RU2849887C2 - Therapeutic agents against neurodegenerative diseases - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE; invention relates to a method of providing neuroprotection in a subject suspected of having or at risk of developing a neurodegenerative disease that is cerebellar ataxia. The method includes administering a therapeutically effective amount of acetylleucine or its pharmaceutically acceptable salt to the subject for a period selected from at least 1 year, at least 2 years, and at least 5 years.

EFFECT; proposed invention can be used in the treatment of cerebellar ataxia.

7 cl, 13 dwg, 35 tbl, 20 ex

Description

Field of technology to which the invention relates

The present disclosure relates to the treatment of non-degenerative diseases comprising the administration of acetylleucine or a pharmaceutically acceptable salt thereof.

This application claims the benefit of priority of UK 1613829.9, filed August 11, 2016, UK 1702555.1, filed February 16, 2017, UK 1705766.2, filed April 10, 2017, and UK 1706867.7, filed April 28, 2017, all of which are incorporated herein by reference in their entirety.

State of the art

Neurodegenerative diseases are diseases that affect neurons. The degenerative process can involve progressive loss of neuronal structure, progressive loss of neuronal function, or progressive death of nerve cells. This progressive neurodegeneration typically leads to disability and mental retardation. Many neurodegenerative diseases are dangerously progressive and persistent, and there are few, if any, curative treatments.

Although the process of neurodegeneration is not fully understood, therapeutic agents shown to be broad-spectrum neuroprotective are believed to be applicable to neurodegenerative diseases. Furthermore, many neurodegenerative diseases are associated with lysosomal dysfunction. This includes neurodegenerative lysosomal storage diseases (LSDs) as well as many other neurodegenerative diseases, such as Alzheimer's disease and Parkinson's disease, which are suspected to be linked to lysosomal deficiencies.

This disclosure addresses the need for the development and widespread use of treatments for neurodegenerative diseases. Specifically, this disclosure describes acetylleucine for the treatment of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease in a subject. A neurodegenerative disease may, but need not, be associated with lysosomal dysfunction.

In one embodiment, acetylleucine or a pharmaceutically acceptable salt thereof is disclosed for use in a method of treating a neurodegenerative disease or one or more symptoms of a neurodegenerative disease associated with a neurodegenerative disease in a subject in need thereof, wherein the neurodegenerative disease is not cerebellar ataxia or Niemann-Pick disease type C.

In one embodiment of the present disclosure, acetylleucine or a pharmaceutically acceptable salt thereof is disclosed for use in a method of treating a neurodegenerative disease in a subject in need thereof, wherein the subject is asymptomatic.

In one embodiment of the present disclosure, acetylleucine or a pharmaceutically acceptable salt thereof is disclosed for use in a method for delaying the onset of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease that might otherwise be expected to occur according to the typical progression of the disease,

In another embodiment, the present disclosure includes acetylleucine or a pharmaceutically acceptable salt thereof for use in a method of treating a neurodegenerative disease or one or more symptoms of a neurodegenerative disease associated with a neurodegenerative disease in a subject in need thereof, wherein the method comprises administering to the subject in need thereof a therapeutically effective amount of acetylleucine for a period selected from at least about 3 months, at least about 6 months, at least about 1 year, at least about 2 years, and at least about 5 years.

In one embodiment, the present disclosure describes acetylleucine or a pharmaceutically acceptable salt thereof for use in a method for delaying the progression over time of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease associated with a neurodegenerative disease, compared to the typical progression of the disease, wherein the method comprises administering to a subject in need thereof a therapeutically effective amount of acetylleucine for a period selected from at least about 3 months, at least about 6 months, at least about 1 year, at least about 2 years, and at least about 5 years.

In another embodiment, acetylleucine or a pharmaceutically acceptable salt thereof is disclosed for use in a method for reversing the progression over time of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease associated with a neurodegenerative disease, wherein the method comprises administering to a subject in need thereof a therapeutically effective amount of acetylleucine for a period selected from at least about 3 months, at least about 6 months, at least about 1 year, at least about 2 years, and at least about 5 years.

In another embodiment, acetylleucine or a pharmaceutically acceptable salt thereof is disclosed for use in a method for improving, over time, a biochemical marker of a neurodegenerative disease in a subject in need thereof, wherein the method comprises administering to the subject in need thereof a therapeutically effective amount of acetylleucine for a period selected from at least about 3 months, at least about 6 months, at least about 1 year, at least about 2 years, and at least about 5 years.

In another embodiment, the present disclosure includes acetylleucine or a pharmaceutically acceptable salt thereof for use in a method for reducing the severity of a neurodegenerative disease or reducing the severity or eliminating one or more symptoms of a neurodegenerative disease associated with a neurodegenerative disease in a subject in need thereof, wherein the neurodegenerative disease is not cerebellar ataxia or Niemann-Pick disease type C.

In another embodiment, the present disclosure includes acetylleucine or a pharmaceutically acceptable salt thereof for use in a method of providing neuroprotection in a subject having, suspected of having, or at risk of developing a neurodegenerative disease, wherein the method comprises administering to the subject a therapeutically effective amount of acetylleucine for a period selected from at least about 3 months, at least about 6 months, at least about 1 year, at least about 2 years, and at least about 5 years.

Other embodiments of the present disclosure include acetylleucine or a pharmaceutically acceptable salt thereof for use in a method for delaying the development of a neurodegenerative disease or a lysosomal storage disease (LSD) in a subject. In another embodiment, acetylleucine or a pharmaceutically acceptable salt thereof is disclosed for use in a method for providing neuroprotection in a subject having, suspected of having, or at risk of developing a neurodegenerative disease or LSD. In one embodiment, acetylleucine is in the form of a racemate, in an enantiomeric excess of the L-enantiomer, or in an enantiomeric excess of the D-enantiomer. In another embodiment, the methods also comprise administering acetylleucine at a dose of 1.5 g to 10 g per day. Furthermore, in one embodiment, the methods comprise administering acetylleucine for treatment over a period of two weeks or more. For example, the methods include administering acetylleucine or a pharmaceutically acceptable salt thereof before the onset of a symptom of the disease or disorder being treated. In another embodiment, the methods also include administering another therapy or agent intended to prevent or treat the disease or disorder being treated. One embodiment of the present disclosure relates to a kit for delaying the development of a neurodegenerative disease or LD in a subject, wherein the kit comprises means for diagnosing or predicting the neurodegenerative disease or LD and acetylleucine or a pharmaceutically acceptable salt thereof. For example, the kit comprises means for diagnosing or predicting the neurodegenerative disease or LD, and acetylleucine or a pharmaceutically acceptable salt thereof. Another embodiment of the present disclosure is the use of acetylleucine or a pharmaceutically acceptable salt thereof as a neuroprotectant for a subject having a neurodegenerative disease or LD. In another embodiment of the methods, kits, or uses, the neurodegenerative disease is associated with lysosomal storage defects. In one embodiment of the methods, kits or uses, the neurodegenerative disease is alcoholism, Alexander disease, Alpers disease, Alzheimer's disease, amyotrophic lateral sclerosis (ALS), ataxia-telangiectasia, neuronal ceroid lipofuscinosis, Batten disease, bovine spongiform encephalopathy (BSE), Canavan disease, cerebral palsy, Cockayne syndrome, corticobasal degeneration, Creutzfeldt-Jakob disease, frontotemporal degeneration, Huntington's disease, HIV-associated dementia, Kennedy disease, dementia with Lewy bodies, neuroborreliosis, Machado-Joseph disease, multiple system atrophy, multiple sclerosis, multiple sulfatase deficiency, mucolipidosis, narcolepsy, Niemann-Pick disease, Parkinson's disease, Pick's disease, Pompe disease, primary lateral sclerosis, prion diseases, progressive supranuclear palsy, Refsum disease, Schilder disease, subacute combined degeneration of the spinal cord secondary to pernicious anemia, Spielmeyer-Vogt-Sjögren-Batten disease, spinocerebellar ataxia, spinal muscular atrophy, Steele-Richardson-Olszewski disease, or tabes dorsalis. In another embodiment of the methods, kits, or uses, the LPN is Niemann-Pick disease type C (NPC ₁ and/or NPC ₂ defect), Smith-Lemli-Opitz syndrome (SLOS), congenital disorder of cholesterol synthesis, Tangier disease, Pelizaeus-Merzbacher disease, neuronal ceroid lipofuscinosis, primary glycosphingolipidosis, Farber disease, or multiple sulfatase deficiency. For example, in the methods, kits, or uses, the primary glycosphingolipidosis is Gaucher disease, Fabry disease, GM gangliosidosis ₁ , GM gangliosidosis ₂ , Krabbe disease, or metachromatic leukodystrophy (MLD). In another example in the methods, kits or uses, the LPN is NPC, Tay-Sachs disease, Sandhoff disease, _GM1 gangliosidosis, Fabry disease, neurodegenerative mucopolysaccharidosis, MPS I, MPS IH, MPS IS, MPS II, MPS III, MPS IIIA, MPS IIIB, MPS IIIC, MPS HID, MPS IV, MPS IV A, MPS IV B, MPS VI, MPS VII, MPS IX, secondary lysosomal injury disease, SLOS or Tangier disease. In another embodiment of the methods, kits, or uses, the neurodegenerative disease is cerebellar ataxia, Niemann-Pick disease, parkinsonism, neuropathic Gaucher disease, Sandhoff disease, Louis-Bar syndrome, Alzheimer's disease, Parkinson's disease, multiple system atrophy, frontotemporal dementia, or lower body Parkinson's syndrome. In another embodiment of the methods, kits, or uses, the neurodegenerative disease is cerebellar ataxia, Niemann-Pick disease, Niemann-Pick disease type C, Niemann-Pick disease type A, Tay-Sachs disease, Sandhoff disease, amyotrophic lateral sclerosis (ALS), multiple system atrophy of the cerebellar type (MSA-C), frontotemporal dementia with parkinsonism, corticobasal degeneration syndrome, progressive supranuclear palsy, or down-beating cerebellar nystagmus. In one embodiment of the methods, kits, or uses, the LBN is Niemann-Pick disease, Niemann-Pick disease type C, Niemann-Pick disease type A, Tay-Sachs disease, Sandhoff disease, or mucolipidosis type II.

Brief description of the figures

Figure 1 shows photographs of treated (Figure A) and untreated (Figure B) Npc1 ^−/− mice at nine weeks of age.

Figures 2A and 2B show weight data for Npc1 ^−/− mice compared to wild-type (Npc1 ^+/+ ) mice with and without acetyl-DL-leucine treatment from weaned mothers.

Figures 3A–3G show gait analysis data for Npc1 ^−/− mice compared to wild-type (Npc1 ^+/+ ) mice with and without acetyl-DL-leucine treatment from weaned mothers. For example, data on diagonal support, rhythm, and step sequence are shown in Figures 3A–3C, respectively. Figures 3D and 3E show data for the forepaws (FP) (midstance and step cycle in panel D; work cycle in panel E). Figures 3F and 3G show data for the hindpaws (HP) (midstance and step cycle in panel F; work cycle in panel G).

Figures 4A–4H show the motor function analysis data of Npc1 ^−/− mice compared to wild-type (Npc1 ^+/+ ) mice treated with and without acetyl-DL-leucine from weaned mice. Center rearing, activity, rearing, and forward-backward (FR) counts are shown in Figures 4A–4D, respectively. Active time, movement time, rearing time, and manual total rearing counts are shown in Figures 4E–4H, respectively.

Figure 5 shows that acetyl-DL-leucine treatment (0.1 g/kg from 3 weeks of age) was associated with a small but statistically significant increase in lifespan in Npc1 ^−/− mice.

Figures 6A and 6B show a decrease in lysosomal volume in non-neuronal NPC cells after treatment with acetyl-DL-leucine. Figures 6C-6H show the effect of acetyl-DL-leucine treatment on lysosomal volume in fibroblasts from patients with NPA, MLII, MPS IIIB, aspartylglucosaminuria, MLIIIA, and MPS VII, respectively.

Figure 7A shows the survival curve representing mortality in untreated and acetylleucine-treated wild-type mice and those with Sandhoff disease. Figure 7B shows the stepping cycle time for untreated and acetylleucine-treated mice in the Sandhoff disease model, assessed at 12 weeks of age.

Figures 8A-8C show the effect of acetyl-DL-leucine treatment on glycosphingolipid (GSL) levels in fibroblasts from a patient with GM gangliosidosis ₂ (Tay-Sachs disease, Sandhoff disease, and AB variant Tay-Sachs disease, respectively).

Figure 9 shows a gait analysis matrix for a 75-year-old patient diagnosed with corticobasal degeneration syndrome before and during treatment with acetylleucine, where smaller dotted areas on the matrix indicate improvement compared to pre-treatment gait.

Figures 10A and 10B show the effect of acetyl-DL-leucine treatment over time on the overall clinical severity score (CSS) and the overall clinical annual severity increase score (ASIS), respectively, in ten patients with NPC.

Figures 11A-11J show the effect of acetyl-DL-leucine treatment over time on CSS subscores for each of the ten NPC patients.

Figures 12A and 12B show the effect of acetyl-DL-leucine treatment of Npc1 ^−/− wild-type mice on the levels of amyloid precursor protein C-terminal fragment (APP-CTF) and microtubule-associated protein 1A/3-phosphatidylethanolamine 1β-light chain conjugate (LC3-II), respectively.

Figures 13A-13C show that after treatment with acetyl-DL-leucine, a patient diagnosed with down-beating cerebellar nystagmus can partially suppress the nystagmus by visual fixation.

Disclosure of invention

Racemic acetylleucine (acetyl-DL-leucine) and its salts are effective in treating dizziness of various origins, particularly Ménière's vertigo and vertigo of inflammatory (vestibular neuritis) or toxic origin. For example, racemic acetylleucine is marketed by Pierre Fabre Medicament as an anti-vertigo medication under the trade name Tanganil®. Clinical results for Tanganil®, reported by various authors, show improvement in dizziness symptoms in over 95% of cases, including the disappearance of vertigo attacks.

Acetyl-DL-leucine has been used in France for the treatment of acute vertigo since 1957 and has an excellent safety profile, but its long-term safety with chronic use has not been determined. Despite many hypotheses, including stabilization of membrane potential, its pharmacological and electrophysiological modes of action remain unclear (Vibert et al. (2001), Eur. J. Neurosci., 13(4): 735–48; Ferber-Viart et al. (2009), Audiol. Neurootol., 14(1): 17–25). An FDG-µPET study in a rat model of acute unilateral labyrinectomy (Zwergal et al. (2016) Brain Struct. Func. 221(1): 159–70) showed a significant effect of the L-enantiomer of N-acetyl-L-leucine on postural compensation by activating the vestibular cerebellum and deactivating the posterolateral thalamus (Gunther et al. (2015) PLoS One 10(3): e0120891). Symptomatic improvement in cerebellar ataxia with acetyl-DL-leucine is seen in a number of patients with cerebellar diseases (Strupp et al. (2013) J. Neurol. 260(10): 2556–61). In another case, no improvement was observed (Pelz et al. (2015) J. Neurol. 262(5): 1373–5). Qualitative gait analysis showed that acetyl-DL-leucine improves transient gait variability in patients with cerebellar ataxia (Schniepp et al. (2015) Cerebellum 3:8). Symptomatic improvement of ataxia was shown in a one-month study of 12 patients with Niemann-Pick disease type C (NPC) (Bremova et al. (2015) Neurology 85(16): 1368–75). Furthermore, a PET study of ataxia patients treated with acetyl-DL-leucine showed increased metabolism in the midbrain and decreased metabolism in the brainstem (Becker-Bense et al. (2015) Abstract EAN).

However, acetylleucine is not known to treat neurodegenerative diseases, which typically develop over decades. The present disclosure unexpectedly reveals that acetylleucine or a pharmaceutically acceptable salt thereof can be used in a method for treating a neurodegenerative disease in a subject in need thereof, for example, by delaying the onset of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease that would otherwise be expected to manifest according to the typical progression of the disease, and/or by delaying or reversing the development of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease for a period of time longer than the typical progression of the disease. These examples of use according to the present disclosure, as well as others described herein, were unexpected, since a beneficial effect was not observed in the prior art and could not be concluded. Based on examples demonstrating the efficacy of acetylleucine in many neurodegenerative diseases, the inventors believe that acetylleucine acts as a neuroprotectant, thereby inhibiting neurodegeneration that might otherwise occur. Furthermore, many neurodegenerative diseases are associated with defects in lysosomal storage, and lysosomal dysfunction, such as aberrantly high levels of lysosomal storage, may be a cause of neuronal dysfunction and death. Based on examples, but without wishing to be bound by any particular theory, the inventors found, among other things, that acetylleucine can reduce cellular dysfunction (e.g., by reducing lysosomal volume to control values) and provide neuroprotection.

Therefore, the present disclosure relates to acetylleucine or a pharmaceutically acceptable salt thereof for use in a method of treating a neurodegenerative disease or one or more symptoms of a neurodegenerative disease in a subject in need thereof.

A "subject," as used herein, may be a vertebrate, mammal, or domestic animal. Therefore, the compositions of the present disclosure can be used to treat any mammal, such as livestock (e.g., horses, cows, sheep, or pigs), companion animals (e.g., cats, dogs, rabbits, or guinea pigs), laboratory animals (e.g., mice or rats), or other veterinary applications. In one embodiment, the subject is a human.

The term "neurodegenerative disease" as used herein refers to any disorder that affects neurons and includes progressive loss of neuronal structure, progressive loss of neuronal function, or progressive death of nerve cells.

As used in this description, the singular forms include reference to the plural.

The terms "approximately" and "approximately" mean approximately the same as the number or quantity stated, including an acceptable degree of error for the quantity measured, given the nature or accuracy of the measurements.

As used herein, the terms "approximately" and "approximately" are generally understood to encompass ±20% of a specified quantity, frequency, or amount. Numerical quantities given herein are approximate unless otherwise specified, meaning that the term "approximately" or "approximately" may be inferred when the quantity is not precisely stated.

The terms "administer," "administering," or "administered," when used herein, refer to (1) the provision, delivery, dispensing, and/or prescribing by or under the direction of a healthcare professional or his authorized representative of a composition according to the disclosure, and (2) the ingestion, receipt, or consumption by a patient or the individual of a composition according to the disclosure.

References to "acetylleucine" throughout the description include its pharmaceutically acceptable salts unless otherwise stated.

Acetylleucine may exist in racemic form, meaning the compound comprises approximately equal amounts of the enantiomers. Alternatively, it may exist in enantiomeric excess of either the L-enantiomer or the D-enantiomer. Acetylleucine may exist in a single enantiomeric form—either the L-enantiomer or the D-enantiomer. In one embodiment, the single enantiomeric form is the L-enantiomer. The racemic and enantiomeric forms can be prepared using procedures known in the art.

"Pharmaceutically acceptable salt" as used herein is any salt that is suitable for use in pharmaceutical application. Pharmaceutically acceptable salts include, but are not limited to, salts of amines such as N,N'-dibenzylethylenediamine, chloroprocaine, choline, ammonia, diethanolamine and other hydroxyalkylamines, ethylenediamine, N-methylglucamine, procaine, N-benzylphenethylamine, 1-para-chlorobenzyl-2-pyrrolidin-1'-ylmethylbenzimidazole, diethylamine and other alkylamines, piperazine, tris(hydroxymethyl)aminomethane, and the like; salts of alkali metals such as lithium, potassium, sodium, and the like; salts of alkaline earth metals such as barium, calcium, magnesium, and the like; salts of transition metals such as zinc, aluminum, and the like; other metal salts such as sodium hydrogen phosphate, disodium phosphate, etc.; salts of inorganic acids such as hydrochlorides, sulfates, etc.; and salts of organic acids such as acetates, lactates, malates, tartrates, citrates, ascorbates, succinates, butyrates, valerates, fumarates, etc.

