Compositions for treating and / or preventing protein aggregation disorders

Abstract

The present invention provides compositions used for the treatment and / or prevention of protein aggregation disorders. [Solution] Proteopathy encompasses a wide range of ailments, including neurodegenerative diseases (e.g., polyglutamine diseases such as huntingtin in Alzheimer's disease, Parkinson's disease, and Huntington's disease, and prion diseases); amyloidosis of other non-neuronal proteins (especially I1-antitrypsin, immunoglobulin light and heavy chains, lactadherin, apolipoprotein, gelzolin, lysozyme, fibrinogen, atrial natriuretic factor, keratin, lactoferrin, and β-2 microglobulin, etc.); sickle cell disease; cataracts; cystic fibrosis; retinitis pigmentosa; and nephrogenic diabetes insipidus. Administration of sulfatase inhibitors is generally suitable for treating and / or preventing protein toxicity associated with proteopathy. Therefore, the present invention provides compositions comprising sulfatase inhibitors for the treatment of proteopathy.

Description

Technical Field

[0001] The present invention is included in the field of medicine and provides a composition for the treatment and / or prevention of protein aggregation diseases.

Background Art

[0002] The proper functioning of organs and cells in an organism depends on the proper functioning of proteins. Proteins are biological entities having a primary amino acid sequence, a secondary structure that forms protein domains and, most importantly, includes α-helices and β-sheets, and a tertiary structure that results from the complex folding of a three-dimensional polypeptide chain involving interactions between the polypeptide chain backbone and amino acid side chains. Some proteins function as multi-subunit complexes, and arranging multiple proteins in a quaternary structure is important for their proper functioning.

[0003] When a protein fails to fold into its correct three-dimensional structure, it leads to diseases called proteopathies ( also sometimes referred to as protein aggregation diseases, protein misfolding diseases, proteinopathies or protein conformational disorders). This can occur. This failure may be due to one or more mutations in the protein's gene, or environmental factors such as oxidative stress, alkalosis, acidosis, pH shift, and osmotic shock. Protein misfolding can lead to clamping or aggregation into amyloid plaques or fibrils, which can worsen the disease. Proteopathies encompass a wide range of ailments, including neurodegenerative diseases (e.g., polyglutamine diseases such as huntingtin in Alzheimer's disease, Parkinson's disease, and Huntington's disease, and prion diseases); amyloidosis of other non-neuronal proteins (especially I1-antitrypsin, immunoglobulin light and heavy chains, lactadherin, apolipoproteins, gelzolin, lysozyme, fibrinogen, atrial natriuretic factor, keratin, lactoferrin, and β-2 microglobulin); sickle cell disease; cataracts; cystic fibrosis; retinitis pigmentosa; and nephrogenic diabetes insipidus.

[0004] Amyloidosis refers to the pathological deposition of proteins in the form of green, birefringent fibrils with affinity for Congo Red, which, when stained with Congo Red, are dispersed or localized amyloidomas. Such depositions are associated with several diseases, such as Alzheimer's disease, inflammation-associated amyloid, type II diabetes, and bovine amyloidosis. These are symptoms of scotomatous encephalopathy (BSE), Creutzfeldt-Jakob disease (CJD), scrapie, and primary amyloidosis.

[0005] Amyloidosis is generally classified into three groups: major systemic amyloidosis, major focal amyloidosis, and miscellaneous amyloidosis. Major systemic amyloidosis includes chronic inflammatory conditions (e.g., tuberculosis, osteomyelitis); non-infectious conditions such as juvenile rheumatoid arthritis, ankylosing spondylitis, and Crohn's disease; familial Mediterranean fever, plasma cell proliferation (primary amyloidosis), and various familial polyneuropathy and cardiomyopathy. Major focal amyloidosis includes dialysis-associated amyloidosis, Alzheimer's disease, Down syndrome, hereditary cerebral hemorrhage (Dutch), and non-traumatic cerebral hemorrhage in the elderly. Various types of amyloidosis include familial polyneuropathy (Iowa), familial amyloidosis (Finland), hereditary cerebral hemorrhage (Iceland), CJD, medullary thyroid carcinoma, atrial amyloid, and diabetes mellitus (insulinoma). Other types of amyloidosis are referenced in Non-Patent Literature 1.

[0006] Transmissible spongiform encephalopathy, which causes CJD and Gerstmann-Sträussler-Scheinker (GSS) disease, is described in Non-Patent Document 2 and Non-Patent Document 3. Many of these diseases are likely to be transmitted by prions, which are infectious proteins. See Non-Patent Document 4 and the references contained therein.

[0007] Attempting to eliminate specific fibrils has been a key objective in research on amyloidosis, but without success. Current treatments for amyloidosis include chemotherapy agents or steroids such as melphalan and dexamethasone. However, such treatments are not suitable for all patients and are often ineffective due to a lack of specificity (Patent Document 1). Similarly, the success of treating other proteopathies not necessarily related to amyloid formation has been limited. Therefore, there is a great need for alternative means that can safely and effectively treat or prevent proteopathies.

[0008] The inventors of this invention were initially interested in elucidating novel factors governing the genetic control of aging in order to improve the understanding of this complex biological process. For this purpose, the inventors isolated a heat-stable mutant and identified an allele of the sul-2 gene (a gene encoding one of the three members of the Caenorhabditis elegans sulfatase family) containing an inactivation site mutation. The inventors found that worms possessing the inactivated version of the sul-2 gene were more resistant than wild-type worms. I discovered that I could live a long life.

[0009] Sulfatases are a large family of proteins involved in various biological processes and affect a wide range of substrates. The collocation of sul-2 in the sulfatase phylogenetic tree is uncertain. However, compared to mammalian sulfatases, the sul-2 cluster is likely closer to the H, F, E, and D type arylsulfatases and C type steroid sulfatases, which probably originate from a common ancestral gene. We hypothesized that sul-2 may exert its activity by modifying sulfated steroid hormones.

[0010] Steroid hormone sulfatase is a conserved protein involved in various processes, including stimulating the growth of hormone-dependent cancers (Non-Patent Literature 5). Specific inhibitors of this type of enzyme have been developed, one of which is STX64 (Non-Patent Literature 6). STX64 is used to treat patients with hormone-dependent cancers (Non-Patent Literature 7). The inventors treated wild-type worms with STX64 and observed a similar longevity effect as in sul-2 mutants. While STX64 did not further increase the longevity of sul-2 deletion mutants, the inventors demonstrated that the mechanism by which STX64 increases longevity is by inhibiting the sulfatase activity of SUL-2 (Patent Literature 2). [Prior art documents] [Patent Documents]

[0011] [Patent Document 1] International Publication No. 2017 / 075540 [Patent Document 2] International Publication No. 2014 / 154927 [Non-patent literature]

[0012] [Non-Patent Document 1] Louis W. Heck, "The Amyloid Diseases" in Cecil's Textbook of Medicine 1504-6 (WB Saunders & Co., Philadelphia, Pa.; 1996) [Non-Patent Document 2] B. Chesebro et al., "Transmissible Spongiform Encephalopathies: A Brief Introduction" in FIELD'S VIROLOGY 2845-49 (3rd Edition; Raven Publishers, Philadelphia, Pa.; 1996) [Non-Patent Document 3] DC Gajdusek in FIELDS VIROLOGY (1996), "Infectious amyloids: Subacute Spongiform Encephalopathies as Transmissible Cerebral Amyloidoses," 2851-2900 [Non-Patent Document 4] SB Prusineri, "Prions" in FIELDS VIROLOGY 2901-50 (1996) [Non-Patent Document 5] Mueller et al., 2015. Endocr Rev. 36(5):526-63 [Non-Patent Document 6] Nussbaumer & Billich, 2004. Med Res Rev. 24(4):529-76 [Non-Patent Document 7] Stanway et al., 2006. Clin Cancer Res. 12(5):1585-92 [Overview of the project]

[0013] Surprisingly, the inventors have found that STX64 can treat or prevent oligomer and / or amyloid formation in C. elegans Parkinson's disease models, C. elegans Huntington's disease models, and C. elegans Alzheimer's disease models (see Figures 1-6). In Parkinson's disease, Huntington's disease, and Alzheimer's disease, pathogenesis is driven by the production and / or deposition of protein aggregates (Murphy & Levine, 2010. J Alzheimers Dis. 19(1):311, Stefanis, 2012. Cold Spring Harb Perspect Med. 2(2): a009399, Daldin et al., 2017. Sci Rep. 7(1):5070). Therefore, the treatment or prevention of aggregate species in these models lends credibility to the idea that STX64 can be used to treat and / or prevent these protein aggregation disorders. This was confirmed in a mouse model of Alzheimer's disease in which STX64 was found to reduce β-amyloid plaque formation and reverse cognitive impairment induced by intrahippocampal administration of β-amyloid oligomers and in transgenic mouse models of Alzheimer's disease (see Figure 7).

