Method for treating amyotrophic lateral sclerosis with pridopidine

Abstract

The present application relates to methods of treating amyotrophic lateral sclerosis with pridopidine. Provided herein is a method for treating a human subject with ALS by administering to the subject a therapeutically effective amount of pridopidine, wherein the pridopidine is used as a monotherapy or as a combination or add-on therapy with riluzole, edaravone, or dextromethorphan / quinidine.

Description

[0001] This application is a divisional application of an application filed on August 13, 2018, with application number 201880065983.3, and invention name “Method for treating amyotrophic lateral sclerosis with pridopidine”.

[0002] CROSS-REFERENCE TO RELATED APPLICATIONS

[0003] This application claims the benefit of U.S. Provisional Application No. 62 / 545,315, filed on August 14, 2017, the entire contents of which are hereby incorporated by reference. Background Art

[0004] Amyotrophic lateral sclerosis

[0005] Amyotrophic lateral sclerosis (ALS) is a devastating degenerative disease characterized by the progressive loss of motor neurons in the motor cortex, brainstem, and spinal cord (Peters 2015). This rapidly fatal disease begins focally and spreads, causing weakness of the limbs, respiratory, and bulbar muscles. Prior to death, there is an almost complete loss of limb and respiratory function, as well as the ability to chew, swallow, and speak.

[0006] ALS is a rare disease with an average incidence of 2.8 / 100,000 in Europe and 1.8 / 100,000 in North America, and an average prevalence of 5.40 / 100,000 in Europe and 3.40 / 100,000 in North America (Bozzoni 2016).

[0007] About 10% of ALS cases are classified as familial (FALS), while the remaining 90% appear to be sporadic (SALS) and occur randomly in the community (Riva 2016). More than 60% of patients die within 3 years of presentation, usually from respiratory failure, and about 10% survive more than 10 years (Zou 2016). There are currently no known disease-modifying therapies for ALS, although riluzole slows the rate of progression of the disease and prolongs survival by 2 or 3 months.

[0008] Clinical manifestations of ALS include muscle weakness and insufficient growth, fasciculations and muscle cramps, with spastic hypertonus and hyperreflexia being the main clinical manifestations. Some patients also present with dysarthria, dysphagia and respiratory weakness. Non-motor symptoms include behavioral disorders, executive dysfunction and frontotemporal dementia.

[0009] Neuropathological hallmarks of ALS include muscle atrophy, loss of anterior horn cells, and sclerosis of the lateral columns of the spinal cord (Martel 2016). Gliosis, defined as the activation of astrocytes and microglia, is also a hallmark of ALS.

[0010] Pridopidine

[0011] Pridopidine (formerly known as ACR16, ) is a unique compound developed for the treatment of patients with Huntington's disease. The chemical name of Pridopidine is 4-(3-(methylsulfonyl)phenyl)-1-propylpiperidine, and its chemical registration number is CAS 346688-38-8 (CSID: 7971505, 2016). The chemical registration number of Pridopidine hydrochloride is 882737-42-0 (CSID: 25948790 2016). The synthesis process of Pridopidine and its pharmaceutically acceptable salts is disclosed in U.S. Patent No. 7,923,459 and PCT Application Publication No. WO 2017 / 015609. U.S. Patent No. RE46,117 discloses Pridopidine for the treatment of a variety of diseases and conditions.

[0012] Pridopidine has been shown to modulate motor activity by suppressing hyperactivity or enhancing hypoactivity. Pridopidine also exhibits neuroprotective properties, which are thought to be due to its high affinity for sigma-1 receptors (S1R, binding IC50 ~ 70-80 nM) (Sahlholm 2013), while the motor activity of pridopidine is thought to be primarily mediated by its low affinity, antagonistic activity at dopamine D2 receptors (D2R) (binding IC50 ~ 10 μM) (Ponten 2010, Sahlholm 2015). Pridopidine exhibits low-moderate affinity binding to additional receptors in the micromolar range.

[0013] S1R is an endoplasmic reticulum (ER) chaperone protein involved in cell differentiation, neuroplasticity, neuroprotection, and cognitive function in the brain. Recently, transcriptomic analysis of the mouse striatum showed that pridopidine treatment activated the expression of BDNF, dopamine receptor 1 (D1R), glucocorticoid receptor (GR), and the serine-threonine kinase protein kinase B (Akt) / phosphoinositide 3-kinase (PI3K) pathway. Pridopidine has been shown to enhance the secretion of neuroprotective brain-derived neurotrophic factor (BDNF) in neuroblastoma cell lines in an S1R-dependent manner (Geva 2016) and rescue spinal cord injury and abnormal calcium signaling by activating S1R (Ryskamp 2017).

[0014] PCT International Patent Application Publication No. WO2016 / 138135 discloses the use of S1R agonists for treating, in particular, familial adult-onset amyotrophic lateral sclerosis (ALS) and juvenile-onset amyotrophic lateral sclerosis (ALS).