Acetylleucine or a pharmaceutically acceptable salt thereof can be incorporated into compositions and administered to a subject according to directions known in the art. For example, acetylleucine or a pharmaceutically acceptable salt thereof can be incorporated into a pharmaceutical composition. The pharmaceutical composition can include acetylleucine or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier. The term "pharmaceutical composition" encompasses a single active agent or a form of the pharmaceutical composition.

A pharmaceutical composition may take any of a number of different forms, depending in particular on its intended use. For example, it may be in the form of a powder, tablet, capsule, liquid, ointment, cream, gel, hydrogel, aerosol, spray, micellar solution, transdermal patch, liposomal suspension, or any other suitable form that can be administered to a human or animal in need of treatment.

A "pharmaceutically acceptable carrier" as used herein is any known compound or combination of known compounds known to those skilled in the art to be useful in formulating pharmaceutical compositions. It should be understood that the carrier in a pharmaceutical composition must be one that is tolerated by the recipient.

In one embodiment, the pharmaceutically acceptable carrier may be a solid, and the composition may be in the form of a powder or a tablet. A solid pharmaceutically acceptable carrier may include, but is not limited to, one or more substances that may also act as flavoring agents, buffers, lubricants, stabilizers, solubilizers, suspending agents, wetting agents, emulsifiers, colorants, fillers, glidants, compression aids, inert binders, sweeteners, preservatives, coatings, or tablet disintegration agents. The carrier may also be an encapsulating material. In powders, the carrier may be a finely divided solid that is in admixture with the finely divided active agents of the invention. In tablets, the active agent may be mixed in suitable proportions with a carrier having the necessary compression properties and compressed into the desired shape and size. Powders and tablets may, for example, contain up to 99% active agents. Suitable solid carriers include, for example, calcium phosphate, magnesium stearate, talc, sugars, lactose, dextrin, starch, gelatin, cellulose, polyvinylpyrrolidone, low-melting waxes, and ion-exchange resins. In another embodiment, the pharmaceutically acceptable carrier may be a gel, and the composition may be in the form of a cream or similar.

The carrier may include, but is not limited to, one or more excipients or diluents. Examples of such excipients include gelatin, acacia, lactose, microcrystalline cellulose, starch, sodium starch glycolate, calcium hydrogen phosphate, magnesium stearate, talc, colloidal silicon dioxide, and the like.

In another embodiment, the pharmaceutically acceptable carrier may be a liquid. In one embodiment, the pharmaceutical composition is in the form of a solution. Liquid carriers are used in the preparation of solutions, suspensions, emulsions, syrups, elixirs, and pressurized compositions. Acetylleucine can be dissolved or suspended in a pharmaceutically acceptable liquid carrier, such as water, an organic solvent, or a mixture of two or more pharmaceutically acceptable oils or fats. The liquid carrier may contain other suitable pharmaceutical additives, such as solubilizers, emulsifiers, buffers, preservatives, sweeteners, flavoring agents, suspending agents, thickeners, colorants, viscosity regulators, stabilizers, or osmoregulators. Suitable examples of liquid carriers for oral and parenteral administration include water (partially containing additives as noted above, such as cellulose derivatives such as sodium carboxymethylcellulose solution), alcohols (including monohydric alcohols and polyhydric alcohols such as glycols) and their derivatives, and oils (such as fractionated coconut oil and peanut oil). For parenteral administration, the carrier may also be a fatty acid ester such as ethyl oleate and isopropyl myristate. Sterile liquid carriers are used in sterile liquid compositions for parenteral administration. The liquid carrier for pressurized compositions may be a halogenated hydrocarbon or other pharmaceutically acceptable propellant.

Liquid pharmaceutical compositions, which are sterile solutions or suspensions, can be administered by, for example, intramuscular, intrathecal, epidural, intraperitoneal, intravenous, and especially subcutaneous injection. The active agent can be prepared as a sterile solid composition that can be dissolved or suspended during administration using sterile water, saline, or another appropriate sterile injection medium.

The compositions can be administered orally in the form of a sterile solution or suspension containing other solutes or suspending agents (e.g., sufficient salt or glucose to render the solution isotonic), bile salts, acacia, gelatin, sorbitan monooleate, polysorbate 80 (sorbitol oleates and its anhydrides copolymerized with ethylene oxide), and the like. The compositions can also be administered orally in the form of either a liquid or solid composition. Compositions suitable for oral administration include solid forms such as pills, capsules, granules, tablets, and powders, and liquid forms such as solutions, syrups, elixirs, and suspensions. Forms suitable for parenteral administration include sterile solutions, emulsions, and suspensions.

On the other hand, acetylleucine and compositions containing it can be administered by inhalation (e.g., intranasally). Topical formulations can also be developed. For example, creams and ointments can be applied to the skin.

Acetylleucine can be incorporated into a sustained- or delayed-release device. Such devices can, for example, be implanted on or under the skin, and the drug can be released over weeks or even months. Such devices may be advantageous when long-term treatment with acetylleucine, as described herein, is required, which typically requires frequent administration (e.g., at least daily).

In one embodiment, the pharmaceutical composition is in tablet form. In tablets, the active agent can be mixed in suitable proportions with a medium, such as a pharmaceutically acceptable carrier, having the necessary compression properties and compressed into the desired shape and size. Tablets can contain up to 99% by weight of active agent.

For example, acetylleucine or a pharmaceutically acceptable salt thereof may be provided in a solid dosage form suitable for oral administration, particularly in the form of a tablet.

Pharmaceutical compositions in solid oral dosage form, such as tablets, can be prepared by any method known in pharmacy. Pharmaceutical compositions are typically prepared by mixing acetylleucine or a pharmaceutically acceptable salt thereof with conventional pharmaceutically acceptable carriers.

The tablet can be prepared as is known in the art. "Tanganil®," for example, includes wheat starch, pregelatinized maize starch, calcium carbonate, and magnesium stearate as excipients. The same or similar excipients, for example, can be used in the present disclosure.

The composition of each 700 mg Tanganyil® tablet is as follows: 500 mg acetyl-DL-leucine, 88 mg wheat starch, 88 mg pregelatinized maize starch, 13 mg calcium carbonate, and 11 mg magnesium stearate. These same tablets, for example, can be used in the present disclosure.

The present disclosure describes acetylleucine, compositions including it, and methods for treating a neurodegenerative disease or one or more symptoms of a neurodegenerative disease in a subject in need thereof. The subject in need thereof may have a genetic, biochemical, or other similar identifiable marker of a neurodegenerative disease. For example, the marker of a neurodegenerative disease may be a cellular marker. The subject in need thereof may be diagnosed as having a neurodegenerative disease. For example, the subject may be diagnosed with a neurodegenerative disease according to a genetic, biochemical, or other similar identifiable marker. The subject in need thereof may be suspected of having or at risk of developing a neurodegenerative disease. For example, the subject may have a genetic predisposition to a neurodegenerative disease (e.g., the subject may have one or more family members with a neurodegenerative disease). A subject in need thereof may be symptomatic (i.e., have one or more symptoms associated with a neurodegenerative disease). A subject in need thereof may be asymptomatic. It should be noted that the terms "symptomatic" and "asymptomatic" are used in connection with the symptoms of a neurodegenerative disease. Subjects having a genetic, biochemical, or other similar identifiable marker of a neurodegenerative disease, such as subjects diagnosed with a neurodegenerative disease based on a genetic, biochemical, or other similar identifiable marker, but who do not have other symptoms of the disease, are included in the "asymptomatic" group for the purposes of this disclosure.

As used herein, the expression "treating a neurodegenerative disease or one or more symptoms of a neurodegenerative disease" and the like refer to delaying the onset of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease that would otherwise be expected to manifest according to the typical progression of the disease, reducing the severity of a neurodegenerative disease or reducing the severity or elimination of one or more existing symptoms associated with a neurodegenerative disease, delaying the progression of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease over time compared to the typical progression of the disease, and/or reversing the progression of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease over time. "Treating a neurodegenerative disease or one or more symptoms of a neurodegenerative disease" may also refer to improving a biochemical marker of a neurodegenerative disease.

As used herein, the terms "typical disease progression," "the disease progression that can usually be expected," and the like refer to the typical or expected progression of a neurodegenerative disease, one or more symptoms associated with a neurodegenerative disease, or a biochemical marker of a neurodegenerative disease if the subject is not treated. The typical or expected progression of a disease may be based, for example, on a known scale, index, rating, or assessment, or other suitable test for assessing the progression of a neurodegenerative disease, one or more symptoms of a neurodegenerative disease, or a biochemical marker of a neurodegenerative disease, such as those described as examples herein. The scale, index, rating, assessment, or other suitable test may correspond to the progression of a neurodegenerative disease as a whole or to the development of one or more symptoms associated with the disease. For example, the typical or expected progression of a disease may be assessed based on the typical or expected onset or severity of a neurodegenerative disease, or the appearance of a symptom or set of symptoms associated with a neurodegenerative disease. A typical or expected disease progression can be determined by comparing individuals or by what is typically observed or experienced by a large number of individuals affected by a neurodegenerative disease, such as a population or subpopulation of individuals. Subpopulations may include, for example, populations of the same sex, the same or similar age, the same or similar time of onset of one or more symptoms, etc.

In one embodiment, "treating a neurodegenerative disease or one or more symptoms of a neurodegenerative disease" refers to delaying the onset of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease that would otherwise be expected to manifest according to the typical progression of the disease. As used herein, the expression "delaying the onset of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease" and the like refer to increasing the time until or preventing the onset of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease. For example, the onset can be said to be delayed when the time until the onset of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease is at least 5% longer than the time observed according to the typical progression of the disease. Also, for example, an increase in time of at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 100% is observed. In one embodiment, the subject is asymptomatic. Administration of acetylleucine at a time when the subject is asymptomatic can cause a delay in the onset of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease that would otherwise be expected to manifest according to the typical progression of the disease. In another embodiment, the subject is symptomatic. Administration of acetylleucine may begin during a period when the subject exhibits several symptoms to delay the onset of one or more additional symptoms of the neurodegenerative disease that would otherwise be expected to manifest according to the typical progression of the disease. The subject in need thereof may continue to receive acetylleucine treatment for the treatment durations described herein. In one embodiment, the treatment prevents the onset of one or more symptoms of the neurodegenerative disease that would otherwise be expected to manifest according to the typical progression of the disease.

In one embodiment, "treating a neurodegenerative disease or one or more symptoms of a neurodegenerative disease" refers to reducing the severity of a neurodegenerative disease or reducing the severity or eliminating one or more symptoms associated with a neurodegenerative disease. The severity of a neurodegenerative disease or existing symptom(s) can be assessed using a known scale, index, rating, or assessment, such as in the examples described as examples herein, or another test suitable for assessing severity. For example, a scale, index, rating, assessment, or other suitable test can correspond to the severity of a neurodegenerative disease as a whole or the severity of one or more symptoms associated with a neurodegenerative disease. In one embodiment, the treatment improves such an assessment from the magnitude or degree characteristic of a symptomatic patient to the magnitude or degree characteristic of an asymptomatic patient.

In one embodiment, "treating a neurodegenerative disease or one or more symptoms of a neurodegenerative disease" refers to delaying the progression of a neurodegenerative disease or one or more symptoms associated with a neurodegenerative disease over time, compared to the typical progression of the disease, or reversing the progression of a neurodegenerative disease or one or more symptoms associated with a neurodegenerative disease over time. The time over which the treatment delays or reverses the progression of the disease may correspond to the duration of treatment described herein. The treatment may delay or reverse progression for, for example, about seven days or longer, about two weeks or longer, about three weeks or longer, about one month or longer, about six weeks or longer, about seven weeks or longer, or about two months or longer. For example, the treatment may delay or reverse progression for about three months or longer, about four months or longer, about five months or longer, or about six months or longer. It can delay or reverse progression over a period of, for example, approximately 1 year or longer, approximately 2 years or longer, approximately 3 years or longer, approximately 4 years or longer, approximately 5 years or longer, or 10 years or longer. Treatment can delay or reverse the progression of a neurodegenerative disease or one or more symptoms associated with a neurodegenerative disease throughout the patient's life.

In one embodiment, "treating a neurodegenerative disease or one or more symptoms of a neurodegenerative disease" refers to delaying the progression of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease over time, compared to the typical progression of the disease. As used herein, the phrase "delaying the progression of a neurodegenerative disease or one or more symptoms associated with a neurodegenerative disease over time" and the like refer to slowing and/or stopping the progression of a disease or one or more symptoms of a disease (e.g., slowing and/or stopping the worsening or increase in the severity of a disease or one or more symptoms of a disease) over time. The progression of a disease can be determined, for example, using a known scale, index, rating, or assessment, such as in the examples described herein, or another test suitable for assessing severity. For example, a scale, index, rating, assessment, or other suitable test can correspond to the progression of the disease as a whole or the development of one or more symptoms associated with the disease. In one embodiment, "delaying the development over time of a neurodegenerative disease or one or more symptoms associated with a neurodegenerative disease" means that the subject's disease severity score (e.g., overall severity or the severity of one or more symptoms), as determined by a known scale, index, rating, assessment, etc., or other suitable test for assessing severity, has not increased significantly (e.g., remains at least substantially constant). In one embodiment, "delaying the development over time of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease" means preventing the subject from achieving or increasing the time it takes for the subject to achieve (e.g., reducing the rate of change in increasing severity) a severity score according to a known scale, index, rating, assessment, etc., or other suitable test for assessing development, compared to the score corresponding to the typical development of the disease. For example, development can be said to be delayed when the time to achieving the severity score becomes at least 5% longer than the time observed according to the typical development of the disease. Also, by way of example, an increase in time of at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 100% is observed. The time during which the treatment delays the development of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease may correspond to the duration of treatment described herein. In one embodiment, the treatment delays the development for at least about three months, at least about four months, at least about five months, or at least about six months. Treatment may delay development for at least about 1 year, at least about 2 years, at least about 3 years, at least about 4 years, at least about 5 years, or at least 10 years. Treatment may delay development for the life of the patient.

In one embodiment, "treating a neurodegenerative disease or one or more symptoms of a neurodegenerative disease" refers to reversing the progression of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease over time. As used herein, the phrase "reversing the progression of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease over time" and the like refer to stopping the progression and reducing the severity of the disease or one or more symptoms of the disease over time. The progression and severity of the disease can be determined, for example, using a known scale, index, rating, or assessment, such as in the examples described herein, or another test suitable for assessing the progression and severity. For example, a scale, index, rating, assessment, or other suitable test can correspond to the progression and severity of the disease as a whole or the progression and severity of one or more symptoms associated with the disease. In one embodiment, "reversal of the progression of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease over time" means that the subject's assessment of the severity of the disease (e.g., overall severity or the severity of one or more symptoms), determined using a known scale, index, rating, assessment, etc., or other suitable test for assessing severity, improves over time (i.e., shows a decrease in severity over time). The time during which the treatment reverses the progression of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease may correspond to the duration of treatment described herein. In one embodiment, the treatment reverses the progression within at least about three months, at least about four months, at least about five months, or at least about six months. In another embodiment, the treatment reverses the progression within at least about 1 year, at least about 2 years, at least about 3 years, at least about 4 years, at least about 5 years, or at least 10 years. Treatment can reverse progression throughout the patient's life.

In one embodiment, "treating a neurodegenerative disease or one or more symptoms of a neurodegenerative disease" refers to improving a biochemical marker of a neurodegenerative disease in a subject (e.g., elevated levels of metabolite(s) accumulation or secondary biochemical changes resulting from primary accumulation). The biochemical marker is a signal of disease activity and can provide ongoing indications of the severity and progression of the disease over time. In one embodiment, the biochemical marker is improved relative to a reference value. In one embodiment, the biochemical marker is selected from increased lysosomal volume, increased glycosphingolipid (GSL) levels, increased levels of macrotubule-associated protein 1A/3-phosphatidylethanolamine 1β-light chain (LC ₃ -II) conjugate, and increased levels of amyloid precursor protein C-terminal fragment (APP-CTF). In one embodiment, the biochemical marker is increased lysosomal volume, and the treatment reduces lysosomal volume in the subject. In one embodiment, the biochemical marker is increased glycosphingolipid (GSL) levels, and the treatment reduces GSL levels in the subject. In one embodiment, the biochemical marker is increased levels of macrotubule-associated protein 1A/3-phosphatidylethanolamine 1β-light chain conjugate (LC ₃ -II), and the treatment reduces LC ₃ -II levels in the subject. In one embodiment, the biochemical marker is increased levels of amyloid precursor protein C-terminal fragment (APP-CTF), and the treatment reduces APP-CTF levels in the subject. In one embodiment, the treatment improves the biochemical marker over time. For example, in one embodiment, improving a biochemical marker over time means that the treatment improves the biochemical marker over time relative to a control value, prevents the development of the biochemical marker over time, and/or delays the development of the biochemical marker over time compared to the typical development in the disease. The time during which the treatment improves the biochemical marker may correspond to the duration of treatment described herein. In one embodiment, the treatment improves the biochemical marker for at least about three months, at least about four months, at least about five months, or at least about six months. The treatment can improve the biochemical marker for at least about 1 year, at least about 2 years, at least about 3 years, at least about 4 years, at least about 5 years, or at least 10 years. The treatment can improve the biochemical marker throughout the patient's life.

A "symptom" of a neurodegenerative disease includes any clinical or laboratory manifestation associated with the neurodegenerative disease and is not limited to what the patient can feel or observe. Symptoms described herein include, but are not limited to, neurological symptoms and psychotic symptoms. Examples of neurological symptoms include ataxia, other movement disorders such as hypokinesia, muscle rigidity, tremor, or dystonia, central oculomotor disorders such as vertical and horizontal supranuclear saccade/gaze palsy, and neuropsychological deficits such as dementia. Examples of psychotic symptoms include depression, behavioral disorders, or psychosis. The onset of symptoms may range from birth to adulthood.

The progression of a neurodegenerative disease over time or during treatment can be monitored, for example, by using one or more known tests at two or more time points and comparing the results. The progression and/or severity of the disease can be assessed, for example, using the Scale for the Assessment and Rating of Ataxia (SARA), the Spinocerebellar Ataxia Functional Index (SCAFI), the International Cooperative Ataxia Rating Scale (ICARS), the Brief Ataxia Rating Scale (BARS), the Modified Disability Rating Scale (mDRS), the EuroQol 5Q-5D-5L (EQ-5D-5L) visual analogue scale (VAS), neuropsychological tests such as the Revised Adult Intelligence Scale (WAIS-R), the Revised Intelligence Scale for Children-IV (WISC-IV), the Montreal Cognitive Assessment (MoCA), as well as scales used in movement disorders such as the Unified Parkinson's Disease Rating Scale (UPRS) or the Unified Multiple System Atrophy Rating Scale (UMSARS), or other suitable tests. For some LDs, such as NPC, separate assessments have been developed and validated over the past decades, such as the Clinical Severity Score (CSS) and the Annual Severity Increasing Score (ASIS) (see Yanjanin et al., “Linear Clinical Progression, Independent of Age of Onset, in Niemann–Pick Disease, Type C,” Am. J. Med. Genet. Part B, 153B: 132–140), and the modified 6-Domain NP-C Disability Rating Scale (mDRS score). For example, the severity of NPC in a patient can be quantified by assessing the CSS, which evaluates various disease parameters (ambulatory ability, seizures, eye movements, etc.) and gives a score of up to 5 for each parameter. A higher score indicates greater severity. The ASIS quantifies the degree of change in the CSS and is calculated by dividing the CSS by the patient’s age. In this regard, some scores in such tests are characteristics of symptomatic patients with neurodegenerative disease and provide evidence of disease progression and/or severity.

