Aminothiols for the treatment of conditions characterized by protein misfolding or aggregation, mitochondrial dysfunction, or chronic inflammation

Abstract

The present invention features aminothiols for treating (1) diseases and conditions characterized by endogenous protein misfolding and / or aggregation, (2) diseases and conditions associated with mitochondrial dysfunction, and (3) diseases and conditions associated with chronic inflammation in a subject in need of such treatment.

Description

[Technical Field]

[0001] Provided herein is technology demonstrating that PrC-210 aminothiol or analogs thereof are highly effective when administered to treat (i) diseases and conditions characterized by endogenous protein misfolding and / or aggregation, (2) diseases and conditions associated with mitochondrial dysfunction, and (3) diseases and conditions associated with chronic inflammation. [Background technology]

[0002] Glutathione (GSH) is the most abundant non-protein thiol in human cells. GSH is synthesized exclusively in the cytoplasm from its constituent amino acids, but distributes to various compartments, including mitochondria, where its concentration in the matrix is ​​equivalent to that in the cytoplasm. GSH plays an important role in detoxifying lipid peroxides and electrophiles. Furthermore, because mitochondria play a central strategic role in the activation and mode of cell death, mitochondrial GSH has been shown to critically regulate the level of mitochondrial membrane permeability and susceptibility to secondary attacks that cause the release of proteins trapped in the intermembrane space. Once in the cytoplasm, it participates in the molecular mechanisms of cell death (see Vicent Ribas et al., Glutathione and mitochondria, Front. Pharmacol., 01 July 2014 Sec. Experimental Pharmacology and Drug Discovery, Volume 5-2014). One of its functions is that GSH can form transient disulfide bonds with cysteines present in many proteins, inhibiting protein misfolding and aggregation. Cysteine ​​is found primarily in functionally and structurally important regions of proteins and plays a critical role in protein folding and, in excess, protein aggregation. GSH binds to ("occupies") cysteine ​​residues in proteins, protecting them and preventing the formation of unnatural disulfide bonds, protecting the surrounding protein structure from damage and loss of function. Importantly, the GSH-cysteine ​​bond is reversible (see, e.g., Koji Aoyama, Int. J. Mol. Sci. 2021, 22(9), 5010). Proteins have been shown to form unnatural intramolecular disulfide bonds, leading to the formation of misfolded proteins (see, e.g., Borges et al., 2014 Jul 20;21(3):511-531). While GSH protects this cysteine ​​residue, the per-cellular pool of GSH is limited and GSH does not cross the blood-brain barrier, so once depleted, the GSH pool cannot be replenished in the event of an imbalance in redox balance in vivo.Because i) GSH pools in brain cells and other body organs are limited and ii) are known to be depleted under conditions of oxidative stress, alternative interventions are needed that provide the stability, biodistribution, and redox performance necessary to protect cellular structures and inhibit protein misfolding and aggregation. There is a need for therapeutic interventions beyond GSH that: i) protect cellular structures, such as proteins, lipids, sugars, DNA, and RNA, from reactive molecules; ii) inhibit protein misfolding and aggregation; and iii) can enter and access all cells in the body, as well as new methods for treating (1) diseases and conditions characterized by endogenous protein misfolding and / or aggregation, (2) diseases and conditions associated with mitochondrial dysfunction, and (3) diseases and conditions associated with chronic inflammation. Summary of the Invention

[0003] The present invention features PrC-210 aminothiol or analogs thereof for treating (i) diseases and conditions characterized by endogenous protein misfolding and / or aggregation, (2) diseases and conditions associated with mitochondrial dysfunction, and (3) diseases and conditions associated with chronic inflammation.

[0004] In a first aspect, the invention features a method of inhibiting protein aggregation in a subject in need thereof, the method comprising administering an effective amount of a compound of formula (I): [ka] or a pharmaceutically acceptable acid addition salt thereof, in an amount sufficient to inhibit protein or peptide aggregation, wherein (i) A is -CHNHR' and B is -CHNHR, or A = -NRR' and B = H; (ii) each of R and R' is independently selected from H, C1-C6 alkyl, and C1-C6 heteroalkyl; With the proviso that when B=H, then R and R' are not both H. In certain embodiments, the compound is [ka] or a pharmaceutically acceptable salt thereof. In some embodiments, the protein whose aggregation is inhibited is selected from TDP-43, alpha-synuclein, tau, amyloid beta, PolyQ htt, islet amyloid polypeptide, and prion, or their non-aggregating precursors. For example, the subject may have a neurodegenerative disease characterized by endogenous protein misfolding and / or aggregation. Neurodegenerative diseases that can be treated using the methods of the present invention include, but are not limited to, Parkinson's disease, prion diseases, Alzheimer's disease, multiple system atrophy, diffuse Lewy body disease, frontotemporal dementia, amyotrophic lateral sclerosis, Huntington's disease, spinocerebellar ataxias and other polyQ diseases, hereditary cerebral amyloid angiopathy, and other neurodegenerative diseases described herein. In certain embodiments, the neurodegenerative disease is a prion disease selected from Creutzfeldt-Jakob disease, variant Creutzfeldt-Jakob disease, hereditary human prion diseases, bovine spongiform encephalopathy (BSE), and scrapie. In some embodiments, the neurodegenerative disease is a synucleinopathy (e.g., Parkinson's disease (PD), dementia with Lewy bodies (DLB), and multiple system atrophy (MSA)). In certain embodiments, the subject has a disease or condition characterized by amyloidosis. The amyloidosis is selected from primary systemic amyloidosis (AL amyloidosis), reactive systemic amyloidosis (AA amyloidosis), type 2 diabetes, injection-limited amyloidosis, beta-2 microglobulin amyloidosis, hereditary non-neuropathic amyloidosis, and hereditary systemic amyloidosis of the Finnish type. In other embodiments, the subject has a disease or condition characterized by ocular protein aggregation. For example, the disease or condition characterized by ocular protein aggregation can be cataract or presbyopia.

[0005] In another aspect, the invention features a method of treating a condition associated with mitochondrial dysfunction in a subject in need thereof, the method comprising administering an effective amount of a compound of formula (I): [ka] or a pharmaceutically acceptable acid addition salt thereof, in an amount sufficient to inhibit protein or peptide aggregation, wherein (i) A is -CHNHR' and B is -CHNHR, or A = -NRR' and B = H; (ii) each of R and R' is independently selected from H, C1-C6 alkyl, and C1-C6 heteroalkyl; With the proviso that when B=H, then R and R' are not both H. In certain embodiments, the compound is [ka] or a pharmaceutically acceptable salt thereof. In some embodiments, the condition associated with mitochondrial dysfunction is a neurodegenerative disease, a neuropsychiatric disorder, diabetes, a metabolic disease, an eye disorder associated with mitochondrial dysfunction, an ischemia-related condition, aging, mitochondrial toxicity associated with a therapeutic agent, or migraine. For example, the condition associated with mitochondrial dysfunction can be a neurodegenerative disease selected from Friedreich's ataxia, amyotrophic lateral sclerosis, mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke (MELAS), myoclonic epilepsy with ragged-red fibers (MERFF), epilepsy, Parkinson's disease, Alzheimer's disease, and Huntington's disease. In some embodiments, the condition associated with mitochondrial dysfunction is a neuropsychiatric disorder selected from bipolar disorder, schizophrenia, depression, addiction, anxiety disorder, attention deficit disorder, personality disorder, autism (i.e., autism spectrum disorder), and Asperger's syndrome. In some embodiments, the method ameliorates mitochondrial toxicity of a therapeutic agent administered to a subject. In certain embodiments, the condition associated with mitochondrial dysfunction is an ophthalmic disorder associated with mitochondrial dysfunction selected from glaucoma, diabetic retinopathy, and age-related macular degeneration, hi certain embodiments, the condition associated with mitochondrial dysfunction is vascular obstruction, arteriosclerosis, valvular heart disease, tachycardia, or an ischemia-related condition resulting from hypotension or hypertension.