[0014] Furthermore, C. elegans containing an inactivating mutation of the sulfatase gene also has a lower tendency to form similarly harmful aggregates, suggesting that this effect is not only related to STX64 but also holds true for any sulfatase inhibitor. The fact that this preventive / therapeutic effect was obtained in models of three different protein aggregation diseases in which the toxic protein was expressed in muscle cells or neuronal cells suggests that sulfatase inhibitors can be used for the treatment and / or prevention of any protein aggregation disease.

[0015] Thus, in a first aspect, the present invention provides a composition comprising a sulfatase inhibitor for use in the treatment and / or prevention of a protein aggregation disease, preferably in a patient and / or an animal.

[0016] In a second aspect, the present invention also provides a kit for use in the manufacture of an agent for treating and / or preventing a protein aggregation disease, the kit comprising (i) a sulfatase inhibitor and (ii) a pharmaceutically acceptable carrier and / or diluent.

Brief Description of the Drawings

[0017] [Figure 1] Reduction of steroid hormone sulfatase (STS) activity improves the symptoms of protein toxicity in the C. elegans Parkinson's disease model strain NL5901 (Van Ham et al., 2010. Cell 142: 601-612). Worms expressing α-synuclein in muscle cells show a decrease in motility (measured as the number of times the head of the worm crosses the body length axis per minute (pitches)) as they age, but (A) the rate of decline is reduced when the worms contain the sul-2(gk187) allele, and (B) the rate of decline is reduced when the worms are treated with STX64. n = 20; *** represents p < 0.001. [Figure 2]Reduction of steroid hormone sulfatase (STS) activity alters the way α-synuclein aggregates in the C. elegans Parkinson's disease model strain NL5901 (Van Ham et al., 2010. Cell 142: 601-612). The graph shows the (A) total number of aggregates, (B) number of aggregates larger than 3 μm in size, and (C) number of aggregates smaller than 1 μm in size in worms expressing α-synuclein fused to YFP, and in worms expressing the same α-synuclein construct that further contains the inactive sul-2 allele, i.e., sul-2(gk187). n = 20; *** represents p < 0.001, and * represents p < 0.05. [Figure 3] Reduction of steroid hormone sulfatase (STS) activity alters the way α-synuclein aggregates in the C. elegans Parkinson's disease model strain NL5901 (Van Ham et al., 2010. Cell 142: 601-612). The graph shows the (A) total number of aggregates, (B) number of aggregates larger than 3 μm in size, and (C) number of aggregates smaller than 1 μm in size in worms expressing α-synuclein fused to YFP treated with DMSO, and in worms expressing the same α-synuclein construct further treated with STX64. n = 20; *** means p < 0.001, and * represents p < 0.05. [Figure 4] Reduction of steroid hormone sulfatase (STS) activity ameliorates the symptoms of protein toxicity in the C. elegans Parkinson's disease model strain UA44 (Cooper et al., 2006. Science 313: 324-328). (A) The survival rate of GFP-labeled dopaminergic neurons expressing human α-synuclein increases when the worms contain the inactive sul-2(gk187) allele. The graph represents the number of neurons surviving in 6-day-old worms when the worms contain the wild-type sul-2 allele or the inactive sul-2(gk187) allele. (B) Representative images of 9-day-old worms containing the wild-type sul-2 allele and worms containing the inactive sul-2(gk187) allele. The black arrows indicate the surviving dopaminergic neurons. n = 37; p < 0.0001. [Figure 5] Reduced steroid hormone sulfatase (STS) activity improves the symptoms of protein toxicity in the C. elegans Huntington's disease model strain AM140 (Morley et al. 2002. PNAS. 99: 10417-10422). The nematodes were modified to express a 35-polyglutamine repeat fused to YFP, and the number of fluorescent aggregates was counted on day 5 of adulthood. (A) Graph shows a comparison between nematodes containing the wild-type allele of sul-2 and nematodes containing the inactive mutant allele of sul-2. (B) Graph shows a comparison between nematodes containing the wild-type allele of sul-2 and nematodes containing the wild-type allele of sul-2 further treated with STX64. n=20; *** represents p<0.001. [Figure 6] Reduced steroid hormone sulfatase (STS) activity improves the symptoms of protein toxicity in the C. elegans Alzheimer's disease model strain CL2006 (Link, 1995. PNAS. 92: 9368-9372). Nematodes expressing β-amyloid in muscle cells develop paralysis as they age, but the rate of paralysis is reduced when (A) the nematodes contain the sul-2(gk187) allele, and when (B) the nematodes are treated with STX64. n=50; p<0.001. [Figure 7]STX64 treatment mitigated β-amyloid deposition and cognitive impairment in an Alzheimer's disease mouse model. (A) Effects of intrahippocampal and oral administration of STX64 in a passive avoidance test of wild-type mice injected with β-amyloid oligomers into the hippocampus. The number of mice in each group was greater than 5. (B) Representative β-amyloid immunoreactivity images were assessed in the frontal cortex and hippocampus of APP-PS1 mice over 15 months of age after 3-4 weeks of ingestion of vehicle or STX64 (0.005 mg / ml in drinking water). (C)-(E) Quantification of β-amyloid area percentage (C), deposition density (D), and mean plaque size (E) in the frontal cortex and hippocampus of APP-PS1 mice over 15 months of age after oral administration of STX64 or vehicle for 3-4 weeks. n=4 mice per group. (F) Time course of β-amyloid deposition in APP-PS1 mice, and the effect of 3-4 weeks of oral STX64 treatment on β-amyloid area in the prefrontal cortex and hippocampus. More than 3 micrographs were used per mouse. (G) Effect of oral administration of STX64 in APP-PS1 mice over 15 months of age, and comparison of APP-PS1 mice over 15 months of age with wild-type mice in passive avoidance tests. More than 5 mice were used in each group. In histological analysis, * indicates a statistically significant difference between APP-PS1 mice administered with vehicle and those administered with STX64. In behavioral tests, * indicates a statistically significant difference between short-term and long-term memory sessions (STM and LTM, respectively) and training sessions in the same experimental group, and + indicates a statistically significant difference between STM and LTM sessions between each experimental group and the β-amyloid group. One symbol represents p < 0.05, two repeating symbols represent p < 0.01, and three repeating symbols represent p < 0.001. [Figure 8] STX64 prevented deterioration of motor activity in a mouse model of Huntington's disease. One-month-old R6 / 1 mice were treated daily with drinking water containing STX64 or the vehicle itself. Motor activity was tested at two months of age. n>5 mice per group. Results are expressed as mean ± SEM. *, p<0.05. [Figure 9]Comparison of STX64 and EMATE in an Alzheimer's disease model, namely the GMC101 strain of C. elegans (McColl et al., 2012. Mol Neurodegener. 7:57). (A) In a background of sul-2(gk187) deletion, the number of paralyzed worms decreased. The graph shows the total number obtained from three biological replicas (worm n=100). Chi-squared, two-tailed, p-value = 0.0161. (B) Treatment with 1 μg / ml STX64 decreased the number of paralyzed worms. The graph shows the total number obtained from three biological replicas (worm n>120). Chi-squared, two-tailed, p-value = 0.0014. (C) Treatment with 1 μg / ml EMATE decreased the number of paralyzed worms. The graph shows the total number obtained from three biological replicas (worm n>117). Chi-squared, two-tailed, p-value = 0.0091. [Modes for carrying out the invention]

