RU2797677C2 - Alpha-synuclein modulators - Google Patents

Abstract

FIELD: pharmaceuticals.

SUBSTANCE: invention relates to the use of an aminopyrazole compound 4-[2-(4-bromophenylsulfanyl)acetylamino]-1-phenethyl-1H-pyrazole-3-carboxylic acid or a pharmaceutically acceptable salt thereof or a composition of the said aminopyrazole compound or a pharmaceutically acceptable salt thereof for the preparation of a medicinal product to treat alpha synucleinopathy: caused by Lewy bodies, Lewy neuritis or aggregates α-synuclein in neurons and glia; or caused by a mutation in the alpha-synuclein (SNCA) gene; or mediated by the activity of FAF1 in relation to the regulation of the amounts of LC-3 and p62, the activity of FAF1 in relation to the regulation of the amount α-synuclein, or mediated by the amount of aggregated α-synuclein, or mediated by the amount α-synuclein, which is phosphorylated at the amino acid S129 in the cells of the specified subject; or caused by accumulation of misfolded alpha synuclein or accumulation of alpha synuclein aggregates in the cells of the said subject.

EFFECT: effective treatment of alpha-synucleinopathy caused by the above reasons.

8 cl, 16 dwg

Description

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. Provisional Application No. 62/521758, filed June 19, 2017, the disclosure of which is incorporated herein by reference in its entirety.

FIELD OF TECHNOLOGY

[0002] This disclosure relates to compositions and methods for modulating alpha - synuclein protein levels via the FAF1 gene and protein, and to inhibitors of FAF1 activity to modulate the level, or aggregation, or modification of alpha - synuclein for the treatment or prevention of alpha - synuclein-mediated synucleinopathies. The disclosure includes appropriate diagnostic methods and methods of treatment using compounds having activity in modulating FAF1 levels or activity.

BACKGROUND OF THE INVENTION

[0003] FAF1 is a Fas apoptosis receptor-associated protein called Fas - associated factor 1. FAF1 binds to Fas when Fas interacts with a ligand called Fas ligand (FasL) and promotes apoptosis. Thus, FAF1 is a pro-apoptotic protein. FAF1 has an N - terminal 200 amino acid region that binds to Fas (Ryu SW et al. 2003, Yu C et al. 2016). FAF1 then activates caspase 8 and the apoptotic signaling pathway by activating the caspase cascade. FAF1 also activates JNK and binds to PARP1 and thus stimulates the necrosis initiation signaling pathway (Yu C et al. 2016). However, in the inactivated state, FAF1 is associated with HSP70 and maintained as an inactive form (Kim HJ et al., 2005, Gao X et al., 2015). FAF1 is reported to be overexpressed in the post-mortem tissue of Parkinson's patients (Betarbet R et al. 2008).

MJ et al. 2003). [0004] Synucleinopathies are a group of neurodegenerative diseases caused by the accumulation of misfolded alpha synuclein and its aggregates, such as inclusions of Lewy bodies (LB) or Lewy neurites (LN). Examples include Parkinson's disease (PD), Lewy body dementia (DLB) and multiple system atrophy (MSA) (Spillantini MG and Goedert M 2000,

MJ et al. 2003).

[0005] Alpha - synuclein, which has the name of the SNCA gene, is a small 14 kDa cytoplasmic protein, mainly located in synaptic vesicles in neuronal cells, and performs the functions of activating the signaling pathway and transmission (Snead D. and Eliezer D., 2014) . The protein consists of three domains: an N - terminal domain with amphipathic helices, a central domain with flexible beta - sheets, and a C - terminal tail with many acidic residues (Gallegos S et al. 2015). Protein is known to exist in soluble form as a monomer or tetramer. However, alpha - synuclein folds and aggregates to form oligomers and other multimers, and may, for various unclear reasons, coalesce into a fibrillar structure called Lewy body and Lewy neuritis. The process includes C - terminal proteolysis, serine phosphorylation at position 129, monoubiquitination, and other modifications (Sato H et al., 2013, Oueslati A, 2016). Causes of misfolding and aggregation of alpha synuclein include protein accumulation due to overexpression of the protein by gene multiplication or decreased degradation or alteration of the properties of the protein itself due to mutation. However, other non-genetic causes are also known, such as environmental toxins, cellular stresses such as cellular toxicity, ER stress, and oxidative stress (Rockenstein E et al. 2014, Ingelsson M, 2016). In addition, the level of alpha - synuclein and its aggregation are affected by polymorphic variants of the SNCA locus (Mata IF et al., 2010).

[0006] The toxicity of misfolded alpha - synuclein multimers and Lewy bodies has been studied and may be associated with various pathogenic processes, in particular with dysfunctions in membrane transport, including exocytosis, transport between the ER and the Golgi apparatus, ER stress, Golgi homeostasis, endocytosis , autophagy and oxidative stress (Wang T and Hay JC 2016, Snead D et al. 2014). In addition, alpha synuclein toxicity can also include mitochondrial toxicity and inflammation (Ingelsson M, 2016). Oligomers of alpha - synuclein and Lewy bodies can interfere with the proteasome pathway and are associated with the destruction of the plasma membrane and the formation of pores, which lead to the activation of apoptosis in cells (Gallegos S et al. 2015).

MJ 2003). Эти заболевания вызваны тельцами Леви или невритами Леви или агрегатами альфа-синуклеина в нейронах и глие. Клинически они характеризуются хроническим и прогрессирующим снижением двигательных, когнитивных и вегетативных функций в зависимости от распределения поражений. [0007] "Synucleinopathies" are a group of neurodegenerative disorders having a range of clinical symptoms such as Parkinson's disease (PD), Parkinson's disease with dementia (PDD), Lewy body dementia (DLB), multiple system atrophy (MSA), atypical parkinsonism , atypical Parkinson's disease and pure autonomic failure (PAF) (Spillantini MG and Goedert M 2000,

MJ 2003). These diseases are caused by Lewy bodies or Lewy neuritis or alpha - synuclein aggregates in neurons and glia. Clinically, they are characterized by a chronic and progressive decline in motor, cognitive, and autonomic functions, depending on the distribution of lesions.

[0008] Parkinson's disease is the second most common neurodegenerative disease of our time (Feng LR et al. 2010), commonly thought of as a movement disorder. The pathological hallmark of PD is selective progressive degeneration and loss of dopaminergic neurons in the substantia nigra (SNc), especially in the compact substantia nigra (SNpc) region (Luk KC et al. 2012) in the midbrain responsible for movement impairment. However, symptoms are often accompanied by autonomic, cognitive and psychiatric problems (Jankovic J 2008).