[0006] In another aspect, the invention features a method for treating a chronic inflammatory condition in a subject in need thereof, the method comprising administering an effective amount of a compound of formula (I): [ka] or a pharmaceutically acceptable acid addition salt thereof, in an amount sufficient to inhibit protein or peptide aggregation, wherein (i) A is —CH2NHR′ and B is - CHNHR, or A=-NRR' and B=H; (ii) each of R and R' is independently selected from H, C1-C6 alkyl, and C1-C6 heteroalkyl; With the proviso that when B=H, then R and R' are not both H. In certain embodiments, the compound is [ka] or a pharmaceutically acceptable salt thereof. In some embodiments, the chronic inflammatory condition is selected from among rhinosinusitis, arthritis, dermatitis, vasculitis, acute malaria, sickle cell disease, a gastrointestinal inflammatory condition, or an inflammatory pulmonary condition. In certain embodiments, the chronic inflammatory condition is an inflammatory pulmonary condition selected from asthma, chronic obstructive pulmonary disease (COPD), a pulmonary condition caused by physical trauma, emphysema, bronchitis, sarcoidosis, histiocytosis, lymphangioleiomyomatosis, acute lung injury, chronic lung disease, bronchopulmonary dysplasia, pneumonia, airway exacerbation, and acute respiratory distress syndrome (ARDS). In some embodiments, the chronic inflammatory condition is a gastrointestinal inflammatory condition selected from inflammatory bowel disease (IBD), ulcerative colitis (UC), Crohn's disease (CD), and colitis. In some embodiments, the chronic inflammatory condition is an autoimmune disease. For example, the methods of the present invention can be used to treat an autoimmune disease selected from multiple sclerosis, type 1 diabetes, rheumatoid arthritis, juvenile rheumatoid arthritis, psoriatic arthritis, pancreatitis, psoriasis, plaque psoriasis, guttate psoriasis, inverse psoriasis, pustular psoriasis, erythrodermic psoriasis, paraneoplastic autoimmune disease, autoimmune hepatitis, bullous pemphigoid, myasthenia gravis, inflammatory bowel disease, Crohn's disease, ulcerative colitis, celiac disease, thyroiditis, Sjögren's syndrome, Guillain-Barré disease, Raynaud's phenomenon, Addison's disease, primary biliary cirrhosis, primary sclerosing cholangitis, nonalcoholic fatty liver disease, nonalcoholic steatohepatitis, and diabetes. In some embodiments, the chronic inflammation is associated with atherosclerosis, hardened or leaky heart valves, arterial endothelial damage, vascular damage during dialysis, and lipid deposition, such as in metabolic syndrome. In certain embodiments, the chronic inflammatory condition is a neuroinflammatory disease. Neuroinflammatory diseases treatable using the methods of the present invention include multiple sclerosis, neuromyelitis optica, anti-myelin oligodendrocyte glycoprotein antibody disease, autoimmune encephalitis, transverse myelitis, optic neuritis, neurosarcoidosis, but also all neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, Huntington's disease, etc.

[0007] definition To facilitate understanding of the present invention, a number of terms are defined below. Terms defined herein have meanings commonly understood by one of ordinary skill in the art relevant to the present invention. Terms such as "a," "an," and "the" are not intended to refer solely to a singular entity, but rather encompass the general type for which a specific example may be used for illustration. While terms herein are used to describe particular embodiments of the present invention, their use does not limit the invention except as defined in the claims.

[0008] As used herein, the term "about" refers to a value that is within 10% above or below the stated value.

[0009] As used herein, any value provided in a range of values ​​includes both the upper and lower limits, and any value subsumed within those limits.

[0010] As used herein, the term "pharmaceutically acceptable salt" refers to those salts of the described compounds that are suitable for use in contact with human and animal tissues without undue toxicity, irritation, allergic response, etc., within the normal scope of sound medical judgment, and commensurate with a reasonable benefit / risk ratio. Pharmaceutically acceptable salts are well known in the art. For example, pharmaceutically acceptable salts are described in Berge et al., J. Pharmaceutical Sciences 66:1-19, 1977, and Handbook of Pharmaceutical Salts: Properties, Selection, and Use, (Eds. P.H. Stahl and C.G. Wermuth), Wiley-VCH, 2008. These salts may be acid addition salts, including inorganic or organic acids. Salts may be prepared in situ during the final isolation and purification of the compounds described herein, or separately, by reacting the free base group with a suitable acid.

[0011] As used herein, the term "effective amount" refers to an amount sufficient to achieve a beneficial or desired result, e.g., a clinical result; therefore, a "therapeutically effective amount" depends on the context in which it is applied. For example, in the context of administering a compound disclosed herein to inhibit protein misfolding, protein aggregation, or peptide aggregation, an effective amount of the compound is, for example, an amount sufficient to ameliorate the condition or progression of a disease characterized by such misfolding or aggregation. In the context of treating a condition characterized by mitochondrial dysfunction, an effective amount of the compound is, for example, an amount sufficient to ameliorate the condition or progression of a disease characterized by such mitochondrial dysfunction. In the context of treating a condition characterized by chronic inflammation, an effective amount of the compound is, for example, an amount sufficient to ameliorate the condition or progression of a disease characterized by chronic inflammation. The methods of the present invention can include systemic administration (e.g., intravenous) or local administration (e.g., topical application or local injection) depending on the nature of the condition being treated.

[0012] As used herein, and as is well understood in the art, "treating" a condition or "treatment" of various diseases or disorders is an effort to achieve a beneficial or desired result, e.g., a clinical outcome. Beneficial or desired results may include, but are not limited to, alleviation of one or more symptoms or conditions; a reduction in the severity of the disease, disorder, or condition; a stable (i.e., not worsening) state of the disease, disorder, or condition; a delay or slowing of the progression of the disease, disorder, or condition; an improvement or palliation of the disease, disorder, or condition; and a remission that may be detectable or undetectable (either partial or complete). "Alleviating" a disease, disorder, or condition means that the severity and / or undesirable clinical signs of the disease, disorder, or condition are reduced and / or the time course of progression is slowed or prolonged compared to the severity or time course in the absence of treatment.

[0013] As used herein, the term "subject" may be a human, a non-human primate, or other mammal, such as, but not limited to, a dog, cat, horse, cow, pig, goat, monkey, rat, mouse, and sheep. In a preferred embodiment, the subject is a human.

[0014] As used herein, the term "pharmaceutical composition" refers to an active compound formulated with one or more pharmaceutically acceptable excipients. In some embodiments, the compounds of the present invention are present in a unit dose suitable for administration in a treatment regimen that exhibits a statistically significant probability of achieving a predetermined therapeutic effect when administered to an appropriate population. In certain embodiments, pharmaceutical compositions may be specially formulated for administration in solid or liquid form, including those suitable for oral administration, such as drenches (aqueous or non-aqueous solutions or suspensions), tablets, or capsules; and parenteral administration, such as parenteral administration by subcutaneous, intramuscular, or intravenous injection.

[0015] As used herein, the term "pharmaceutically acceptable excipient" refers to any inert ingredient (e.g., a vehicle capable of suspending or dissolving an active compound) that is biocompatible and suitable for administration to a subject. Representative excipients include, for example, anti-adherents, antioxidants, binders, coating agents, compression aids, disintegrants, dyes, humectants, emulsifiers, diluents, film-forming or coating agents, flavors, fragrances, glidants, lubricants, preservatives, printing inks, adsorbents, suspending or dispersing agents, sweeteners, or hydration water. Excipients include, but are not limited to, optionally substituted butylated hydroxytoluene (e.g., BHT), calcium carbonate, calcium phosphate (dibasic), calcium stearate, croscarmellose, cross-linked polyvinylpyrrolidone, citric acid, crospovidone, cysteine, ethylcellulose, gelatin, optionally substituted hydroxypropylcellulose, optionally substituted hydroxypropylmethylcellulose, lactose, magnesium stearate, maltitol, mannitol, methionine, methylcellulose, methylparaben, microcrystalline cellulose, polyethylene glycol, polyvinylpyrrolidone, povidone, pregelatinized starch, propylparaben, retinyl palmitate, shellac, silicon dioxide, sodium carboxymethylcellulose, sodium citrate, sodium starch glycolate, sorbitol, starch, stearic acid, stearic acid, sucrose, talc, titanium dioxide, vitamin A, vitamin E, vitamin C, and xylitol. Those skilled in the art are familiar with the variety of agents and substances useful as excipients.

[0016] As used herein, the term "alkyl" refers to a branched or straight-chain monovalent saturated aliphatic radical that, when unsubstituted, contains only C and H. A monovalent alkyl group does not include any substituents on the alkyl group. For example, when an alkyl group is attached to a compound, the monovalent alkyl means that it is attached to the compound and does not include any additional substituents that may be present on the alkyl group. In some embodiments, an alkyl group can contain, for example, 1 to 6, 1 to 4, or 1 to 2 carbon atoms (e.g., C1-C6, C1-C4, or C1-C2). Examples include, but are not limited to, methyl, ethyl, isobutyl, sec-butyl, tert-butyl, 2-methylpropyl, and 2,2-dimethylpropyl.