Thus, "treating a neurodegenerative disease or one or more symptoms of a neurodegenerative disease," for example, can be equated to achieving an improved score, such as those described herein, a score on the SARA, SCAFI, ICARS, BARS, mDRS, EQ-5D-5L VAS, WAIS-R, WISC-IV, CSS, UPRS, UMSARS and/or MoCA, or the result of another test suitable for characterizing a patient with a neurodegenerative disease. For example, in one embodiment, "reducing the severity of a neurodegenerative disease or reducing the severity or eliminating one or more existing symptoms of a neurodegenerative disease" means improving the score on the SARA, SCAFI, ICARS, BARS, mDRS, EQ-5D-5L VAS, WAIS-R, WISC-IV, CSS, UPRS. UMSARS and/or MoCA or another test result suitable for assessing severity, such as an improvement in the score or result from the severity score characteristic of a symptomatic subject to the value characteristic of an asymptomatic subject. In another embodiment, "delay in the development of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease" means that the subject's score on the SARA, SCAFI, ICARS, BARS, mDRS, EQ-5D-5L VAS, WAIS-R, WISC-IV, CSS, UPRS, UMSARS and/or MoCA or another test result suitable for assessing development does not increase significantly (e.g., at least remains substantially constant). In another embodiment, "delaying the development of a neurodegenerative disease or one or more symptoms associated with a neurodegenerative disease" means preventing the subject's SARA, SCAFI, ICARS, BARS, mDRS, EQ-5D-5L VAS, WAIS-R, WISC-IV, CSS, UPRS, UMSARS and/or MoCA score or the result of another test suitable for assessing development from reaching, or increasing the time taken to reach, a value comparable to the value during the typical development of the disease. In another embodiment, "reversing the development of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease over time" means that the subject's SARA, SCAFI, ICARS, BARS, mDRS, EQ-5D-5L VAS, WAIS-R, WISC-IV, CSS, UPRS score. The UMSARS and/or MoCA or other test result appropriate for assessing development improves over time (i.e., shows a decrease in severity over time).

For example, to assess overall neurological status using the mDRS, a four-item scale (ambulation, manipulation, speech, and swallowing) can be used. Cerebellar function can be assessed using the SARA, an eight-item clinical rating scale (gait, standing, sitting, speech, fine motor function, and taxis; range 0–40, where 0 is the best neurological status and 40 is the worst), and the SCAFI, which includes the 8-m-Walking-Time (8MW; performed by having patients walk as quickly as possible twice from one line to another, including turning), the 9-Hole-Peg-Test (9HPT), and a series of “PATA” repetitions for 10 seconds. Subjective improvement and quality of life can be assessed using the EQ-5D-5L questionnaire and VAS. To assess oculomotor function, 3-D videooculography (EyeSeeCam) can be used to measure peak saccade velocity, smooth pursuit improvement, peak slow-phase velocity of gaze-evoked nystagmus (gaze-holding function), peak slow-phase velocity of optokinetic nystagmus, and horizontal vestibulo-ocular reflex improvement. To assess cognitive status, the WAIS-R or WISC-IV and MoCA can be used, assessing various cognitive domains, including attention and concentration, executive functions, memory, language, visuoconstructive skills, conceptual thinking, calculation, and orientation with a maximum of 30 points and a cutoff score of 26. A specialist will be familiar with how to perform these and other similar tests.

Acetylleucine or a pharmaceutically acceptable salt thereof can be administered, for example, at a dose ranging from about 500 mg to about 15 g per day, or ranging from about 500 mg to about 10 g per day, such as ranging from about 1.5 g to about 10 g per day, optionally orally in solid or liquid form. Acetylleucine or a pharmaceutically acceptable salt thereof can be administered, for example, at a dose corresponding to Tanganil®, which is prescribed for adults at a dose of 1.5 g to 2 g per day, 3-4 tablets in two doses morning and evening.

If a single enantiomer is administered, the doses may be reduced accordingly. For example, if only acetyl-L-leucine is administered, or if only acetyl-D-leucine is administered, the dose may range from about 250 mg to about 15 g per day, range from about 250 mg to about 10 g per day, or range from about 250 mg to about 5 g per day, such as from about 0.75 g to about 5 g per day.

In one embodiment, the administered dose ranges from about 1 g to about 15 g per day, from about 1 g to about 10 g per day, or from about 1.5 g to about 7 g per day. It can be from about 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14 g to about 15 g per day. It can be from about 2, 3, 4, 5, 6, 7, 8, or 9 g to about 10 g per day. It can be more than about 1.5 g per day, but less than about 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, or 5 g per day. In one embodiment, the dose ranges from about 4 g to about 6 g per day. In one embodiment, the dose ranges from about 4 g to about 5 g per day. In one embodiment, the dose is about 4.5 g per day. In one embodiment, the dose is about 5 g per day. In one embodiment, such doses are administered in solid oral dosage form, especially in tablets. In another embodiment, such doses are doses in which acetylleucine is in its racemic form. Doses of acetylleucine in enantiomeric excess may be lower than the doses described herein, for example, about 50% lower. Thus, the above-mentioned dose ranges, when divided in half, are also expressly encompassed by the present disclosure.

The total daily dose can be divided into several administrations, i.e., administration can occur two or more times per day to achieve the total daily dose. For example, the required number of tablets to provide the full daily dose of acetylleucine can be divided into two administrations (e.g., morning and evening) or three administrations (e.g., morning, noon, and evening). Each dose can be administered with or without food, as appropriate. For example, acetylleucine can be taken about 1 or about 2 hours before a meal, such as at least about 20 minutes, at least about 30 minutes, at least about 40 minutes, or at least about 1 hour before a meal, or can be taken about 1, about 2, or about 3 hours after a meal, such as by waiting at least about 20 minutes, at least about 30 minutes, at least about 1 hour, at least about 1.5 hours, at least about 2 hours, or at least about 2.5 hours after a meal. For example, a total daily dose of 4.5 g of acetyl-DL-leucine can be taken as three ^Tanganyil® tablets (or equivalent) before, with, or after breakfast, three other tablets before, with, or after lunch, and three other tablets before, with, or after dinner.

The administration of acetylleucine according to the present disclosure can be started before or after the subject is found to have a genetic, biochemical, or other suitable identifiable marker of a neurodegenerative disease, for example, in the first instance when it is suspected that the subject has or is at risk of developing a neurodegenerative disease. The administration can be started at or around the time when the subject is found to have a genetic, biochemical, or other suitable identifiable marker of a neurodegenerative disease. Similarly, the administration can be started before or around the time or after the subject is diagnosed with a neurodegenerative disease, for example, before or around the time or after the subject is found to have a genetic, biochemical, or other suitable identifiable marker of a neurodegenerative disease. The administration of acetylleucine can be started when the subject is symptomatic or asymptomatic. In particular, one advantage of the acetylleucine treatment according to the present disclosure is that acetylleucine administration can be initiated early, at a time after a subject is detected to have a genetic and/or biochemical marker for a neurodegenerative disease, but before the subject exhibits symptoms of a neurodegenerative disease (other than the genetic and/or biochemical marker for a neurodegenerative disease, i.e., the subject is asymptomatic), or before the subject exhibits one or more symptoms considered indicative of the disease. The treatment can delay the onset of a neurodegenerative disease or one or more symptoms associated with a neurodegenerative disease, as described herein. The treatment can also be continued long-term, as described herein.

As discussed herein, an advantage of acetylleucine treatment according to the present disclosure is that acetylleucine can be administered over an extended period of time in order to, for example, delay or even reverse the progression of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease in a subject compared to the typical progression of the disease. The duration of treatment can be, for example, about seven days or longer, about two weeks or longer, about three weeks or longer, about one month or longer, about six weeks or longer, about seven weeks or longer, or about two months or longer. In one embodiment, the duration of treatment is about three months or longer, about four months or longer, about five months or longer, or about six months or longer. The duration of treatment can be about 1 year or longer, about 2 years or longer, about 4 years or longer, about 5 years or longer, or about 10 years or longer. The duration of treatment can be the entire life of the patient.

Any and all combinations of dosage form, dosage amount, administration regimen, and treatment duration are contemplated and encompassed by the invention. In one embodiment, the dose is from about 4 g to about 10 g per day, administered in one, two, or three administrations per day, for a treatment duration of about two months or longer. In another embodiment, the dose is greater than 4 g but less than 5 g per day, administered in one, two, or three administrations per day, for a treatment duration of about six months or longer. The dosage form may be a solid oral dosage form, especially tablets.

The pharmaceutical composition can be used as monotherapy (e.g., using a single active substance) to treat a neurodegenerative disease in a subject. Alternatively, the pharmaceutical composition can be used as an adjunct to or in combination with other known therapies, for example, to treat a neurodegenerative disease in a subject.

Neurodegenerative disease may, but need not, be associated with lysosomal dysfunction (eg, lysosomal storage defect). Neurodegenerative diseases, according to the present disclosure, not associated with lysosomal dysfunction include, but are not limited to, Alexander disease, Alpers disease, cerebral palsy, Cockayne syndrome, corticobasal degeneration, HIV-associated dementia, Kennedy disease, neuroborreliosis, primary lateral sclerosis, Refsum disease, Schilder disease, subacute combined degeneration of the spinal cord secondary to pernicious anemia, hereditary motor and sensory neuropathy with proximal dominance, wandering hedgehog syndrome (WHS), progressive muscular atrophy (Duchenne-Aran muscular atrophy), progressive bulbar palsy, pseudobulbar palsy, HIV-associated neurocognitive impairment (HAND), vascular parkinsonism, lower body Parkinson syndrome, cerebellar Downward-beating nystagmus and cerebellar ataxia, which includes spinocerebellar ataxia (SCA) 4, spinocerebellar ataxia (SCA) 5 (Lincoln ataxia), spinocerebellar ataxia (SCA) 8, spinocerebellar ataxia (SCA) 10, spinocerebellar ataxia (SCA) 11, spinocerebellar ataxia (SCA) 12, spinocerebellar ataxia (SCA) 13, spinocerebellar ataxia (SCA) 14, spinocerebellar ataxia (SCA) 15/16, spinocerebellar ataxia (SCA) 18 (sensory/motor neuropathy with ataxia), spinocerebellar ataxia (SCA) 19/22, spinocerebellar ataxia (SCA) 20, spinocerebellar ataxia (SCA) 21, spinocerebellar ataxia (SCA) 23, spinocerebellar ataxia (SCA) 25, spinocerebellar ataxia (SCA) 26, spinocerebellar ataxia (SCA) 27, spinocerebellar ataxia (SCA) 29, spinocerebellar ataxia (SCA) 30, spinocerebellar ataxia (SCA) 31, spinocerebellar ataxia (SCA) 32, spinocerebellar ataxia (SCA) 35, spinocerebellar ataxia (SCA) 36, episodic ataxia (EA) 1, episodic ataxia (EA) 2, episodic ataxia (EA) 3, episodic ataxia (EA) 4, episodic ataxia (EA) 5, episodic ataxia (EA) 6, episodic ataxia (EA) 7, spinocerebellar ataxia (SCA) 28, spinocerebellar ataxia (SCA) 24 (spinocerebellar ataxia type 4 (SCAR4); spinocerebellar ataxia with saccadic intrusions), tabes dorsalis, ataxia with oculomotor apraxia type 1 (AOA1), ataxia with oculomotor apraxia type 2 (AOA2), ataxia with oculomotor apraxia type 4 (AOA4), spinocerebellar ataxia type 10 (SCAR 10), mitochondrial recessive ataxia syndrome (MIRAS), myoclonic epilepsy, myopathy, and sensory ataxia (MEMSA), sensory ataxic neuropathy, dysarthria, ophthalmoparesis (SANDO), childhood-onset spinocerebellar ataxia, hereditary spastic paraplegia 7 (HSP gene SPG7), mitochondrial myopathy, encephalopathy, lactic acidosis, stroke syndrome (MELAS), myoclonic epilepsy with ragged red fibers (MERRF), neurogenic muscle weakness, ataxia, and retinitis pigmentosa (NARP), Kerse-Sayre syndrome (KSS), fragile X-associated tremor/ataxia (FXTAS), Art syndrome, X-linked spinocerebellar ataxia 1, X-linked spinocerebellar ataxia 2, X-linked spinocerebellar ataxia 3, X-linked spinocerebellar ataxia 4, or X-linked spinocerebellar ataxia 5, X-linked syndromic intellectual disability, Christiansen type, X-linked sideroblastic anemia, late-onset idiopathic cerebellar ataxia, sporadic adult ataxia of unknown etiology (SAOA), and cerebellar ataxia, neuropathy, vestibular areflexia syndrome (CANVAS). In one embodiment, the neurodegenerative disease not associated with lysosomal dysfunction is corticobasal degeneration, SCA 28, and AOA4.

As mentioned above, many neurodegenerative diseases are associated with lysosomal dysfunction, which includes both neurodegenerative lysosomal storage diseases (LSDs) and many other neurodegenerative diseases where links to lysosomal defects are suggested. See, e.g., Boman et al., Journal of Parkinson’s Disease, vol. 6, no. 2, pp. 307–315 (May 2016); Makioka et al., Neurorepor. t, 23(5): 270–276 (March 2012); Orr et al., Alzheimer’s Research & Therapy, 5:53 (Oct. 2013); Barlow et al., Proc. Nat’l. Acad. Sci. USA, 18;97(2): 871–6 (2000).

In one embodiment, the neurodegenerative disease is associated with lysosomal dysfunction (e.g., a lysosomal storage defect). Neurodegenerative diseases according to the present disclosure associated with lysosomal dysfunction include, but are not limited to, alcoholism, Alzheimer's disease, amyotrophic lateral sclerosis (ALS), Canavan disease, frontotemporal lobar degeneration, Huntington's disease, dementia with Lewy bodies, multiple system atrophy (MSA-P/MSA-C), multiple sclerosis, narcolepsy, Parkinson's disease, Smith-Lemli-Opitz syndrome (SLOS) (inborn error of cholesterol synthesis), Tangier disease, Pelizaeus-Merzbacher disease, Pick's disease, frontotemporal dementia and parkinsonism linked to chromosome 17, prion diseases including scrapie, transmissible mink encephalopathy, chronic wasting disease, bovine spongiform encephalopathy (BSE), feline spongiform encephalopathy, exotic ungulate encephalopathy, Creutzfeldt-Jakob disease, Gerstmann-Straussler-Scheinker syndrome and fatal familial insomnia, progressive supranuclear palsy, spinal muscular atrophy, neurodegenerative LPNs, and cerebellar ataxias, which include spinocerebellar ataxia (SCA) 1, spinocerebellar ataxia (SCA) 2, spinocerebellar ataxia (SCA) 3 (Machado-Joseph disease), spinocerebellar ataxia (SCA) 6, spinocerebellar ataxia (SCA) 7, spinocerebellar ataxia (SCA) 17, dentatoturbopallidoluysian atrophy, Autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS), autosomal recessive cerebellar ataxia type 1 (recessive ataxia of Boes (RAB), SYNE-1 mutation), autosomal recessive cerebellar ataxia type 2 (spinocerebellar ataxia autosomal recessive 9, SCAR 9), ataxia with vitamin E deficiency (AVED), ataxia-telangiectasia (Louis-Bar disease), Freidrech ataxia (FRDA), and ataxia with coenzyme Q10 deficiency. In one embodiment, the neurodegenerative disease associated with lysosomal dysfunction is selected from alcoholism, Alzheimer's disease, amyotrophic lateral sclerosis (ALS), Canavan disease, frontotemporal lobar degeneration, Huntington's disease, multiple system atrophy (MSA-P/MSA-C), multiple sclerosis, narcolepsy, Parkinson's disease, Smith-Lemli-Opitz syndrome (SLOS) (inborn error of cholesterol synthesis), Tangier disease, Pelizaeus-Merzbacher disease, Pick's disease, frontotemporal dementia, frontotemporal dementia with parkinsonism, prion diseases, progressive supranuclear palsy, and spinal muscular atrophy. In one embodiment, the neurodegenerative disease associated with lysosomal dysfunction is selected from ALS, MSA-P, MSA-C, frontotemporal dementia with parkinsonism, progressive supranuclear palsy, SCA 28, SCA 1, and Alzheimer's disease.

Neurodegenerative LBNs are characterized by the accumulation of undigested or partially digested macromolecules, leading to cellular dysfunction that is typically progressive, resulting in disability and/or intellectual impairment. They tend to appear in the first few years of life, and their severe progression leads to frequent hospitalization. If left untreated, patients often die in adolescence. Patients with adult-onset disease have also been described. Neurodegenerative LPNs according to the present disclosure include, but are not limited to, neuronal ceroid lipofuscinosis (types 1-10), Gaucher disease, type 2/3 (neuropathic), Krabbe disease, multiple sulfatase deficiency, mucolipidoses including mucolipidosis I, mucolipidosis II, mucolipidosis IV, Niemann-Pick disease type A, Niemann-Pick disease type B, Niemann-Pick disease type C, infantile Pompe disease, late-onset Pompe disease, Tay-Sachs disease, Sandhoff disease, Farber disease, galactosialidosis, Fabry disease, Schindler disease, GM gangliosidosis ₁ , GM gangliosidosis AB variant ₂ , metachromatic leukodystrophy (MLD), mucopolysaccharidoses including MPS IH, MPS IS, MPS IH-S, MPS II, MPS IIIA, MPS IIIB, MPS IIIC, MPS IIID and MPS VII, beta-mannosidosis, aspartylglucosaminuria, fucosidosis, Salla disease, infantile free sialic acid storage disease (ISSD) and Danon disease. In one embodiment, the neurodegenerative LPN is selected from NPC, NPA, mucolipidosis II, MPS IIIB, aspartylglucosaminuria, mucolipidosis IIIA, MPS VII, Sandhoff disease, Tay-Sachs disease, Tay-Sachs variant _AB and GM gangliosidosis. In one embodiment, the neurodegenerative disease is not selected from neurodegenerative LPN.

In one embodiment, the neurodegenerative disease is a motor neuron disease. In one embodiment, the motor neuron disease is selected from primary lateral sclerosis, progressive muscular atrophy, progressive bulbar palsy, pseudobulbar palsy, ALS, Alzheimer's disease, Canavan disease, frontotemporal lobar degeneration, Huntington's disease, multiple sclerosis, narcolepsy, Parkinson's disease, Pelizaeus-Merzbacher disease, and spinal muscular atrophy.