[0009] Another hallmark of PD is the presence of cytoplasmic inclusions called Lewy bodies (LB) and Lewy neurites (LN) in dopaminergic neuronal cells, which are observed in post-mortem tissues in more than 90 percent of patients with Parkinson's disease. The main protein component of LB is misfolded and aggregated alpha - synuclein (Angot E et al. 2012).

[0010] Certain genetic changes, point mutations, and gene multiplication of the SNCA gene are believed to cause familial Parkinson's disease (PD) or a causative reaction for PD. The dominant inheritance of these genetic mutations suggests a mechanism for enhancing function, such as aggregation due to changes in protein properties or increased protein levels (Devine MJ et al. 2011, Appel - Cresswell S et al. 2013, Kasten M and Klein C 2013, Petrucci S et al. 2016). Certain polymorphisms in the SNCA gene or mutations in the distal enhancer region can increase its expression and hence aggregation, and have been reported to be associated with PD (Mata IF et al. 2010, Soldner F et al. 2016).

[0011] In addition to mutations in the SNCA gene, many mutations are associated with Parkinson's disease and associated synucleinopathies (Wang C - D and Chan P 2014, Kalinderi K et al. 2016, Federoff M 2015, Nussbaum R 2017). Some of these genes are thought to be involved in alpha - synuclein processing and accumulation. For example, a mutation in GBA (the gene for glucocerebrosidase) accounts for 23% of dementia with Lewy bodies and 21% with PD (Sidransky E et al. 2009, Jesus S et al. 2016). GBA mutants have been reported to increase SNCA accumulation by altering alpha - synuclein processing (Cullen V et al. 2011, Fernandes HJ 2016). Other known genes that are associated with these diseases are Parkin, LRRK2, DJ - 1 and PINK1. Although the role of these genes in the pathogenesis of parkinsonism and other synucleinopathies is largely unknown, these genes perform post-translational modification and protein processing functions such as ubiquitination, membrane traffic, autophagy, and protein folding, and alpha - synuclein overexpression is due to mutations in some of the these genes (Walden H and Muqit MMK 2017, Galegos S 2015).

[0012] Dementia with Lewy bodies (DLB) is the second most common form of dementia, with clinical symptoms of progressive cognitive decline with dementia and fluctuating cognition, particularly visual hallucination. Post-mortem tissues show numerous Lewy bodies (LBs) in the cerebral cortex except in the substantia nigra, and these LBs include alpha - synuclein (Gaser F et al. 2005, Goeder M et al. 2017).

[0013] Multiple system atrophy (MSA) is a fatal neurodegenerative disease with symptoms of autonomic failure, parkinsonism and cerebellar ataxia in various combinations (Daniela Kuzdas - Wood et al. 2014). It is a rare disease with a prevalence of 1.9 - 4.9/100,000 and a survival rate of 6 - 9 years (Stefanova et al. 2009), and develops in adults at a mean age of 57 years. In terms of pathology, there are widespread cytoplasmic inclusions (GCIs) in glia throughout the CNS (Trojanowski JQ and Revesz T 2007), which are composed of filamentous alpha - synuclein. Newly proposed criteria are predominant cerebellar ataxia with predominant motor features and neuropathological demonstration of alpha - synuclein - positive CNS glioplasitic inclusions with neurodegenerative changes in striatonigral or olivopontocerebellar structures (Gilman S, 2008).

BRIEF DESCRIPTION OF THE FIGURES

[0014] Figure 1 illustrates overexpression in SH - SY5Y cells of the transfected FAF1 - FLAG transgene.

[0015] Figure 2 shows α - synuclein aggregation in SH - SY5Y cells that overexpress the FAF1 - FLAG transgene.

[0016] Figures 3A and 3B show the formation of Lewy body-like forms in SH - SY5Y cells overexpressing the transfected FAF1 - FLAG transgene. Figure 3A shows cells transfected with vector control. 3B shows cells transfected with the FAF1 - FLAG transgene. DAPI - DAPI staining; α - syn - staining of anti - α - synuclein antibodies; Ps129 - α - syn - staining with anti - phosphorylated α - synuclein staining; combined - superposition of three images.

[0017] Figure 4 illustrates FAF1 overexpression in the midbrain of mice having a FAF1 - adenovirus expression construct microinjected into midbrain tissue.

[0018] Figure 5 shows the attenuation of FAF1 expression in SH - SY5Y cells co - transfected with the α - synuclein gene and the corresponding siRNA.

[0019] Figure 6 shows changes in p62 and LC3 protein expression in SH - SY5Y cells transfected with the FAF1 gene.

[0020] Figure 7 shows KM - 819 binding to FAF1 in cells transfected with the FAF1 - FLAG construct.

[0021] Figure 8 shows the inhibition of FAF1 - caspase 8 and FAF1 - JNK complexes by KM - 819 in cell extracts.

[0022] Figure 9 shows inhibition of caspase 8 activity in caspase 3 cleavage and JNK phosphorylation by KM - 819 in cell extracts.

[0023] Figure 10 shows a decrease in p62 protein and an increase in LC3 protein in SH - SY5Y cells overexpressing FAF1 and treated with KM - 819.

[0024] Figure 11 shows the reduction of α - synuclein protein in cells co - transfected with the FAF1 - FLAG expression construct and the α - synuclein - GFP (green fluorescent protein) expression construct.

[0025] Figure 12 shows the reduction in α - synuclein and the amount of S129 phosphorylated α - synuclein in midbrain tissue of FAF1-overexpressing transgenic mice following administration of KM - 819.

[0026] Figure 13 shows the decrease in the amount of α - synuclein in the hippocampus in transgenic mice overexpressing FAF1 after administration of KM - 819.

[0027] Figure 14 shows the decrease in the amount of α - synuclein in the cerebral cortex of transgenic mice overexpressing FAF1 after administration of KM - 819.

[0028] Figure 15 shows the decrease in the amount of α - synuclein in the cerebellum of transgenic mice overexpressing FAF1 after administration of KM - 819.

[0029] Figure 16 shows the decrease in the amount of α - synuclein in the striatum of transgenic mice overexpressing FAF1 after administration of KM - 819.