[0018] definition The terms “individual,” “patient,” or “subject” are used indiscriminately in this application to identify a human being and are not intended to limit it in any way. “Individual,” “patient,” or “subject” may be of any age, sex, and physical condition. The term “animal” as used in this application refers to any multicellular eukaryotic heterotroph that is not human. In preferred embodiments, the animal is selected from the group consisting of cats, dogs, pigs, ferrets, rabbits, gerbils, hamsters, guinea pigs, horses, worms, rats, mice, cattle, sheep, goats, alpacas, camels, donkeys, llamas, yaks, giraffes, elephants, meerkats, lemurs, lions, tigers, kangaroos, koalas, bats, monkeys, chimpanzees, gorillas, bears, dugongs, manatees, seals, and rhinoceroses.

[0019] As used herein, “pharmaceutically acceptable carrier” or “pharmaceutically acceptable diluent” means any solvent, dispersion, coating agent, antibacterial and antifungal agent, isotonic agent and absorption retarder that is compatible with the administration of a pharmaceutical product. The term “pharmaceutically acceptable excipient” refers to any substance that is compounded with the active ingredient of a pharmaceutical product and is included for the purpose of long-term stabilization, increasing the volume of a solid formulation containing a small amount of potent active ingredient, or enhancing the therapeutic effect of the active ingredient in the final dosage form, such as by promoting drug absorption, reducing viscosity, or improving solubility. Excipients may also be useful in the manufacturing process, assisting in the handling of the active substance in question, such as by promoting the flowability or non-stickiness of powders, in addition to assisting in vitro stability, such as preventing denaturation or aggregation over the expected shelf life. The use of such media and active ingredients for pharmaceutical active ingredients is well known in the art. Acceptable carriers, excipients, or stabilizers are non-toxic to the recipient at the dosage and concentration used and do not limit the scope of the present invention, but include additional buffers, preservatives, cosolvents, antioxidants including ascorbic acid and methionine, chelating agents such as EDTA, metal complexes (e.g., Zn-protein complexes), biodegradable polymers such as polyesters, salt-forming counterions such as sodium and polyhydric sugar alcohols, amino acids such as alanine, glycine, glutamine, asparagine, histidine, arginine, lysine, ornithine, leucine, 2-phenylalanine, glutamic acid, and threonine, lactitol, stachyose, mannose, sorbose, xylose, ribose, ribitol, myoinisitose, myoinisitol, galactose, galactitol, glycerol, cyclitols (e.g.) For example, inositol, organic sugars or sugar alcohols such as polyethylene glycol, urea, Examples include sulfur-containing reducing agents such as glutathione, thioctic acid, sodium thioglycolate, thioglycerol, α-monothioglycerol, and sodium thiosulfate; low molecular weight proteins such as human serum albumin, bovine serum albumin, gelatin, or other immunoglobulins; and hydrophilic polymers such as polyvinylpyrrolidone. (Remington: The Science and Practice of Pharmacy 22) nd This edition contains other pharmaceutically acceptable carriers, excipients, or stabilizers as described in Pharmaceutical Press (2012), ISBN-13: 9780857110626. It's okay to be there.

[0020] As used in this application, the terms “treatment” and “therapy” refer to a set of sanitary, pharmacological, surgical, and / or physical means used with the aim of improving a health problem, as well as for the purpose of curing and / or alleviating a disease and / or symptom. The terms “treatment” and “therapy” include preventive and curative methods, for both are directed toward maintaining and / or restoring the health of an individual or animal. Regardless of the cause of the symptoms, disease, and disability, the administration of a medicine suitable for alleviating and / or curing a health problem should be interpreted as a form of treatment or therapy in the context of this application.

[0021] As used in this application, the term “prevention” refers to a set of sanitary, pharmacological, surgical, and / or physical means used to prevent the onset and / or development of a disease and / or symptoms. The term “prevention” encompasses preventive methods, as these are used to maintain the health of animals or individuals.

[0022] The term "sulfatase inhibitor" refers to any substance that can reduce the activity of an esterase class enzyme that catalyzes the hydrolysis of sulfate esters. This substance may be any of the following: a gene encoding a sulfatase enzyme, a transcription factor of said gene, an expression product of said gene, for example, but not limited to these, a molecule that binds to either messenger RNA or a sulfatase enzyme, thereby reducing or inhibiting the expression and activity of the molecule it binds to, and / or its intracellular or extracellular signaling, resulting in complete or partial inhibition of the activity of the sulfatase enzyme. The inhibitor can be selected from a list of, but not limited to, antagonists against sulfatase enzymes (preferably chemicals), silencing RNA, or specific antibodies against sulfatase enzymes (preferably monoclonal antibodies), and in this invention, such antibodies may be defined as neutralizing antibodies against the effects of sulfatase enzymes. Examples of chemical inhibitors of sulfatase enzyme activity include, but are not limited to, a series of alternative substrates such as 2-(hydroxyphenyl)indol sulfate and 5-androsten-3β,17β-di Synthetic or natural steroids exhibiting inhibitory activity against STS, such as all-3 sulfates; competitive inhibitors such as E1-MTP or EMAT; non-estrogen inhibitors such as DU-14 (CAS number: 186303-55-9), COUMATE (4-methylcoumarin-7-O-sulfamate), or STX64 (i.e., compounds of formula (II)); or IC against sulfatase enzymes. 50 Various studies (Purohit & Foster, 2012, J. Endocrinol., 212(2):99-110) have requested KW-2581 or STX213, etc. Other items.

[0023] The term “steroid hormone sulfatase” (“STS”) refers to any sulfatase enzyme involved in steroid metabolism. In particular, the enzyme catalyzes the conversion of sulfated steroid precursors to free steroids. Exemplary human STSs have been sequenced and characterized, and the data are deposited in the UniProtKB database under accession number P08842. The term “steroid hormone sulfatase inhibitor” refers to any substance that can reduce the activity of steroid hormone sulfatase. The substance may be one of the following: a gene encoding an STS enzyme, a transcription factor of the gene, or the gene itself. A molecule that binds to either of the expression products of the STS enzyme, for example, a molecule that binds to either messenger RNA or the STS enzyme, and thereby reduces or inhibits the expression and activity of the molecule it binds to, and / or its intracellular signaling, resulting in complete or partial inhibition of the activity of the STS enzyme. The inhibitor can be selected from a list of antagonists (preferably chemical substances), silencing RNA, or specific antibodies (preferably monoclonal) against the STS enzyme, for example, a molecule that binds to either messenger RNA or the STS enzyme, for example, a molecule that binds to either messenger RNA or the STS enzyme, and / or inhibits the activity of and / or inhibits the activity of the STS enzyme, and / or inhibits the activity of the STS enzyme, and / or inhibits the activity of the STS enzyme, and / or inhibits the expression and activity of the molecule it binds to, and / or inhibits its intracellular signaling, and / or inhibits its intracellular signaling, and for example, a molecule that binds to either messenger RNA or the STS enzyme, and / or inhibits the expression and activity of the molecule it binds to, and / or inhibits its intracellular signaling, and / or inhibits its intracellular signaling, and for example, a molecule that binds to either messenger RNA or the STS enzyme, and for example, a molecule that binds to either messenger RNA Synthetic or natural steroids exhibiting inhibitory activity against STS, such as 3β,17β-diol-3 sulfate; competitive inhibitors such as E1-MTP or EMAT; non-estrogen inhibitors such as DU-14, COUMATE (4-methylcoumarin-7-O-sulfamate) or STX64 (i.e., compounds of formula (II)); or IC against sulfatase enzymes. 50 However, various studies (Purohit & Foster, 2012, J. Endocrinol., 212(2):99-110) have identified other options such as KW-2581 or STX213.

