatively, the subject may receive a starting dose of 2000 mg, followed by subsequent doses of 2000 mg or more but less than 4500 mg and subsequent doses of 4500 mg. In some regimens, subjects receive doses of 3000-5000 mg of antibody intravenously every 4 weeks for at least 52 weeks. In subjects receiving a multiple dose regimen with doses within a specified range such as 3500-5000 mg, the subject may receive the same or different doses within a specified range at each administration. In some regimens, subjects receive the same dose within a specified range at each administration.

In another exemplary regimen, a dose of 1300-1700 mg of antibody is administered intravenously to a subject at intervals of 3-5 weeks. An exemplary dose is 1500 mg. Subjects can receive a single fixed dose or two or more different doses within this range based, for example, on the subject's weight. Some subjects dosed within this range receive 18-25 mg/kg, eg, 20 mg/kg of antibody. As in other methods, intervals can be 3-5 weeks, eg, every 4 weeks or every calendar month. A subject may receive at least 6, at least 9, at least 12, or at least 18 doses, or may be administered at such intervals for the duration of symptoms or for the remainder of the subject's life.

For any of the therapeutic regimens described above, the area under the curve can be calculated to indicate the amount of antibody delivered over time. Alternative treatment regimens can be devised (eg, using different routes of administration, frequencies or doses) to deliver substantially the same area under the curve (eg, within 25, 20 or 15%). Preferably, such alternative treatment regimens do not substantially exceed the Cmax of the designated regimen (eg, 25, 20 or 15% or less). Other routes of administration include topical, oral, subcutaneous, intraarterial, intracranial, intrathecal, intraperitoneal, intranasal or intramuscular.

Any treatment regimen may involve monitoring the subject undergoing treatment for changes in motor and/or cognitive impairment. Preferably, such monitoring includes at least one assessment before and after initiation of treatment. Preferably, the monitoring indicates a reduction in motor and/or cognitive deficits in response to treatment compared to before treatment was initiated, or a previous rate of decline in a subject or in a control patient not receiving any immunotherapy At least a reduction in the rate of decline relative to the rate of decline is indicated. Subjects may also be monitored for changes in autonomic dysfunction, gastrointestinal dysfunction, visual hallucinations or one or more psychological symptoms, among other signs or symptoms.

A subject's symptoms, for example motor symptoms such as tremor, stiffness and slowness of movement, may be monitored. A wearable system or on-body sensor may be used to assess and quantify a subject's motor symptoms. "On-body sensors" may be used in laboratory settings or in free-living conditions [S. Del Din, et al., J. of NeuroEngineering and Rehabilitation, 2016 13:46].

Subjects may be monitored using mobile device-based monitoring. A mobile device may be a smart phone, smart watch, wearable sensor, portable multimedia device or tablet computer. A subject's daily activities may be recorded using built-in mobile device sensors. Subjects may carry mobile devices to record their daily activities. Mobile device-based assessment and sensors may be used for remote passive monitoring of ambulation and mobility in subjects undergoing treatment, eg, for Parkinson's disease. [e.g., Lipsmeier, F., et al.. Mov Disord. 2017; 32 (suppl 2); W. Y. Cheng et al., 2017 IEEE/ACM International Conference on Connected Health: Applications, Systems and Engineering Technologies (CHASE), Philadelphia , PA, 2017, pp. 249-250]. Sensor data may be analyzed by machine learning-based activity profiling. Gait and mobility can be correlated with the MDS-UPDRS used in the clinic to assess the severity of Parkinson's disease.