DESCRIPTION

[0030] the Invention came about due to the unexpected discovery that FAF1 increases the accumulation of alpha - synuclein in neuronal cells and its aggregation. After this study, the authors confirmed that the FAF1 inhibitor based on an aminopyrazole derivative indeed reduces the accumulation of alpha - synuclein in neuronal cells and its aggregation. The invention also came to light through the discovery that FAF1 inhibits autophagy by increasing the p62 protein and decreasing LC3. And the inventors' study confirmed that the aminopyrazole derivative FAF1 inhibitor indeed inverted FAF1 activity, enhancing autophagy by reducing p62 protein and increasing LC3. These results are unexpected since FAF1 is a pro-apoptotic protein that promotes cell death, and the autophagy activity of FAF1 is a novel finding.

[0031] The present invention relates to compositions and methods for modulating alpha - synuclein protein levels with a FAF1 gene and protein, and FAF1 inhibitors for modulating alpha - synuclein levels or aggregation or modification to treat or prevent alpha - synuclein mediated synucleinopathies. The invention includes appropriate diagnostic methods and methods of treatment using compounds having activity in relation to the modulation of FAF1 levels.

[0032] The present invention relates generally to compositions and methods for using antibodies, peptides, nucleic acids such as aptamers or antisense nucleic acids or siRNA or the like, or small molecules that alter FAF1 expression or FAF1 protein activity in human cells that provide blocking or slowing the progression of synucleinopathies. Therefore, compositions and methods are provided for providing modified gene sequences and altered FAF1 protein expression and activities that affect disease progression. Also described are the use of such compositions and methods for detecting and quantifying the level of alpha - synuclein and/or characterizing the form of synuclein in a sample for diagnostic or prognostic purposes.

[0033] Surprisingly, using a transfected cell assay, FAF1 was found to increase the level of alpha - synuclein in cells in a dose-dependent manner. In addition, overexpression of FAF1 in animals by infection with recombinant FAF1-expressing adenovirus increases alpha - synuclein levels. In addition, the reduction of FAF1 due to RNAi transfection attenuates alpha synuclein levels. The authors also found that FAF1 inhibits autophagy by increasing LC - 3II and p62, suggesting that alpha - synuclein is, at least in part, transmitted by autophagy pathways in cells in vitro and in animals.

[0034] The inventors have developed the FAF1 inhibitor KM - 819 (KR - 88493, US Pat. No. 7,939,550, You SE et al., 2016, which is incorporated by reference in its entirety and for all purposes). This document describes that the compound aminopyrazole KM - 819 with the chemical name 4 - [2 - (4 - bromophenylsulfanyl)acetylamino] - 1 - phenethyl - 1H - pyrazole - 3 - carboxylic acid is an inhibitor of FAF1. The present document describes that the aminopyrazole compound KM - 819 reverses the effects of FAF1 overexpression by reducing alpha - synuclein in cells and animals. Taken together, the experiments in the examples below show that FAF1 siRNA or antisense or related gene constructs, peptides and antibodies and small molecules that modulate FAF1 can be used to modulate alpha - synuclein amounts or phosphorylation state and thus to treatment of alpha - synuclein-induced synucleinopathies.

[0035] One aspect of the present invention relates to a method of treating a disease or condition resulting from synucleinopathies, comprising administering to a subject having symptoms of or at risk of said synucleinopathies, an inhibitor of FAF1 activity in relation to increasing the amount of alpha - synuclein or regulating the amount of LC - 3 and p62 in subject's cells. An effective amount administered is an amount that reduces the amount of alpha - synuclein or increases the amount of LC - 3 and/or reduces the amount of p62 in the subject's cells, respectively.

[0036] In some embodiments of this method, the FAF1 activity inhibitor may be an aminopyrazole compound of formula 1 or a pharmaceutically acceptable salt thereof:

,

,

where R1 is - CO2R3, - CH2OR3, - CONR3R4 or

,

,

where R3 and R4 are independently H or straight, branched or cyclic C1 - C6 alkyl; R2 is - (CH2)2 - phenyl; B is H, phenyl or C1 - C3 alkyl or halogenated phenyl; n is an integer from 0 to 2; Y is S, O, CH2, SO, SO2 or NR3R4, where R3 and R4 are independently H or straight, branched or cyclic C1 - C6 alkyl; Z is H, halogen, OCH3, NO2, NH2 or straight, branched or cyclic C1 - C3 alkyl; and A is CH or N.

[0037] In some instances, the aminopyrazole compound may be 4- [ 2- (4 - bromophenylsulfanyl)acetylamino] -1 - phenethyl - 1H - pyrazole - 3 - carboxylic acid, or an ester or amide thereof.

[0038] Another aspect of the present invention relates to a method of treating a disease or condition resulting from synucleinopathies, comprising administering to a subject having symptoms of or at risk of said synucleinopathies, an amount of a compound that binds to the FAF1 protein, which is effective to reduce the amount of α - synuclein, which is phosphorylated at the amino acid S129 in the cells of said subject.

[0039] In some embodiments of this aspect, the FAF1 activity inhibitor may be an aminopyrazole compound of Formula 1, or a pharmaceutically acceptable salt thereof:

,

,

where R1 is - CO2R3, - CH2OR3, - CONR3R4 or

,

,

where R3 and R4 are independently H or straight, branched or cyclic C1 - C6 alkyl; R2 is - (CH2)2 - phenyl; B is H, phenyl or C1 - C3 alkyl or halogenated phenyl; n is an integer from 0 to 2; Y is S, O, CH2, SO, SO2 or NR3R4, where R3 and R4 are independently H or straight, branched or cyclic C1 - C6 alkyl; Z is H, halogen, OCH3, NO2, NH2 or straight, branched or cyclic C1 - C3 alkyl; and A is CH or N.

[0040] In some instances, the aminopyrazole compound may be 4- [ 2- (4 - bromophenylsulfanyl)acetylamino] -1 - phenethyl - 1H - pyrazole - 3 - carboxylic acid, or an ester or amide thereof.

[0041] In any aspect of the treatment of the present invention, the FAF1 inhibitor may be administered orally or by injection into the brain or cerebrospinal fluid or blood, or through the skin, or by other conventional practices such as implantation of a mini - pump. The most convenient method is one in which the FAF1 binding compound is administered orally.

[0042] The aminopyrazole derivatives of the present invention may be not only in the form of their pharmaceutically acceptable salts, but also in the form of their solvates, hydrates, and enantiomers which may be derived from them.