[0024] The terms "protein aggregation disorders," "proteopathies," "proteinopathy," or "protein misfolding disorders" refer to any disease in which a particular protein malfunctions structurally, thereby interfering with the function of the body's cells, tissues, and organs. In many cases, proteins are unable to fold into their normal shape, and in this misfolded state, the protein may become toxic in some way or lose its normal function. Non-exclusive examples of protein aggregation disorders include systemic AL amyloidosis, Alzheimer's disease, type 2 diabetes, Parkinson's disease, transmissible spongiform encephalopathy (e.g., bovine spongiform encephalopathy), fatal familial insomnia, Huntington's disease, medullary thyroid carcinoma, cardiac arrhythmias, atherosclerosis, rheumatoid arthritis, aortic medial amyloidosis, prolactinoma, familial amyloid polyneuropathy, hereditary nonneuropathic systemic amyloidosis, dialysis-associated amyloidosis, Finnish amyloidosis, and lattice corneal disease. Stroke, cerebral amyloid angiopathy, cerebral amyloid angiopathy (Icelandic type), sporadic inclusion body myositis, amyotrophic lateral sclerosis (ALS), prion-related or cavernous encephalopathy such as Creutzfeldt-Jakob, dementia with Lewy bodies, frontotemporal dementia with parkinsonism, spinocerebellar ataxia, spinocerebellar ataxia, bulbar spinal muscular atrophy, hereditary dentatorubral-pallidoluysian atrophy, familial dementia in the UK, familial dementia in Denmark, non-neuropathic localized diseases such as type II diabetes, medullary thyroid carcinoma, atrial amyloidosis, hereditary cerebral hemorrhage with amyloidosis, pituitary prolactinoma, injection-localized amyloidosis, aortic medial amyloidosis, hereditary lattice corneal dystrophy, eyelashes Examples include corneal amyloidosis associated with trichiasis, cataracts, odontogenic calcifying epithelioma, alveolar proteinosis, inclusion body myositis, lichenoid amyloidosis, and non-neuropathic systemic amyloidosis such as AL amyloidosis, AA amyloidosis, familial Mediterranean fever, senile systemic amyloidosis, familial amyloid polyneuropathy, hemodialysis-associated amyloidosis, ApoAI amyloidosis, ApoAII amyloidosis, ApoAIV amyloidosis, Finnish hereditary amyloidosis, lysozyme amyloidosis, fibrinogen amyloidosis, Icelandic hereditary cerebral amyloid angiopathy, familial amyloidosis, and systemic amyloidosis occurring in multiple tissues such as light chain amyloidosis, as well as various other neurodegenerative disorders.

[0025] The term "protein aggregate" refers to any accumulation of abnormally folded proteins that cause and / or are associated with the negative progression of protein aggregation disorders.

[0026] The term "amyloid" refers to a morphology of protein aggregates that form unbranched fibers binding to Congo Red, and subsequently exhibit green birefringence when viewed between crossed polarizers (see, for example, Eisenberg & Jucker, 2012. Cell. 148(6):1188-203 and Sipe et al., 2012. Amyloid. 19(4):167-70).

[0027] The term "oligomer" refers to any accumulation of abnormally folded proteins that cause and / or are associated with the negative progression of protein aggregation disorders and do not meet the definition of amyloid. For example, polyglutamine oligomers cause and / or are associated with the negative progression of Huntington's disease (see Hoffner & Dijan, 2014. Brain Sci. 4(1): 91-122).

[0028] Compositions and kits In a first embodiment, the present invention provides a composition comprising a sulfatase inhibitor used for the treatment and / or prevention of protein aggregation disorders.

[0029] In a second embodiment, the present invention provides a kit for use in the manufacture of a drug for treating and / or preventing protein aggregation disorders, comprising (i) a sulfatase inhibitor and (ii) a pharmaceutically acceptable carrier and / or diluent. In a preferred embodiment, the kit further comprises a pharmaceutically acceptable excipient.

[0030] Preferred embodiments of the kit and composition of the present invention are provided below.

[0031] Sulfatase inhibitors In preferred embodiments, the sulfatase inhibitor is a steroid hormone sulfatase inhibitor. Preferably, the steroid hormone sulfatase inhibitor is 2-(hydroxyphenyl)indole sulfate, DU- The antibody is selected from a list consisting of 14,5-androsten-3β,17β-diol-3 sulfate, E1-MTP, Emate, Coumate, STX64, KW-2581, STX213, morpholine, silencing RNA, and specific antibodies against STS enzymes.

[0032] In a preferred embodiment, the sulfatase inhibitor or steroid hormone sulfatase inhibitor is of formula (I): [ka] (In the formula, (a) R1 to R6 are independently selected from hydrogen, halogens (fluorine, chlorine, bromine, iodine, or astatine), hydroxyl, sulfamates (OSO2NH2), alkyls, and their salts. (b) At least one of R1 to R6 is a sulfamate group, It is a compound in which two or more R1-R6 groups are linked together to form an additional cyclic structure.

[0033] In a preferred embodiment, R1 and R2 form an additional cyclic structure containing 3 to 10 carbon atoms. 、 R6 is OSO2NH2. In a preferred embodiment, Alkyl groups are C1-C6 alkyl groups.

[0034] In a preferred embodiment, the sulfatase inhibitor is of formula (II): [ka] It is a compound of [the compound].

[0035] The synthesis methods for the compounds of formulas (I) and (II) described above are disclosed in International Publication No. 97 / 30041.

[0036] Protein aggregation disorders Protein aggregates, such as amyloids and oligomers, are associated with many diseases. In some cases, these protein aggregates can be toxic and cause significant damage to cells and tissues. This toxicity is considered one of the contributing factors that cause and / or contribute to the pathogenesis of protein aggregation disorders.

[0037] Furthermore, abnormal protein processing and folding that lead to protein aggregation disorders may begin decades before the visible symptoms of the disorder can be observed (Jack et al., 2010. Lancet Neurol. 9(1):119-28). Therefore, In preferred embodiments, amyloid and / or oligomers are removed and / or their formation is prevented in patients and / or animals as a result of administration of any one of the compositions of the present invention. Furthermore, in preferred embodiments, sulfatase inhibitors treat and / or prevent protein toxicity in protein aggregation disorders. The term "protein toxicity" refers to any dysfunction of cellular function caused by protein misfolding.

[0038] By directly targeting the formation of protein aggregates, the compositions and kits of the present invention can treat and / or prevent protein aggregation disorders in patients and / or animals that are in the early stages of the disorder but do not yet show any outward symptoms. Furthermore, the compositions and kits of the present invention can also treat advanced stages of protein aggregation disorders, as shown in Example 2, and in Figures 7F and 7G.

[0039] In a preferred embodiment, the patient and / or animal has undergone pathophysiological changes that cause protein aggregation, but has not yet reached a stage of disease where outward symptoms are observable. In other words, the patient and / or animal is in the early stages of the disease. The term "outward symptoms" refers to any symptoms that can be observed by a physician using any non-invasive method.