[0017] As used herein, the term "C1-C6 heteroalkyl" refers to a branched or straight-chain monovalent saturated alkyl radical containing C, H, and 1 to 3 N atoms.

[0018] Other features and advantages of the invention will be apparent from the following detailed description, the drawings and the claims. [Brief explanation of the drawings]

[0019] [Figure 1] Figures 1(A) and 1(B) show the H NMR spectra of PrC-210 after 24 hours in deuterium oxide at pH 3.0 (Figure 1a) or pH 9.1 (Figure 1b). These spectra show that PrC-210 does not change charge conformation and is stable in this pH range for at least 24 hours. [Figure 2] 1H NMR spectra of PrC-210 in deuterium oxide at pH 9.0 are shown at 1 hour, 24 hours, and 72 hours. These spectra show that PrC-210 exhibits some instability at high pH after 72 hours. [Figure 3] 1 is a graph showing that the pKa of two amines of PrC-210 is much higher (ie, 3 pH units) than physiological pH (7.2). [Figure 4]Two H NMR spectra of PrC-210 in deuterium oxide containing ammonium acetate at pH 7.0 are shown at 1 hour and 24 hours. This figure shows that PrC-210 begins a substantial conformational change within a few hours, likely indicating the formation of a disulfide bond between two PrC-210 molecules in the presence of acetate anions. [Figure 5] H NMR spectra of PrC-210 in deuterium oxide containing ammonium acetate and the thiol blocker TCEP at pH 7.0 at 1 hour and 24 hours are shown. This figure shows that PrC-210 in the presence of TCEP is stable for 24 hours in the presence of acetate anions. [Figure 6] Figure 6(A): A graph showing the half-life of PrC-210 in acetate buffer at pH 7.2 is 3.5 hours, whereas the half-life of PrC-210 in water was previously observed to be several days. Figure 6(B): A reaction scheme illustrating the reaction between anionic ammonium acetate and PrC-210 thiol, which triggers the formation of a PrC-210-PrC-210 disulfide bond. Briefly, (1) the negatively charged free electron pair of acetate attacks the thiol proton on PrC-210, (2) the deprotonated activated thiolate anion on the PrC-210 molecule can then (3) react with the PrC-210 thiol molecule to form a PrC-210 dimer. Figure 6(C): An image showing the disulfide-forming reaction of Figure 6(B) in the presence of a protein with one or more cysteine ​​residues, such as TDP-43. [Figure 7] This table shows the recovery of

[14] C-labeled PrC-210 after intravenous administration to male rats. After 48 hours, only 1.07% of PrC-210 was metabolized. [Figure 8] This image shows that the consumption of PrC-210 thiol by added 0.5 mM hydrogen peroxide is significantly increased ("accelerated") in a dose-dependent manner by adding ammonium acetate a few seconds before the hydrogen peroxide. [Figure 9]Figure 9 shows the levels of markers (A) 8-oxo-2'-deoxyguanosine (8-oxo-dG), (B) p53, (C) cytochrome C, (D) caspase 8, and (E) caspase 3 / 7 in the brains of mice irradiated with 8.68 Gy. (F) Caspase 3 / 7 in mouse plasma after 8.68 Gy irradiation. Caspases are increased without PrC-210 treatment and are suppressed to background levels by administration of PrC-210 with 0.5 MTD PrC-210 30 minutes before 8.68 Gy irradiation. [Figure 10] Figure 1 shows the time course of body weight in three SOD1G93A mice. Weight loss progressed with the onset of ALS motor symptoms and the appearance of hindlimb paralysis. Systemic administration of PrC-210 (0.1 MTD, IP) was associated with an immediate plateau of weight loss. [Figure 11] Figure 11 (A) shows the levels of cytochrome C and (B) shows the levels of caspase 3 / 7 markers in mouse hearts after 8.68 Gy of irradiation. PrC-210 (0.3 MTD) was administered 24 hours after 8.68 Gy of irradiation. [Figure 12] Figure 12 shows (A) caspase 1, (B) complement 3, and (C) caspase 3 / 7 marker levels in mouse brains after 8.68 Gy of irradiation. PrC-210 (0.3 MTD) was administered 24 hours after 8.68 Gy of irradiation. [Figure 13] Figure 13(A): Paralysis scores in EAE mouse model, where intraperitoneal PrC-210 administration was initiated twice weekly 24 hours before immunization with MOG peptide. Figure 13(B): Intraperitoneal PrC-210 administration was initiated twice weekly when the first paralysis was observed. [Figure 14] Paw swelling scores are shown in DBA / 1J mice, where i) twice-weekly intraperitoneal PrC-210 administration was initiated with initial paw status, and ii) topical application of 370 mM PrC-210 was initiated when paw swelling scores exceeded 10. DETAILED DESCRIPTION OF THE INVENTION

[0020] This invention features the use of aminothiols to protect cellular structures from damage and inhibit protein misfolding and aggregation in subjects in need thereof. Applicant has discovered that the aminothiols of the invention have an accelerated ability to form disulfide bonds with cysteine ​​residues of endogenous proteins in the presence of negative charges, inhibiting the formation of Cys-Cys disulfide bonds in intracellular proteins and the associated protein misfolding and aggregation, among many other cellular proteins.

[0021] Aminothiol Aminothiols useful in the methods of the present invention can be synthesized, for example, as described in US Pat. No. 7,314,959.

[0022] Administration The dosage of the compounds disclosed herein depends on factors such as the route of administration, the disease to be treated, and the age, weight, and general health of the subject. Typically, the amount of the compounds disclosed herein (e.g., PrC-210) can be an amount that effectively treats the disease without causing serious toxicity, whether administered as a single dose or multiple times over a long period of time. The dosage can be adjusted by a clinician based on conventional factors, such as the extent of the disease and various parameters of the subject. Generally, the pharmaceutical compositions disclosed herein can be administered in an amount of about 0.001 mg to about 500 mg / kg / day of an aminothiol, such as PrC-210.

[0023] treatment method The present invention relates to the systemic administration of PrC-210 aminothiol or its analogs, which protect proteins, lipids, sugars, RNA, and DNA, and protect cysteines located within proteins, thereby preventing protein misfolding and aggregation and significantly inhibiting various disease states. The present invention demonstrates that the two amines of the aminothiol PrC-210 are both positively charged within a very wide physiological pH range, which indicates that they are strongly attracted to the negative charges of proteins, lipids, sugars, RNA, and DNA through electrostatic interactions. DNA, RNA, lipids, sugars, proteins, lipoproteins, and glycoproteins all have several negative charges on their surfaces. The two positive charges of PrC-210 are attracted to the negative charges on these cellular biomolecules by charge complementarity. The thiol group of PrC-210 exhibits very low reactivity in water, where there is no negative charge, and is very stable as a monomeric aminothiol. However, in the present invention, we have shown that electrostatic interactions between PrC-210 and nearby negative charges significantly alter the stability of the aminothiol, increasing ("activating") the thiol group of PrC-210. This "activated thiol" reacts very readily with other thiols or reactive structures. Furthermore, the activated aminothiol of the present invention forms disulfide bonds with cysteine ​​residues of endogenous proteins, indicating that PrC-210 reacts with other cellular proteins, suppressing the formation of Cys-Cys disulfide bonds within proteins and preventing the misfolding and aggregation of associated proteins.

[0024] The present invention features the administration of PrC-210 aminothiol or an analog thereof to treat (i) diseases and conditions characterized by misfolding and / or aggregation of endogenous proteins, (2) diseases and conditions associated with mitochondrial dysfunction, and (3) diseases and conditions associated with chronic inflammation.

[0025] Neurodegenerative diseases The methods of the present invention can be used to treat neurodegenerative diseases characterized by the misfolding and / or aggregation of endogenous proteins, such as Parkinson's disease, prion diseases, Alzheimer's disease, multiple system atrophy, diffuse Lewy body disease, frontotemporal dementia, amyotrophic lateral sclerosis, Huntington's disease, spinocerebellar ataxias and other polyQ diseases, and hereditary cerebral amyloid angiopathy.