[0043] The pharmaceutically acceptable salts of the aminopyrazole derivatives of the present invention include acid addition salts derived from pharmaceutically acceptable free acids. Free acids can be inorganic or organic. Examples of organic acids include citric acid, maleic acid, fumaric acid, gluconic acid, methanesulfonic acid, acetic acid, glycolic acid, succinic acid, tartaric acid, 4 - toluenesulfonic acid, galacturonic acid, embonic acid, glutamic acid, and aspartic acid. In addition, hydrochloric acid, hydrobromic acid, sulfuric acid, sulfurous acid or phosphoric acid, preferably methanesulfonic acid and hydrochloric acid, can be used as inorganic acids.

[0044] Acid addition salts in accordance with the present invention can be obtained using a conventional method, for example, by dissolving the aminopyrazole derivatives of formula 1 in a water-miscible organic solvent such as acetone, methanol, ethanol and acetonitrile, adding an excess of an organic acid or an aqueous solution of an inorganic acid for precipitating or crystallizing salts, evaporating the solvent or excess acids from the resulting mixture and drying or filtering with suction the precipitated salts. The aminopyrazole derivatives of the present invention, their pharmaceutically acceptable salts, and a pharmaceutical composition containing them can be clinically administered in oral or non-oral forms. They are usually prepared in combination with a diluent or adjuvant such as a filler, thickener, binder, wetting agent, disintegrant or surfactant , etc.

[0045] Solid agents intended for oral administration can be prepared by mixing at least one aminopyrazole derivative of the present invention with at least one excipient such as starch, calcium carbonate, sucrose, lactose, or gelatin. In addition, a lubricant such as magnesium stearate, talc and the like can also be added.

[0046] Liquid agents intended for oral administration include suspensions, oral solutions, emulsions, syrups, and the like. In addition to a simple diluent such as water or liquid paraffin, various adjuvants such as wetting agents, sweeteners, flavoring agents, preservatives and the like can be used in liquid agents for oral administration of the compound of the present invention.

[0047] In addition, the compound of the present invention can be administered by a non-oral route. For this, sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilic agents, suppositories, and the like can be used. For non-aqueous solvents and suspensions, an injectable vegetable oil such as propylene glycol, polyethylene glycol or olive oil and an ester such as ethylolate may be suitable. Suppository bases include witsol, macrogol, tween 61, cocoa paper, lauric paper, glycerin, and gelatin.

[0048] Depending on the condition of the patients, including age, body weight, gender, route of administration, health status, and severity of the disease, the doses for administering to humans the aminopyrazole derivatives of the present invention, their pharmaceutically acceptable salts, and the pharmaceutical composition containing them, may vary. Typically, the compound of the present invention is administered at a dose of 0.1 to 1000 mg per day for a 70 kg adult human, and preferably at a dose of 1 to 500 mg per day. The compound may be administered in a single dose or in divided doses per day.

[0049] The 4- [ 2- (4 - bromo-phenylsulfanyl)-acetylamino] -1 - phenethyl - 1H - pyrazole - 3 - carboxylic acid aminopyrazole compound has an EC50 of 1 nM to modulate LC - 3 and p62 levels in cells and an _{EC50 of} 20 μM to reduce the amount of alpha - synuclein in cells. Accordingly, an effective amount of this compound to modulate the amount of LC - 3 and p62 in brain tissue cells is an amount that provides a concentration of at least 0.5 nM, preferably at least 1 nM, at least 2 nM, at least 5 nM or at least 10 nM in brain tissue cells, and an effective amount of this compound to reduce the amount of alpha - synuclein in brain tissue cells is an amount that provides a concentration of at least 10 nM, preferably at least 20 mM, at least 40 nM, at least 100 nM, or at least 200 nM in brain tissue cells.

[0050] Another aspect of the invention relates to a method for diagnosing synucleinopathy or assessing the risk of synucleinopathy in a subject, comprising measuring the amount of FAF1 protein or the level of FAF1 activity in relation to inhibition of autophagy mediated by LC - 3 and p62 proteins in cells of the subject or in a sample taken from a subject where the level of activity or the level of FAF1 protein is higher than in normal people, indicating the development of synucleinopathy or the subject is at risk of developing synucleinopathy.

[0051] Another aspect of the invention combines diagnosis and treatment to provide a method of treating a disease or condition resulting from synucleinopathies in a subject, comprising administering to a subject in which the cells of said subject have an amount of FAF1 protein or a level of FAF1 activity that inhibit autophagy mediated by LC proteins - 3 and p62 in a subject's cells is higher than in normal humans, that amount of an inhibitor of FAF1 activity in regulating the amounts of LC - 3 and p62 that is effective to increase the amount of LC3 and/or decrease the amount of p62 in the subject's cells, or that amount of a compound that binds to the FAF1 protein, which is effective to reduce the amount of α - synuclein that is phosphorylated at amino acid S129 in the subject's cells.

[00052] In some embodiments of this aspect of the invention, the compound that binds to FAF1 or inhibits the activity of FAF1 in regulating the amounts of LC - 3 and p62 may be an aminopyrazole compound of formula 1 or a pharmaceutically acceptable salt thereof:

,

,

where R1 is - CO2R3, - CH2OR3, - CONR3R4 or

,

,

where R3 and R4 are independently H or straight, branched or cyclic C1 - C6 alkyl; R2 is - (CH2)2 - phenyl; B is H, phenyl or C1 - C3 alkyl or halogenated phenyl; n is an integer from 0 to 2; Y is S, O, CH2, SO, SO2 or NR3R4, where R3 and R4 are independently H or straight, branched or cyclic C1 - C6 alkyl; Z is H, halogen, OCH3, NO2, NH2 or straight, branched or cyclic C1 - C3 alkyl; and A is CH or N.

[0053] Cerebrospinal fluid (CSF) can be taken from a subject to determine levels of FAF1 and alpha - synuclein. Time sampling with an indwelling spinal catheter can be used to collect CSF samples for FAF1 or alpha - synuclein analysis, for example, as described by TH Langenickel et al., Br. J. Clin Pharmacol., vol. 81, pp. 878 - 890 (2016) (herein incorporated by reference in its entirety).