[0040] In preferred embodiments, a sulfatase inhibitor slows the progression of protein aggregation disorders by inhibiting the formation of protein aggregates, and / or delays the onset of protein aggregation disorders by inhibiting the formation of protein aggregates.

[0041] In preferred embodiments, protein aggregation disorders include systemic AL amyloidosis, Alzheimer's disease, type 2 diabetes, Parkinson's disease, and transmissible spongiform encephalopathy, such as bovine spongiform encephalopathy. Fatal familial insomnia, Huntington's disease, medullary thyroid carcinoma, cardiac arrhythmia, atherosclerosis, rheumatoid arthritis, aortic medial amyloidosis, prolactinoma, familial amyloid polyneuropathy, hereditary nonneuropathic systemic amyloidosis, dialysis-associated amyloidosis, Finnish amyloidosis, lattice keratitis, cerebral amyloid angiopathy, cerebral amyloid angiopathy (Icelandic type), sporadic inclusion body myositis, amyotrophic lateral sclerosis ALS, prion-related or cavernous encephalopathy such as Creutzfeldt-Jakob syndrome, dementia with Lewy bodies, frontotemporal dementia with parkinsonism, spinocerebellar ataxia, spinocerebellar ataxia, bulbar spinal atrophy, hereditary dentatorubral-pallidoluysian atrophy, familial dementia in the UK, familial dementia in Denmark, nonneuropathic focal disorders in type II diabetes, medullary thyroid carcinoma, atrial amyloidosis, hereditary cerebral hemorrhage with amyloidosis, pituitary prolamydia Cutinoma, injection-specific amyloidosis, aortic medial amyloidosis, hereditary lattice corneal dystrophy, corneal amyloidosis associated with trichiasis, cataract, odontogenic calcifying epithelioma, pulmonary alveolar proteinosis, inclusion body myositis, lichenoid amyloidosis, and AL amyloidosis, AA amyloidosis, familial Mediterranean fever, senile systemic amyloidosis, familial amyloid polyneuropathy, hemodialysis-associated amyloidosis, ApoAI amyloidosis The selection is made from a list of non-neuropathic systemic amyloidosis such as loidosis, ApoAII amyloidosis, ApoAIV amyloidosis, Finnish hereditary amyloidosis, lysozyme amyloidosis, fibrinogen amyloidosis, Icelandic hereditary cerebral amyloid angiopathy, familial amyloidosis, and systemic amyloidosis occurring in multiple tissues such as light chain amyloidosis, as well as various other neurodegenerative disorders. Preferably, the protein aggregation disorder is selected from a list of Alzheimer's disease, Parkinson's disease, and Huntington's disease. In preferred embodiments, the protein aggregation disorder is not Alzheimer's disease and / or a type of cancer.

[0042] In a preferred embodiment, the protein aggregation disorder is selected from a list consisting of Alzheimer's disease, Parkinson's disease, and Huntington's disease. In a preferred embodiment, amyloid and / or oligomers are removed and / or their formation is prevented in patients with Alzheimer's disease, Parkinson's disease, or Huntington's disease.

[0043] In a preferred embodiment, the protein aggregation disorder is a central nervous system-localized protein aggregation disorder. In a preferred embodiment, the protein aggregation disorder is also a neurodegenerative disorder. The term “neurodegenerative disorder” refers to any disorder characterized by progressive loss of neuronal structure or function, including neuronal death. For example, Alzheimer’s disease is an example of both a protein aggregation disorder and a neurodegenerative disorder.

[0044] Embodiments of combinations of sulfatase inhibitors and protein aggregation disorders In preferred embodiments, the sulfatase inhibitor is 2-(hydroxyphenyl)indol sulfate, 5-androsten-3β, A sulfatase inhibitor may be selected from a list consisting of DU-14, 17β-diol-3 sulfate, E1-MTP, Emate, Coumate, STX64, KW-2581, STX213, morpholine, silencing RNA, and specific antibodies against STS enzymes, or the sulfatase inhibitor may be formulated with formula (I): [ka] (In the formula, (a) R1 to R6 are independently selected from hydrogen, halogens (fluorine, chlorine, bromine, iodine, or astatine), hydroxyl, sulfamates (OSO2NH2), alkyls, and their salts. (b) At least one of R1 to R6 is a sulfamate group, The compound is one in which two or more R1-R6 groups are linked to form an additional cyclic structure, and the protein aggregation disease is selected from the list consisting of Alzheimer's disease, Huntington's disease, and Parkinson's disease.

[0045] In a preferred embodiment, the sulfatase inhibitor is STX64 (i.e., a compound of formula (II)), and the protein aggregation disorder is selected from a list consisting of Alzheimer's disease, Huntington's disease, and Parkinson's disease. In a preferred embodiment, the sulfatase inhibitor is STX64, and the protein aggregation disorder is Alzheimer's disease. In a preferred embodiment, the sulfatase inhibitor is STX64, and the protein aggregation disorder is Huntington's disease. In a preferred embodiment, the sulfatase inhibitor is STX64, and the protein aggregation disorder is Parkinson's disease.

[0046] Pharmaceutical composition In preferred embodiments, the composition is a pharmaceutical composition further comprising a pharmaceutically acceptable carrier and / or diluent. Preferably, the pharmaceutical composition may further comprise a pharmaceutically acceptable excipient.

[0047] The pharmaceutical compositions described herein may also contain other substances. These substances include, but are not limited to, cryoprotectants, freeze-drying protectants, surfactants, fillers, antioxidants, and stabilizers. In some embodiments, the pharmaceutical compositions may be freeze-dried.

[0048] As used herein, the term “freeze protectant” includes substances that provide stability to a composition against freeze-induced stress. Freeze protectants may also provide protection during primary and secondary drying and long-term product storage. Non-limiting examples of freeze protectants include sugars such as sucrose, glucose, trehalose, mannitol, mannose, and lactose; polymers such as dextran, hydroxyethyl starch, and polyethylene glycol; surfactants such as polysorbates (e.g., PS-20 or PS-80); and amino acids such as glycine, arginine, leucine, and serine. Freeze protectants with low toxicity in biological systems are generally used.

[0049] In one embodiment, a lyophilization protectant is added to the pharmaceutical composition described herein. As used herein, the term “lyophilization protectant” includes substances that provide stability to the composition during the lyophilization or dehydration process (primary and secondary lyophilization cycles). The lyophilization protectant helps to improve long-term product stability by minimizing product degradation during the lyophilization cycle. Non-limiting examples of lyophilization protectants include sugars such as sucrose or trehalose, amino acids such as monosodium glutamate, amorphous glycine or histidine, methylamines such as betaine, lyotropic salts such as magnesium sulfate, trivalent or higher sugar alcohols, polyols such as glycerin, erythritol, glycerol, arabitol, xylitol, sorbitol and mannitol, propylene glycol, polyethylene glycol, Pluronic® compounds and combinations thereof. The amount of lyophilization protectant added to the pharmaceutical composition is generally such that it does not result in an unacceptable amount of degradation when the pharmaceutical composition is lyophilized.

[0050] In some embodiments, a volume extender is included in the pharmaceutical composition. The term "extender" includes active ingredients that contribute to the structure of a lyophilized product without directly interacting with the pharmaceutical product. In addition to providing a pharmaceutically refined cake, extenders can also provide useful qualities in terms of altering the decay temperature, providing freeze-thaw protection, and improving stability over long-term storage. Non-limiting examples of extenders include mannitol, glycine, lactose, and sucrose. Extenders may be crystalline (e.g., glycine, mannitol, or sodium chloride) or amorphous (e.g., dextran, hydroxyethyl starch) and are generally used in formulations at concentrations of 0.5% to 10%.