Some mobile device-based monitoring (a) receives and receives data obtained from sensors internal and/or external to the mobile device regarding movement disorders in a subject having or suspected of having a synucleinopathic disease; A subject is provided with a mobile device programmed to transmit, the subject then performs a series of exercises to reveal a motor impairment, if any, and sensors internal or external to the device (b) collecting data transmitted from the mobile device; and (c) comparing the data obtained from the subject to control data to assess the presence or extent of movement disorders in the subject. may include In some mobile device-based monitoring, the mobile device is programmed to receive and transmit data from at least two external sensors attached to the subject's upper and lower extremities. In some mobile device-based monitoring, the mobile device acquires data from sensors on the subject's upper and lower extremities. In some mobile device-based monitoring, the mobile device is carried by the subject and acquires data from internal sensors. For some mobile device-based monitoring, the motion sequence includes tapping the device, sitting and standing.

A regimen of the invention may be administered in combination with another agent effective in treating or preventing the disease being treated. Other agents can be another immunotherapeutic agent described herein or other agents for treating Parkinson's disease, such as levodopa, benzaseride, carbidopa, dopamine agonists, non-ergot dopamine agonists, catechol -O-methyl ("COMT") inhibitors, such as entacopone or tolcopone; monoamine oxidase ("MAO") inhibitors, such as rasagaline, amantadine; Cholinergic agents may be used in combination with the regimens of the invention. Some such other agents reduce one or more symptoms of the disease without affecting the causative factor.

All publications (including GenBank accession numbers, UniProtKB/Swiss-Prot accession numbers, etc.), patents and patent applications cited are hereby incorporated by reference in their entirety. are incorporated herein for all purposes in their entireties by reference to the same extent as if specifically and individually indicated to be incorporated for the purposes of. In the event of any variation in the sequence or website associated with Genbank and UniProtKB/Swiss-Prot accession numbers, etc., or in the organization's disease criteria, this application refers to what is valid as of its effective filing date.

VIII. Example

Phase II Clinical Trial for Alpha-Synuclein Antibodies Study Design: The Phase II trial will examine the heavy chain variable region designated as SEQ ID NO: 10 and the light chain designated as SEQ ID NO: 5 in subjects with Parkinson's disease. An alpha-synuclein antibody with variable regions is performed. The study has two treatment groups and one control group. Subjects are randomized 1:1:1 into groups (N=300). The initial phase of the study is 52 weeks of double-blind treatment. During the initial phase of the study, subjects receive no other treatment for Parkinson's disease (including symptomatic treatment). Subjects in one treatment group receive a fixed dose of antibody of 1500 mg intravenously every 4 weeks. Subjects in the other treatment groups received 3500 mg or 4500 mg of antibody intravenously every 4 weeks depending on body weight, with subjects under 65 kg receiving lower doses and subjects weighing 65 kg or greater receiving higher doses. receive. Subjects in the second group receive a loading dose of 2000 mg and, as appropriate, additional titrating doses of 2000 mg or more until a target dose of 3500 mg or 4500 mg is reached. Dosing is continued for one year. The trial then had an extension period in which subjects in the placebo group initially received one of the two treatment regimens from the early phase and subjects from the early phase treatment group received the same treatment as before. continue. During the extension phase of the study, subjects may receive systematic treatment with the antibody subject in the study in addition to levodopa, but no other treatment for Parkinson's disease.

the purpose:
The primary objective was to compare the heavy chain variable region designated SEQ ID NO: 10 and SEQ ID NO: 5 using the Movement Disorder Society (MDS)-sponsored revision of the Unified Parkinson's Disease Rating Scale (MDS-UPDRS). To evaluate the efficacy of alpha-synuclein antibodies with designated light chain variable regions.
A secondary purpose is
- to evaluate the effects of antibodies on signs and symptoms of Parkinson's disease, including changes in DaT-SPECT signaling and motor and cognitive deficits; - to evaluate the safety and tolerability of antibodies for up to 104 weeks; To evaluate the pharmacokinetics of

Passive Monitoring of Mobility in Early-Stage Parkinson's Disease Patients in a Phase I Alpha-Synuclein Antibody Clinical Trial Using Smartphone Sensors