[0054] EXAMPLES

1. Increasing alpha synuclein by FAF1 in cells

FAF1 is overexpressed in SH - SY5Y cells by transfection with the FAF1 gene tagged with Flag. Overexpression is confirmed by Western blotting of cell extracts using anti - Flag antibodies and anti - FAF1 antibodies. Rabbit anti-FAF1 antibody is available from Proteintech, Chicago, IL, USA, cat. No. 10271 - 1 - AR. The cells were co - transfected with the alpha - synuclein genome. Alpha - synuclein levels are monitored by Western blotting using an anti - alpha - synuclein antibody. Clearly and unexpectedly, alpha - synuclein levels are elevated when FAF1 is overexpressed (Figure 1). Attention is drawn to the fact that the level of beta - actin is used as an internal control. Attention is drawn to the fact that there is a certain level of endogenous FAF1 protein in cells.

[0055] 2. Increasing the aggregated form of alpha synuclein in cells by FAF1

FAF1 is overexpressed in SH - SY5Y cells by transfection with the FAF1 gene tagged with Flag. The cells were co - transfected with the alpha - synuclein genome. The level of the aggregated form of alpha - synuclein is monitored by Western blotting using an anti - alpha - synuclein antibody with long film exposure. Clearly and unexpectedly, the level of the aggregated form of alpha - synuclein is increased when FAF1 is overexpressed (Figure 2).

[0056] 3. Production of the Lewy body-like form of alpha - synuclein in cells by FAF1

FAF1 is overexpressed in SH - SY5Y cells by transfection with the FAF1 gene tagged with Flag. The cells were co - transfected with the alpha - synuclein genome. The LB - like form of alpha - synuclein is monitored by immunofluorescent staining using an anti - alpha - synuclein antibody. Clearly and unexpectedly, LB - like form of alpha - synuclein is observed when FAF1 is overexpressed (FIG. 3B, top, arrow - marker). However, the LB - like form is not observed in the vector control (FIG. 3A, top), demonstrating that FAF1 induces LB - like formation.

[0057] Alpha - synuclein is known to be phosphorylated at the serine residue at position 129, which is a likely pathological process leading to the production of Lewy bodies. The phosphorylated form of alpha synuclein at position 129 is monitored by immunofluorescent staining using an anti - Ps129 alpha synuclein antibody. It is clear that a phosphorylated form of S129 alpha - synuclein is observed when FAF1 is overexpressed in an LM - like form (Figure 3B below, arrow mark). However, this phosphorylation was not observed with the vector control (Figure 3A bottom), demonstrating that FAF1 induces alpha - synuclein phosphorylation at the serine residue at position 129.

[0058] 4. Increasing alpha synuclein in mice with FAF1

FAF1 is overexpressed in mice by injection of the stereotype in the usual way with a recombinant adenovirus containing the FAF1 gene into the midbrain region. Overexpression is confirmed by Western blotting of midbrain tissue extracts using an anti - FAF1 antibody. The midbrain is a tissue that contains substantial dopaminergic neurons known to influence movement in synucleinopathies. Two mice were injected with FAF1 adenovirus and two uninjected mice were used as controls. Alpha - synuclein levels are monitored by Western blotting using an anti - alpha - synuclein antibody. It can be seen that the increased level of alpha - synuclein correlates with overexpression of FAF1 (figure 4).

[0059] 5. Decreased alpha synuclein siRNA FAF1 in cells

FAF1 expression is attenuated by treatment of cells with SH - SY5Y siRNA (sequence: GUGUUGUGGCACAAACCAU) and FAF reduction is confirmed by Western blotting of cell extracts using an anti - FAF1 antibody. The cells were co - transfected with the alpha - synuclein genome. Alpha - synuclein levels are monitored by Western blotting using an anti - alpha - synuclein antibody. It can be seen that the level of alpha - synuclein is reduced by processing siRNA FAF1 (figure 5). On the other hand, treatment with control siRNA (scRNA, degenerate sequence) does not reduce alpha - synuclein levels.

[0060] 6. FAF1 is an autophagy suppressor

Alpha - synuclein is known to be cleaved in cells by various pathways, including the proteasome, autophagy, and chaperone-mediated autophagy (Sampaio - Marques B and Ludovico P 2015). FAF1 is overexpressed by transfection in SH - SY5Y cells, and various proteins involved in protein degradation pathways are analyzed by Western blotting (Figure 6).

[0061] As shown in Figure 6, when FAF1 is overexpressed, the p62 protein increases and LC3 decreases, indicating that FAF1 suppresses autophagy. It should be noted that all other proteins are unchanged. This change mediates autophagy as described (Sampaio - Marques B and Ludovico P, 2015). These data demonstrate that FAF1 increases alpha synuclein by suppressing the autophagy pathway.

[0062] 7. KM - 819 (KR - 33493) is an inhibitor of FAF1

KM - 819 (KR - 33493, 4 - [2 - (4 - bromophenylsulfanyl) acetylamino] - 1 - phenethyl - 1H - pyrazole - 3 - carboxylic acid) is an aminopyrazole derivative described in the prior art for ischemic cell death inhibitor ( US patent No. 7939550, Yoo SE et al. 2016). Inhibition of FAF1 by KM - 819 is confirmed by binding of KM - 819 to FAF1 as described and shown in Figure 7). In addition, the inhibition of FAF1 KM - 819 is confirmed by the inhibition of downstream targets of FAF1 in cells, including the apoptosis pathway proteins caspase 8 and JAK (figure 8). The compound showed 70 % inhibition of apoptosis at 10 μM in ischemic cell death assays caused by deoxyglucose-induced hypoxia-mediated apoptosis (US Pat. No. 7,939,550).

[0063] The binding of KM - 819 to FAF1 is confirmed by dropout analysis in SH - SY6H cell extracts using biotin-labeled KM - 819 as described (Yoo SE et al. 2016). Cells are transfected with flag labeled FAF1. The resulting FAF1 - KM - 819 complex is precipitated using avidin-coated agarose. Precipitated FAF1 in the complex is measured by Western blotting using an anti - Flag antibody. It appears that FAF1 binds to KM - 819 when cells are stressed with hydrogen peroxide and thus FAF1 is activated. It should be noted that the total level of FAF1 in whole cell extracts (WCL) is measured as a control.

[0064] FAF1 activates apoptotic stress with various stimuli through activation of the caspase cascade and necrosis as described (Yu C et al. 2016). Direct targets for FAF1 in these two pathways are caspase 8 and JNK, respectively. And the stage of this activation is the formation of these complexes with subsequent activation of downstream targets.