[0051] Remington's Pharmaceutical Sciences 16th edition, Osol, A. Ed. (1980) or Remington: The Science and Practice of Pharmacy 22nd Other pharmaceutically acceptable carriers, excipients, or stabilizers, such as those described in edition, Pharmaceutical Press (2012), ISBN-13: 9780857110626, may also be included in the pharmaceutical compositions described herein, provided they do not adversely affect the desired characteristics of the pharmaceutical composition.

[0052] For solid pharmaceutical compositions, conventional non-toxic solid carriers can be used, such as pharmaceutical-grade mannitol, lactose, starch, magnesium stearate, sodium saccharin, talc, cellulose, glucose, sucrose, and magnesium carbonate. For injectable solutions, the pharmaceutical composition may further contain cryoprotectants, freeze-drying protectants, surfactants, fillers, antioxidants, stabilizers, and pharmaceutically acceptable carriers. For aerosol administration, the pharmaceutical composition is generally supplied in a micronized form together with a surfactant and a propellant. The surfactant must, of course, be non-toxic and generally soluble in the propellant. Typical examples of such active ingredients include caproic acid, octanoic acid, lauric acid, palmitic acid, stearic acid, linoleic acid, linolenic acid, olesteric acid, and oleic acid. These are esters or partial esters of fatty acids containing 6 to 22 carbon atoms and aliphatic polyhydric alcohols or their cyclic anhydrides. Mixed esters, such as those of natural glycerides, can be used. A carrier may be included as needed, for example, lecithin for intranasal delivery. For suppositories, conventional binders and carriers may include, for example, polyalkalene glycols or triglycerides.

[0053] In preferred embodiments, the compositions of the present invention are prepared for administration mediated by oral, sublingual, buccal, intranasal, intravenous, intramuscular, intraperitoneal, and / or inhalation.

[0054] Administration The compositions of the present invention may be administered by any route known to those skilled in the art. In preferred embodiments, the compositions of the present invention are administered transdermally, sublingually, intravenously, intranasally, intraventricularly, intraarterially, intracerebrally, intramuscularly, intraperitoneally, orally, or by inhalation.

[0055] In preferred embodiments, the compositions of the present invention are administered transdermally, sublingually, intravenously, intraperitoneally, orally, or by inhalation. When the compositions are administered by inhalation, they may be aerosolized and administered, for example, via an anesthetic mask.

[0056] In preferred embodiments, the composition of the present invention is administered transdermally, sublingually, intravenously, subcutaneously, orally, or by inhalation. Preferably, the composition is administered orally or sublingually.

[0057] In a preferred embodiment, the composition contains a therapeutically effective dose of a sulfatase inhibitor. The term "therapeutically effective dose" refers to the amount of sulfatase inhibitor in the composition that has a therapeutic effect and can treat and / or prevent protein aggregation disorders. In a preferred embodiment, a dose of 0.01 mg / kg to 100 mg / kg of sulfatase inhibitor is administered to the patient. Preferably, a dose of 0.01 mg / kg to 10 mg / kg is administered to the patient. More preferably, a dose of 0.05 mg / kg to 1 mg / kg is administered to the patient.

[0058] In preferred embodiments, the composition is used in combination therapy with any other treatment or therapy commonly used for the treatment and / or prevention of protein aggregation disorders. In preferred embodiments, the composition is used in combination therapy with donepezil, rivastigmine, galantamine, memantine, levodopa, carbidopa and / or tetrabenazine.

[0059] The compositions of the present invention may be administered once or more times. Those skilled in the art can determine the most effective dosing regimen for a patient. For example, the most effective dosing regimen may involve administering the composition to the patient twice daily, once daily, once every three days, once weekly, once monthly, once every three months, once every six months, or once annually.

[0060] The following provisions are included in the present invention. [1] A composition comprising a sulfatase inhibitor used for the treatment and / or prevention of protein aggregation disorders, preferably, (i) In patients and / or animals, amyloid and / or oligomers are removed and / or their formation is prevented and / or (ii) A composition wherein the sulfatase inhibitor described above treats and / or prevents protein toxicity in protein aggregation disorders. [2] The composition for use according to item [1], wherein the sulfatase inhibitor slows the progression of a protein aggregation disorder by inhibiting the formation of protein aggregates, and / or delays the onset of a protein aggregation disorder by inhibiting the formation of protein aggregates. [3] The composition for use according to item [1] or [2], wherein the protein aggregation disorder is a central nervous system localized protein aggregation disorder. [4] A composition for use according to any one of the items [1] to [3], wherein amyloid and / or oligomers are removed and / or their formation is prevented in patients with Alzheimer's disease, Parkinson's disease or Huntington's disease, or the protein aggregation disorder is Alzheimer's disease, Parkinson's disease or Huntington's disease. [5] The above sulfatase inhibitors are given by formula (I): [ka] (In the formula, (a) R1 to R6 are independently selected from hydrogen, halogen, hydroxyl, sulfamate, alkyl and their salts. (b) At least one of R1 to R6 is a sulfamate group, (c) A composition for use according to any one of the items [1] to [4], which is a compound of (c) two or more R1 to R6 linked together to form an additional cyclic structure. [6] The composition for use according to item [5], wherein R1 and R2 form an additional cyclic structure containing 3 to 10 carbon atoms. [7] A composition for use as described in item [5] or [6], wherein R6 is OSO2NH2. [8] The above sulfatase inhibitor is given by formula (II): [ka] A composition for use as described in any one of the items [5] to [7], which is a compound of [the compound]. [9] A composition for use according to any one of the items [1] to [8], wherein the composition is a pharmaceutical composition comprising a sulfatase inhibitor according to any one of the items [1] to [8] and a pharmaceutically acceptable carrier and / or diluent.

[10] A composition for use according to any one of the items [1] to [9], wherein the composition is administered orally.

[11] A composition for use according to any one of items [1] to

[10] , wherein a dose of 0.01 mg / kg to 100 mg / kg, preferably 0.01 mg / kg to 10 mg / kg, and more preferably 0.05 mg / kg to 1 mg / kg is administered to the patient.

[12] The composition for use according to any one of the items [1] to

[11] , used in combination therapy with donepezil, rivastigmine, galantamine, memantine, levodopa, carbidopa and / or tetrabenazine.

[13] A kit for use in the manufacture of a drug for the treatment and / or prevention of protein aggregation disorders, comprising (i) a sulfatase inhibitor and (ii) a pharmaceutically acceptable carrier and / or diluent.

[14] The above sulfatase inhibitors are given by formula (I): [ka] (In the formula, (a) R1 to R6 are independently selected from hydrogen, halogen, hydroxyl, sulfamate, alkyl and their salts. (b) At least one of R1 to R6 is a sulfamate group, A compound having two or more R1-R6 groups linked together to form an additional cyclic structure, as described in section

[13] , for use.

[15] The above sulfatase inhibitor is given by formula (II): [ka] A compound of the kit for use described in item

[14] . [Examples]

[0061] C. elegans containing the sul-2(gk187) knockout allele (wormbase ID: WBVar00145594) was generated by the C. elegans Deletion Mutant Consortium (2012. G3: GENES, GENOMES, GENETICS. 2(11):1415-1425) and deposited with the Caenorhabditis Gene Center (https: / / cbs.umn.edu / cgc / home). The sul-2(gk187) allele consists of a deletion of 131 amino acids, including the catalytic core of sulfatase. The deletion generates a frameshift, with only the first four amino acids of the original sequence being conserved. Therefore, we consider sul-2(gk187) to be a null mutant allele of sul-2. Mutant lesions are available in the wormbase (http: / / www.wormbase.org / species / c_elegans / variation / WBVar00145594#02-45-3).