[0026] In some embodiments, the prion disease to be treated is selected from Creutzfeldt-Jakob disease, variant Creutzfeldt-Jakob disease, inherited human prion diseases, bovine spongiform encephalopathy (BSE), and scrapie.

[0027] In certain embodiments, the neurodegenerative disease to be treated is a synucleinopathy. Synucleinopathy is characterized by the intracellular accumulation of protein aggregates, oligomers, profibrils, and fibrils, primarily containing α-synuclein. In synucleinopathy, the pathological effects on neurons are thought to be caused by the formation of α-synuclein oligomeric aggregates and subsequent membrane pore formation. Examples of synucleinopathy include Parkinson's disease (PD), dementia with Lewy bodies (DLB), and multiple system atrophy (MSA).

[0028] The methods of the present invention can be used to treat conditions associated with amyloidosis, a general term that refers to several diseases characterized by the presence of a pathological form of amyloid protein, often involving the extracellular deposition of protein fibrils, forming numerous "amyloid deposits" or "amyloid plaques" that can occur locally or systemically.

[0029] In some embodiments, the amyloidosis being treated is selected from primary systemic amyloidosis (AL amyloidosis), reactive systemic amyloidosis (AA amyloidosis), type 2 diabetes, injection-limited amyloidosis, beta-2 microglobulin amyloidosis, hereditary non-neuropathic amyloidosis, and hereditary systemic amyloidosis of the Finnish type. Ocular protein aggregation disease

[0030] The methods of the present invention can be used to treat ocular diseases and conditions characterized by endogenous protein misfolding and / or aggregation, including cataracts and presbyopia.

[0031] Presbyopia refers to the loss of the eye's ability to accommodate, resulting in the inability to focus on nearby objects. Stiffening of the ocular lens and changes in the elasticity of the lens capsule are common causes of presbyopia. Therefore, pharmacological treatments that can prevent or reverse crystallin stiffening offer promising new, non-invasive treatments for presbyopia. At the molecular level, a protein known as crystalline plays a key role in the stiffening of the ocular lens. Lens crystallins, including three isoforms—a, b, and g—account for 90% of the ocular lens's protein content. Crystallin (AC), an ATP-independent chaperone and a member of the small heat shock protein (sHsp) family, accounts for 40% of the crystallin protein content. It exists as a hetero-oligomer of two subunits, αA-crystallin (AAC) and αB-crystallin (ABC), whose expression is primarily restricted to the ocular lens. It recognizes exposed conformational features of partially unfolded lens proteins and segregates them from one another, thereby reducing the number of aggregation-prone species that would otherwise cause various age-related vision defects. The conversion of soluble AC into HMW aggregates significantly increases lens stiffness. Mitochondrial dysfunction

[0032] The methods of the present invention can be used to treat diseases and conditions characterized by mitochondrial dysfunction. Conditions associated with mitochondrial dysfunction can result in the progressive appearance of a mosaic of cells with defective electron transport activity in muscle, with cells largely lacking cytochrome c oxidase (COX) activity randomly interspersed among cells with normal activity, resulting in a high incidence of COX-negative cells in biopsies (e.g., in elderly individuals). During the aging process or under various conditions associated with mitochondrial dysfunction, organisms face the need to preserve and maintain some degree of function in irreplaceable post-mitotic cells (e.g., neurons, skeletal muscle, cardiac muscle), which inevitably face a progressive decline in mitochondrial respiratory chain function. Neurons with dysfunctional mitochondria become increasingly sensitive to external stimuli such as excitotoxicity. Mitochondrial dysfunction significantly contributes to age-related degenerative diseases, particularly neurodegeneration. Congenital mitochondrial diseases often involve early-onset neurodegeneration, the underlying mechanisms of which are similar to disorders that occur with aging in people born with normal mitochondria. The methods and compositions of the present invention allow for the amelioration of dysfunctional mitochondria in these irreplaceable post-dividing cells, allowing for the rescue or improvement of mitochondrial function in these cells.

[0033] Conditions associated with mitochondrial dysfunction include those in which defects in the activity of the mitochondrial respiratory chain contribute to the pathophysiology of these diseases or disorders in mammals, including (1) congenital genetic defects in the activity of one or more components of the mitochondrial respiratory chain or electron transport chain, and (2) acquired defects in the level or activity of one or more components of the mitochondrial respiratory chain, caused by (a) oxidative damage due to aging, (b) increased intracellular calcium, (c) exposure of the target cells to nitric oxide, (d) hypoxia or ischemia, (e) microtubule-associated defects in axonal transport of mitochondria, or (f) expression of mitochondrial uncoupling proteins.

[0034] Common conditions of mitochondrial dysfunction include cardiomyopathy, muscle weakness and atrophy, developmental delay (related to motor, language, cognitive, or executive function), ataxia, epilepsy, renal tubular acidosis, peripheral neuropathy, optic neuropathy, autonomic neuropathy, neurogenic bowel dysfunction, sensorineural hearing loss, neurogenic bladder dysfunction, dilated cardiomyopathy, migraine, liver failure, lactic acidosis, and diabetes. Neuropsychiatric disorders

[0035] The brain requires ten times more energy than the rest of the body on average. Many neuropsychiatric disorders may be specifically related to abnormalities in energy production or mitochondrial dysfunction. Neuropsychiatric disorders include, but are not limited to, bipolar disorder (BD), schizophrenia, depression, anxiety disorders, attention deficit disorder, addiction, personality disorders, autism, and Asperger's syndrome. The methods and compositions of the present invention can be used to treat neuropsychiatric disorders. Neurodegenerative diseases

[0036] The methods and compositions of the present invention can be used to treat neurodegenerative diseases. Many progressive neurological diseases are caused by the death of neurons due to mitochondrial apoptosis. Friedreich's ataxia is caused by a genetic defect in the frataxin gene, which is involved in mitochondrial iron transport (Babcock et al., Science 276:1709 (1997)). Human deafness dystonia is caused by a defect in a small component of the mitochondrial protein import apparatus (Koehler et al., Proc. Natl. Acad. Sci. USA 96:2141 (1999)). One of the characteristic causes of amyotrophic lateral sclerosis is a deficiency of Cu-Zn superoxide dismutase, which is present in the mitochondrial intermembrane space and cytoplasm (Deng et al., Science 261:1047 (1993)). The discovery that multiple environmental toxins cause Parkinsonism by inhibiting respiratory chain complex I and promoting the production of reactive oxygen species has made this complex the focus of research into the basis of Parkinson's disease (Dawson et al., Science 302:819 (2003)). Recently, the mitochondrial protein encoded by PINK1 provided a direct link between mitochondria and Parkinson's disease (Valente et al., Science 304:1158 (2004)). Alzheimer's disease is also associated with mitochondrial toxicity through the mitochondrial protein ABAD, a target of amyloid (Lustbader et al., Science 304:448 (2004)). Huntington's disease is associated with defects in energy metabolism, which are widespread and affect both the brain and peripheral tissues, resulting from mitochondrial dysfunction (Leegwater-Kim et al., NeuroRx 1:128 (2004)). Fundamental abnormalities involved in the pathogenesis of bipolar disorder (BD) are thought to involve energy production, particularly mitochondrial activity.Evidence from a variety of sources, including postmortem, genetic, brain imaging, and peripheral cell studies, supports the idea that energy deficiency and mitochondrial dysfunction are important factors in the development of BD (see Hough et al., Bipolar Disord. 2:145 (2000); Fattal et al., Psychosomatics 47:1 (2006); and Kato et al., Bipolar Disord. 2:180 (2000)).

[0037] Diabetes and metabolic diseases The methods and compositions of the present invention can be used to treat diabetes and metabolic diseases. The central role of mitochondria in carbohydrate and fatty acid metabolism confers a critical function for this organelle in diabetes (Maechler et al., Nature 414:807 (2001)). Knockout mice of abundant mitochondrial transcription factors have provided a model of 13-cell loss in childhood diabetes (Silva et al., Nat. Genet. 26:335 (2000)). Mutations in PPARγ, the master regulator of mtDNA and mitochondrial biogenesis, correlate with type 2 diabetes. Insulin release depends on mitochondrial function, which is affected by the expression of the membrane transporter UCP2 (Petersen et al., Science 300:1140 (2003); Zhang et al., Cell 105:745 (2001)). The antidiabetic action of thiazolidinediones appears to depend on their ability to function as ligands for PPARγ and its coactivator PGC-1, which regulates the expression of nuclear genes for mitochondrial gene products (Mootha et al., Nature Genet 34:267 (2003); Puigserver et al., Endocr. Rev. 24:78 (2003)).