[0065] Inhibition of complexation of FAF1 - caspase 8 and FAF1 - JNK by KM - 819 is confirmed by conventional immunoprecipitation assays using caspase 8 DED antibody and JNK antibody, and bound and precipitated FAF1 is measured using Western blot using anti - FAF1 antibody . It is clear that KM - 819 effectively inhibits the formation of the complex, the inhibition is about 80% at 0.1 μm (figure 8).

[0066] Inhibition of caspase 8 and JNK activation by KM - 819 is confirmed by analysis of caspase 8 substrate cleavage, caspase 3 and JNK autophosphorylation, respectively (Figure 9). Cells are treated with various concentrations of KM - 819 as indicated, and levels of cleaved caspase 3 and autophosphorylated form of JNK are measured by Western blotting using an anti - caspase 3 antibody and an anti - phospho - JNK antibody, respectively. It can be seen that KM - 819 inhibits the activation activity of caspase 8 and JNK (figure 9). IC ₅₀ activation of caspase 8 KM - 819 is 0.73 μm, and IC ₅₀ activation of JNK KM - 819 is 0.25 μm.

[0067] 8. KM - 819 enhances autophagy

Alpha - synuclein is known to be cleaved in a variety of ways in cells, including through proteasomes, autophagy, and chaperone-mediated autophagy (Sampaio - Marques B and Ludovico P, 2015). FAF1 suppresses autophagy.

[0068] To confirm that the FAF1 inhibitor inhibits the activity of FAF1 - mediated autophagy suppression, SH - SY5Y cells overexpressing FAF1 are treated with KM - 819, as indicated, and the level of LC3 and p62 is measured by Western blot (figure 10). KM - 819 reduces p62 protein and increases LC3, demonstrating that KM - 819 promotes the autophagy pathway. EC ₅₀ to stimulate autophagy KM - 819 is 1 nm.

[0069] 9. Decreasing alpha - synuclein in cells with a FAF1 inhibitor

To confirm that the FAF1 inhibitor reduces the level of alpha - synuclein in cells, SH - SY5Y cells are treated with the FAF1 inhibitor KM - 819 (KR - 88493, US patent 7939550, Yoo SE et al. 2016). Cells are co-transfected with genes encoding FAF1 tagged with Flag and alpha - synuclein tagged with GFP. Alpha synuclein levels are measured by Western blotting of cell extracts using an anti - GFP antibody. FAF1 levels are measured by Western blotting of cell extracts using an anti - Flag antibody. It can be seen that the level of alpha - synuclein is reduced when treated with an inhibitor of FAF1 (figure 11). The IC ₅₀ is 20 nM and the inhibition is over 95% at 10 μM. This is a much higher efficiency compared to the apoptosis inhibition activity measured in the prior art, 70 % inhibition at 10 μM. It should be noted that the level of FAF1 itself does not change when treated with a FAF1 inhibitor.

[0070] 10. Decreased midbrain alpha synuclein in mice with a FAF1 inhibitor

To see the effect of a FAF1 inhibitor on alpha synuclein levels in animals, transgenic mice overexpressing alpha synuclein are treated with the FAF1 inhibitor KM - 819. , bearing human synuclein (Lee MK et al. 2002).

[0071] Three-month-old mice are orally administered KM - 819 at 1, 5, and 10 mg/kg/day for 9 days. Two non-transgenic mice (non - Tg) are used as controls and three transgenic mice (alpha - syn A53T Tg) are treated with vehicle alone without KM - 819. And two mice are used for each dosing of KM - 819. Animals are sacrificed and midbrain tissues isolate and make extracts. Alpha synuclein, S129 the phosphorylated form of alpha synuclein, and FAF1 are measured by Western blotting using an anti - alpha synuclein antibody, P - alpha syn antibody, and FAF1 antibody, respectively. It can be seen that both the levels of alpha - synuclein and its phosphorylated form are reduced by treatment with the FAF1 inhibitor KM - 819, demonstrating that the FAF1 inhibitor reduces the accumulation of alpha - synuclein in animals (Figure 12). It should be noted that the level of FAF1 itself does not change when treated with a FAF1 inhibitor. It should be noted that the level of beta - actin is used as an internal control.

[0072] 11. Reduction of alpha - synuclein in the hippocampus in mice with a FAF1 inhibitor

To see the effect of a FAF1 inhibitor on alpha synuclein levels in animals, transgenic mice overexpressing alpha synuclein are treated with the FAF1 inhibitor KM - 819. , bearing human synuclein (Lee MK et al. 2002).

[0073] Three-month-old mice are orally administered KM - 819 at 1, 5, and 10 mg/kg/day for 9 days. Three non-transgenic (non-Tg) mice are used as controls, and two transgenic mice (alpha - syn A53T Tg) are injected with vehicle alone without KM - 819. And two mice are used for each dosing of KM - 819. Animals are sacrificed and hippocampal tissues are isolated and make extracts. Alpha synuclein and FAF1 are measured by Western blotting using an anti - alpha synuclein antibody and FAF1 antibody, respectively. It can be seen that the level of alpha - synuclein is reduced by treatment with the FAF1 inhibitor KM - 819, demonstrating that the FAF1 inhibitor reduces the accumulation of alpha - synuclein in animals (figure 13).

[0074] 12. Reduction of alpha - synuclein in the cerebral cortex in mice with a FAF1 inhibitor

To see the effect of a FAF1 inhibitor on alpha synuclein levels in animals, transgenic mice overexpressing alpha synuclein are treated with the FAF1 inhibitor KM - 819. , carrying human synuclein (Lee MK

[0075] Three-month-old mice are administered KM - 819 orally at a dose of 1, 5 and 10 mg/kg/day for 9 days. Two non-transgenic mice (non - Tg) are used as controls, and two transgenic mice (alpha - syn A53T Tg) are injected with vehicle alone without KM - 819. And two mice are used for each dosing of KM - 819. Animals are sacrificed and cortical tissues the brain is isolated and extracts are made. Alpha synuclein is measured by Western blotting using an anti - alpha synuclein antibody. It can be seen that the level of alpha - synuclein is reduced by treatment with the FAF1 inhibitor KM - 819, demonstrating that the FAF1 inhibitor reduces the accumulation of alpha - synuclein in animals (figure 14).