In one embodiment, the neurodegenerative disease is cerebellar ataxia. In one embodiment, the neurodegenerative disease is Niemann-Pick disease. In one embodiment, the neurodegenerative disease is Niemann-Pick disease type C. In one embodiment, the neurodegenerative disease is Niemann-Pick disease type A. In one embodiment, the neurodegenerative disease is parkinsonism. In one embodiment, the neurodegenerative disease is Gaucher's disease. In one embodiment, the neurodegenerative disease is Tay-Sachs disease. In one embodiment, the neurodegenerative disease is Sandhoff disease. In one embodiment, the neurodegenerative disease is Fabry disease. In one embodiment, the neurodegenerative disease is _GM gangliosidosis. In one embodiment, the neurodegenerative disease is Louis-Bar syndrome. In one embodiment, the neurodegenerative disease is Alzheimer's disease. In one embodiment, the neurodegenerative disease is Parkinson's disease. In one embodiment, the neurodegenerative disease is multiple system atrophy. In one embodiment, the neurodegenerative disease is multiple system atrophy type C (MSA-C). In one embodiment, the neurodegenerative disease is multiple system atrophy type P (MSA-P). In one embodiment, the neurodegenerative disease is frontotemporal dementia. In one embodiment, the neurodegenerative disease is frontotemporal dementia with parkinsonism. In one embodiment, the neurodegenerative disease is lower body Parkinson's disease. In one embodiment, the neurodegenerative disease is amyotrophic lateral sclerosis (ALS). In one embodiment, the neurodegenerative disease is corticobasal degeneration syndrome. In one embodiment, the neurodegenerative disease is progressive supranuclear palsy. In one embodiment, the neurodegenerative disease is cerebellar downstroke nystagmus. In one embodiment, the neurodegenerative disease is SCA 28. In one embodiment, the neurodegenerative disease is ataxia-telangiectasia. In one embodiment, the neurodegenerative disease is SCA 1. In one embodiment, the neurodegenerative disease is AOA 4.

The primary symptoms of Parkinson's disease (PD) include rigidity, tremor, and slow movement. Other disorders also have predominantly these symptoms. These disorders, along with PD itself, fall under the broader term "parkinsonism." PD can be called primary parkinsonism. Other examples of parkinsonism include multiple system atrophy, progressive supranuclear palsy, normal-pressure hydrocephalus, and vascular or arteriosclerotic parkinsonism. Disorders that can be classified as parkinsonism but are PD can also be referred to as "parkinsonism-plus syndromes." Unlike patients with PD, individuals with parkinsonism-plus syndromes do not respond to L-dopa. The term "parkinsonism," as used herein, can be used to refer to a movement disorder syndrome whose primary symptoms are resting tremor, rigidity, slow movement, and postural instability. Parkinsonian syndromes can be divided into four subtypes according to their origin: primary or idiopathic, secondary or acquired, hereditary parkinsonism, and parkinsonism-plus syndromes or multisystem degeneration.

In one embodiment, parkinsonism is parkinsonism-plus syndrome or multisystem degeneration.

In one embodiment, parkinsonism is vascular (arteriosclerotic) parkinsonism, lower body parkinsonism, multiple system atrophy with dominant parkinsonism (MSA-P), multiple system atrophy with cerebellar atrophy features (MSA-C; sporadic olivopontocerebellar atrophy (OPCA)), Shy-Drager syndrome, progressive supranuclear palsy (Steele-Richardson-Olszewski syndrome), dementia with Lewy bodies, Pick disease or frontotemporal dementia and parkinsonism linked to chromosome 17.

Niemann-Pick diseases (NPDs) represent a diverse group of autosomal recessive LSDs. Common cellular features include abnormal accumulation of sphingomyelin (SM) in mononuclear phagocytes and parenchymal tissue, as well as (hepato)splenomegaly. Among these three major Niemann-Pick (AC) subgroups, NPC (previously classified as NPC and NPD, and now recognized as a single disease) is classified as a fatal neurovisceral LSD caused by abnormal intracellular cholesterol trafficking—caused by the accumulation of unesterified cholesterol in the late endosome/lysosome compartments. Outside the CNS, cellular characteristics of NPC include abnormal accumulation of unesterified cholesterol and other lipids (e.g., GSL) in the late endosome/lysosome compartments. In contrast, there is no net increase in cholesterol in the CNS (although there is an altered distribution), but there are greatly elevated GSL levels. Progressive neurodegeneration is clearly characterized by subsequent degeneration of GABAergic Purkinje neurons, which occurs in parallel with the onset and development of cerebellar ataxia and other aspects of neurological dysfunction seen in NPC. Genetic studies have shown that NPC disease is caused by mutations in the Npc ₁ or Npc ₂ genes. The precise mechanistic relationship between these two genes remains unknown, and the functional roles of these proteins remain enigmatic. NPC ₁ encodes a multispanning membrane protein of the late endosome/lysosome limiting membrane, while NPC ₂ is a soluble cholesterol-binding protein of the lysosome. When NPC ₁ is inactivated, sphingosine is the first lipid to accumulate, suggesting that NPC ₁ plays a role in sphingosine transport out of the lysosome, where it is normally generated as part of sphingolipid catabolism. Elevated sphingosine, in turn, causes a defect in calcium entry into acidic stores, leading to significantly reduced calcium release from the lysosome. This further prevents late endosome-lysosome fusion, a calcium-dependent process, and causes secondary accumulation of lipids (cholesterol, sphingomyelin, and glycosphingolipids), which are transporters in transit through the late endocytic pathway. Other secondary consequences of inhibiting NPC ₁ function include imperfect endocytosis and a lack of clear autophagic vacuoles. The NPC ₁ /NPC ₂ cellular pathway has been shown to be targeted by pathogenic mycobacteria, promoting their survival in late endosomes.

The NPC mouse model shares several pathological features with, for example, Alzheimer's disease (AD). It has previously been reported that levels of the microtubule-associated protein 1A/3-phosphatidylethanolamine 1B-light chain conjugate (LC3-II) were assessed in NPC mice. LC3-II is a marker of autophagosome formation, and elevated LC3-II levels may reflect impaired clarity of autophagic vacuoles. Autophagosomes are formed but are not clearly defined. Autophagy is impaired in AD, and AD brains exhibit elevated LC3-II levels. Furthermore, amyloid precursor protein (APP) is a precursor molecule whose proteolysis generates beta-amyloid (Aβ). Aβ plaques are a hallmark of AD brains and are thought to be a causal factor in the disease pathology. Amyloid protein precursor protein C-terminal fragments (APP-CTFs), which are intermediates in the proteolysis of APP to Aβ, accumulate in AD brain and also progressively accumulate in the brain of NPC ₁ mice.

Tay-Sachs disease is a fatal inherited disorder of lipid metabolism, characterized primarily in central nervous system tissue due to deficiency of the A isozyme of β-hexosaminidase. Mutations in the HEXA gene, which encodes the α-subunit of β-hexosaminidase, cause deficiency of the A isozyme. Tay-Sachs disease is the prototype of a group of disorders, the _GM2 gangliosidoses, characterized by incomplete breakdown of the _GM2 ganglioside. _GM2 ganglioside (monosialylated ganglioside 2) accumulates in neurons, beginning in fetal life.

Sandhoff disease results from deficiency of both the A and B isozymes of β-hexosaminidase. Mutations in the HEXEB gene, which encodes the β-subunit of β-hexosaminidase, cause deficiency of the B isozyme.

GM gangliosidosis ₁ is caused by a deficiency of β-galactosidase, which results in lysosomal accumulation of ganglioside ₁ (monosialylated ganglioside 1).

Fabry disease is caused by a deficiency of α-galactosidase, which leads to lysosomal accumulation of ceramide trihexoside.

In one embodiment, the neurodegenerative disease is not cerebellar ataxia. In one embodiment, the neurodegenerative disease is not Niemann-Pick disease. In one embodiment, the neurodegenerative disease is not Niemann-Pick disease type C. In one embodiment, the neurodegenerative disease is not cerebellar ataxia or Niemann-Pick disease (e.g., Niemann-Pick disease type C).

In one embodiment, acetylleucine or a pharmaceutically acceptable salt thereof is used to treat weight loss, gait impairment, and/or functional impairment associated with Niemann-Pick disease (e.g., Niemann-Pick disease type C or A) or mucolipidosis type II. For example, acetylleucine or a pharmaceutically acceptable salt thereof can delay the onset, reduce the severity, or eliminate, or delay or reverse, the progression of weight loss, gait impairment, and/or functional impairment associated with Niemann-Pick disease (e.g., Niemann-Pick disease type C or A) or mucolipidosis type II. In one embodiment, the weight loss, gait impairment, and/or functional impairment are associated with Niemann-Pick disease type A or mucolipidosis type II.

In one embodiment, acetylleucine or a pharmaceutically acceptable salt thereof is used to treat gait impairment, motor function impairment, and/or decreased mobility associated with Sandhoff disease. For example, acetylleucine or a pharmaceutically acceptable salt thereof can delay the onset, reduce the severity, eliminate, or delay or reverse the progression of gait impairment, motor function impairment, and/or decreased mobility associated with Sandhoff disease.

In one embodiment, acetylleucine or a pharmaceutically acceptable salt thereof treats decreased coordination, tremors, decreased mobility, cognitive impairment, and/or gait impairment associated with Tay-Sachs disease. For example, acetylleucine or a pharmaceutically acceptable salt thereof can delay the onset, reduce the severity, or eliminate, or delay or reverse, the development of decreased coordination, tremors, decreased mobility, cognitive impairment, and/or gait impairment associated with Tay-Sachs disease.

In one embodiment, acetylleucine or a pharmaceutically acceptable salt thereof treats speech impairment (e.g., speech fluency and/or voice modulation), gait impairment, decreased mobility, decreased swallowing functions, and/or paresis associated with amyotrophic lateral sclerosis (ALS). For example, acetylleucine or a pharmaceutically acceptable salt thereof can delay the onset, reduce the severity, or eliminate, or delay or reverse the development of speech impairment (e.g., speech fluency and/or voice modulation), gait impairment, decreased mobility, decreased swallowing functions, and/or paresis associated with ALS. In another embodiment, acetylleucine or a pharmaceutically acceptable salt thereof treats decreased sleep quality associated with ALS. For example, acetylleucine or a pharmaceutically acceptable salt thereof can delay the onset, reduce the severity, or eliminate, or delay or reverse the development of decreased sleep quality associated with ALS.

In one embodiment, acetylleucine or a pharmaceutically acceptable salt thereof is used to treat speech impairment, gait impairment, and/or increased tendency to fall associated with multiple system atrophy, cerebellar type (MSA-C). For example, acetylleucine or a pharmaceutically acceptable salt thereof can delay the onset, reduce the severity, eliminate, or delay or reverse the progression of speech impairment, gait impairment, and/or increased tendency to fall associated with MSA-C.

In one embodiment, acetylleucine or a pharmaceutically acceptable salt thereof is used to treat gait impairment, increased tendency to fall, and/or speech impairment associated with frontotemporal dementia with parkinsonism. For example, acetylleucine or a pharmaceutically acceptable salt thereof can delay the onset, reduce the severity, eliminate, or delay or reverse the progression of gait impairment, increased tendency to fall, and/or speech impairment associated with frontotemporal dementia with parkinsonism.

In one embodiment, acetylleucine or a pharmaceutically acceptable salt thereof is used to treat increased falling tendency and/or gait impairment associated with corticobasal degeneration syndrome. For example, acetylleucine or a pharmaceutically acceptable salt thereof can delay the onset, reduce the severity, or eliminate, or delay or reverse, the development of increased falling tendency and/or gait impairment associated with corticobasal degeneration syndrome.

In one embodiment, acetylleucine or a pharmaceutically acceptable salt thereof is used to treat gait impairment associated with progressive supranuclear palsy. For example, acetylleucine or a pharmaceutically acceptable salt thereof can delay the onset, reduce the severity, or eliminate, or delay or reverse, the progression of gait impairment associated with progressive supranuclear palsy.

In one embodiment, acetylleucine or a pharmaceutically acceptable salt thereof is used to treat oscillopsia, impaired spatial orientation, decreased visual acuity, and/or increased postural tremor associated with cerebellar downbeat nystagmus. For example, acetylleucine or a pharmaceutically acceptable salt thereof can delay the onset, reduce the severity, eliminate, or delay or reverse the progression of oscillopsia, impaired spatial orientation, decreased visual acuity, and/or increased postural tremor associated with cerebellar downbeat nystagmus.

The present disclosure also relates to a method for treating a neurodegenerative disease or one or more symptoms of a neurodegenerative disease in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of acetylleucine or a pharmaceutically acceptable salt thereof.

A "therapeutically effective amount" of an agent is any amount that, when administered to a subject, is the amount of the agent necessary to produce the desired effect, which, in the case of the present disclosure, may be therapeutic and/or prophylactic. The dose can be determined according to various parameters, such as the specific form of acetylleucine used; the age, weight, and condition of the patient being treated; the type of disease; the route of administration; and the desired administration regimen. A physician will be able to determine the desired route of administration and dosage for any given patient. For example, a daily dose can be from about 10 to about 225 mg/kg, from about 10 to about 150 mg/kg, or from about 10 to about 100 mg/kg of body weight.

The disclosure also relates to a kit for treating a neurodegenerative disease in a subject, comprising means for diagnosing or predicting the disease and acetylleucine or a pharmaceutically acceptable salt thereof.

The means for diagnosing or predicting a neurodegenerative disease may include a specific binding agent, probe, primer, primer pair or combination, enzyme, or antibody, including an antibody fragment, capable of detecting or facilitating the detection of a neurodegenerative disease as defined herein. The kit may include LysoTracker®, a fluorescent marker commercially available from both Invitrogen and Lonza. LysoTracker® may be blue, blue-white, yellow, green, or red.

The kit also includes acetylleucine or a pharmaceutically acceptable salt thereof, as defined herein. The kit may also include buffers or aqueous solutions. The kit may also include instructions for using acetylleucine or a pharmaceutically acceptable salt thereof in the method of the invention.

In another embodiment, acetylleucine or a pharmaceutically acceptable salt thereof is disclosed for use in a method of providing neuroprotection in a subject having, suspected of having, or at risk of developing a neurodegenerative disease.

"Neuroprotection" and similar terms as used herein refer to the prevention, slowing, and/or reversal of neurodegeneration, including, but not limited to, the progressive loss of neuronal structure and/or the progressive death of neurons. Providing neuroprotection may result in a delay in the onset of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease that would otherwise be expected to manifest according to the typical progression of the disease, a reduction in the severity of a neurodegenerative disease or a reduction in the severity of one or more existing symptoms associated with a neurodegenerative disease, a delay in the progression of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease over time compared to the typical progression of the disease, and/or a reversal of the progression of a neurodegenerative disease or one or more symptoms of a neurodegenerative disease over time. The duration of neuroprotection may coincide with the duration of treatment. The treatment may provide neuroprotection for a period of time such as about seven days or longer, about two weeks or longer, about three weeks or longer, about one month or longer, about six weeks or longer, about seven weeks or longer, or about two months or longer. The treatment may also provide neuroprotection for, for example, about three months or longer, about four months or longer, about five months or longer, or about six months or longer. It may provide neuroprotection for, for example, about one year or longer, about two years or longer, about three years or longer, about four years or longer, about five years or longer, or about 10 years or longer. The treatment may provide neuroprotection for the life of the patient.

In one embodiment, a method for providing neuroprotection in a subject in need thereof (e.g., having, suspected of having, or at risk of developing a neurodegenerative disease) comprises administering to the subject a therapeutically effective amount of acetylleucine or a pharmaceutically acceptable salt thereof.

The disclosure also relates to a kit for providing neuroprotection in a subject in need thereof (e.g., having, suspected of having, or at risk of developing a neurodegenerative disease), comprising means for diagnosing or predicting the disease/disorder and acetylleucine or a pharmaceutically acceptable salt thereof.

The present disclosure also relates to the use of acetylleucine or a pharmaceutically acceptable salt thereof as a neuroprotective agent for a subject in need thereof (e.g., having, suspected of having, or at risk of developing a neurodegenerative disease).

All features described in this description (including any accompanying claims, abstract and drawings) and/or all steps of any method disclosed thereby may be combined with any of the aspects described above in any combination, except for combinations in which at least some of such features and/or steps are mutually exclusive.

Examples

The invention will now be explained in more detail in the following examples, which demonstrate the utility of acetylleucine in treating a neurodegenerative disease in a subject and providing neuroprotection in said subject.

Example 1

In vivo mouse study methods

Mouse model

This study uses an authentic mouse model of NPC, the Npc1 ^−/− mouse (BALB/cNctr-Npc1 ^m1N /J), which is null for the NPC1 protein and displays all clinical features of the disease (Loftus, 1997).

This mutant strain arose spontaneously and has a lifespan of 10-14 weeks, resulting in a more acute course of disease, as seen in a large number of patients. The mutant mouse has been successfully used not only to determine the ontogenesis of the disease and underlying pathogenic mechanisms but also to evaluate experimental therapies. Assays using these mice are accepted for all animal models at the cellular and molecular levels (Baudry, 2003; Smith, 2009; Cologna, 2014; Cologna, 2012). The NPC animal model is the most intensively studied.

Until approximately 4–5 weeks of age, Npc1 ^−/− mice show no noticeable behavioral signs of the disease, which distinguishes them from their wild-type counterparts. The first signs of behavioral disturbances, such as tremors and an ataxic gait, appear at 5–6 weeks; at 7–8 weeks, motor coordination impairments become more pronounced, and at 9–11 weeks, ataxia progresses, accompanied by increased weight loss and poor coat condition, with eating and drinking becoming difficult (humane endpoint used) (Smith, 2009).

Wild-type Npc1 ^+/+ pups are used as controls.

Processing protocol

A group of Npc1 ^−/− mice and a group of Npc1 ^+/+ weaned mice (three weeks old) were treated with 0.1 g/kg acetyl-DL-leucine mixed into the mouse chow. Separate groups of Npc1 ^−/− and Npc1 ^+/+ mice were left untreated as controls.

Condition of the coat

The coat condition of Npc1 ^−/− mice treated and untreated with acetyl-DL-leucine was compared by simple observation of mice at nine weeks of age.

Mass data

Animals are weighed twice a week. Weights are averaged (mean calculated) across all mice in each group and compared.

Gait analysis

Gait analysis was performed on eight-week-old mice using the CatWalk® 15.0 system according to the manufacturer's instructions (Noldus, Nottingham, UK). Five walks per animal were recorded.

The measurable parameters of the CatWalk® are:

1) Stand Mean: average duration (s) of contact of paws with the glass platform;

2) Step Cycle: duration (s) between two consecutive contacts of the same paw;

3) Duty Cycle: percentage of time the feet are in contact with the platform compared to the full step cycle;

4) Step Sequence (AB): percentage of time spent walking in the LF-RH-RF-LH pattern (LF: left front; RH: right rear; RF: right front; LH left rear);

5) Cadence: step per second in the test;

6) Diagonal Support: percentage of time that the diagonal paws are in simultaneous contact with the glass platform (RF and LH or RH and LF).

Motor function analysis

Motor function analysis is performed on mice aged eight and nine weeks using an Open Field Activity Monitor according to the manufacturer's instructions (Linton Instruments, Amlogger software). Each mouse is placed in a plastic cage with bedding, and the analysis is performed for five minutes. Risings are counted manually.

The measured parameters of motor function are:

1) Center Rearing: mice stand on their hind legs without support;

2) Rearing: mice stand on their hind legs, leaning or not leaning on the walls of the cage;

3) activity: constant movement of the animal, including walking;

4) Front to Back (FR) count: the animal's movement from the front wall of the cage to the back;

5) active time: duration (sec/min) of activity regardless of movement;

6) mobile time: duration (sec/min) of mobility;

7) Rearing Time: the duration of any standing up.

Results

Condition of the coat

Figure 1B shows an untreated Npc1 ^−/− pup at the appropriate age. Npc1 ^−/− mice are observed to have poor coat condition at nine weeks of age, when eating and drinking become difficult (see Figure 1B).