[0062] Example 1: Effects of STX64 on a nematode model of neurodegenerative disease C. elegans model of Parkinson's disease: The inventors have developed a model of sul-2 mutation or treatment with 1 μg / ml concentration of STX64 (Sigma catalog number S1950) dissolved in DMSO. However, we tested whether it could improve symptoms caused by overexpression of human α-synuclein in muscle cells (see the following references for information on the model used: van Ham et al., 2008. PLoS Genet. 4(3):e1000027, Gidalevitz et al., 2009. PLoS Genet. 5(3):e1000399, van Ham et al., 2010. Cell. 142(4):601-12). Specifically, nematodes were synchronized at 20°C and monitored every two days. On each monitoring day, a nematode was placed in one drop of M9 buffer and allowed to adapt for 30 seconds, after which the number of pitches per minute was counted, assuming that pitching occurred when the nematode's head crossed the axial axis. N=20 for each day and condition assayed (this protocol was adapted from Van Ham et al., 2010. Cell. 142: 601-612). As seen in Figures 1A and 1B, treatment with sul-2 mutation or STX64 significantly improved motility.

[0063] Furthermore, the inventors investigated the effects of sul-2 deletion and STX64 administration on α-synuclein aggregation. To quantify the aggregates, animals were transferred to a pad containing 2% agarose and fixed with 25 mM levamisole. The number of α-synuclein aggregates was counted by counting fluorescent aggregates at 40x magnification from the first bulb pharyngeal to the second bulb. This was determined by the following: Loss or inhibition of SUL-2 function increased the number of larger α-synuclein aggregates while decreasing the number of smaller aggregates (see Figures 2 and 3), suggesting better handling of protein aggregates in animals with reduced STS activity (Roberts & Brown, 2015. Biomolecules. 5(2):282-305, Moll et al., 2016. FASEB J. 30(4):1656-69).

[0064] In a surrogate C. elegans model of Parkinson's disease in which GFP-labeled dopaminergic neurons express α-synuclein, dopaminergic neurons were found to die due to α-synuclein toxicity (Cooper et al., 2006. Science. 313(5785):324-8). Consistent with previous findings, the sul-2 mutant showed increased neuronal survival. (See Figure 4) It exhibits a neuroprotective effect that reduces STS activity.

[0065] C. elegans model of Huntington's disease: In a Huntington's disease model expressing 35 polyglutamine (polyQ) repeats fused to a fluorescent protein, the inventors found that both sul-2 mutation and administration of 1 μg / ml STX64 dissolved in DMSO reduced the total number of aggregates (see Figure 5). To quantify the aggregates, animals were transferred to a pad containing 2% agarose and fixed with 25 mM levamisole. The number of polyQ aggregates was determined by counting fluorescent aggregates at a 10x magnification from the first to the second pharyngeal bulb.

[0066] C. elegans model of Alzheimer's disease: The inventors also found that as the nematode ages... We tested an Alzheimer's disease model expressing β-amyloid protein in muscle cells that cause paralysis. Nematodes were synchronized at 20°C. Paralysis was monitored from day 1 of the adult stage by observing whether the nematodes became immobile after stimulation with a platinum pick. In addition, to track paralyzed nematodes, we visualized which nematodes had a bacterial halo around their heads, as paralysis hinders food movement. Consistent with our previous data, sul-2 mutation and STX64 treatment delayed paralysis (see Figure 6).

[0067] Example 2: Effects of STX64 on a mammalian model of neurodegenerative disease Mouse strains and conditions: The male Swiss (CD1) and APP-PS1 (Blanchard et al., 2003. Exp Neurol. 184(1):247-63) mice used in this study were certified. The mice were purchased from a supplier (University of Seville, Spain) and acclimatized to standard animal housing conditions (12-hour light-dark cycle, temperature, and humidity) for 2-3 weeks. Behavioral studies were conducted using 8-week-old Swiss mice and APP-PS1 mice over 15 months old in a C57 Black background. Histological studies used male APP-PS1 mice from 2 months to over 15 months old. All experiments were conducted in accordance with the European Union guidelines (2010 / 63 / EU) and the Spanish regulations on the use of laboratory animals in chronic experiments (RD53 / 2013:BOE08 / 02 / 2013 on the care of laboratory animals), and approval was obtained from the Animal Care Committee of Pablo de Olavide University prior to the conduct of this study.

[0068] Local drug injection in mice: Mice were anesthetized with 4% chloral hydrate (10 μL / kg body weight, ip), and once fully anesthetized, the mice were placed in a stereotactic frame. To injure the hippocampus, 0.5 μl of a 5 μM solution of β-amyloid oligomer was injected bilaterally on the dorsal side of the hippocampus of the mice at the following stereotactic coordinates: AP = -2.2 mm, ML = ±1.5 mm, V = -1.5 mm from the cruciate suture. The mice were then allowed to recover for at least 15 days. In addition, 0.5 μl of a 1 mg / ml solution of STX64 was administered to the same mice that had been given STX64, 20 minutes before administering β-amyloid oligomer. STX64 was injected at the same rostral hippocampal coordinates. STX64 and β-amyloid oligomer were delivered at a rate of 0.2 μl / min using an injection syringe (Hamilton) and left for 2.5 minutes after injection.

[0069] Oral STX administration in mice: STX64 was dissolved in drinking water at a concentration of 0.005 mg / ml. Mice were exposed to the STX solution for 3 to 4 weeks, and daily water intake was recorded during treatment. The estimated daily dose of STX64 was 1 mg / kg to 2 mg / kg.

[0070] Step-through passive avoidance (PA) test: Mice have an innate preference for dark, enclosed environments. During the habituation phase, mice were handled and allowed to move freely for one minute in a chamber (47cm × 18cm × 26cm, Ugo Basile) symmetrically divided into one light and one dark compartment (each measuring 28.5cm × 18cm × 26cm). During the training phase, mice were temporarily confined to the light compartment, and then 30 seconds later, the door separating the dark-light compartments was opened. The door automatically closed when the mouse entered the dark compartment, and the mouse received an electric shock delivered through a metal floor (0.5mA for 5 seconds and 0.3mA for 5 seconds for Swiss mice and C57Black mice, respectively). In the retention test performed at specified time points, mice that recalled the experience of the electric shock when moved to the light compartment avoided entering the dark compartment or took at least a long time to enter it. Therefore, the waiting time before entering the dark compartment (escape latency) is a measure of information learning or memory retention, depending on how much time has passed since the training session before the test. Escape latency (seconds) represents the training, short-term and long-term memory (STM and LTM) sessions.

[0071] Immunohistochemistry and histological analysis: In immunohistochemistry, antibodies against β-amyloid (1 Antibody staining was performed using Sigma-Aldrich (3000). Antibody staining was performed with H2O2 and diaminobenzidine. Visualization and nickel enhancement were performed. To minimize variability, at least five sections were analyzed per mouse under a bright-field DMRB RFY HC microscope (Leica). Each section In this study, the percentage area occupied by β-amyloid, the density of β-amyloid accumulation, and the average size were quantified using Image-J software (public domain: downloaded as a free software package from http: / / rsb.info.nih.gov / ij / download.html).

[0072] Results: Since STX64 showed a significant therapeutic effect in a C. elegans neurodegeneration model, the inventors tested the effect of this drug on cognitive changes induced by intrahippocampal β-amyloid oligomer injection in an acute Alzheimer's disease (AD) mouse model (Figure 7A). Both topical and systemic STX64 treatment reversed the cognitive impairment induced by intrahippocampal administration of β-amyloid oligomers.

[0073] To evaluate the effect of oral STX64 treatment on amyloid pathology in a chronic AD mouse model, the inventors evaluated the effect of 3-4 weeks of oral STX64 administration on neocortical (cerebral cortex and hippocampal) amyloid deposition in APP-PS1 mice older than 15 months of age (Figure 7B). Analysis of β-amyloid immunoreactivity area, plaque density, and size in older mice (15 months of age and above), which are also related to the later stages of neocortical amyloid deposition in the APP-PS1 model, revealed a significant reduction in STX64-treated mice, except for plaque size in the hippocampus (Figures 7C-7E). Interestingly, when the inventors compared the β-amyloid deposition of older (over 15 months old) APP-PS1 mice treated with STX64 to the normal amyloid deposition rate of untreated APP-PS1 mice, they observed that STX64 reduced β-amyloid deposition in APP-PS1 mice older than 15 months compared to APP-PS1 mice aged 10 to 12 months (Figure 7F).