[0076] 13. Reduction of alpha - synuclein in the cerebellum in mice with a FAF1 inhibitor

To see the effect of a FAF1 inhibitor on alpha synuclein levels in animals, transgenic mice overexpressing alpha synuclein are treated with the FAF1 inhibitor KM - 819. , bearing human synuclein (Lee MK et al. 2002).

[0077] Three-month-old mice are administered KM - 819 orally at 1, 5, and 10 mg/kg/day for 9 days. Three non-transgenic mice (non - Tg) are used as controls and two transgenic mice (alpha - syn A53T Tg) are injected with vehicle alone without KM - 819. And two mice are used for each dosing of KM - 819. Animals are sacrificed and cerebellar tissues are isolated and make extracts. Alpha synuclein and FAF1 are measured by Western blotting using an anti - alpha synuclein antibody and FAF1 antibody, respectively. It can be seen that both alpha - synuclein levels are reduced by treatment with the FAF1 inhibitor KM - 819, demonstrating that the FAF1 inhibitor reduces the accumulation of alpha - synuclein in animals (Figure 15).

[0078] 14. Reducing alpha - synuclein in the striatum in mice with a FAF1 inhibitor

To see the effect of a FAF1 inhibitor on alpha synuclein levels in animals, transgenic mice overexpressing alpha synuclein are treated with the FAF1 inhibitor KM - 819. , bearing human synuclein (Lee MK et al. 2002).

[0079] Three-month-old mice are administered KM - 819 orally at a dose of 1, 5 and 10 mg/kg/day for 9 days. Three non-transgenic (non-Tg) mice are used as controls and two transgenic mice (alpha - syn A53T Tg) are injected with vehicle alone without KM - 819. And two mice are used for each dosing of KM - 819. Animals are sacrificed, striatal tissues are isolated and make extracts. Alpha synuclein and FAF1 are measured by Western blotting using an anti - alpha synuclein antibody and FAF1 antibody, respectively. It can be seen that the level of alpha - synuclein is reduced by treatment with the FAF1 inhibitor KM - 819, demonstrating that the FAF1 inhibitor reduces the accumulation of alpha - synuclein in animals (figure 16).

EMBODIMENTS FOR CARRYING OUT THE INVENTION

[0080] Embodiment 1: A method of treating a disease or condition resulting from synucleinopathies, comprising administering to a subject having symptoms of or at risk of said synucleinopathies an effective amount of an inhibitor of FAF1 activity that increases alpha - synuclein, or an effective amount of an inhibitor of FAF1 activity against adjusting the amounts of LC - 3 and p62.

[0081] Embodiment 2: The method of Embodiment 1 wherein the FAF1 activity inhibitor is an aminopyrazole compound of Formula 1 or a pharmaceutically acceptable salt thereof:

,

,

where R1 is - CO2R3, - CH2OR3, - CONR3R4 or

,

,

where R3 and R4 are independently H or straight, branched or cyclic C1 - C6 alkyl; R2 is - (CH2)2 - phenyl; B is H, phenyl or C1 - C3 alkyl or halogenated phenyl; n is an integer from 0 to 2; Y is S, O, CH2, SO, SO2 or NR3R4, where R3 and R4 are independently H or straight, branched or cyclic C1 - C6 alkyl; Z is H, halogen, OCH3, NO2, NH2 or straight, branched or cyclic C1 - C3 alkyl; and A is CH or N.

[0082] Embodiment 3: The method of Embodiment 1 wherein the aminopyrazole compound is 4- [ 2- (4 - bromophenylsulfanyl)acetylamino] -1 - phenethyl - 1H - pyrazole - 3 - carboxylic acid or an ester thereof or amide.

[0083] Embodiment 4: A method of treating a disease or condition resulting from synucleinopathies, comprising administering to a subject having symptoms of or at risk of said synucleinopathies an amount of a FAF1 protein-binding compound that is effective to reduce the amount of α - synuclein, which is phosphorylated at the amino acid S129 in the cells of said subject.

[0084] Embodiment 5: The method of Embodiment 4 wherein the compound that binds to the FAF1 protein is an aminopyrazole compound of Formula 1 or a pharmaceutically acceptable salt thereof:

,

,

where R1 is - CO2R3, - CH2OR3, - CONR3R4 or

,

,

where R3 and R4 are independently H or straight, branched or cyclic C1 - C6 alkyl; R2 is - (CH2)2 - phenyl; B is H, phenyl or C1 - C3 alkyl or halogenated phenyl; n is an integer from 0 to 2; Y is S, O, CH2, SO, SO2 or NR3R4, where R3 and R4 are independently H or straight, branched or cyclic C1 - C6 alkyl; Z is H, halogen, OCH3, NO2, NH2 or straight, branched or cyclic C1 - C3 alkyl; and A is CH or N.

[0085] Embodiment 6: The method of Embodiment 4 wherein the aminopyrazole compound is 4- [ 2- (4 - bromophenylsulfanyl)acetylamino] -1 - phenethyl - 1H - pyrazole - 3 - carboxylic acid or an ester thereof or amide.

[0086] Embodiment 7: The method of any one of Embodiments 1 to 3, wherein the FAF1 inhibitor is administered orally, or by injection into the brain or blood, or through the skin.

[0087] Embodiment 8: The method of any one of embodiments 4 to 6, wherein the compound that binds to the FAF1 protein is administered orally, or by injection into the brain or blood, or through the skin, or by other well-known practices, in which the FAF1 binding compound is administered orally.

[0088] Embodiment 9: A method for diagnosing synucleinopathy or assessing the risk of synucleinopathy in a subject, comprising measuring FAF1 activity or FAF1 levels for autophagy inhibition, including LC - 3 and p62 proteins in the subject's cells or in any other sample from the subject, wherein the level of activity or protein level of FAF1 is higher than in normal people, indicating the development of synucleinopathy or the subject is at risk of developing synucleinopathy.

[0089] Embodiment 10: A method of treating a disease or condition arising from synucleinopathies in a subject, comprising administering to a subject that exhibits FAF1 activity or FAF1 levels in the cells of said subject in relation to inhibition of autophagy mediated by LC - 3 and p62 proteins in cells a subject where the level of activity or the level of FAF1 protein is higher than in normal individuals, indicating the development of synucleinopathies or the subject is at risk of developing synucleinopathies, such an amount of a compound that binds to the FAF1 protein that is effective to reduce the amount of α - synuclein that is phosphorylated at the amino acid S129 in the cells of the specified subject.