In stark contrast, Figure 1A shows the Npc1 mouse^−/−, processed acetyl-DL-leucine from weaned Npc1 mice^−/−, treated with DL-leucine, have a smooth and shiny coat resembling that of wild-type (Npc1) pups^+/+) (see Figure 1A).

Mass data

As can be seen in Figure 2A, wild-type (Npc1 ^+/+ ) mice steadily gained weight throughout the study, i.e., from 3 weeks to 10 weeks of age. Furthermore, Figure 2A shows the mean weight per group of mice at each time point (Npc1 ^−/− – untreated, n = 1; Npc1 ^−/− – acetyl-DL-leucine 0.1 g/kg, n = 3; Npc1 ^+/+ – untreated, n = 3; Npc1 ^+/+ – acetyl-DL-leucine 0.1 g/kg, n = 2).

Acetyl-DL-leucine treatment has little effect on this mass gain.

Npc1 ^-/- mice initially gain weight at a similar rate to control Npc1 ^+/+ mice. However, Npc1 ^-/- mice then begin to lose weight at six weeks of age. At the end of the study (at 10 weeks of age), the mice weighed almost as little as they did at four weeks of age.

Treatment with acetyl-DL-leucine delayed these symptoms of weight loss by two weeks compared to the untreated group.

A comparison of weight changes in Npc1 ^−/− mice with and without acetyl-DL-leucine treatment is shown in Figure 2B. Specifically, Figure 2B shows the weight change (%) per group of mice at each time point for Npc1 ^−/− mice only. The beneficial effect of acetyl-DL-leucine treatment—delayed weight loss—is clearly evident from this figure.

Gait analysis

The results of the gait analysis are shown in Figure 3. The data for diagonal support, rhythm, and step sequence are shown in Figures 3A–3C, respectively. Figures 3D and 3E show the data for the forepaws (FP) (mid-stance and step cycle in Figure 3D; work cycle in Figure 3E). Figures 3F and 3G show the data for the hindpaws (HP) (mid-stance and step cycle in Figure 3F; work cycle in Figure 3G). Data are given as mean ± SEM; n=3 for untreated Npc1 ^+/+ , n=2 for treated Npc1 ^+/+ , n=1 for untreated Npc1 ^−/− (therefore statistical analysis was not performed), n=3 for treated Npc1 ^−/− .

The first bar in each diagram shows the gait properties of wild-type (Npc1 ^+/+ ) mice.

The second column in each diagram shows the gait properties of wild-type mice (Npc1 ^+/+ ) treated with acetyl-DL-leucine. There is no significant difference in gait properties between these mice and the same untreated pups.

The third column in each diagram shows the gait properties of the Npc1 ^−/− mouse. Overall, this mouse exhibits a poor gait compared to Npc1 ^+/+ mice. The mouse spends very little time, if any, maintaining a diagonal (Figure 3A) or a step sequence (Figure 3C), and its hind paw functions in midstance (Figure 3F) and the work cycle (Figure 3G) are also radically impaired.

The fourth column in each diagram shows the gait properties of Npc1 ^-/- mice treated with acetyl-DL-leucine. These mice exhibit significantly improved gait compared to the same untreated pups. In fact, they exhibit gait properties similar to Npc1 ^+/+ mice.

Motor function analysis

Analysis in 8-week-old mice showed no difference in motor function properties between Npc1 ^−/− and wild-type (Npc1 ^+/+ ) mice (data not shown).

However, at nine weeks of age, problems with motor coordination appeared.

The results of the motor function analysis at nine weeks are shown in Figure 4. Focused stand-up, stand-up, and back-and-forth (FR) counts are shown in Figures 4A–4D, respectively. Active time, mobile time, stand-up time, and total number of manual stand-ups are shown in Figures 4E–4H, respectively. Data are presented as mean ± SEM; n=3 for untreated Npc1 ^+/+ , n=2 for treated Npc1 ^+/+ , n=1 for untreated Npc1 ^−/− (therefore statistical analysis was not performed), n=3 for treated Npc1 ^−/− .

The first column in each diagram shows the motor function properties of wild-type (Npc1 ^+/+ ) mice.

The second column in each diagram shows the motor performance of wild-type (Npc1 ^+/+ ) mice treated with acetyl-DL-leucine. There is no significant difference in motor performance between these mice and the same untreated pups.

The third column in each diagram shows the motor function properties of the Npc1 ^−/− mouse. Overall, this mouse exhibits poor motor function compared to Npc1 ^+/+ mice. The mouse spends very little time, if any, standing up (panel H), especially on its unsupported hind legs (panel A).

The fourth column in each diagram shows the motor performance of Npc1 ^-/- mice treated with acetyl-DL-leucine. These mice exhibit significantly improved motor performance compared to similar untreated pups. In fact, they exhibit motor performance similar to that of Npc1 ^+/+ mice.

Life expectancy

It was also observed that treatment of Npc1-/- mice with acetyl-DL-leucine (0.1 g/kg from 3 weeks of age) was associated with a statistically significant increase in lifespan (Figure 5). These data also demonstrate the effect of acetyl-DL-leucine on delaying the onset of the disease.

Conclusions

When Npc1 mice^−/−have a distinctive disease signature that distinguishes them from their wild-type counterparts at 5-6 weeks of age, Npc1 pups^−/−, treated with acetyl-DL-leucine, from weaned mice do not display such symptoms for more than two weeks or longer. Treatment of mice Npc1^−/−acetyl-DL-leucine delays the onset and development of symptoms NPC and shows evidence of neuroprotection.

It is reasonable to expect that since acetyl-DL-leucine provides general neuroprotection, the results observed in NPC will also be observed in other neurodegenerative disorders and neurodegenerative disorders that are associated with lysosomal storage defects.

Example 2

Methods

A fibroblast cell line from a patient with NPC is treated with N-acetyl-DL-leucine (1 mM) for 3 days, and the relative lysosome volume is quantified using LysoTracker, a fluorescent dye that accumulates in organelles with acidic contents. More intense LysoTracker fluorescence indicates an increase in lysosome size and/or number and is a hallmark of NPC cells.

In addition, fibroblasts obtained from patients with Niemann-Pick A (NPA), mucolipidosis type II (MLII), mucopolysaccharidosis type IIIB (MPS IIIB), aspartylglucosaminuria, mucolipidosis type IIIA (MLIIIA), and mucopolysaccharidosis type VII (MPS VII) were treated with acetyl-DL-leucine (1 mM) for 6 days, and lysosomal volume was quantified using LysoTracker.

Results

Treatment of fibroblasts from a patient with moderate clinical NPC with 1 mM N-acetyl-DL-leucine was associated with a significant decrease in LysoTracker fluorescence intensity, indicating a decrease in lysosomal volume over time (Figure 6A). These findings were replicated in fibroblasts from patients with other NPCs of varying clinical severity treated with 1 mM N-acetyl-DL-leucine for 72 hours (Figure 6B).

Fibroblasts obtained from patients with NPA, MLII, MPS IIIB, aspartylglucosaminuria, MLIIIA, and MPS VII were observed to have elevated LysoTracker fluorescence levels relative to age-matched wild-type controls (Figures 6C-6H). This is indicative of enlarged lysosomes resulting from lipid accumulation compared to fibroblasts from healthy individuals. Treatment with acetylleucine was associated with a statistically significant decrease in LysoTracker fluorescence relative to control in NPA, MLII, and MPS IIIB fibroblasts relative to untreated NPA, MLII, and MPS IIIB fibroblasts, respectively (Figures 6C-6E), and was associated with a tendency to decrease LysoTracker fluorescence relative to control in aspartylglucosaminuria, MLIIIA, and MPS VII fibroblasts relative to untreated aspartylglucosaminuria, MLIIIA, and MPS VII fibroblasts, respectively (Figures 6F-6H). The decrease in LysoTracker fluorescence is indicative of decreased lysosomal volume (Figures 6C-6H and 6D). Data presented in Figures 6A–6D show the results after 6 days of treatment with 1 mM acetylleucine for each cell line, respectively, for lysosomal volume expressed as a fold change relative to untreated wild-type fibroblasts. Asterisks (*/****) indicate p-values (<0.05/0.001) compared to untreated fibroblasts in disease.

Conclusion

Treatment with N-acetyl-DL-leucine is associated with clearance of impaired lysosomal storage by reducing lysosomal volume, thus directly correcting the phenotype of these lysosomal storage diseases. These diseases represent various classes of LDs, and thus, these results also support the beneficial effects of acetyl-leucine against a wide range of lysosomal storage diseases.

Example 3

Sandhoff disease is a disorder that can result from autosomal recessive inheritance of mutations in the HEXEB gene, which encodes the beta subunit of beta-hexosaminidase. As a result, ganglioside _GM2 is unable to be broken down and accumulates in lysosomes in cells of the peripheral and central nervous system (CNS).

This study uses the Hexb ^−/− mouse model of Sandhoff disease as described in Jeyakumar et al. (Jeyakumar M. et al. (1999) Proc. Natl. Acad. Sci. USA, 96: 6388–6393).

Wild-type mice (Hexb ^+/+ ) are used as controls.

Life expectancy

Treatment with acetyl-DL-leucine is associated with a statistically significant increase in lifespan in mice with Sandhoff disease (Figure 7A). In Figure 7A, acetyl-DL-leucine-treated mice were treated with 0.1 g/kg acetyl-leucine starting at 3 weeks of age. The asterisk (*) indicates a p-value <0.05 vs. untreated mice with Sandhoff disease. Data represent the mean of n=6 mice per group. Without treatment, the median survival time of mice with Sandhoff disease is 112 days. Treatment with acetyl-DL-leucine (0.1 g/kg body weight starting at 3 weeks of age) increased the median lifespan to 120 days.

Motor function

Treatment of mice with Sandhoff disease with acetylleucine results in improvements in motor function as demonstrated by bar crossing and step cycle studies.

Test of transition on a block

The bar-walking test is a method for assessing motor function in mice. In this test, a mouse is suspended by its forelimbs from the center of a horizontal bar. A wild-type mouse with normal motor function will be able to hook its hindlimbs and thereby move toward one of the platforms at either end of the bar, thus completing the test.

Untreated mice with Sandhoff disease are able to perform the test until approximately 11 weeks of age. After this point, motor function and hindlimb mobility/gripping deteriorate to the point where the mouse is unable to perform the test and will fall off the bar onto the soft surface below.

Treatment of a mouse model of Sandhoff disease with acetyl-DL-leucine (0.1 g/kg body weight starting at 3 weeks of age) is associated with improved motor function and hindlimb mobility/gripping as assessed by the bar walk test (Figure 7B). Figure 7B shows treatment with acetyl-DL-leucine at 0.1 g/kg body weight starting at 3 weeks of age. Acetyl-leucine-treated Sandhoff disease mice retain the ability to perform the test up to and including 13 weeks of age. Data represent the mean of 6 mice per group. Treated Sandhoff disease mice retain the ability to perform the test up to and including 13 weeks of age.

Step cycle

The step cycle is the interval of time occupied by a limb during locomotion, from the time it leaves the ground to the time it leaves the ground on the next occasion.

The pacing cycle is assessed at 12 weeks of age in untreated and acetylleucine-treated mice with Sandhoff disease. Acetylleucine treatment consists of 0.1 g of acetylleucine per kg of body weight starting at 3 weeks of age.

Acetylleucine treatment of the Sandhoff disease mouse model was associated with a significantly shorter anterior step cycle time (p<0.05 vs. untreated SH mouse), a significantly shorter posterior step cycle time (p<0.01 vs. untreated SH mouse), and a significantly shorter mean step cycle time (p<0.001 vs. untreated SH mouse) (Figure 7C). In Figure 7C, acetylleucine treatment at 0.1 g/kg body weight is shown from 3 weeks of age. The term “anterior step cycle” refers to the raising of the mouse forelimbs, “posterior step cycle” refers to the raising of the mouse hindlimbs, and the mean step cycle includes all limbs of the mouse. Asterisks (*/**/***) indicate p-values \<0.05/0.01/0.001 vs. untreated Sandhoff disease mouse. Data are presented as mean ± SEM.

Thus, treatment with acetylleucine in a mouse model of Sandhoff disease is associated with a faster step cycle, which may indicate improvement in motor function.

Conclusions

These studies show that treatment of a mouse model of Sandhoff disease with acetylleucine can produce improvements in motor function when assessed in two independent experiments, and also significantly increases lifespan.

Example 4

_GM2 gangliosidoses are a group of lysosomal storage diseases that result from deficiencies in β-hexosaminidase activity. This group includes Tay-Sachs disease, Sandhoff disease, and variant AB of Tay-Sachs disease.

Fibroblasts obtained from patients with _GM2 (Tay-Sachs disease, Sandhoff disease, and variant AB Tay-Sachs disease) and healthy controls were treated with acetyl-DL-leucine (1 mM for 6 days) before extraction and quantification of glycosphingolipid (GSL) levels by high-performance liquid chromatography (HPLC).

In the absence of treatment, fibroblasts from patients with all three _GM2 gangliosidosis variants exhibit elevated GSL levels compared to untreated wild-type controls. In all three cases, treatment with acetyl-DL-leucine (1 mM for 6 days) is associated with decreased GSL accumulation. For Tay-Sachs disease, this reduction is statistically significant (p<0.05). For Sandhoff disease and Tay-Sachs variant AB, there is a trend toward decreased GSL levels in association with treatment. Data presented in Figures 8A–8C show the results of treatment for each cell line, respectively, with GSL levels adjusted for protein content and expressed as fold change relative to levels in untreated wild-type fibroblasts.

Example 5

Patient 1

The patient in this study is a 28-year-old man with a genetic diagnosis of Tay-Sachs disease. He exhibits dystrophony, tremor, ataxia in standing and gait, paraparesis, and muscle atrophy. Specifically, the patient is unable to stand or walk, can take individual steps with good support, has marked postural instability, impaired eye movement, dysphagia, dysarthria, and moderate cognitive impairment. The first symptoms were observed at age 16.

Before treatment, the patient's Severe Ataxia Assessment and Rating Scale (SARA) score showed 15.5/40. Additionally, the patient's Spinocerebellar Ataxia Functional Index (SCAFI) score was obtained:

test average 8-meters Walking (8MW): 21.6 s;

MW 9-Hole Pegboard Test Dominant (9HPTD) (right): 48.3 sec;

MW 9-Hole Pegboard Test Non-Dominant (9HPTND): 44.9 sec;

MW PATA Word Test: 20;

Montreal Cognitive Assessment (MoCA): 18/30.

A video of the patient is also recorded for later comparison.

The day after this check, the patient was started on acetylleucine therapy at a dose of 3 g per day for the first week and then 5 g per day for the second week progressively.

After one month and four months, respectively, the patient was re-evaluated, continuing treatment. After one month, the patient's fine motor skills improved and hand tremors, such as those experienced while eating or drinking, decreased. Walking remained unchanged. After four months, the patient was stable, with slightly improved cognitive function but impaired standing, gait, and fine motor skills. The patient's SARA scores and SCAFI test results are presented below, compared to baseline.

Table 1. Patient assessment parameters

Base After one month with acetyl-DL-leucine After 4 months with acetyl-DL-leucine SARA 15.5/40 15.5/40 17/40 8 MWT 21.6 s 7 s 25.49 s 9HPTD 48.3 s 45.9 s 48.67 s 9HPTND 44.9 s 40.1 s 47.09 s PATAPPLICATION 20 22 21 MoCA 18/30 21/30 22/30

Overall, the patient shows improvement in symptoms after treatment with acetylleucine.

Patient 2

The patient in this study is a 32-year-old woman with a genetic diagnosis of Tay-Sachs disease. She exhibits standing and gait ataxia, fine motor impairment, lower extremity paraparesis, and muscle atrophy. Specifically, the patient is unable to walk without support and suffers from dysphagia and speech impairment, eye movement impairment, and moderate cognitive impairment. The first symptoms were observed at age 7.

Before treatment, the patient's Severe Ataxia Assessment and Rating Scale (SARA) score showed 10.5/40. Additionally, the patient's Spinocerebellar Ataxia Functional Index (SCAFI) results were obtained:

test average 8-meters Walking (8MW): 12.5 s;

MW 9-Hole Pegboard Test Dominant (9HPTD) (right): 21.5 sec;

MW 9-Hole Pegboard Test Non-Dominant (9HPTND): 35.5 sec;

MW PATA Word Test: 18;

Montreal Cognitive Assessment (MoCA): 21/30.

A video of the patient is also recorded for later comparison.

On the day of the examination, the patient was started on acetylleucine therapy at a dose of 3 g per day for the first week and then 5 g per day for the second week progressively.

After one month, the patient was re-evaluated while continuing treatment, and improved pronunciation, postural stability, and cognitive function were noted. Standing and walking became possible without support. The patient's SARA scores and SCAFI results are presented below, compared to baseline.

Table 2. Patient assessment parameters

Base After one month with acetyl-DL-leucine SARA 10.5/40 5/40 8 MWT 12.5 s 9.55 s 9HPTD 21.5 s 34.97 s 9HPTND 35.5 s 39.34 s PATA 18 17 MoCA 21/30 25/30

Patient 3

The patient in this study is an 8-year-old boy with a genetic diagnosis of Tay-Sachs disease. He experiences epileptic seizures (grand, approximately 10 seconds long, self-limited) almost daily before falling asleep, along with impaired eye movement, anarthria, various cognitive and concentration problems (neurological testing not possible), an inability to stand or walk independently, and very limited daily activities (eating, washing, or dressing independently). The first symptoms were observed at 9 months of age.

Before treatment, the patient's SARA score was 36/40, mRDS 18/24, EQ-5D-5L visual scale 50, and 8MWT 18.1 (only with good support).

The patient was started on acetylleucine therapy at a dose of 1.5 g per day for the first week and then 3 g per day for the second week progressively.

After one month, the patient is re-evaluated while continuing treatment. Improved fine motor skills (able to pick up small objects), increased mobility (more often attempting to walk independently), improved postural stability, gait, and standing, as well as the ability to pronounce individual words, are noted. The patient's SARA, mRDS, EQ-5D-5L visual scale, and 8MWT scores are presented below, compared to baseline.

Table 3. Patient assessment parameters

Base After one month on acetyl-DL-leucine SARA 36/40 33/40 mRDS 18/24 16/24 Visual scale
EQ-5D-5L 50 60 8 MWT 18.1 (only with good support) 11.75 (with support on one arm)

Example 6

The patient in this study is a 13-year-old boy genetically diagnosed with _GM1 gangliosidosis. He is unable to stand or walk independently, has limited daily activities (eating, washing, or dressing independently), and has impaired eye movement, anarthria, and various cognitive and concentration problems (neurological testing is not possible). The first symptoms were observed at age 2.

Before treatment, the patient's SARA score was 35/40, mRDS score 15, and EQ-5D-5L visual scale score 50.

The patient was started on acetylleucine therapy at a dose of 1.5 g per day for the first week and then 3 g per day for the second week progressively.

After one month, the patient is re-evaluated while continuing treatment, and overall stability, improved gait (smoother), and stable standing in a natural position are noted. The patient's SARA, mRDS, and EQ-5D-5L visual scale scores are presented below, compared to baseline.

Table 4. Patient assessment parameters

Base After one month on acetyl-DL-leucine SARA 35/40 35/40 mRDS 15 16 Visual scale EQ-5D-5L 50 60

Example 7

Patient 1

The patient in this study is a 73-year-old man previously diagnosed with amyotrophic lateral sclerosis (ALS).

The patient's symptoms are characterized by progressive dysarthria (nasal and slurred speech) and right dorsiflexor weakness with foot drop resulting from a history of foot drop over the previous three years.