1. Summary—Gait and mobility in patients with early stage Parkinson's disease (PD) were measured using smartphone-based passive monitoring. 44 PD patients and 35 age and gender-matched in a multiple escalating dose clinical trial of an alpha-synuclein antibody having a heavy chain variable region designated as SEQ ID NO: 10 and a light chain variable region designated as SEQ ID NO: 5 healthy individuals underwent smartphone-based assessments of up to 24 weeks and up to 6 weeks, respectively [Lipsmeier, F., et al.. Mov Disord. 2017; 32 (suppl 2); W. Y. Cheng et al., 2017 IEEE/ ACM International Conference on Connected Health: Applications, Systems and Engineering Technologies (CHASE), Philadelphia, PA, 2017, pp. 249-250].

For 'passive monitoring', subjects carried a smartphone as part of their daily routine and continuously recorded movement data using the smartphone's sensors. A total of over 30,000 hours of passive monitoring data were collected. To classify sensor signals into activity profiles, we constructed a Human Activity Recognition (HAR) model using a deep neural network (DNN) trained on previously published data. Activity profiles of participants determined by the HAR model showed significant differences between PD patients and healthy controls in the percentage of walking time and the frequency with which controls changed postures (sitting and standing sitting and standing). .

2. Method

A. Data Collection Analysis focused only on examining differences between the HC and PD cohorts and did not examine antibody-related effects. In total, 24,104 hours of passive monitoring data were recorded for the PD cohort and 8,614 hours for the HC cohort. Consistent with the approach of Rai, A. et al. (MobiCom'12, August 22-26, 2012), we filtered out accelerometer data where the standard deviation of the Euclidean norm was less than 0.03 m/s2 and longer than 30 min. This step removed 14% of the passive monitoring data, since it appears that no smartphones were carried by the subjects during these spans.

B. HAR
A 9-layer neural network model structure was used. Similar structures have been used previously for HAR and have been shown to outperform traditional machine learning methods (FJ Ordonez and D. Roggen, Sensors 2016, 16, 115). Set (GM Weiss and JW Lockhart, Proceedings of the AAAI-12 Workshop on Activity Context Representation: Techniques and Languages, Toronto, CA. 2012; A. Stisen, et al., 13th ACM Conference on Embedded Networked Sensor Systems, Seoul, Korea , 2015) classified six activities: walking, stairs, jogging, sitting, standing, and recumbent.The continuous accelerometer data were downsampled to 20 Hz, and 75% overlap with adjacent 4 Segmented into second windows.

3. result

A. Verification of human activity recognition performance:
To ensure that the HAR model can accurately translate sensor data into activity profiles, model performance was first analyzed on a holdout validation set. The HAR model was able to correctly distinguish ambulatory activity (walking, stairs, jogging) from resting activity (sitting, standing, lying) with greater than 98% accuracy. Additional validation on the labeled ambulation and resting data from the test data also showed that the HAR model was able to successfully profile the walking segment with 96.9% accuracy and the resting segment with 99.5% accuracy. showed that.

B. Activity Profile Comparison Mobility was quantified for each subject by calculating the percentage of time the subject was engaged in ambulatory activity (walking, stairs, jogging) relative to the patient's total passive monitoring coverage time. The overall proportion of different ambulatory activities to total coverage for the PD and HC cohorts was calculated. A median gait span of 9.7% for all coverage spans was detected in the PD cohort, in contrast to 15.1% in the HC cohort. The HC cohort had significantly higher levels of ambulatory activity per subject than the PD cohort, with a Mann-Whitney test P-value of 2.43E-8.

C. Comparison of the number of sitting-to-standing and standing-to-sitting switches (A. Zijlstra, et al., J. NeuroEngineering and Rehabilitation 2012, 9:75]. From activity profiles, coverage-normalized STS events were A median number of STSs per hour was observed for PD patients of 1.44, which was significantly lower than 1.74 for HC subjects.The P-value of the Mann-Whitney test between the two groups was 1.60E-8.