[0038] migraine The methods and compositions of the present invention can be used to treat migraines. Metabolic studies on patients with recurrent migraines have shown that defects in mitochondrial activity are commonly associated with this disorder, manifesting as impaired oxidative phosphorylation and excessive lactate production. Such defects are not necessarily due to genetic defects in mitochondrial DNA. Migraine patients are hypersensitive to nitric oxide, an endogenous inhibitor of cytochrome c oxidase. Furthermore, patients with mitochondrial cytopathies, such as MELAS, often experience recurrent migraines.

[0039] Eye disorders associated with mitochondrial dysfunction The methods and compositions of the present invention can be used to treat ocular disorders such as glaucoma, diabetic retinopathy, and age-related macular degeneration (AMD). Retinal damage is due to reactions caused by free radicals in glaucoma, diabetic retinopathy, and age-related macular degeneration (AMD). The eye is part of the central nervous system and has limited regenerative capacity. The retina is composed of numerous nerve cells that are subject to oxidation and have the highest concentration of polyunsaturated fatty acids (PFAs). Free radicals are generated by mitochondria in rods and cones, which generate the energy necessary to convert ultraviolet light entering the eye into visual impulses. Free radicals cause peroxidation of PFAs by hydroxyl radicals or superoxide radicals, resulting in the propagation of additional free radicals. Free radicals can cause temporary or permanent damage to retinal tissue.

[0040] Glaucoma is typically considered a disorder in which intraocular pressure (IOP) is elevated, causing permanent damage to retinal nerve fibers; however, one-sixth of all glaucoma cases do not result in elevated IOP. The disorder is now recognized as a combination of decreased vascular perfusion and increased neurotoxic factors. Recent studies suggest that elevated levels of glutamate, nitrogen dioxide, and paroxynitrite in the eye may be responsible for retinal ganglion cell death.

[0041] Diabetic retinopathy occurs when the underlying blood vessels develop microvascular abnormalities, primarily consisting of microaneurysms and intraretinal hemorrhages. Oxidative metabolites are directly involved in the pathogenesis of diabetic retinopathy, and free radicals enhance the production of growth factors, leading to increased proliferative activity. Nitric oxide produced by vascular endothelial cells can relax smooth muscle cells and cause segmental dilation of blood vessels. Retinal ischemia and hypoxia occur after thickening of the arterial basement membrane, endothelial proliferation, and loss of pericytes. Inadequate oxygen supply leads to capillary blockage or no blood flow, arteriovenous shunting, sluggish blood flow, and a reduced ability of red blood cells to release oxygen. Lipid peroxidation of retinal tissue also occurs as a result of free radical damage.

[0042] ischemia-related conditions The methods and compositions of the present invention can be used to treat ischemia-related conditions. Oxygen deprivation directly inhibits mitochondrial respiratory chain activity by depriving cells of the terminal electron acceptor for the reoxidation of cytochrome c at complex IV, and indirectly affects the activity of the mitochondrial respiratory chain through secondary deoxidative excitotoxicity and nitric oxide production, particularly in the nervous system. In conditions such as cerebral deoxidation and angina pectoris, tissues are relatively hypoxic. In such cases, increased mitochondrial activity protects affected tissues from the harmful effects of hypoxia, inhibits secondary delayed cell death, and accelerates recovery from hypoxic tissue stress and injury. The methods and compositions of the present invention can be useful, for example, in preventing delayed cell death (apoptosis, which occurs in regions such as the hippocampus or cortex approximately 2 to 5 days after an episode of cerebral ischemia) following ischemia or hypoxia in the brain.

[0043] Muscle function The methods and compositions of the present invention can be used to improve muscle performance. For example, the methods and compositions of the present invention may be useful for improving physical endurance (e.g., the ability to perform physical tasks such as exercise, manual labor, sports activities, etc.), inhibiting or delaying physical fatigue, improving blood oxygen levels, increasing energy in healthy individuals, improving work capacity and endurance, reducing muscle fatigue, reducing stress, improving cardiac and cardiovascular function, improving sexual performance, increasing muscle ATP levels, and / or reducing blood lactate.

[0044] Improved athletic performance, muscle strength, speed, and endurance are typically measured by increased muscle contractile force, increased contraction amplitude, decreased muscle reaction time between stimulation and contraction, the ability to overcome muscle fatigue, and the ability to sustain prolonged activity. Apart from muscle performance during endurance exercise, parameters of free radical and oxidative stress are affected in pathological conditions. Substantial data now suggest that oxidative stress contributes to muscle wasting or atrophy in pathophysiological conditions (see Clarkson, Crit. Rev. Food Sci. Nutr. 35:31 (1995); and Powers et al., Proc. Nutr. Soc. 58:1025 (1999)). For example, in muscular dystrophies, defects in the dystrophin-glycoprotein complex (DGC) suggest that one mechanism of cellular injury is functional ischemia associated with alterations in cellular NOS and disruption of NO's normal protective role. Rando (Microsc. Res. Tech. 55:223 (2001)) has shown that oxidative damage precedes pathological changes and that muscle cells with DGC defects are more susceptible to oxidant challenge. Excessive lipid peroxidation by free radicals has been shown to be a contributing factor in muscle diseases such as McArdle disease (see Russo et al., Med. Hypotheses. 39:147 (1992)). Furthermore, mitochondrial dysfunction is a well-known correlate of age-related muscle decline (sarcopenia), and free radical damage has been proposed as a contributing factor but is less frequently investigated (see Navarro et al., Front. Biosci. 6:D26 (2001)). Other indications include acute sarcopenia, e.g., muscle wasting and / or cachexia associated with burns, bed rest, limb immobilization, or major thoracic, abdominal, and / or orthopedic surgery. The methods of the present invention can be effective in treating muscle-related pathological conditions.

[0045] Aging The methods and compositions of the present invention can be used to treat aging and its associated conditions. During normal aging, there is a progressive decline in mitochondrial respiratory chain function. Beginning at approximately age 40, the accumulation of mitochondrial DNA defects in humans increases exponentially, accompanied by a concomitant decline in nuclear regulatory elements of mitochondrial respiratory activity. Many mitochondrial DNA lesions confer a selective advantage during mitochondrial turnover, particularly in postmitotic cells. The proposed mechanism is that mitochondria with defective respiratory chains incur less oxidative damage to themselves than mitochondria with normally functioning respiratory chains (mitochondrial respiration is the primary source of free radicals in the body). Thus, normally functioning mitochondria accumulate oxidative damage to membrane lipids more quickly than defective mitochondria, and are therefore "tagged" for degradation by autophagy and the lysosomal system. The relentless decline in mitochondrial function with age contributes to aging-related conditions associated with neurodegeneration and type 2 diabetes. Oxidative stress is thought to contribute to general aging as well as to underlie some of these specific diseases (Harman, Proc. Natl. Acad. Sci. USA 78:7124 (1981)). Mutations in C. elegans and D. melanogaster that reduce mitochondrial oxidative stress have been shown to extend lifespan in these organisms (Hekimi et al., Science 299:1351 (2003)). Furthermore, mammals maintained on a calorie-restricted diet experience a reduced metabolic rate, which may contribute to a significant increase in lifespan. Numerous studies document an increase in point mutations and deletions in mtDNA with aging. Furthermore, Trifunovic et al. recently showed that mice engineered to express an error-prone mitochondrial DNA polymerase serve as an excellent model of premature aging (Nature 429:417 (2004)). The methods of the present invention can be used to treat progeria, Werner's syndrome, Wiedemann-Rautenstrauch syndrome, and conditions of aging (e.g., wrinkles, forgetfulness, arthropathy, age-related muscle wasting).