Clinically, the patient exhibits bulbar speech, paresis of three-fifths of the right dorsiflexors and the left thumb, generalized exaggerated reflexes, and spastic increased tone in the right lower extremity. EMG shows spontaneous activity, and cMRI reveals no abnormalities.

The patient began treatment with riluzole around the time of his ALS diagnosis. However, the clinical symptoms remain unchanged.

The patient then begins therapy with acetyl-DL-leucine at a dose of 3 g per day for the first week and then 5 g per day for the second week, gradually increasing. The results are recorded on video.

After 15 days of treatment, a follow-up assessment is conducted, during which the patient reports a significant improvement in speech. The patient is able to speak more fluently and modulate their voice better than before treatment (as confirmed by video).

After another 20 days of treatment, a follow-up examination is performed, during which the patient reports further improvement in speech. Furthermore, the patient reports an improvement in gait. The right dorsiflexor palsy and foot drop have significantly improved and are now almost clinically undetectable. Furthermore, the patient reports improved sleep, falling asleep more quickly, staying asleep longer, and feeling distinctly more rested in the morning.

The patient continues treatment for approximately 30 more days. Approximately 7 days after stopping treatment, a follow-up assessment is performed, during which the patient reports no further subjective improvement in speech or relief of right dorsiflexor palsy. Sleep also worsens. After approximately 1-2 additional weeks without acetylleucine treatment, the patient reports worsening speech. At this time, the patient resumes treatment, and approximately two months later, reports stable symptoms. Compared to when acetylleucine treatment was first initiated, a slight worsening of speech is observed.

Since the patient's speech had not improved, he requested discontinuation of the medication. Approximately 2-3 weeks later, the patient again reported worsening speech after discontinuing acetylleucine therapy. He resumed treatment and reported improvement in his symptoms, particularly his speech.

Overall, the patient shows improvement in symptoms after treatment with acetylteicin.

Patient 2

The patient in this study is a 74-year-old man previously diagnosed with ALS.

The patient's symptoms are characterized by progressive dysarthria (nasal and slurred speech) with accompanying dysphagia and weakness when walking for a year, and paresis of the left upper limb for approximately four months. EMG shows generalized multiphasic activity and chronic neurogenic deterioration in the bulbar, cervical, and lumbar segments.

Clinical examination of the patient reveals severe dysarthria, tongue hypokinesia, paresis of two-fifths to three-fifths of the left hand with impaired fine motor skills, generalized exaggerated reflexes, and fasciculations. Treatment with riluzole had been initiated one month earlier.

The patient is started on acetyl-DL-leucine at a dose of 3 g per day for the first week and then 5 g per day for the second week progressively.

Approximately two months later, the patient is re-examined and reports a progressive deterioration in motor function in his left leg, but a discrete improvement in walking. Swallowing function also remains stable.

Patient 3

The patient in this study is a 66-year-old man previously diagnosed with ALS.

The patient's symptoms are characterized by progressive weakness and atrophy of both proximal upper extremities, mild deterioration of fine motor skills, and generalized fasciculations and seizures. An EMG shows abnormal spontaneous activity, and an MRI of the brain and cervical spine reveals no abnormalities. Treatment with riluzole is initiated.

Approximately two months later, a medical examination reveals paresis of 3/5-4/5 of both shoulders and the proximal parts of the arms, along with slowed fine motor skills, generalized fasciculations, and normal reflexes. The patient is started on acetyl-DL-leucine at a dose of 3 g per day for the first week and then 5 g per day for the second week, gradually increasing.

After one month, the patient reports no improvement in symptoms or upper limb muscle strength. Medication is discontinued, and the patient is asked to report any worsening symptoms.

Patient 4

The patient in this study is a 66-year-old man diagnosed with ALS. His symptoms include progressive weakness and atrophy of both proximal upper extremities, mild fine motor impairment, and generalized fasciculations and seizures. An EMG reveals abnormal spontaneous activity and chronic neurogenic changes. An MRI of the brain and cervical spine reveals no abnormalities. Treatment is initiated with riluzole.

Physical examination reveals paresis of 3/5-4/5 of both shoulders and proximal arms, slow fine motor skills, generalized fasciculations and normal reflexes, and an ALS-FRS score of 44/48.

The patient is started on acetyl-DL-leucine at a dose of 3 g per day for the first week and then 5 g per day for the second week progressively.

After approximately one month, the patient reports subjective improvement in dysphagia and reduced hypersalivation. A comparison of their results reveals an improved and more lively facial expression. Limb weakness remains unchanged. Therapy is suspended until the patient reports worsening symptoms, particularly subjective worsening of dysphagia and hypersalivation. The patient resumes continuous treatment.

The patient is re-evaluated approximately eight weeks later, and his symptoms remain stable. His ALS-FRS score is 44/48. Compared to his symptoms at approximately the time of diagnosis, there is only slight progression of weakness in gait and the upper extremities.

Example 8

Treatment with acetylleucine shows improvement in 3 patients diagnosed with multiple system atrophy, cerebellar type (MSA-C).

Patient 1

In this study, the patient is a woman in her late 60s who developed progressive ataxia with speech problems and walking difficulties over the previous three years.

A medical examination revealed central cerebellar gastromotor symptoms, moderate dysarthrophonia, mild limb ataxia, and moderate ataxia in standing and gait. Furthermore, an MRI revealed atrophy of the cerebellum and brainstem, particularly the pons and midbrain. The patient was therefore diagnosed with MSA-C.

The patient is started on acetyl-DL-leucine at a dose of 5 g per day (2 g after sleep, 1.5 g before lunch and 1.5 g before dinner).

After one week of treatment, the patient already shows significant improvement in speech.

Patient 2

In this study, the patient is a 77-year-old man diagnosed with MSA-C.

The patient's symptoms are characterized by progressive difficulty walking and an unsteady gait with a tendency to fall (the patient falls approximately 10 times per month). The patient has exhibited dizziness, hypokinetic-rigid syndrome, saccadic eye movements, dysmetria in coordination tests, and autonomic dysfunction, such as erectile dysfunction, orthostatic hypotension, and incomplete bladder emptying, for the past four years.

Before treatment, the patient's symptoms had remained unchanged for at least three months.

The patient is started on acetyl-DL-leucine at a dose of 3 g per day for the first week and then 5 g per day.

After three weeks of treatment, a follow-up examination is performed. The patient and his wife report a significant improvement in their gait: they walk more confidently and have completely stopped falling. Furthermore, the dizziness they experienced has significantly decreased.

The patient is instructed to discontinue medication, and after one week, the patient reports a worsening of gait and dizziness. The patient also reports a feeling of increased unsteadiness in their gait, with a strong tendency to fall.

The patient is then instructed to resume medication for 40 days, after which it is discontinued again. Following a medical examination seven days after discontinuing medication, the patient reports progressive gait deterioration and dizziness two days after discontinuing treatment, and a severe tendency to fall five days after discontinuing treatment. The patient is then returned to continuous treatment.

Patient 3

In this study, the patient is a 76-year-old man diagnosed with asymptomatic MSA-C.

The patient's symptoms are characterized by progressive difficulty walking and an unsteady gait (without falling), as well as dizziness.

Clinically, the patient exhibits saccadic eye movements and dysmetria in coordination tests. cMRI reveals midbrain atrophy, and FDC-PET of the brain reveals decreased metabolism in the striatum and cerebellum. The patient's posturography results are abnormal, with a high tendency to fall.

Before treatment, the patient's clinical symptoms remained unchanged for at least one year.

A gait analysis is performed, revealing an ataxic gait, reduced speed, and increased track width compared to normal gait spacing and fluctuations. The patient is then treated with acetyl-DL-leucine at a dose of 3 g per day for the first week and then 5 g per day for the second week, gradually increasing.

Following treatment, a follow-up examination is performed within one month. Gait analysis reveals improved walking speed and reduced gait width and fluctuations.

Table 5. Gait analysis parameters

Soon after starting treatment After 27 days of treatment Normal range (± SD) Speed (cm/s) 72 106 110.81 (18.33) Max. speed (cm/s) 183 208 158.30 (22.66) Rhythm (steps/minute) 101 113 109.19 (12.75) Track width (cm) 16.8 14.6 9.06 (1.94) Length of step cycle (cm) 87 113 121.81 (11.52) Double standing (%) 32.5 27.3 20.73 (2.55) Coefficient of variation (temporal) 3.3 3.1 1.94 (0.85) Functional gait assessment 21/30 20/30 24.9 (3.6)

The patient is instructed to discontinue drug therapy and reports progressive gait deterioration and dizziness approximately two to three weeks after discontinuing treatment.

The patient then returns to continuous treatment, and symptoms improve again. Treatment is suspended again, and the patient is assessed three weeks later. The patient reports worsening symptoms, particularly dizziness. Gait analysis reveals increased stride width compared to pre-therapy.

Table 6. Gait analysis parameters

20 days after repeated interruption of treatment Speed (cm/s) 106 Max. speed (cm/s) 197 Rhythm (steps/minute) 112 Track width (cm) 17.5 Length of step cycle (cm) 115 Double standing (%) 27.1 Coefficient of variation (temporal) 2.5 Functional gait assessment 22/30

Example 9

The patient in this study is a 59-year-old man with a progressive personality disorder characterized by apathy, lethargy, and indifference. Furthermore, the patient exhibits hypokinetic-rigid syndrome, primarily on the left side, with impaired fine motor skills and decreased resonation in the left hand. Furthermore, the patient exhibits generalized bradykinesia and gait disturbance with small steps and 2-3 falls per month. The patient also exhibits slurred speech and cognitive impairments, including psychomotor retardation and reduced semantic fluency.

The patient is diagnosed with frontotemporal dementia with Parkinsonism, and Datscan detects decreased dopamine receptors, supporting the diagnosis. FDC-PET of the brain shows primarily decreased metabolism in the frontal lobe.

The patient shows slight improvement during treatment with L-dopa and ropinirole.

The patient is started on acetyl-DL-leucine at a dose of 3 g per day for the first week and then at a dose of 5 g per day for 4 weeks.

After treatment with acetylleucine for approximately one month, drug treatment is stopped and the patient is examined again after 13 days.

The patient, his wife, and daughter report a significant improvement in their gait after acetylleucine therapy, and the patient has also stopped falling. The patient also shows improvement in speech, which is less slurred, more understandable, and subjectively significantly more controlled. After discontinuing treatment, the symptoms worsen.

Example 10

The patient in this study is a 75-year-old man with progressively unsteady gait and dizziness, leading to backward falls. He also has a predominantly left-sided hypokinetic-rigid syndrome with apraxia and "alien" limb phenomenon.

The patient was diagnosed with corticobasal syndrome. Datscan revealed decreased dopamine receptors, and MRI showed atrophic motor cortex in the right hemisphere, supporting the diagnosis.

The patient shows no improvement during treatment with L-dopa.

The patient is started on acetyl-DL-leucine at a dose of 3 g per day for the first week and then 5 g per day. A gait analysis is performed before starting treatment.

After 20 days of treatment with acetylleucine, the patient was re-examined. No improvement in dizziness symptoms or significant reduction in the frequency of falls was observed.

Table 7. Gait analysis parameters

Before treatment After 20 days of treatment Mean (± SD) Speed (cm/s) 63 92 110.81 (18.33) Max. speed (cm/s) 131 152 158.30 (22.66) Rhythm (steps/minute) 90 106 109.19 (12.75) Track width (cm) 12.1 12.3 9.06 (1.94) Length of step cycle (cm) 84 104 121.81 (11.52) Double standing (%) 29.8 26.6 20.73 (2.55) Coefficient of variation (temporal) 3.9 3.4 1.94 (0.85) Functional gait assessment 13/30 18/30 24.9 (3.6)

There is an objective improvement in gait analysis parameters such as speed, maximum speed, rhythm and reduced double stance (Table 2 and Figure 9).

After 8 weeks of treatment with acetylleucine, drug treatment is stopped and the patient is examined again after 6 days.

The patient reports an increase in dizziness symptoms two days after stopping treatment (feeling like a drunk).

The patient then resumes continuous treatment.

Example 11

Patient 1

The patient in this study is a 76-year-old woman with dizziness, primarily occurring while walking. No falls were reported. The patient also exhibited gait disturbance with small steps and generalized bradykinesia, as well as vertical gaze palsy with impaired fine motor skills.

The patient is diagnosed with progressive supranuclear palsy. Datscan reveals decreased dopamine receptors, and an FDC-PET scan of the brain shows primarily decreased metabolism in the frontal lobe, supporting the diagnosis.

The patient shows slight improvement during treatment with L-dopa.

The patient is started on acetyl-DL-leucine at a dose of 3 g per day for one week and then 5 g per day for four weeks. After 27 days of treatment, the medication is discontinued, and the patient is re-evaluated after 60 days.

The patient reported a significant reduction in dizziness and a slight improvement in gait after treatment with acetylleucine. After discontinuing treatment, the symptoms worsened.

The patient was re-evaluated approximately two months later, and was reported to have persistent symptoms of the underlying progressive supranuclear palsy without clinical progression. Her PSSRS score remained stable, and the dizziness relief was still significant.

Patient 2

The patient in this study is a 66-year-old man with symmetrical hypokinetic-rigid syndrome, a gait disorder characterized by unsteady and small steps (a strong tendency to fall), and vertical gaze palsy, with impaired fine motor skills. The patient is diagnosed with progressive supranuclear palsy. Datscan reveals decreased dopamine receptors, and FDC-PET of the brain shows primarily decreased frontal lobe metabolism, supporting the diagnosis. He is unresponsive to levodopa.

The patient is started on acetyl-DL-leucine at a dose of 3 g per day for the first week and then 5 g per day. A gait analysis is performed before treatment. After 17 days of treatment, medication is discontinued, and the patient is re-evaluated after 4 days. The patient reports no significant improvement in gait or hypokinetic-rigid syndrome.

Table 8. Gait analysis parameters

Before treatment After 17 days of treatment Normal values (± SD) Speed (cm/s) 51 68 119.12 (17.27) Max. speed (cm/s) 123 98 176.78 (19.10) Rhythm (steps/minute) 93 99 113.06 (10.38) Track width (cm) 10.3 9.3 9.49 (3.56) Length of step cycle (cm) 66 82 126.71 (13.06) Double standing (%) 34.5 28.6 20.35 (3.21) Coefficient of variation (temporal) 8.5 6.6 1.76 (0.73) Functional gait assessment 15/30 15/30 27.1 (2.3)

The patient is re-evaluated after approximately two months and reports no worsening of symptoms after stopping drug treatment.

Patient 3

The patient in this study is a 56-year-old man with symmetrical hypokinetic-rigid syndrome, an unsteady gait with falls, vertical gaze palsy, and impaired fine motor skills. The patient is diagnosed with progressive supranuclear palsy. Datscan reveals decreased dopamine receptors, and FDC-PET of the brain shows decreased metabolism in the frontomesial and temporoparietal regions, supporting the diagnosis. He is unresponsive to levodopa.

The patient is started on acetyl-DL-leucine at a dose of 3 g per day for the first week and then 5 g per day. A gait analysis is performed before treatment. After 17 days of treatment, medication is discontinued, and the patient is re-evaluated after 4 days. The patient reports no significant improvement in gait or hypokinetic-rigid syndrome.

Table 9. Gait analysis parameters

Before treatment After 17 days of treatment Normal values (± SD) Speed (cm/s) 103 120 125.34 (20.66) Max. speed (cm/s) 163 194 180.07 (26.57) Rhythm (steps/minute) 106 112 115.27 (12.02) Track width (cm) 15.5 15.1 9.12 (2.97) Length of step cycle (cm) 118 129 130.34 (13.01) Double standing (%) 25.8 24.3 19.05 (2.75) Coefficient of variation (temporal) 3.6 3.1 1.77 (1.04) Functional gait assessment 28/30 27/30 28.4 (1.6)

The patient is re-evaluated after approximately two months and reports no worsening of symptoms after stopping drug treatment.

Patient 4

The patient in this study is a 76-year-old man with a progressive gait disturbance, including unsteady and small steps (a strong tendency to fall), camptocormia, slow and hypometric saccades, and impaired fine motor skills. The patient is diagnosed with progressive supranuclear palsy. An MRI reveals mild midbrain atrophy (Mickey Mouse sign). He has a poor response to levodopa.

The patient is started on acetyl-DL-leucine at a dose of 3 g per day for the first week and then 5 g per day. A gait analysis is performed before treatment. After three weeks of treatment, medication is discontinued, and the patient is re-examined. The patient reports increased subjective confidence in gait and a reduced incidence of falls. Gait analysis shows improved gait with increased speed and step cycle length and decreased stance width, double stance, and coefficient of variation.

Table 10. Gait analysis parameters

Before treatment After 3 weeks of treatment Normal values (± SD) Speed (cm/s) 39 65 110.81 (18.33) Max. speed (cm/s) 70 93 158.30 (22.66) Rhythm (steps/minute) 109 125 109.19 (12.75) Track width (cm) 15.4 13.6 9.06 (1.94) Length of step cycle (cm) 43 63 121.81 (11.52) Double standing (%) 45.4 36 20.73 (2.55) Coefficient of variation (temporal) 8.2 6.8 1.94 (0.85) Functional gait assessment 14/30 24.9 (3.6)

After three months without medication, the patient developed hypokinetic-rigid syndrome. His gait worsened, with an increased frequency of falls.

Example 12

Patient 1

In this study, the patient is a 42-year-old engineer who has been suffering from dizziness and postural imbalance for almost one year.

The patient was diagnosed with downward swaying nystagmus: his condition worsened significantly due to blurred vision (oscillopsia) due to the nystagmus and difficulty reading and writing. His visual acuity is as follows: right 0.75, left 0.67, binocular 0.83, and the downward swaying nystagmus is confirmed by video-oculography. The patient also exhibits a body swaying motion, which is confirmed by posturography.

Treatment with 4-aminopyridine (Fampyra, 10 mg twice daily) for four weeks does not produce any beneficial effect.

The patient is started on acetyl-DL-leucine at a dose of 3 g per day (1 g upon waking, 1 g before lunch, and 1 g before dinner) for one week and then at a dose of 5 g per day (2 g upon waking, 1.5 g before lunch, and 1.5 g before dinner).

After 10 days, the patient reports significant improvement and that the effect is gradually developing. The patient continues treatment at the prescribed dosage and experiences no side effects. Temporary interruption of treatment leads to significant deterioration.

The patient was seen again approximately 14 weeks after starting treatment with acetylleucine, during which time the patient was very satisfied with the beneficial effect.

The patient's reading and writing have improved significantly, as the oscillopathy has subsided: the visual image is stable. The patient is able to suppress nystagmus by fixating his gaze. Furthermore, his spatial orientation has improved.

Clinical examination by two independent examiners revealed a reduction in nystagmus, and video-oculography showed that the patient could suppress nystagmus with gaze fixation. The patient's visual acuity was: right 0.83, left 1.0, binocular 1.

Posturography shows a reduction in postural sway.

Overall, the study in this case shows a reduction in the patient's symptoms for this indication.

Patient 2

The patient in this study was diagnosed with downward swaying nystagmus. He exhibited postural imbalance and dizziness. Fampira® did not improve the patient's symptoms.