4. Conclusion

Results from this study show that it is feasible to measure ambulation and mobility in early-stage PD patients using smartphone-based passive monitoring. Sensor data collected during passive monitoring provide previously inaccessible, ecologically relevant insights into a patient's daily behavior and function. A significant difference was observed between PD patients and healthy controls (HC).

Claims (107)

A method of treating or preventing a subject having or at risk of having a synucleinopathic disease, wherein the subject is administered a dose of 3000-5000 mg of an antibody against alpha-synuclein intravenously at intervals of 3-5 weeks. or administering to the subject another regimen that delivers the antibody in substantially the same area under the curve. 2. The method of claim 1, wherein the dose is 3500-4500 mg. 2. The method of claim 1, wherein multiple subjects with or at risk for synucleinopathic disease are administered, subjects receiving one or two fixed doses. 4. The method of claim 3, wherein subjects weighing less than 65 kg receive a dose of 3500 mg of the antibody and subjects weighing 65 kg or greater receive a dose of 4500 mg. 2. The method of claim 1, wherein the dose is 45-75 mg/kg. 2. The method of claim 1, wherein the dose is 50-70 mg/kg. 2. The method of claim 1, wherein the interval is 28 days. 2. The method of claim 1, wherein the subject receives at least 6 doses of the antibody at 3-5 week intervals. 2. The method of claim 1, wherein the subject receives at least 12 doses of the antibody at 3-5 week intervals. 2. The method of claim 1, wherein the subject receives at least 18 doses of the antibody at intervals of 3-5 weeks. 11. The method of any one of claims 8-10, wherein the interval is 4 weeks. 2. The method of claim 1, wherein the subject receives the antibody every 4 weeks for at least 52 weeks. 2. The method of claim 1, further comprising monitoring the subject for changes in movement, cognitive impairment, autonomic dysfunction, gastrointestinal dysfunction, visual hallucinations, or psychological symptoms. to monitor
(a) a mobile device programmed to receive and transmit data obtained from sensors internal and/or external to the mobile device regarding movement disorders in a subject having or suspected of having a synucleinopathic disease; providing to a subject, after which the subject performs a series of exercises to reveal a movement disorder, if any, and sensors internal or external to the device acquire data on the movements;
(b) collecting data transmitted from the mobile device; and (c) comparing data obtained from the subject to control data to assess the presence or extent of movement disorders in the subject. 13. The method according to 13. 15. The method of claim 14, wherein the mobile device is programmed to receive and transmit data from at least two external sensors attached to the subject's upper and lower extremities. 16. The method of claim 14 or 15, wherein the mobile device acquires data from sensors on the subject's upper and lower extremities. 16. The method of claim 14 or 15, wherein a mobile device is carried by the subject and acquires data from internal sensors. 16. The method of claim 14 or 15, wherein the sequence of movements includes tapping the device, sitting and standing. 19. The method of claims 13-18, wherein monitoring indicates a reduction in motor impairment or a reduction in cognitive impairment in response to administering the antibody. 2. The method of claim 1, wherein the antibody binds within residues 115-130 of alpha-synuclein. 2. The method of claim 1, wherein the antibody binds within residues 118-126 of alpha-synuclein. 2. The method of claim 1, wherein the antibody binds within residues 117-123 of alpha-synuclein. 20. The method of any of claims 1-19, wherein the antibody is NI-202.21D11. 20. The method of any of claims 1-19, wherein the antibody comprises three heavy chain CDRs designated in SEQ ID NOs: 139-141, respectively, and three light chain CDRs designated in SEQ ID NOs: 143-145, respectively. 20. The method of any of claims 1-19, wherein the antibody comprises a heavy chain variable region designated as SEQ ID NO:138 and a light chain variable region designated as SEQ ID NO:142. 