[0074] All of these results demonstrated that STX64 treatment in APP-PS1 mice reduced β-amyloid deposition. To assess whether histological improvement correlated with improvement in cognitive and behavioral impairment, we compared the cognitive abilities of APP-PS1 mice over 15 months of age treated with either a vehicle or STX64 for 3–4 weeks. Vehicle-treated APP-PS1 mice showed defects in the passive avoidance test (Figure 7G), while those mice treated with STX64 completely reversed their cognitive impairment, reaching levels similar to wild-type mice under 15 months of age. All of these results suggest that STX64-induced changes in β-amyloid metabolism reduce cognitive and behavioral impairment caused by β-amyloid accumulation in acute and chronic AD mouse models, and that STX64 and other sulfatase inhibitors may be useful for treating proteopathy.

[0075] Example 3: Effect of STX64 on a mouse model of Huntington's disease Mouse strains and conditions: The R6 / 1 mice used in this study (Yi Li et al., 2005. NeuroRX. 2(3): 447-464, Mangiarini et al., 1996. Cell. 87(3):493-506) were purchased from certified suppliers and acclimatized to standard animal housing conditions (12-hour light-dark cycle, temperature, and humidity) for 2-3 weeks. Motor activity studies were conducted on 2-month-old R6 / 1 mice. All experiments were conducted in accordance with the European Union guidelines (2010 / 63 / EU) and the Spanish regulations on the use of laboratory animals in chronic experiments (RD53 / 2013:BOE08 / 02 / 2013 on the care of laboratory animals), and approval was obtained from the Animal Care Committee of Pablo de Olavide University prior to the conduct of this study.

[0076] Oral STX administration to mice: STX64 was dissolved in drinking water at a concentration of 0.005 mg / ml. One-month-old mice were exposed to the STX solution for one month, and their daily water intake was recorded during treatment. The estimated daily dose of TX64 was 1 mg / kg body weight to 2 mg / kg body weight.

[0077] Motor activity test: Spontaneous motor activity was tested for 15 minutes in a non-stress open field (56cm x 40cm x 40cm). The distance moved by the mice was estimated using Smart video-tracking software (Panlab).

[0078] Results: R6 / 1 mice exhibit decreased activity at 2 months of age. This decrease in motor activity can be avoided by oral administration of STX64 when the mice reach 1 month of age (see Figure 8). This example provides further evidence that the composition of the present invention is suitable for the treatment of protein aggregation disorders.

[0079] Example 4: Comparison of STX64 and EMATE in a C. elegans model of Alzheimer's disease Introduction: To determine the effects of different steroid sulfatase (STS) inhibitors on protein aggregation, the inventors used the GMC101 strain (McColl et al., 2012. Mol Neurodegener. 7:57), an AD-improved model of C. elegans, to study EMAT (CAS number: The assay (148672-09-7) was performed. EMATE, also known as estrone sulfamate, is a potent, irreversible inhibitor of estrone sulfatase (E1-STS) and dehydroepiandrosterone sulfatase (DHA-STS), but it also possesses strong estrogenic activity.

[0080] Strain: GMC101, Genotype: dvIs100 [pCL354(unc-54:DA-Aβ1-42)+pCL26(mtl-2):GFP].

[0081] Paralysis assay: Worms were grown at an unrestricted temperature of 16°C until they reached the L4 young adult stage. The temperature was then shifted to 25°C. The number of paralyzed and non-paralyzed animals was counted after 18 hours of incubation at the restricted temperature of 25°C.

[0082] Statistical analysis: Data was analyzed using a two-tailed chi-squared test on a contingency table. Statistical analysis was performed using GraphPad Prism software (version 7.00).

[0083] Results: The inventors assayed the temperature-dependent phenotype of paralysis under different backgrounds and conditions. The absence of sul-2 improved paralysis symptoms in the GMC101 strain (see Figure 9A). Similar effects were observed with STX64 (see Figure 9B) and EMAT (see Figure 9C). However, STX64 appeared to have the greatest effect.

Claims

1. A composition comprising a sulfatase inhibitor used for the treatment and / or prevention of protein aggregation disorders, wherein amyloid and / or oligomers are removed and / or their formation is prevented in patients and / or animals. The aforementioned sulfatase inhibitor is defined by formula (I): 【Chemistry 1】 (In the formula, (d) R １ ~R ６ These are independently selected from hydrogen, halogens, hydroxyl, sulfamates, alkyls and their salts, (e) R １ ~R ６ At least one of them is a sulfamate group, Two or more R １ ~R ６ A composition which is a compound of (which are linked together to form an additional cyclic structure).

2. The composition according to claim 1, wherein the sulfatase inhibitor treats and / or prevents protein toxicity in protein aggregation disorders.

3. The composition according to claim 1 or 2, wherein the sulfatase inhibitor slows the progression of protein aggregation disease by inhibiting the formation of protein aggregates, and / or delays the onset of protein aggregation disease by inhibiting the formation of protein aggregates.

4. The composition according to any one of claims 1 to 3, wherein the protein aggregation disorder is a central nervous system-localized protein aggregation disorder.

5. The composition according to any one of claims 1 to 4, wherein the formation of the amyloid and / or oligomers is prevented in patients with Alzheimer's disease, Parkinson's disease, or Huntington's disease, or the protein aggregation disorder is Alzheimer's disease, Parkinson's disease, or Huntington's disease.

6. R １ and R ２ The composition according to any one of claims 1 to 5, wherein the carbon atoms form an additional cyclic structure containing 3 to 10 carbon atoms.

7. R ６ is OSO ２ NH ２ The composition according to any one of claims 1 to 6, wherein

8. The aforementioned sulfatase inhibitor is given by formula (II): 【Chemistry 2】 A composition according to any one of claims 1 to 7, wherein the compound is [the compound].

9. The composition according to any one of claims 1 to 8, wherein the composition is a pharmaceutical composition comprising a sulfatase inhibitor according to any one of claims 1 to 8 and a pharmaceutically acceptable carrier and / or diluent.

10. The composition according to any one of claims 1 to 9, wherein the composition is administered orally.

11. The composition according to any one of claims 1 to 10, wherein a dose of 0.01 mg / kg to 100 mg / kg, preferably 0.01 mg / kg to 10 mg / kg, and more preferably 0.05 mg / kg to 1 mg / kg, is administered to the patient.

12. The composition according to any one of claims 1 to 11, wherein the composition is used in combination therapy with donepezil, rivastigmine, galantamine, memantine, levodopa, carbidopa and / or tetrabenazine.

13. A drug used for the treatment and / or prevention of protein aggregation disorders, wherein the drug is manufactured using a kit comprising (i) a sulfatase inhibitor and (ii) a pharmaceutically acceptable carrier and / or diluent. The aforementioned sulfatase inhibitor is defined by formula (I): 【Transformation 3】 (In the formula, (c) R １ ~R ６ These are independently selected from hydrogen, halogens, hydroxyl, sulfamates, alkyls and their salts, (d) R １ ~R ６ At least one of them is a sulfamate group, Two or more R １ ~R ６ It is a compound in which (these are linked to form an additional cyclic structure), A drug that, in the aforementioned patient and / or animal, removes amyloid and / or oligomers and / or inhibits their formation.

14. The aforementioned sulfatase inhibitor is given by formula (II): 【Chemistry 4】 The agent according to claim 13, which is a compound of the above.

Images