The patient is started on acetyl-DL-leucine (3 g/day for the first week, then 5 g/day) and subsequently shows an improvement in gait with the ability to walk longer distances (one hour) and improved agility. The patient also has a decrease in downswing nystagmus (confirmed by videooculography). The patient can partially suppress nystagmus by fixating his gaze when assessed using a target center (a dot placed in the center of the display for 30 seconds, Figure 13A) and in complete darkness using protective glasses coated with special lenses for 45 seconds (Figure 13B). The results (mean slow phase velocity, SPV) are as follows: target center - horizontal -0.02°/s, vertical 2.41°/s; complete darkness - horizontal 0.05°/s, vertical 3.27°/s (Figure 13C). The patient is able to minimize eye movement by applying fixation force as shown in Figure 13A.

Gait analysis reveals an increase in self-selected walking speed from 56 to 85 cm/s and maximum walking speed from 122 to 155 cm/s. Drug therapy is then discontinued. Approximately one month after discontinuing acetyl-DL-leucine, the patient's symptoms worsen. Gait analysis reveals a decrease in self-selected walking speed from 85 to 72 cm/s and maximum walking speed from 155 cm/s to 113 cm/s.

Example 13

Patient 1

In this study, the patient is a 70-year-old woman with kinetic rigid syndrome mainly on the right side and tremor, antecollis, frequent falls, orthostatic dysfunction and incontinence.

The patient was diagnosed with multiple system atrophy with parkinsonism (MSA-P). Datscan reveals a predominantly left-sided decrease in dopamine receptors, and FDC-PET of the brain shows decreased metabolism primarily in the parieto-occipital region. There is a poor response to levodopa (100/25 mg 3 times daily).

The patient begins taking acetyl-DL-leucine (3 g/day for the first week; then 5 g/day). After 3 weeks on acetyl-DL-leucine, the patient is examined and no significant improvement in gait, falls, or kinetic rigidity is noted. Medication is discontinued.

After 6 weeks, the patient reported no worsening of symptoms after discontinuing drug treatment.

Patient 2

The patient in this study is a 78-year-old man diagnosed with multiple system atrophy with parkinsonism (MSA-P). The patient's symptoms are characterized by progressive kinetic rigidity syndrome, orthostatic dysfunction, and subsequent dizziness and balance disturbances. The patient exhibits saccadic eye movements and symmetrical muscle rigidity in both upper extremities. Balancing on an imaginary rope is associated with insecurity and loss of balance. FDC-PET of the brain reveals decreased metabolism in both the parietal and occipital cortex, reminiscent of dementia with Lewy bodies.

The patient is started on acetyl-DL-leucine at a dose of 3 g per day for the first week, then 5 g per day. Before starting treatment, the patient is examined and noted to exhibit the clinical symptoms described above, including very pronounced gait instability and dizziness.

After one month of treatment, medication is discontinued and the patient is evaluated. The patient reports a subjective improvement in dizziness, and clinical testing reveals improved balance on an imaginary rope, which the patient performs without any difficulty compared to the previous test. Gait analysis is performed.

After one month of untreated treatment, the patient reports stable symptoms. No worsening of dizziness or gait instability is reported. A gait analysis is performed.

After two months without treatment, a gait analysis is performed, revealing a decrease in walking speed, shortening of stride length, and a worsening of the FGA score. The patient's overall symptoms worsen, including progressive leg weakness and increased gait instability.

Table 11. Gait analysis parameters

After 1 month of treatment After 1 month without treatment After 2 months without treatment Normal values (± SD) Speed (cm/s) 109 114 94 110.81 (18.33) Max. speed (cm/s) 214 196 177 158.30 (22.66) Rhythm (steps/minute) 109 109 106 109.19 (12.75) Track width (cm) 3.8 2.3 3.4 9.06 (1.94) Length of step cycle (cm) 120 126 107 121.81 (11.52) Double standing (%) 25.6 22.5 26.7 20.73 (2.55) Coefficient of variation (temporal) 2.7 1.7 2.1 1.94 (0.85) Functional gait assessment 24/30 25/30 22/30 24.9 (3.6)

Patient 3

The patient in this study is a 78-year-old man diagnosed with multiple system atrophy with parkinsonism (MSA-P). The patient's symptoms include progressive kinetic rigidity syndrome, urinary incontinence, emerging cognitive dysfunction, and gait disturbance with small steps and 2-3 falls per month. Cognitive impairments include psychomotor retardation and intermittent confusion. Datscan reveals decreased dopamine receptors, supporting the diagnosis. FDC-PET of the brain reveals decreased metabolism, primarily in the striatum. Levodopa therapy is discontinued due to adverse effects.

The patient is started on acetyl-DL-leucine at a dose of 3 g per day for the first week, then 5 g per day. After one month on acetyl-DL-leucine, the patient is assessed. The patient's wife reports a significant improvement in cognitive function. Episodes of confusion have completely disappeared. The patient's cognitive structure appears clearer and more accurate. There is no improvement in walking ability. The patient's wife supports continued medication.

Example 14

The patient in this study is a 45-year-old man diagnosed with spinocerebellar ataxia type 28 (SCA 28). Genetic testing reveals a known pathogenic variant in AFC3L2. The patient's symptoms include a progressive cerebellar syndrome since age 30, characterized by slurred speech, unsteady gait, impaired balance, and dizziness. The patient's father and grandmother suffered from similar symptoms. The patient exhibits saccadic eye movements and dysmetria in coordination tests, an ataxic gait, slurred speech, exaggerated lower extremity reflexes, lower extremity spasticity, and a positive Babinski sign on the left. cMRI reveals marked cerebellar atrophy.

The patient is started on acetyl-DL-leucine at a dose of 5 g per day. A gait analysis is performed before treatment. After approximately one month of treatment, medication is discontinued, and the patient is assessed. The patient reports an improvement in symptoms, specifically a decrease in dizziness (almost eliminated) and a more stable gait. The patient reports no longer walking like a robot and can climb stairs without using a handrail. A gait analysis is performed, which shows improvement.

Table 12. Gait analysis parameters

Before treatment After treatment one month Standard values (± SD) Self-selected speed (cm/s) 101 118 126.61 (21.43) Maximum speed (cm s) 188 201 182.94 (25.37) Rhythm (steps/minute) 100 105 115.52 (9.36) Track width (cm) 14.5 11.7 9.43 (2.27) Length of step cycle (cm) 122 135 131.55 (17.98) Double standing (%) 24.9 23.8 18.76 (3.46) Coefficient of variation (temporal) 4.1 2.1 1.88 (0.79) Functional gait assessment 24/30 23/30 28.9 (1.5)

Example 15

Patients 1 and 2

The patients in this study are two sisters, aged 24 (patient 1) and 19 (patient 2), respectively. Both patients suffer from ataxia-telangiectasia.

Patient 1 exhibits developmental delay. She did not walk until age 2 and developed signs and symptoms of cerebellar ataxia, including paroxysms with generalization, distal marked hypertension, and telangiectasia affecting the eyes, ears, and chest. She was diagnosed at age 9. Her oculomotor function includes downward swaying nystagmus with gaze directed straight ahead and more pronounced leftward than rightward, upward gaze holding nystagmus, vertical and horizontal saccadic smooth pursuit, and hypometric horizontal and vertical saccades with limited mobility during upward gaze.

Before treatment, Patient 1's score on the Scale for Ataxia Assessment and Rating (SARA) showed 22/40. The results of the Spinocerebellar Ataxia Functional Index (SCAF1) analysis were as follows:

test average 8-meters Walking (8MW): 21.8 s;

MW 9-Hole Pegboard Test Dominant (9HPTD) (right): 90.2 sec;

MW 9-Hole Pegboard Test Non-Dominant (9HPTND): 125.8 sec;

MW PATA Word Test: 12.5;

Visual analogue scale (patient assessment): 99.

The results of videooculography are given below.

Table 13. Videooculography parameters

Velocity of the slow phase of gaze holding nystagmus Horizontal Vertical Center 0.54 6.9 To the right -1.16 6.2 To the left 12.2 12.6 Down -0.14 6.9 Up -0.39 5.95

Following evaluation, Patient 1 begins treatment with acetyl-DL-leucine (5 g/day). After one month of treatment, the patient is re-evaluated. Caregivers report improvement in speech and gait. The patient herself does not perceive any changes. A scale assessment and rating scale (SARA) score of 15.5/40 is obtained. The results of the spinocerebellar ataxia functional index (SCAF1) are as follows:

test average 8-meters Walking (8MW): 18.5 s;

MW 9-Hole Pegboard Test Dominant (9HPTD) (right): 77.9 sec;

MW 9-Hole Pegboard Test Non-Dominant (9HPTND): 101.3 sec;

MW PATA Word Test: 13;

Visual analogue scale (patient assessment): 85.

The results of videooculography are given below.

Table 14. Videooculography parameters

Velocity of the slow phase of gaze holding nystagmus Horizontal Vertical Center 0.4 4.77 To the right -0.95 4.83 To the left 12.2 12.6 Down -0.44 5.29 Up -0.72 2.67

Patient 1's SARA and SCAF1 scores improved after treatment, and video-oculography shows significant improvement in fixation stability and a decrease in the intensity of downstroke nystagmus.

Patient 2 presented with developmental delay, paroxysms at 1 year of age, generalized hypotonia, lower limb contractures with bilateral equinovarus deformities of the feet, areflexia, acute lymphoblastic leukemia at 3 years of age, mild spleen enlargement, hypercholesterolemia, hypochromic microcytic anemia, pigmented nevus, and vitiligo. The parents first noticed symptoms at 15 months of age. Patient 2's oculomotor function showed square-wave jerks, left-gaze nystagmus with a vertical component, downward slash nystagmus, saccadic smooth pursuit, vertical upward gaze palsy with a vertical component, and impaired convergence.

Before treatment, Patient 2's Severe Ataxia Assessment and Rating Scale (SARA) score showed 28.5/40. The patient's Spinocerebellar Ataxia Functional Index (SCAF1) results were as follows:

Average 8-meter Walking (8MW) test: cannot perform without support;

MW 9-Hole Pegboard Test Dominant (9HPTD) (right): 300 sec;

MW 9-Hole Pegboard Test Non-Dominant (9HPTND): 299.2 sec;

MW PATA Word Test: 13.5;

Visual analogue scale (patient assessment): 45.

The results of videooculography are given below.

Table 15. Videooculography parameters

Velocity of the slow phase of gaze holding nystagmus Horizontal Vertical Center -2 6.18 To the right -3.97 7.6 To the left -0.24 9.49 Down -2.86 5.58 Up -1.11 4.07

Following evaluation, Patient 2 begins treatment with acetyl-DL-leucine (5 g/day). After one month of treatment, the patient is re-evaluated. Caregivers report improvements in fine motor function, hand tremors, and speech. The patient herself does not perceive any beneficial effects. A scale assessment and rating scale (SARA) score of 23.5/40 is available. The patient's spinocerebellar ataxia functional index (SCAF1) results are as follows:

Average 8-meter Walking (8MW) test: cannot perform without support;

MW 9-Hole Pegboard Test Dominant (9HPTD) (right): 300 sec;

MW 9-Hole Pegboard Test Non-Dominant (9HPTND): 300 s;

MW PATA Word Test: 14;

Visual analogue scale (patient assessment): 80.

The results of videooculography are given below.

Table 16. Videooculography parameters

Velocity of the slow phase of gaze holding nystagmus Horizontal Vertical Center -1.45 5.6 To the right -3.45 6.56 To the left 3.58 7 Down -2.5 4.69 Up -0.51 3.45

Video-oculography shows improved fixation stability and a significant reduction in the intensity of downward-beating nystagmus in patient 2.

Patient 3

In this study, the patient is a 19-year-old woman suffering from ataxia-telangiectasia since early childhood, with

- delayed motor development, signs and symptoms of cerebellar ataxia, marked axial hypotonia with peripheral hypertonia, severe contractures of the feet with bilateral equine and transversoplanus deformities and therefore limited to a wheelchair, dysdiadochokinesia and areflexia of the lower extremities with decreased proprioceptive sensitivity; and

- non-Hodgkin's lymphoma, MTHFR polymorphism (C677T), lymphangioma of the lower lip, cholecystolithiasis, hereditary cardiomyopathy, pigmented nevus, thoracolumbar kyphoscoliosis and scleral telangiectasia in both eyes.

The patient's oculomotor function shows nystagmus of gaze holding to the right and left, saccadic eye movements, slow saccades in all directions, especially horizontally, pathological vestibulo-ocular reflex with corrective catch-up saccades, and pathological suppression of the vestibulo-ocular reflex by gaze fixation.

Before treatment, the patient's SARA score showed 23/40. The patient's Spinocerebellar Ataxia Functional Index (SCAF1) results were as follows:

Average 8-meter Walking (8MW) test: cannot perform;

MW 9-Hole Pegboard Test Dominant (9HPTD) (right): 150 sec;

MW 9-Hole Pegboard Test Non-Dominant (9HPTND): 161.6 sec;

MW PATA Word Test: 14;

Visual analogue scale (patient assessment): 80.

The results of videooculography are given below.

Table 17. Videooculography parameters

Slow phase velocity of gaze holding nystagmus [°/s] Horizontal Vertical Center 3 -1.5 To the right -0.4 -1 To the left 7.3 2.5 Down 2.4 -0.2

The patient begins treatment with acetyl-DL-leucine (5 g/day) approximately six months after the initial assessment. After nearly seven months of treatment, the patient is reassessed. Caregivers and the patient report general improvement in health, but no specific details are needed. The Scale for Ataxia Assessment and Rating (SARA) score is 21.5/40. The patient's Spinocerebellar Ataxia Functional Index (SCAF1) results are as follows:

Average 8-meter Walking (8MW) test: cannot be performed;

MW 9-Hole Pegboard Test Dominant (9HPTD) (right): 124.5 sec;

MW 9-Hole Pegboard Test Non-Dominant (9HPTND): 147.5 sec;

MW PATA Word Test: 10;

Visual analogue scale (patient assessment): 80.

The results of videooculography are given below.

Table 18. Videooculography parameters

Slow phase velocity of gaze holding nystagmus [°/s] Horizontal Vertical Center -0.1 0.4 To the right -4.2 1.7 To the left 1.7 0.1 Down -0.6 1.2

The patient shows a slight improvement in SARA and the SCAF1 9HPT subset and a significant improvement in fixation stability and a decrease in the intensity of gaze holding nystagmus in all positions.

Patient 4

The patient in this study is a 15-year-old girl who has suffered from ataxia-telangiectasia since age 4. Since age 7, the patient has exhibited signs and symptoms of severe cerebellar ataxia, fine motor impairment, muscle hypotonia with arelexia, muscle atrophy, and plantar flexion with discrete contractures. She is wheelchair-bound but is able to walk with constant support. The patient has severe hemolytic anemia, hypogammaglobulinemia, telangiectasia on the sclera and chest wall, secondary Cushing's syndrome due to corticosteroid use, and is suspected of having non-Hodgkin's lymphoma of the central nervous system.

The patient's oculomotor function shows a slight upward deviation of the eye, a "flapping" nystagmus to the left in the central position, nystagmus of maintaining the gaze in all directions, horizontal with a downward component, and fright with a sudden movement of the head.

Before treatment, the patient's Severe Ataxia Assessment and Rating Scale (SARA) score showed 23.5/40. The patient's Spinocerebellar Ataxia Functional Index (SCAF1) score was as follows:

Average 8-meter Walking (8MW) test: cannot perform;

MW 9-Hole Pegboard Test Dominant (9HPTD) (right): 124.5 sec;

MW 9-Hole Pegboard Test Non-Dominant (9HPTND): 52.3 sec;

MW PATA Word Test: 14;

Visual analogue scale (patient assessment): 70.

The results of videooculography are given below.

Table 19. Videooculography parameters

Slow phase velocity of gaze holding nystagmus [°/s] Horizontal Vertical Center -0.6 1.76 To the right 3.7 -0.1 To the left -6 0.6 Down 5.7 2.7

The patient begins treatment with acetyl-DL-leucine (5 g/day) after evaluation. After nearly one month of treatment, the patient is re-evaluated. The patient and her mother report improved handwriting, particularly due to reduced hand tremors and improved fine motor skills. The patient also reports easier drinking and no longer requires a straw. Family members describe an improved gait with increased stability and reduced need for support. A scaled ataxia assessment and rating (SARA) score of 18.5/40 was obtained. The patient's spinocerebellar ataxia functional index (SCAF1) results are as follows:

Average 8-meter Walking (8MW) test: cannot perform;

MW 9-Hole Pegboard Test Dominant (9HPTD) (right): 93.5 sec;

MW 9-Hole Pegboard Test Non-Dominant (9HPTND): 101.7 sec;

MW PATA Word Test: 15.5;

Visual analogue scale (patient assessment): 70.

The results of videooculography are given below.

Table 20. Videooculography parameters

Slow phase velocity of gaze holding nystagmus [°/s] Horizontal Vertical Center -0.2 4.8 To the right -1 0.3 To the left -0.8 1.9 Down -0.7 -2.3

The patient shows slight improvement in SARA and the SCAF1 9HPT subset of her dominant hand. Video-oculography shows an overall improvement in fixation stability and a decrease in the intensity of spontaneous and gaze-sustaining nystagmus in all positions.

Patient 5

In this study, the patient is a 10-year-old boy suffering from ataxia-telangiectasia since early childhood, having

- psychomotor delay, unsteady gait from 14 months with increased frequency of falls, signs and symptoms of severe cerebellar ataxia, dysarthria and dyslalia, deterioration of fine motor skills, infrequent head tremor, slow psychomotor tempo, hypotonia with muscle atrophy and hyporeflexia, anteflexion of the head with thoracic kyphosis, pedes tranversoplani, winged scapulae, parasomnia with night terrors and autism; and

- significant immunosuppression, telangiectasia on the soft palate, sclera, incontinence and asthenic physique.

The patient is confined to a wheelchair but is also capable of taking a few steps with strong, constant support. Oculomotor function shows oculomotor apraxia with marked head anteflexion, en bloc head and eye movements when looking right and left, vertical gaze palsy with slow vertical saccades and smooth saccadic pursuit, slow horizontal saccades to the left, saccade palsy when pursuing rightward, limited ocular motility, especially vertical, and fixation instability in all directions.

Before treatment, the patient's Severe Ataxia Assessment and Rating Scale (SARA) score was 24.5/40. The patient's Spinocerebellar Ataxia Functional Index (SCAF1) score was as follows:

Average 8-meter Walking (8MW) test: cannot walk without constant support;

MW 9-Hole Pegboard Test Dominant (9HPTD) (right): 102.7 sec;

MW 9-Hole Pegboard Test Non-Dominant (9HPTND): 116.8 sec;

MW PATA Word Test: 6;

visual analog scale (patient assessment): 90.

The results of videooculography are given below.

Table 21. Videooculography parameters

Slow phase velocity (SPV) of fixation and gaze holding nystagmus [°/s]

Horizontal Vertical Center 0.96 1.53 To the right 0.21 2 To the left 2.1 3.64 Down 0.71 3.91 Up 0.31 1.69

The patient begins treatment with acetyl-DL-leucine (5 g/day) after a follow-up evaluation. After approximately one month of treatment, the patient is re-evaluated. The patient's mother describes a significant improvement in gait stability; prior to therapy, he would fall frequently and required partial "transportation." After treatment, he is able to walk only by holding a caregiver's hand.

Fine motor skills and body balance improved, and hand tremors decreased. Improved fine motor skills are reflected in daily activities, such as eating and drinking independently. The patient gained 1.5 kg and has an improved appetite. A scale assessment and rating scale (SARA) showed a score of 20.5/40. The patient's spinocerebellar ataxia functional index (SCAF1) results are as follows:

Average 8-meter Walking (8MW) test: cannot walk without support;

MW 9-Hole Pegboard Test Dominant (9HPTD) (right): 103.6 sec;

MW 9-Hole Pegboard Test Non-Dominant (9HPTND): 88.8 sec;

MW PATA Word Test: 7.5;

visual analog scale (patient assessment): 80.