2. The method of claim 1, wherein the antibody comprises the three light chain Kabat CDRs of SEQ ID NO:5 and the three heavy chain Kabat CDRs of SEQ ID NO:10. The antibody comprises a heavy chain variable region of any of SEQ ID NOs:8-11 and a light chain variable region of any of SEQ ID NOs:3-5, preferably the heavy chain variable region is the heavy chain variable region of SEQ ID NO:10 and the light chain variable region is the light chain variable region of SEQ ID NO:5. 28. The method of any of claims 1-27, wherein the antibody is of the human IgGl isotype. 29. The method of claim 28, wherein the antibody comprises a heavy chain constant region of SEQ ID NO:35 and a light chain constant region of SEQ ID NO:30, optionally without the C-terminal lysine of SEQ ID NO:35. 29. The method of claim 28, wherein the antibody comprises a heavy chain constant region of SEQ ID NO:35, optionally absent the C-terminal lysine of SEQ ID NO:35, and a light chain constant region of SEQ ID NO:13. 22. The method of any of claims 1-21, wherein the antibody is 9E4. 22. The method of any of claims 1-21, wherein the antibody comprises three heavy chain CDRs designated in SEQ ID NOs:146-148, respectively, and three light chain CDRs designated in SEQ ID NOs:149-151, respectively. 22. Any of claims 1-21, wherein the antibody comprises a heavy chain designated in SEQ ID NO:37 and a light chain designated in SEQ ID NO:32, wherein the C-terminal lysine of SEQ ID NO:37 may be absent. the method of. 2. The method of claim 1, wherein the antibody binds within residues 1-20 of alpha-synuclein. 35. The method of claim 34, wherein the antibody binds within residues 4-15 of alpha-synuclein. 20. The method of any of claims 1-19, wherein the antibody is NI-202.12F4. 20. The method of any of claims 1-19, wherein the antibody comprises three heavy chain CDRs designated in SEQ ID NOs: 131-133, respectively, and three light chain CDRs designated in SEQ ID NOs: 135-137, respectively. 20. The method of any of claims 1-19, wherein the antibody comprises a heavy chain variable region designated as SEQ ID NO:130 and a light chain variable region designated as SEQ ID NO:134. 39. The method of any of claims 1-38, wherein the subject weighs less than 65 kg and is administered a dose of 3500 mg of antibody every 4 weeks. Administering a loading dose of 2000 mg of antibody prior to administering an antibody within the range of 3000-5000 mg, and one or more subsequent doses of 2000 mg or more but less than 3500 mg until a dose of 3500 mg is reached. 40. The method of any of claims 1-39, wherein incremental titrations may be performed and all doses are separated by intervals of 3-5 weeks. 41. The method of claim 40, wherein the 3500 mg dose is administered in the next interval after the 2000 mg dose. 41. The method of claim 40, wherein the dose is gradually increased over multiple intervals prior to administration of the 3500 mg dose. 39. The method of any one of claims 1-38, wherein the subject has a body weight greater than 65 kg and is administered a dose of 4500 mg of antibody every 4 weeks. Prior to administering antibody at 4500 mg, a loading dose of 2000 mg of antibody was administered followed by escalating titration with one or more subsequent doses greater than or equal to 2000 mg but less than 4500 mg until a dose of 4500 mg was reached. 39. The method of any of claims 1-38, wherein all doses are separated by intervals of 3-5 days. 45. The method of claim 44, wherein the 4500 mg dose is administered in the next interval after the 2000 mg dose. 45. The method of claim 44, wherein the dose is gradually increased over multiple intervals prior to administration of the 4500 mg dose. 47. The method of any of claims 44-46, further comprising monitoring the subject for changes in movement, cognitive impairment, autonomic dysfunction, gastrointestinal dysfunction, visual hallucinations, or psychological symptoms. to monitor
(a) a mobile device programmed to receive and transmit data obtained from sensors internal and/or external to the mobile device regarding movement disorders in a subject having or suspected of having a synucleinopathic disease; providing to a subject, after which the subject performs a series of exercises to reveal a movement disorder, if any, and sensors internal or external to the device acquire data on the movements;