[0015] There remains an unmet need to provide effective treatments for ALS, particularly sporadic ALS. Summary of the invention

[0016] The present invention is based, at least in part, on the surprising experimental findings that pridopidine treatment ameliorates axonal transport deficits, enhances ERK activation and restores neuromuscular junction (NMJ) activity in SOD1-impaired muscle cell co-cultures, reduces mutant SOD1 aggregates in the spinal cord and mitigates NMJ disruption and consequent muscle wasting in SOD1-impaired mice.

[0017] The present invention provides a method for treating a subject suffering from amyotrophic lateral sclerosis (ALS), the method comprising: periodically administering to the subject an amount of pridopidine effective to treat the human subject.

[0018] The present invention also provides pridopidine for use in treating a human subject suffering from ALS.

[0019] The present invention also provides a method for treating ALS, comprising: regularly administering to a subject in need thereof an amount of pridopidine effective for treating ALS.

[0020] In some embodiments, the subject has sporadic ALS.

[0021] The present invention also provides a pharmaceutical composition for treating a human subject suffering from ALS, the pharmaceutical composition comprising an effective amount of pridopidine.

[0022] The present invention also provides use of pridopidine in preparing a medicament for treating ALS.

[0023] Further provided is a package comprising pridopidine and, optionally, instructions for using pridopidine to treat ALS.

[0024] In one embodiment, pridopidine is administered to a subject with ALS as a monotherapy. In other embodiments, a subject with ALS is treated with pridopidine and one or more additional agents.

[0025] Further provided is a method for treating a subject having ALS, the method comprising: periodically administering to the subject an amount of riluzole and an amount of pridopidine.

[0026] Further provided is a method for treating a subject having ALS, the method comprising: periodically administering to the subject an amount of edaravone and an amount of pridopidine.

[0027] Further provided is a method for treating a subject having ALS, the method comprising: periodically administering to the subject an amount of dextromethorphan / quinidine and an amount of pridopidine.

[0028] A package is further provided, comprising:

[0029] a) a first pharmaceutical composition, the first pharmaceutical composition comprising an amount of an agent, the agent being riluzole, edaravone or dextromethorphan / quinidine; and

[0030] b) a second pharmaceutical composition, wherein the second pharmaceutical composition comprises a certain amount of pridopidine and a pharmaceutically acceptable carrier.

[0031] In yet another embodiment, the package further comprises:

[0032] c) instructions for using the first pharmaceutical composition and the second pharmaceutical composition together to treat a subject suffering from ALS.

[0033] In some embodiments, the second pharmaceutical composition is riluzole, edaravone, a combination of dextromethorphan / quinidine, or laquinimod.

[0034] Further provided is pridopidine for use as an add-on therapy to riluzole or in combination with riluzole for treating a subject with ALS. Further provided is pridopidine for use as an add-on therapy to edaravone or in combination with edaravone for treating a subject with ALS. Further provided is pridopidine for use as an add-on therapy to dextromethorphan / quinidine or in combination with dextromethorphan / quinidine for treating a subject with ALS.

[0035] The subject invention also provides a pharmaceutical composition, which comprises a certain amount of riluzole, edaravone or dextromethorphan / quinidine and a certain amount of pridopidine, and at least one pharmaceutically acceptable carrier.

[0036] The subject invention also provides the use of:

[0037] a) a dose of riluzole, edaravone, or dextromethorphan / quinidine; and

[0038] b) a certain amount of Pridopidine,

[0039] Wherein, a certain amount of riluzole, edaravone or dextromethorphan / quinidine and a certain amount of pridopidine are administered simultaneously or synchronously.

[0040] The subject invention also provides a pharmaceutical composition comprising an amount of riluzole, edaravone or dextromethorphan / quinidine for use in combination with an amount of pridopidine to treat a subject suffering from movement disorders by regularly administering the pharmaceutical composition and an amount of pridopidine to the subject.

[0041] The subject invention also provides a pharmaceutical composition comprising an amount of pridopidine for use in combination with an amount of riluzole, edaravone, or dextromethorphan / quinidine to treat a subject suffering from movement disorders by periodically administering the pharmaceutical composition and an amount of riluzole, edaravone, or dextromethorphan / quinidine, respectively, to the subject.

[0042] The subject invention also provides riluzole and pridopidine for use in treating a subject suffering from a movement disorder, wherein said riluzole and said pridopidine are administered simultaneously or synchronously.

[0043] The subject invention also provides a product comprising an amount of riluzole and an amount of pridopidine for simultaneous or concurrent use in treating a subject suffering from ALS.

[0044] In another aspect, the present invention provides a combination of pridopidine and riluzole, edaravone or dextromethorphan / quinidine for use as a medicament for treating, preventing or alleviating ALS.

[0045] In another aspect, the present invention provides a combination of pridopidine and riluzole, edaravone, laquinimod or dextromethorphan / quinidine for use as a medicament.