The results of videooculography are given below.

Table 22. Videooculography parameters

Slow phase velocity (SPV) of fixation and gaze holding nystagmus [°/s]

Horizontal Vertical Center 0.96 1.53 To the right 0.21 2 To the left 2.1 3.64 Down 0.71 3.91 Up 0.31 1.69

After almost seven months of treatment, the patient was re-evaluated. The patient's mother described a significantly more stable gait; this result has remained consistent since the initial post-treatment assessment. The patient can stand up independently and has generally become more independent in daily activities. The patient, with an improved appetite, has gained another 3 kg and demonstrates improved stability.

A scale assessment and rating scale (SARA) showed a score of 17.5/40. The patient's spinocerebellar ataxia functional index (SCAF1) score was as follows:

Average 8-meter Walking (8MW) test: 15.3 s (holding hand);

MW 9-Hole Pegboard Test Dominant (9HPTD) (right): 92.4 sec;

MW 9-Hole Pegboard Test Non-Dominant (9HPTND): 98.3 sec;

MW PATA Word Test: 11;

visual analog scale (patient assessment): 100.

The patient was re-evaluated after treatment for almost a year and demonstrated improved social interaction, activity, and mobility. The patient's parents reported improvement in incontinence. The patient developed recurring localized frontal pain with morning vomiting. Based on the family history, it is suspected that the localized pain may be due to childhood migraines.

A scale assessment and rating scale (SARA) showed a score of 16.5/40. The patient's spinocerebellar ataxia functional index (SCAF1) score was as follows:

Average 8-meter Walking (8MW) test: 13.9 sec (can walk independently, holds mother's hand);

MW 9-Hole Pegboard Test Dominant (9HPTD) (right): 95.6 sec;

MW 9-Hole Pegboard Test Non-Dominant (9HPTND): 127.6 sec;

MW PATA Word Test: 14.5;

visual analog scale (patient assessment): 95.

When treated with acetyl-DL-leucine, the patient shows improved SARA and SCAF1 scores, improved quality of life, improved overall fixation stability, and a reduction in the intensity of spontaneous and gaze-holding nystagmus, especially in the vertical plane (after 1 month).

Patient 6

The patient in this study is a 10-year-old girl with ataxia-telangiectasia, characterized by an ataxic gait and stance, impaired fine motor skills with hand tremors, dysphagia and speech impairment, oculomotor impairment, and problems with cognitive function and concentration. The first symptoms were observed at the age of one year.

Following a baseline assessment, the patient began treatment with acetyl-DL-leucine at 1.5 g/day for the first week and 3 g/day for the second and subsequent weeks. The patient was assessed after one month and six months of treatment, respectively. After one month of treatment, the patient demonstrated improved fine motor skills with reduced hand tremors, improved postural stability and gait, improved pronunciation, and increased self-confidence. After six months of treatment, the patient has a stable general condition, stable gait and standing, and improved handwriting. However, the patient suffers from anxiety, which might be expected to negatively impact her reactions.

The patient's SARA and SCAFI scores are provided below.

Table 23. Patient assessment parameters Basic After one month of treatment with acetyl-DL-leucine After six months of treatment with acetyl-DL-leucine SARA 11/40 8.5/40 11/40 8 MWT 8.95 s 8.45 sec 7.53 s 9HPTD 85.38 s 81.95 s 136.27 s 9HPTND 71.2 s 70.39 s 80.32 s PATA 14.5 15.5 13

Example 16

The patient in this study is a woman in her early 60s who was genetically diagnosed with spinocerebellar ataxia (SCA) 1. Before treatment, the patient had severe speech and swallowing problems, tremors in both hands, spasticity, and moderate ataxia of standing and gait. The patient also has sleep problems.

After three months of treatment with acetyl-DL-leucine (5 g/day), all symptoms were significantly improved as shown by clinical examination, including spasticity and deterioration of oculomotor function.

After three months, drug treatment is discontinued. After two weeks, the intensity of signs and symptoms has returned to pre-treatment levels. Treatment is restarted, and the patient remains on the same dose for two years without any side effects.

The patient's daughter reports significant progression of the disease over time with persistent symptomatic effects that are still present, which is unlikely given the long-term symptomatic effectiveness of the treatment.

Example 17

The patient in this study is a 70-year-old woman with an unsteady gait and frequent falls, nocturnal visual hallucinations, and REM sleep disorder. The patient has a symmetrical hypokinetic-rigid syndrome with impaired fine motor skills and fluctuations in attention and awareness.

The patient was diagnosed with dementia with Lewy bodies. An FDG-PET scan of the brain revealed synaptic dysfunction in the parietal and occipital lobes, and a DATscan revealed degeneration of presynaptic dopamine transport, supporting the diagnosis. Treatment with levodopa, 100 mg four times daily, and quetiapine, 25 mg at night, improved symptoms.

The patient is started on acetyl-DL-leucine (3 g/day for one week; then 5 g/day), and her condition is assessed after four weeks. She reports increased fatigue and impaired balance and speech. The medication is reduced to 3 g/day, and the patient is instructed to discontinue the medication after two weeks.

The patient is re-examined approximately one month after stopping drug treatment, and no improvement in symptoms is observed with a reduced dose and no worsening of symptoms after stopping drug treatment.

Table 24. Gait parameters

After 1 week on acetyl-DL-leucine After 4 weeks on acetyl-DL-leucine Normal value (± SD) Speed (cm/s) 55 56 119.12 (17.27) Max. speed (cm/s) 92 74 176.78 (19.10) Rhythm (steps/minute) 81 85 113.06 (10.38) Track width (cm) 4.6 6.7 9.49 (3.56) Length of step cycle (cm) 81 79 126.71 (13.06) Double standing (%) 36.1 38.7 20.35 (3.21) Coefficient of variation (temporal) 9.1 8.8 1.76 (0.73) Functional gait assessment 11/30 27.1 (2.3)

Example 18

In this study, four patients (brothers) suffered from, and were later diagnosed with, ataxia with oculomotor apraxia type 4. The three older brothers were 12, 11, and 10 years old, respectively, at the onset of the disease. Before starting treatment with acetyl-DL-leucine at age 15/16, the three older brothers walked with a walking aid, according to their mother. The older brothers began treatment with acetyl-DL-leucine at the ages of 25, 23, and 19, respectively, and remained on treatment for approximately four years. No long-term clinical data are available for these three patients.

The youngest brother was 11 years old at the onset of the disease. He began treatment with acetyl-DL-leucine at age 13. While on treatment, the youngest brother did not walk with a walking aid until he was about 18 years old, according to the patient's mother. The patient's mother also reported that the youngest brother had improved fine motor skills and speech compared to his older brothers. No long-term clinical data are available for the youngest brother.

Example 19

The severity of NPC in a patient can be quantified using the clinical severity score (CSS), which assesses various disease parameters and assigns a score to each parameter out of 5 (higher score = greater severity). See Yanjanin et al., “Linear Clinical Progression, Independent of Age of Onset, in Niemann–Pick Disease, Type C,” Am. J. Med. Genet., Part B, 153B: 132–140. For an untreated patient, it is usually possible to predict how an individual’s CSS will change over time, as disease progression appears to be linear. For example, if patient A progresses from a CSS of 8 to a CSS of 12 from 0 to 12 months, it can be predicted that at 36 months the patient will have a CSS of 20. The annualized severity increase score (ASIS) quantifies the rate of change in CSS per year, calculated by dividing the patient’s CSS by the patient’s age. For example, if patient B, who is not treated, has a CSS of 8 at age 2, the patient's ASIS might be 4. Each year, the patient's CSS might be expected to progress by a factor of 4, so that at age 4, the patient's CSS might be 16. If a therapeutic intervention slows or stops the progression of the disease, the patient might be expected to have a lower ASIS score than the baseline after such intervention.

Ten patients with NPC are administered 4.5 g/day of acetylleucine for a long period. CSS is determined at baseline and at various time points for eye movements, ambulation, speech, swallowing, fine motor skills, cognition, memory, and paroxysms. The overall CSS is calculated for baseline and at each time point by summing the individual CSS values for each parameter (eye movements, ambulation, etc.). The number of days after therapy initiation when the CSS is determined varies for each patient, as shown in Table 25.

Table 25. Days after the start of acetylleucine administration when CSS was assessed

Patient, I.D. Base (days) Time point 2 (days) Time point 3 (days) Time point 4 (days) 1 0 126 231 2 0 119 200 297 3 0 91 240 4 0 107 196 5 0 78 238 414 6 0 184 238 414 7 0 81 165 8 0 90 217 9 0 400 644 10 0 83

Each of Tables 26-34 below shows the CSS overall for eye movement, ambulation, speech, swallowing, fine motor skills, cognition, memory, and paroxysms, respectively.

Table 26. General CSS

Clinical Severity Score (CSS) Patient, I.D. Base Time point 2 Time point 3 Time point 4 1 11 11 10 2 33 33 33 33 3 13 12 11 4 13 13 10 5 12 12 12 12 6 21 23 21 21 7 19 19 19 8 13 12 11 9 22 22 21 10 14 11

Table 27. CSS for eye movement

Clinical Severity Score (CSS) Patient, I.D. Base Time point 2 Time point 3 Time point 4 1 3 3 3 2 3 3 3 3 3 3 3 3 4 2 2 2 5 3 3 3 3 6 3 3 3 3 7 3 3 3 8 3 3 2 9 3 3 3 10 3 3

Table 28. CSS for ambulation Clinical Severity Score (CSS) Patient, I.D. Base Time point 2 Time point 3 Time point 4 1 2 2 1 2 5 5 5 5 3 1 1 1 4 2 2 1 5 1 1 1 1 6 2 4 2 2 7 2 2 2 8 1 1 1 9 2 2 2 10 2 2

Table 29. CSS for speech Clinical Severity Score (CSS) Patient, I.D. Base Time point 2 Time point 3 Time point 4 1 1 1 1 2 2 2 2 2 3 1 1 1 4 2 2 1 5 1 1 1 1 6 2 2 2 2 7 1 1 1 8 1 1 1 9 2 2 2 10 1 1

Table 30. CSS for swallowing Clinical Severity Score (CSS) Patient, I.D. Base Time point 2 Time point 3 Time point 4 1 0 0 0 2 4 4 4 4 3 2 2 2 4 2 2 2 5 2 2 2 2 6 3 3 3 3 7 3 3 3 8 2 2 2 9 3 3 3 10 2 2

Table 31. CSS for fine motor skills

Clinical Severity Score (CSS) Patient, I.D. Base Time point 2 Time point 3 Time point 4 1 1 1 1 2 5 5 5 5 3 2 1 1 4 1 1 1 5 1 1 1 1 6 4 4 4 4 7 2 2 2 8 2 1 1 9 4 4 4 10 1 1

Table 32. CSS for Cognition

Clinical Severity Score (CSS) Patient, I.D. Base Time point 2 Time point 3 Time point 4 1 3 3 3 2 5 5 5 5 3 3 3 3 4 3 3 3 5 3 3 3 3 6 4 4 4 4 7 4 4 4 8 3 3 3 9 4 4 4 10 3 2

Table 33. CSS for Memory

Clinical Severity Score (CSS) Patient, I.D. Base Time point 2 Time point 3 Time point 4 1 1 1 1 2 4 4 4 4 3 1 1 0 4 1 1 0 5 1 1 1 1 6 3 3 3 3 7 4 4 4 8 1 1 1 9 4 4 3 10 2 0

Table 34. CSS for paroxysms

Clinical Severity Score (CSS) Patient, I.D. Base Time point 2 Time point 3 Time point 4 1 0 0 0 2 5 5 5 5 3 0 0 0 4 0 0 0 5 0 0 0 0 6 0 0 0 0 7 0 0 0 8 0 0 0 9 0 0 0 10 0 0

The ASSIS for the baseline and each time point is calculated using each patient's CSS and age at the time of assessment. The overall ASSIS for each patient at each time point is presented below in Table 35.

Table 35. General ASSIS

Annual Severity Increasing Estimates (ASIS) Base Time point 2 Time point 3 Time point 4 0.381371618 0.376864272 0.339262493 1.94125463 1,904748736 1,880675612 1.852636028 0.65 0.592617631 0.53250497 0.481909063 0.476731928 0.363469002 0.433188377 0.429874461 0.423232908 0.416160273 0.561964246 0.607297766 0.552333117 0.545420607 0.536675431 0.533334614 0.529913714 0.486750384 0.445200609 0.402903129 0.738624874 0.71243018 0.665646967 0.595597228 0.406038403

As can be seen from Table 26 and Figure 10A, none of the ten patients shows an overall increase in CSS over the course of the experiment. Patient 6 shows an increased CSS between baseline and time 2, but returns to the baseline value at time 3 and remains there at time 4. Four of the ten patients (patients 2, 5, 6, and 7) have a constant CSS over the course of the experiment, indicating that the disease is not progressing in these individuals. Six of the ten patients (patients 1, 3, 4, 8, 9, and 10) show a decrease in CSS over the course of the experiment, indicating that the disease is not progressing and is actually becoming less severe. Improvements are seen across various subscores: in patient 1 for ambulation; in patient 3 for fine motor skills; in patient 4 for ambulation and speech; in patient 8 for eye movements and fine motor skills; in patient 9 for memory; in patient 10 for cognition. The data presented in Figures 11A-11J show the subscores for each patient, respectively, in bar chart form.

As shown in Table 35 and Figure 10B, all ten patients show a decrease in ASIS during treatment relative to the baseline ASIS. In patients 2, 5, 6, and 7, CSS remains the same with age, resulting in a slight decrease in ASIS. In patients 1, 3, 4, 8, 9, and 10, the decrease in ASIS is greater due to a decrease in CSS with age.

Example 20

A mouse model of Niemann-Pick disease type C (NPC) shares several pathological features with Alzheimer's disease (AD), as described herein. Wild-type NPC1 ^-/- mice are treated with acetyl-dl-leucine (0.1 g/kg body weight per day) starting at 3 weeks of age. Mice are sacrificed at 8 weeks of age. Levels of amyloid precursor protein C-terminal fragments (APP-CTF) relative to total amyloid precursor protein (APP) levels are assessed in the cerebellum of wild-type, untreated wild-type NPC1 ^-/- mice, and AL-treated wild-type NPC1 ^-/- mice. Microtubule-associated protein 1A/3-phosphatidylethanolamine 1β-light chain (LC3-II) conjugate levels were also assessed relative to tubulin-loaded control levels for wild-type, untreated wild-type NPC1 ^-/- mice, and AL-treated wild-type NPC1 ^-/- mice.

APP-CTF data are shown in Figure 12A. The data replicate the previously observed accumulation of APP-CTF in the brains of NPC1-infected mice. Treatment with acetyl-dl-leucine is associated with a decrease in APP-CTF.

LC3-II data are shown in Figure 12B. The data replicate the previously observed LC3-II accumulation in the brains of NPC1-infected mice. Treatment with acetyl-dl-leucine is associated with a decrease in LC3-II, indicating a partial restoration of autophagic flux.

Conclusion

Acetylleucine treatment is associated with improvement in AD pathology in the brains of NPC1-positive mice.

Claims (7)

1. A method for providing neuroprotection in a subject having a neurodegenerative disease, suspected of having a neurodegenerative disease, or at risk of developing a neurodegenerative disease that is cerebellar ataxia, comprising administering a therapeutically effective amount of acetylleucine or a pharmaceutically acceptable salt thereof to the subject for a period selected from at least 1 year, at least 2 years, and at least 5 years. 2. The method according to claim 1, wherein the cerebellar ataxia is selected from spinocerebellar ataxia (SCA) 4, spinocerebellar ataxia (SCA) 5 (Lincoln ataxia), spinocerebellar ataxia (SCA) 8, spinocerebellar ataxia (SCA) 10, spinocerebellar ataxia (SCA) 11, spinocerebellar ataxia (SCA) 12, spinocerebellar ataxia (SCA) 13, spinocerebellar ataxia (SCA) 14, spinocerebellar ataxia (SCA) 15/16, spinocerebellar ataxia (SCA) 18 (sensorimotor neuropathy with ataxia), spinocerebellar ataxia (SCA) 19/22, spinocerebellar ataxia (SCA) 20, spinocerebellar ataxia (SCA) 21, spinocerebellar ataxia (SCA) 23, spinocerebellar ataxia (SCA) 25, spinocerebellar ataxia (SCA) 26, spinocerebellar ataxia (SCA) 27, spinocerebellar ataxia (SCA) 29, spinocerebellar ataxia (SCA) 30, spinocerebellar ataxia (SCA) 31, spinocerebellar ataxia (SCA) 32, spinocerebellar ataxia (SCA) 35, spinocerebellar ataxia (SCA) 36, episodic ataxia (EA) 1, episodic ataxia (EA) 2, episodic ataxia (EA) 3, episodic ataxia (EA) 4, episodic ataxia (EA) 5, episodic ataxia (EA) 6, episodic ataxia (EA) 7, spinocerebellar ataxia (SCA) 28, spinocerebellar ataxia (SCA) 24 (spinocerebellar ataxia autosomal recessive type 4 (SCAR4); spinocerebellar ataxia with saccadic intrusions), Tabes dorsalis, ataxia with oculomotor apraxia type 1 (AOA1), ataxia with oculomotor apraxia type 2 (AOA2), spinocerebellar ataxia autosomal recessive type 10 (SCAR 10), mitochondrial recessive ataxia (MIRAS), myoclonic epilepsy with myopathy and sensory ataxia (MEMSA), sensory ataxic neuropathy with dysarthria and ophthalmoparesis (SANDO), infantile spinocerebellar ataxia, hereditary spastic paraplegia 7 (HSP SPG7 gene), mitochondrial myopathy, encephalopathy, lactic acidosis with stroke-like episodes (MELAS), myoclonic epilepsy with ragged fibers (MERRF), neurogenic muscle weakness, ataxia, and retinitis pigmentosa (NARP), Kearns-Sayre syndrome (KSS), fragile X syndrome with tremor/ataxia (FXTAS), Arts syndrome, X-linked spinocerebellar ataxia 1, X-linked spinocerebellar ataxia 2, X-linked spinocerebellar ataxia 3, X-linked spinocerebellar ataxia 4 or X-linked spinocerebellar ataxia 5, X-linked syndromic intellectual disability Christianson type, X-linked sideroblastic anemia, late-onset idiopathic spinocerebellar ataxia, and cerebellar ataxia, neuropathy, vestibular areflexia syndrome (CANVAS). 3. The method according to any one of claims 1, 2, wherein the method comprises initiating the administration of a therapeutically effective amount of acetylleucine to a subject in need thereof when the subject is asymptomatic. 4. The method of claim 3, wherein the administration is initiated after a genetic and/or biochemical marker of a neurodegenerative disease has been identified in the subject. 5. The method according to any one of claims 1-4, wherein the acetylleucine is acetyl-DL-leucine. 6. The method according to any one of claims 1-5, wherein acetylleucine has an enantiomeric excess of the L-enantiomer or the D-enantiomer. 7. The method according to any one of claims 1-4, wherein the method comprises administering acetylleucine to a subject in need thereof in a therapeutically effective amount of 1 to 15 g per day, 1 to 10 g per day, 1.5 to 7 g per day, 4 to 6 g per day, 4 to 5 g per day.