[0090] Embodiment 11. The method of Embodiment 10 wherein an aminopyrazole compound of formula 1 or a pharmaceutically acceptable salt thereof is administered:

where R1 is - CO2R3, - CH2OR3, - CONR3R4 or

where R3 and R4 are independently H or straight, branched or cyclic C1 - C6 alkyl; R2 is - (CH2)2 - phenyl; B is H, phenyl or C1 - C3 alkyl or halogenated phenyl; n is an integer from 0 to 2; Y is S, O, CH2, SO, SO2 or NR3R4, where R3 and R4 are independently H or straight, branched or cyclic C1 - C6 alkyl; Z is H, halogen, OCH3, NO2, NH2 or straight, branched or cyclic C1 - C3 alkyl; and A is CH or N.

[0091] Embodiment 12: A method for preparing a medicament comprising mixing an aminopyrazole compound of Formula 1 or a pharmaceutically acceptable salt thereof:

,

,

where R1 is - CO2R3, - CH2OR3, - CONR3R4 or

,

,

where R3 and R4 are independently H or straight, branched or cyclic C1 - C6 alkyl; R2 is - (CH2)2 - phenyl; B is H, phenyl or C1 - C3 alkyl or halogenated phenyl; n is an integer from 0 to 2; Y is S, O, CH2, SO, SO2 or NR3R4, where R3 and R4 are independently H or straight, branched or cyclic C1 - C6 alkyl; Z is H, halogen, OCH3, NO2, NH2 or straight, branched or cyclic C1 - C3 alkyl; and A is CH or N; and a pharmaceutically acceptable carrier.

[0092] Embodiment 13: The method of Embodiment 12, wherein the medicament is for treating a disease or condition resulting from synucleinopathies in a subject.

[0093] Embodiment 14: Use of an aminopyrazole compound of Formula 1 or a pharmaceutically acceptable salt thereof:

,

,

where R1 is - CO2R3, - CH2OR3, - CONR3R4 or

,

,

where R3 and R4 are independently H or straight, branched or cyclic C1 - C6 alkyl; R2 is - (CH2)2 - phenyl; B is H, phenyl or C1 - C3 alkyl or halogenated phenyl; n is an integer from 0 to 2; Y is S, O, CH2, SO, SO2 or NR3R4, where R3 and R4 are independently H or straight, branched or cyclic C1 - C6 alkyl; Z is H, halogen, OCH3, NO2, NH2 or straight, branched or cyclic C1 - C3 alkyl; for treating a disease or condition resulting from synucleinopathies in a subject.

[0094] Embodiment 15: The use of Embodiment 14, wherein the aminopyrazole compound is effective to inhibit FAF1 activity to increase the amount of alpha - synuclein in a subject's cell or in a sample cell from a subject, or is effective to increase autophagy mediated by proteins LC - 3 and p62, in the cell of the subject or in the cell of the sample from the subject, or is effective to reduce the amount of α - synuclein, which is phosphorylated at amino acid S129 in the cell of the subject or in the cell of the sample from the subject.

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SEQUENCE LIST

<110> Kainos Medicine

<120> Alpha-synuclein modulators

<130> 400514-20005

<140> PCT/IB2018/000766

<141> 2018-06-19

<160> 1

<170>PatentIn version 3.5

<210> 1

<211> 19

<212> RNA

<213> Artificial sequence

<220>

<223> alpha-synuclein antisense RNA

<400> 1

guguuguggc acaaaccau 19

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Claims (20)

1. Use of an aminopyrazole compound 4-[2-(4-bromophenylsulfanyl)acetylamino]-1-phenethyl-1H-pyrazole-3-carboxylic acid or a pharmaceutically acceptable salt thereof or a composition of said aminopyrazole compound or a pharmaceutically acceptable salt thereof; to obtain a medicinal product for the treatment of alpha synucleinopathy: caused by Lewy bodies, Lewy neuritis, or α-synuclein aggregates in neurons and glia; or caused by a mutation in the alpha-synuclein (SNCA) gene; or mediated activity of FAF1 to regulate the amount of LC-3 and p62, activity of FAF1 to regulate the amount of α-synuclein, or mediated by the amount of aggregated α-synuclein, or mediated by the amount of α-synuclein, which is phosphorylated at the amino acid S129 in the cells of the specified subject; or caused by the accumulation of misfolded alpha-synuclein or the accumulation of alpha-synuclein aggregates in the cells of said subject. 2. Use according to claim 1, wherein the aminopyrazole compound, or a pharmaceutically acceptable salt or composition thereof, is formulated for oral administration, or for injection into the brain, or for injection into the blood. 3. Use according to claim 2, wherein the formulation provides 1 to 10 mg/kg/day to a subject upon administration. 4. Application according to any one of paragraphs. 1-3, where alpha synucleinopathy is Parkinson's disease with dementia (PDD), Lewy body dementia, multiple system atrophy, atypical Parkinson's disease, and pure autonomic failure. 5. Use of an aminopyrazole compound 4-[2-(4-bromophenylsulfanyl)acetylamino]-1-phenethyl-1H-pyrazole-3-carboxylic acid or a pharmaceutically acceptable salt thereof or a composition of said aminopyrazole compound or a pharmaceutically acceptable salt thereof; for the treatment of alpha synucleinopathy: caused by Lewy bodies, Lewy neuritis, or α-synuclein aggregates in neurons and glia; or caused by a mutation in the alpha synuclein (SNCA) gene, or mediated activity of FAF1 to regulate the amount of LC-3 and p62, activity of FAF1 to regulate the amount of α-synuclein, or mediated by the amount of aggregated α-synuclein, or mediated by the amount of α-synuclein, which is phosphorylated at the amino acid S129 in the cells of the specified subject; or caused by the accumulation of misfolded alpha-synuclein or the accumulation of alpha-synuclein aggregates in the cells of said subject. 6. Use according to claim 5, wherein the aminopyrazole compound, or a pharmaceutically acceptable salt or composition thereof, is formulated for oral administration, or for injection into the brain, or for injection into the blood. 7. The use of claim 6 wherein the formulation provides 1 to 10 mg/kg/day to a subject upon administration. 8. Application according to any one of paragraphs. 5-7, where alpha synucleinopathy is Parkinson's disease with dementia (PDD), Lewy body dementia, multiple system atrophy, atypical Parkinson's disease, and pure autonomic failure.

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