(b) collecting data transmitted from the mobile device; and (c) comparing data obtained from the subject to control data to assess the presence or extent of movement disorders in the subject. 47. The method according to 47. 49. The method of claim 48, wherein the mobile device is programmed to receive and transmit data from at least two external sensors attached to the subject's upper and lower extremities. 50. The method of claim 48 or 49, wherein the mobile device acquires data from sensors on the subject's upper and lower extremities. 50. The method of claim 48 or 49, wherein a mobile device is carried by the subject to acquire data from internal sensors. 50. The method of claim 48 or 49, wherein the sequence of movements includes tapping the device, sitting and standing. 53. The method of any of claims 47-52, wherein monitoring indicates a reduction in motor impairment or a reduction in cognitive impairment in response to administering the antibody. 54. The method of any of claims 1-53, wherein the synucleinopathy is Parkinson's disease. 54. The method of any of claims 1-53, wherein the synucleinopathy is dementia with Lewy bodies. 54. The method of any of claims 1-53, wherein the synucleinopathy is multiple system atrophy. 54. The method of any of claims 1-53, wherein the synucleinopathy is progressive supranuclear palsy. 54. The method of any of claims 1-53, wherein the synucleinopathy is REM sleep behavior disorder. 54. The method of any of claims 1-53, wherein the synucleinopathy is Alzheimer's disease with Lewy bodies in the amygdala. 60. The method of any of claims 1-59, wherein the subject has said disease. 61. The method of any of claims 1-60, wherein the subject is not receiving symptomatic therapy for Parkinson's disease in combination with the antibody. 61. The method of any of claims 1-60, wherein the antibody is administered in combination with levodopa. A method of treating or preventing a subject having Lewy body disease comprising administering to the subject a dose of 1300-1700 mg of an antibody against alpha-synuclein intravenously at intervals of 3-5 weeks; administering another regimen that delivers the antibody at approximately the same area under the curve. 64. The method of claim 63, wherein the dose is 1500 mg. 64. The method of claim 63, administered to a plurality of subjects with Lewy body disease, wherein the subjects receive one or two fixed doses. 64. The method of claim 63, wherein the subject receives a dose of 18-25 mg/kg. 67. The method of claim 66, wherein the subject receives a dose of 20 mg/kg. 64. The method of claim 63, wherein the interval is 28 days. 64. The method of claim 63, wherein the subject receives at least 6 doses of the antibody at 3-5 week intervals. 64. The method of claim 63, wherein the subject receives at least 12 doses of the antibody at 3-5 week intervals. 64. The method of claim 63, wherein the subject receives at least 18 doses of the antibody at 3-5 week intervals. 71. The method of any one of claims 68-70, wherein the interval is 4 weeks. 64. The method of claim 63, wherein the subject receives the antibody every 4 weeks for at least 52 weeks. 64. The method of claim 63, further comprising monitoring the subject for changes in movement, cognitive impairment, autonomic dysfunction, gastrointestinal dysfunction, visual hallucinations, or psychological symptoms. to monitor
(a) programmed to receive and transmit data obtained from sensors internal and/or external to the mobile device regarding movement disorders in a subject having or suspected of having Lewy body disease; providing a mobile device to a subject, after which the subject performs a series of exercises to reveal a motor impairment, if any, and sensors internal or external to the device acquire data regarding the exercise;