[0046] In certain embodiments, the methods of the invention are used to treat a disease characterized by mitochondrial dysfunction selected from a neurodegenerative disease (e.g., Friedreich's ataxia, amyotrophic lateral sclerosis, mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke (MELAS), myoclonic epilepsy with red fibers (MERFF), epilepsy, Parkinson's disease, Alzheimer's disease, or Huntington's disease), a neuropsychiatric disease (e.g., bipolar disorder, schizophrenia, depression, addiction, anxiety disorder, attention deficit disorder, personality disorder, autism, or Asperger's syndrome), diabetes, a metabolic disease, an eye disorder associated with mitochondrial dysfunction (e.g., glaucoma, diabetic retinopathy, or age-related macular degeneration), an ischemia-related condition (e.g., a condition resulting from vascular occlusion, arteriosclerosis, valvular heart disease, tachycardia, or hypotension), aging, therapeutic drug-associated mitochondrial toxicity, or migraine. chronic inflammation

[0047] The methods and compositions of the present invention can be used to treat chronic inflammation and conditions associated therewith. Exemplary inflammatory conditions treatable using the methods of the present invention include asthma (e.g., aspirin-sensitive / exacerbated asthma, atopic asthma, severe asthma, mild asthma, moderate-to-severe asthma, corticosteroid-naive asthma, chronic asthma, corticosteroid-resistant asthma, corticosteroid-refractory asthma, newly diagnosed and untreated asthma, smoking-related asthma, and corticosteroid-uncontrolled asthma), airway hyperresponsiveness, and airway inflammation. Hypersensitivity, sinusitis, sinusitis with polyps, nasal polyposis, arthritis (e.g., osteoarthritis, rheumatoid arthritis, arthritis as a result of trauma, etc.), seronegative enthesopathy and arthropathy (SEA) syndrome, acute malaria, sickle cell disease, osteoporosis, eosinophilic esophagitis, dermatitis, atopic dermatitis, allergic rhinitis, bullous pemphigoid, chronic urticaria, cartilage inflammation, polymyalgia rheumatica, polyarteritis nodosa nodossa, Wegener's granulomatosis, Behçet's disease, myositis, polymyolitis, dermatomyositis, vasculitis, arteritis, diabetic nephropathy, interstitial cystitis, gastrointestinal inflammatory conditions (e.g., inflammatory bowel disease (IBD), ulcerative colitis (UC), Crohn's disease (CD)), colitis (e.g., colitis caused by environmental insults)insults) (e.g., caused by or associated with therapeutic regimens such as chemotherapy, radiation therapy, etc.), chronic hepatitis (e.g., viral hepatitis, alcoholic hepatitis, chronic hepatitis (e.g., viral hepatitis, alcoholic hepatitis, non-alcoholic steatohepatitis, etc.), ischemic colitis, collagen disease or lymphocytic colitis, colitis in conditions such as chronic granulomatous disease or celiac disease, food allergy, gastritis, inflammatory lung disease (e.g., chronic obstructive pulmonary disease (COPD))), eosinophilic lung inflammation, infection-induced lung conditions (e.g., viral (e.g., influenza, parainfluenza, rotavirus, human metapneumococcal disease, etc.), Examples of chronic inflammatory conditions include those associated with viral, fungal (e.g., Aspergillus), parasitic, or prion infections, allergen-induced pulmonary conditions, pollutant-induced pulmonary conditions (e.g., asbestosis, silicosis, or beryllium disease), gastric aspiration-induced pulmonary conditions, immune dysregulation, genetically predisposed inflammatory conditions such as cystic fibrosis, physical trauma-induced pulmonary conditions (e.g., respiratory injury), emphysema, bronchitis, sarcoidosis, histiocytosis, lymphangioleiomyomatosis, acute lung injury, chronic lung disease, bronchopulmonary dysplasia, pneumonia (e.g., community-acquired pneumonia, hospital-acquired pneumonia, ventilator-associated pneumonia, and severe pneumonia), airway exacerbation, and acute respiratory distress syndrome (ARDS). Other examples of chronic inflammatory conditions include diseases involving neuroinflammation, such as multiple sclerosis and neurodegenerative disorders.

[0048] autoimmune disease The methods and compositions of the present invention can be used to treat autoimmune diseases, including multiple sclerosis, type 1 diabetes, rheumatoid arthritis, juvenile rheumatoid arthritis, psoriatic arthritis, pancreatitis, psoriasis, plaque psoriasis, guttate psoriasis, inverse psoriasis, pustular psoriasis, erythrodermic psoriasis, paraneoplastic autoimmune diseases, autoimmune hepatitis, bullous pemphigoid, myasthenia gravis, inflammatory bowel disease, Crohn's disease, ulcerative colitis, celiac disease, thyroiditis (e.g., Graves' disease), Sjogren's syndrome, Guillain-Barré disease, Raynaud's phenomenon, Addison's disease, liver diseases (e.g., primary biliary cirrhosis, primary sclerosing cholangitis, non-alcoholic fatty liver disease, and non-alcoholic steatohepatitis), and diabetes (e.g., type 1 diabetes). Related States

[0049] The methods and compositions of the present invention can be used to treat conditions complicated by one or more of encephalitis, meningitis, nephritis, myocarditis, pneumonia, necrosis, non-healing wounds, chronic hepatitis, intrinsic liver or organ failure, acne, inflammatory bowel disease, ulcerative colitis, Crohn's disease, psoriasis, vasculitis, glomerulonephritis, pancreatitis, tendinopathy, thyroiditis, myasthenia gravis, pernicious anemia, dermatomyositis, Sjogren's syndrome, aplastic anemia, celiac disease, eczema, arteriosclerosis, hypertension, myocardial ischemia, neuronal ischemia, renal ischemia, chronic heart failure and cardiac hypertrophy, cardiomyopathy, chronic kidney disease, coronary heart disease, stroke, aortic disease, varicose veins, angina pectoris, organ failure (e.g., renal, liver, and / or pulmonary failure), fatty streak formation, atheroma, atherosclerotic plaques, fetal growth restriction, and hypertension of pregnancy.

[0050] The following examples are intended to illustrate the invention, but are not intended to limit the invention in any way. [Example]

[0051] Example 1: Mode of action of the invention We established the chemical structure profile of PrC-210 using nuclear magnetic resonance (NMR) spectroscopy experiments. When titrated in water from pH 3 to 9.1, PrC-210 remained stable and did not change its charge structure (see Figures 1a and 1b). The structural stability of the monothiol form of PrC-210 in this pH range persisted for at least the observed 24 h. Significant changes were observed after 72 h at pH 9.0 (Figure 2). Addition of the thiol blocker TCEP to water did not affect the stability of PrC-210 at pH 9.0 for 72 h (data not shown), implying that the thiol group is stable and unreactive (or "unactivated") under these conditions. Upon pH titration between 9.0 and 13.0, distinct structural changes were observed in PrC-210. From these gradual changes, we calculated that the pKas of the two amines in PrC-210 are all significantly above physiological pH (pH 7.2) (Figure 3).

[0052] Interestingly, upon addition of ammonium acetate, which contains an anionic charge, at pH 7, PrC-210 initiated a significant conformational change within a few hours (Figure 4), indicating the formation of a disulfide bond between two PrC-210 molecules. We confirmed this hypothesis by adding the thiol blocker TCEP to the ammonium acetate incubation, and single PrC-210 was again stable for 24 hours (Figure 5). In Figure 6A, when ammonium acetate was added to an aqueous PrC-210 monothiol solution, the half-life at pH 7.2 was established to be 3.1 hours. These observations indicate that when anionic ammonium acetate molecules are added to an aqueous PrC-210 monothiol solution, the reactivity of the thiol is significantly increased, leading to the formation of PrC-210 disulfide molecules; i.e., 1) the free electron pair of the negatively charged acetate attacks the proton of the PrC-210 thiol, 2) the deprotonated activated thiolate anion on the PrC-210 molecule, 3) develops an affinity to react with another PrC-210 thiol to form a PrC-210 disulfide dimer (Figure 6B).

[0053] Thiol activation in the presence of negatively charged ammonium acetate is expected to reflect the reaction that occurs when a single PrC-210 molecule is brought into close proximity with a negatively charged molecule or site. The thiol is activated to react with several reactive sites, including electrophilic molecules, other thiol residues on the PrC-210 molecule, or other thiols on cysteine ​​residues of cellular proteins, forming disulfides (Figure 6C), carbon structures (thioethers), oxygen structures to generate sulfoxides (1xO), sulfenic ((1xO)H), sulfinic ((2xO)H), and sulfonic acids ((3xO)H), and nitrogen structures to generate nonradicals such as S-nitrosothiols or hydrogen peroxide.