(b) collecting data transmitted from the mobile device; and (c) comparing data obtained from the subject to control data to assess the presence or extent of movement disorders in the subject. 74. The method according to 74. 76. The method of claim 75, wherein the mobile device is programmed to receive and transmit data from at least two external sensors attached to the subject's upper and lower extremities. 77. The method of claim 75 or 76, wherein the mobile device acquires data from sensors on the subject's upper and lower extremities. 77. The method of claims 75 or 76, wherein a mobile device is carried by the subject and acquires data from internal sensors. 77. The method of claim 75 or 76, wherein the sequence of movements includes tapping the device, sitting and standing. 80. The method of any of claims 74-79, wherein monitoring indicates a reduction in motor impairment or a reduction in cognitive impairment in response to administering the antibody. 64. The method of claim 63, wherein the antibody binds within residues 115-130 of alpha-synuclein. 64. The method of claim 63, wherein the antibody binds within residues 118-126 of alpha-synuclein. 64. The method of claim 63, wherein the antibody binds within residues 117-123 of alpha-synuclein. 81. The method of any of claims 63-80, wherein the antibody is NI-202.21D11. 81. The method of any of claims 63-80, wherein the antibody comprises three heavy chain CDRs designated in SEQ ID NOS:139-141, respectively, and three light chain CDRs designated in SEQ ID NOS:143-145, respectively. 81. The method of any of claims 63-80, wherein the antibody comprises a heavy chain variable region designated as SEQ ID NO:138 and a light chain variable region designated as SEQ ID NO:142. 64. The method of claim 63, wherein the antibody comprises the three light chain Kabat CDRs of SEQ ID NO:5 and the three heavy chain Kabat CDRs of SEQ ID NO:10. The antibody comprises a heavy chain variable region of any of SEQ ID NOs:8-11 and a light chain variable region of any of SEQ ID NOs:3-5, preferably the heavy chain variable region is the heavy chain variable region of SEQ ID NO:10 and the light chain variable region is the light chain variable region of SEQ ID NO:5. 89. The method of any of claims 63-88, wherein the antibody is of the human IgGl isotype. 90. The method of claim 89, wherein the antibody comprises a heavy chain constant region of SEQ ID NO:35, optionally absent a C-terminal lysine, and a light chain constant region of SEQ ID NO:30. 90. The method of claim 89, wherein the antibody comprises a heavy chain constant region of SEQ ID NO:35, optionally free of C-terminal lysine, and a light chain constant region of SEQ ID NO:13. 83. The method of any of claims 63-82, wherein the antibody comprises three heavy chain CDRs designated in SEQ ID NOs:146-148, respectively, and three light chain CDRs designated in SEQ ID NOs:149-151, respectively. 83. Any of claims 63-82, wherein the antibody comprises a heavy chain designated in SEQ ID NO:37 and a light chain designated in SEQ ID NO:32, wherein the C-terminal lysine of SEQ ID NO:37 may be absent. the method of. 64. The method of claim 63, wherein the antibody binds within residues 1-20 of alpha-synuclein. 95. The method of claim 94, wherein the antibody binds within residues 4-15 of alpha-synuclein. 81. The method of any of claims 63-80, wherein the antibody is NI-202.12F4. 81. The method of any of claims 63-80, wherein the antibody comprises three heavy chain CDRs designated in SEQ ID NOs:131-133, respectively, and three light chain CDRs designated in SEQ ID NOs:135-137, respectively. 81. The method of any of claims 63-80, wherein the antibody comprises a heavy chain variable region designated as SEQ ID NO:130 and a light chain variable region designated as SEQ ID NO:134. 99. The method of any of claims 63-98, wherein the synucleinopathy is Parkinson's disease. 99. The method of any of claims 63-98, wherein the synucleinopathy is dementia with Lewy bodies. 99. The method of any of claims 63-98, wherein the synucleinopathy is multiple system atrophy. 99. The method of any of claims 63-98, wherein the synucleinopathy is progressive supranuclear palsy. 99. The method of any of claims 63-98, wherein the synucleinopathy is REM sleep behavior disorder. 99. The method of any of claims 63-98, wherein the synucleinopathy is Alzheimer's disease with Lewy bodies in the amygdala. 105. The method of any of claims 63-104, wherein the subject has said disease. 106. The method of any of claims 63-105, wherein the subject is not receiving symptomatic therapy for Parkinson's disease in combination with the antibody. 106. The method of any of claims 63-105, wherein the antibody is administered in combination with levodopa.