[0054] We are also interested in whether PrC-210-cysteine ​​disulfide formation is transient or permanent. Permanent PrC-210 heterodisulfide formation with cellular proteins is expected to affect protein function and potentially lead to unwanted toxic side effects, which should be considered when PrC-210 residues appear as urinary or fecal metabolites. Several experimental results show that there is no clear evidence of permanent PrC-210-cysteine / protein disulfide formation, which supports i) the absence of toxic side effects within the therapeutic window of PrC-210 (see, e.g., Soref et al., Int J Radiation Oncol Biol Phys, Vol. 82, No. 5, pp. e701ee707, 2012), and ii) the absence of toxic side effects within the therapeutic window of PrC-210. Pharmacokinetic studies using C-labeled PrC-210 showed that only 1.07% of PrC-210 was metabolized after 48 hours (Figure 7, unpublished data). It is well documented that i) disulfide bonds are reversible (see, e.g., Koji Aoyama, Int. J. Mol. Sci. 2021, 22(9), 5010), and ii) two highly efficient thiol reduction systems exist within cells (see, e.g., Cumming et al., The Journal of Biological Chemistry vol. 279, No. 21, Issue of May 21, pp. 21749-21758, 2004), which reduce disulfide bonds between glutathione and proteins in the absence of oxidative stress (see, e.g., Koji Aoyama, Int. J. Mol. Sci. 2021, 22(9), 5010). Based on our experimental results and the published literature, it is logical that the binding of the PrC-210 disulfide to proteins is governed by the same chemical and enzymatic processes.

[0055] We believe that increased thiol reactivity in the presence of negative charges, such as ammonium acetate, also occurs when PrC-210-thiol is in close proximity to other reactive structures, such as hydrogen peroxide. We studied hydrogen peroxide because it is a nonradical oxygen species with a much longer half-life than superoxide. We added hydrogen peroxide to an aqueous solution of 5 mM PrC-210-thiol at pH 7. Hydrogen peroxide caused a dose-dependent consumption of free PrC-210-thiol (Figure 8). Adding 5 mM ammonium acetate to an aqueous solution of 5 mM PrC-210-thiol at pH 7 a few seconds before the addition of hydrogen peroxide significantly increased the hydrogen peroxide consumption of PrC-210-thiol (the "accelerating effect").

[0056] To examine the protective effect of PrC-210 in an animal model, we irradiated mice with 8.68 Gy and administered 0.5 MTD of PrC-210 30 minutes before irradiation. Normally, lethal radiation of 8.68 Gy induces massive DNA damage and activation of the intrinsic apoptotic pathway. When we examined the brains of irradiated mice (Figure 9A-C), we found no significant elevation in the levels of 8-oxo-dG, cytochrome C, or p53 damage biomarkers. This suggests that radiation insult did not significantly activate the intrinsic apoptotic pathway or cause mitotic arrest through p53 activation. The most likely reason for this is that neurons are already in the most advanced stage of mitosis, and thus DNA is highly protected from radiation insult. Interestingly, caspase-8 and caspase-3 / 7 were activated without PrC-210 and were suppressed to background levels when PrC-210 was administered systemically 24 h before radiation exposure, indicating that the presence of PrC-210 protected the proteins from damage (Figure 9D, E). This apoptosis suppression and protection continued throughout the 6-day observation period, indicating that PrC-210 was still present after 6 days. When plasma from mice in the same experiment was examined, the same 6-day suppression of apoptosis was observed, as were caspase-3 / 7 levels in the brain (Figure 9F).

[0057] Within cells, the positively charged amines of PrC-210 generally: i) cause PrC-210 molecules to concentrate and "hover" around negatively charged biomolecules; ii) the negative charges on biomolecules increase the reactivity of PrC-210 thiols, facilitating their reaction with reactive structures such as hydrogen peroxide and other thiol groups on cellular proteins. We conclude that, with regard to disease treatment, i) the amount of PrC-210 required at a given time depends on disease activity; ii) the amount of PrC-210 required at steady state may be relatively small, and therefore the PrC-210 present within cells functions as a reservoir; and iii) once inside the cells, PrC-210 remains there for long periods as a stable molecule with low reactivity until activated by the described mechanisms. Therefore, PrC-210 dosing regimens must take into account the acute and chronic nature of the disease and the severity of the injury.

[0058] Example 2: Use of the invention to reduce the progression of diseases associated with protein aggregation, such as amyotrophic lateral sclerosis This experiment was designed to determine whether PrC-210 could halt the progression of protein aggregation-based diseases such as amyotrophic lateral sclerosis (ALS). G93A The time course of mouse body weight is shown. Weight loss was progressive and exponential with the onset of ALS motor status. Systemic administration of PrC-210 (0.1 MTD, IP) was associated with an immediate plateau in body weight, i.e., cessation of weight loss in ALS mice. Because ALS pathology is based on protein aggregation, the results of this experiment support our inventive concept that PrC-210 can inhibit protein misfolding and thus suppress the pathology of these diseases. This invention applies to all neuron-specific and systemic diseases caused by protein aggregation, including neurodegenerative diseases, local and systemic amyloidosis, type 2 diabetes, and cataracts.

[0059] Example 3: Use of the invention to protect mitochondrial membranes Figure 11A shows a second cytochrome C peak in mouse myocardium 48 hours after 8.68 Gy of radiation. Unlike the first peak at 12 hours, this peak is not due to DNA damage or activation of the intrinsic apoptotic pathway; there is no associated increase in p53 or 8-oxo-dG (data not shown) 48 hours after radiation, nor is there activation of the intrinsic apoptotic pathway (Figure 11B). This peak represents extensive damage to complex IV of the respiratory chain, which is located in the intrinsic mitochondrial membrane. PrC-210 administered 24 hours after irradiation after the first cytochrome C release can penetrate mitochondria and suppress cytochrome C release to background levels (Figure 11A). This finding indicates that PrC-210 enters mitochondria and protects mitochondrial membrane proteins and lipids from attack, which would trigger cytochrome C release. The present invention applies to all diseases caused by mitochondrial damage, such as aging and all diseases associated with aging, neurodegenerative diseases, cardiovascular diseases such as arteriosclerosis, damage to heart valves and major arteries, and diseases generally caused by damage to mitochondrial membranes, mitochondrial DNA or proteins.

[0060] Example 4: Use of the invention to directly protect the brain from neuroinflammation and directly suppress inflammation from chronic inflammation and autoimmune diseases To visualize the effects of radiation on immune responses in the mouse brain, ICR mice received 8.68 Gy total-body radiation at time "0" (Figure 10). At 0.3 MTD, ipPrC-210 was administered 24 h after irradiation. As shown in Figure 12A, the inflammatory marker caspase-1 began to be significantly suppressed 8 days after irradiation. Similarly, the inflammatory marker complement 3 (C3) was similarly suppressed (Figure 12B). Treatment with PrC-210 significantly suppressed both inflammatory markers. Most importantly, this suppression was a direct suppression of the immune response, not an indirect suppression via protection from protein or lipid damage. Radiation-induced protein damage triggered an apoptotic response, as seen by the second peak of caspase-3 / 7 in Figure 12C. Therefore, this experiment demonstrates that PrC-210 can enter the brain and directly suppress inflammation after radiation.

[0061] To demonstrate that PrC-210 suppresses neuroinflammation caused by the autoimmune induction of disease, C57 mice were immunized with MOG peptide 24 hours before PrC-210 administration, immediately after the first paralysis from the autoimmunized demyelination (Fig. 13A). PrC-210 significantly suppressed the development and severity of paralysis in both administration models.

[0062] Furthermore, to demonstrate that this suppression of chronic inflammation generally applies to other organs besides the brain, we administered PrC-210 systemically and locally to DBA / 1J mice immunized against collagen III. The typical paw swelling induced in this mouse model was significantly reduced by twice-weekly intraperitoneal injections of 0.075 MTD PrC-210 and by topically applied 370 mM PrC-210.

[0063] The present invention is applicable to all diseases caused by neuroinflammation and inflammation, such as i) the treatment of neuroinflammation by applying Pr-210 after neurodegenerative diseases and multiple sclerosis, and brain damage caused by trauma, radiation, neurotoxins, ischemia and ischemia-reperfusion injury, and microbial diseases, and ii) all diseases caused by inflammation, such as the treatment and prevention of inflammation caused by lipid deposition in autoimmune diseases including, but not limited to, type 1 diabetes, rheumatoid arthritis, colitis, psoriasis, lupus erythematosus, pancreatitis, and hepatitis, atherosclerosis, damage to heart valves or major arteries, vascular damage during dialysis, and metabolic syndrome.

[0064] Other embodiments Various modifications and variations of the described compositions, methods, and uses of the invention will be apparent to those skilled in the art without departing from the scope and spirit of the invention. Although the invention has been described in connection with specific embodiments, it should be understood that the invention as claimed should not be unduly limited to such specific embodiments. Indeed, various modifications to the description of the detailed description that are obvious to those skilled in the art are intended to be within the scope of the invention.

[0065] Other embodiments are found in the claims.

Claims

1. 1. A method of inhibiting protein aggregation in a subject in need thereof, said method comprising administering an effective amount of a compound of formula (I): 【Chemistry 1】 or a pharmaceutically acceptable acid addition salt thereof to said subject in an amount sufficient to inhibit aggregation of said protein or peptide, wherein (i) A is -CH ２ NHR' and B is -CH ２ NHR or A=-NRR' and B=H, and (ii) each of R and R′ is independently H, C １ ~C ６ Alkyl, and C １ ~C ６ heteroalkyl; The above method, with the proviso that when B=H, then R and R' are not both H.

2. The compound is 【Chemistry 2】 or a pharmaceutically acceptable salt thereof.

3. 3. The method of claim 1 or 2, wherein the protein is selected from TDP-43, alpha synuclein, tau, amyloid beta, PolyQ htt, islet amyloid polypeptide, and prion.

4. 3. The method of claim 1 or 2, wherein the subject has a neurodegenerative disease characterized by misfolding and / or aggregation of endogenous proteins.

5. 5. The method of claim 4, wherein the neurodegenerative disease is Parkinson's disease, prion disease, Alzheimer's disease, multiple system atrophy, diffuse Lewy body disease, frontotemporal dementia, amyotrophic lateral sclerosis, Huntington's disease, spinocerebellar ataxia and other polyQ diseases, or hereditary cerebral amyloid angiopathy.

6. 6. The method of claim 5, wherein the neurodegenerative disease is a prion disease selected from Creutzfeldt-Jakob disease, variant Creutzfeldt-Jakob disease, inherited human prion diseases, bovine spongiform encephalopathy (BSE), and scrapie.

7. 5. The method of claim 4, wherein the neurodegenerative disease is a synucleinopathy.

8. 8. The method of claim 7, wherein the synucleinopathy is selected from Parkinson's disease (PD), dementia with Lewy bodies (DLB), and multiple system atrophy (MSA).

9. 3. The method of claim 1 or 2, wherein the subject has a disease or condition characterized by amyloidosis.

10. 10. The method of claim 9, wherein the amyloidosis is selected from primary systemic amyloidosis (AL amyloidosis), reactive systemic amyloidosis (AA amyloidosis), type 2 diabetes, injection-limited amyloidosis, beta-2 microglobulin amyloidosis, hereditary non-neuropathic amyloidosis, and hereditary systemic amyloidosis of the Finnish type.

11. 3. The method of claim 1 or 2, wherein the subject has a disease or condition characterized by ocular protein aggregation.

12. 12. The method of claim 11, wherein the disease or condition is selected from cataracts and presbyopia.

13. 1. A method of treating a condition associated with mitochondrial dysfunction in a subject in need thereof, said method comprising administering an effective amount of a compound of formula (I): 【Transformation 3】 or a pharmaceutically acceptable acid addition salt thereof to said subject in an amount sufficient to inhibit aggregation of said protein or peptide, wherein (i) A is -CH ２ NHR' and B is -CH ２ NHR or A=-NRR' and B=H, and (ii) each of R and R′ is independently H, C １ ~C ６ Alkyl, and C １ ~C ６ heteroalkyl; The above method, with the proviso that when B=H, then R and R' are not both H.

14. The compound is 【Chemistry 4】 or a pharmaceutically acceptable salt thereof.

15. 14. The method of claim 13, wherein the condition is a neurodegenerative disease, a neuropsychiatric disease, diabetes, a metabolic disease, an eye disorder associated with mitochondrial dysfunction, an ischemia-related condition, aging, mitochondrial toxicity associated with a therapeutic agent, or migraine.

16. 16. The method of claim 15, wherein the condition is a neurodegenerative disease selected from Friedreich's ataxia, amyotrophic lateral sclerosis, mitochondrial myopathy, encephalopathy, lactic acidosis, stroke (MELAS), myoclonic epilepsy with ragged-red fibers (MERFF), epilepsy, Parkinson's disease, Alzheimer's disease, and Huntington's disease.

17. 16. The method of claim 15, wherein the condition is a neuropsychiatric disorder selected from bipolar disorder, schizophrenia, depression, addiction, anxiety disorder, attention deficit disorder, personality disorder, autism, and Asperger's syndrome.

18. 16. The method of claim 15, wherein the condition is an eye disorder associated with mitochondrial dysfunction selected from glaucoma, diabetic retinopathy, and age-related macular degeneration.

19. 16. The method of claim 15, wherein the condition is an ischemia-related condition resulting from vascular obstruction, arteriosclerosis, valvular heart disease, tachycardia, hypertension, and hypotension.

20. 1. A method of treating a chronic inflammatory condition in a subject in need thereof, said method comprising administering an effective amount of a compound of formula (I): 【Transformation 5】 or a pharmaceutically acceptable acid addition salt thereof to said subject in an amount sufficient to inhibit aggregation of said protein or peptide, wherein (i) A is -CH ２ NHR' and B is -CH ２ NHR or A=-NRR' and B=H, and (ii) each of R and R′ is independently H, C １ ~C ６ Alkyl, and C １ ~C ６ heteroalkyl; The above method, with the proviso that when B=H, then R and R' are not both H.

21. The compound is 【Transformation 6】 or a pharmaceutically acceptable salt thereof.

22. 21. The method of claim 20, wherein the chronic inflammatory condition is selected from among rhinosinusitis, arthritis, dermatitis, vasculitis, acute malaria, sickle cell disease, a gastrointestinal inflammatory condition, or an inflammatory pulmonary condition.

23. 23. The method of claim 22, wherein the chronic inflammatory condition is an inflammatory pulmonary condition selected from asthma, chronic obstructive pulmonary disease (COPD), pulmonary conditions caused by physical trauma, emphysema, bronchitis, sarcoidosis, histiocytosis, lymphangioleiomyomatosis, acute lung injury, chronic lung disease, bronchopulmonary dysplasia, pneumonia, airway exacerbation, and acute respiratory distress syndrome (ARDS).

24. 23. The method of claim 22, wherein the chronic inflammatory condition is a gastrointestinal inflammatory condition selected from inflammatory bowel disease (IBD), ulcerative colitis (UC), Crohn's disease (CD), colitis, and chronic hepatitis such as viral hepatitis, alcohol-related hepatitis, and non-alcoholic steatohepatitis.

25. 21. The method of claim 20, wherein the chronic inflammatory condition is an autoimmune disease.

26. 26. The method of claim 25, wherein the autoimmune disease is selected from multiple sclerosis, type 1 diabetes, rheumatoid arthritis, juvenile rheumatoid arthritis, psoriatic arthritis, pancreatitis, psoriasis, plaque psoriasis, guttate psoriasis, inverse psoriasis, pustular psoriasis, erythrodermic psoriasis, paraneoplastic autoimmune diseases, autoimmune hepatitis, bullous pemphigoid, myasthenia gravis, inflammatory bowel disease, Crohn's disease, ulcerative colitis, celiac disease, thyroiditis, Sjogren's syndrome, Guillain-Barré disease, Raynaud's phenomenon, Addison's disease, primary biliary cirrhosis, primary sclerosing cholangitis, nonalcoholic fatty liver disease, nonalcoholic steatohepatitis, and diabetes.

27. 21. The method of claim 20, wherein the chronic inflammatory condition is a neuroinflammatory disease.

28. 28. The method of claim 27, wherein the neuroinflammatory disease is selected from multiple sclerosis, type 1 diabetes, neuromyelitis optica, anti-myelin oligodendrocyte glycoprotein antibody disease, autoimmune encephalitis, transverse myelitis, optic neuritis, and neurosarcoidosis, or a neurodegenerative disease selected from Friedreich's ataxia, amyotrophic lateral sclerosis, mitochondrial myopathy, encephalopathy, lactic acidosis, stroke (MELAS), myoclonic epilepsy with ragged-red fibers (MERFF), epilepsy, Parkinson's disease, Alzheimer's disease, and Huntington's disease.

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