[0046] In another aspect, the present invention provides a pharmaceutical composition comprising a therapeutically effective amount of pridopidine and a therapeutically effective amount of riluzole, edaravone, laquinimod or dextromethorphan / quinidine, and one or more adjuvants, excipients, carriers and / or diluents.

[0047] In another aspect, the present invention provides a method for treating ALS in a living animal, including a human, comprising the steps of: administering to the living animal in need thereof a therapeutically effective amount of pridopidine; in combination therapy with riluzole, edaravone, laquinimod or dextromethorphan / quinidine.

[0048] In another aspect, the invention provides a kit of parts comprising: at least two separate unit dosage forms (A) and (B), wherein (A) comprises pridopidine; and (B) comprises riluzole, edaravone, or dextromethorphan / quinidine; and optionally (C) instructions for simultaneously or synchronously administering (A) pridopidine and (B) riluzole, edaravone, or dextromethorphan / quinidine to a patient in need thereof.

[0049] In another aspect, the present invention provides a method for treating a subject having ALS, the method comprising administering to the subject a therapeutically effective amount of pridopidine and a therapeutically effective amount of riluzole, edaravone, laquinimod, or a combination of dextromethorphan / quinidine, wherein the amounts, when taken together, are effective to treat the human patient.

[0050] The present invention also provides the following items:

[0051] 1. A method for treating a subject suffering from amyotrophic lateral sclerosis (ALS), the method comprising: periodically administering to the subject an amount of pridopidine effective to treat the subject.

[0052] 2. The method of claim 1, wherein the ALS is sporadic ALS.

[0053] 3. The method of item 1 or 2, wherein the amount of pridopidine is effective for inhibiting or reducing the progression of ALS symptoms in the subject.

[0054] 4. The method of item 3, wherein the symptoms are muscle stiffness, muscle weakness, muscle wasting, muscle cramps, difficulty speaking, difficulty swallowing, difficulty breathing, difficulty chewing, difficulty walking, fasciculations and / or worsening of posture.

[0055] 5. The method of any one of items 1 to 4, wherein the amount of pridopidine is effective in the subject suffering from ALS to: enhance BDNF axonal transport in motor neurons, enhance ERK activation in motor neurons, improve NMJ formation and retention, preserve NMJ structure, preserve NMJ function, increase the innervation rate of muscle tissue, enhance motor neuron axon growth, reduce axon degeneration, reduce motor neuron axon degeneration, enhance muscle cell survival, enhance muscle fiber diameter and function, reduce SOD1 aggregation, reduce pseudobulbar disease progression, reduce muscle fiber loss progression and / or improve muscle contraction.

[0056] 6. The method of any one of items 1 to 5, wherein the amount of pridopidine is administered daily, or more than once daily.

[0057] 7. The method of any one of items 1 to 6, wherein the amount of pridopidine is administered twice daily.

[0058] 8. The method of any one of items 1 to 7, wherein the amount of pridopidine is administered orally.

[0059] 9. The method of any one of items 1 to 8, wherein the amount of pridopidine administered is 10 mg to 225 mg per day; or 45 mg to 180 mg per day; or 10 mg, 22.5 mg, 45 mg, 67.5 mg, 90 mg, 100 mg, 112.5 mg, 125 mg, 135 mg, 150 mg or 180 mg per day.

[0060] 10. The method of any one of items 1 to 9, wherein the pridopidine is pridopidine hydrochloride.

[0061] 11. The method of any one of items 1 to 10, wherein the subject is a human subject.

[0062] 12. The method of any one of items 1 to 11, further comprising administering to the subject a therapeutically effective amount of a second compound, wherein the second compound is riluzole, edaravone, or dextromethorphan / quinidine.

[0063] 13. The method of claim 12, wherein the second compound is administered before administering pridopidine.

[0064] 14. The method of item 12, wherein pridopidine is administered before administering the second compound.

[0065] 15. The method of claim 12, wherein the pridopidine is administered as an adjunct to the second compound.

[0066] 16. The method of claim 12, wherein the second compound is administered to assist with the pridopidine.

[0067] 17. A therapeutic package for or use in administering a drug to a subject, the therapeutic package comprising:

[0068] a) one or more unit doses, each such unit dose comprising an amount of pridopidine therein, wherein upon administration to said subject, the amount of pridopidine in said unit dose is effective for treating ALS in said subject, and

[0069] b) a pharmaceutical container complete therefor, said container containing one or more said unit doses, said container further containing or comprising a label directing the use of said package in treating a subject suffering from ALS.

[0070] 18. A package, comprising:

[0071] a) a first pharmaceutical composition comprising a certain amount of pridopidine and a pharmaceutically acceptable carrier;

[0072] b) a second pharmaceutical composition comprising an amount of a second compound and a pharmaceutically acceptable carrier, wherein the second compound is riluzole, edaravone or dextromethorphan / quinidine; and optionally

[0073] c) instructions for using the first pharmaceutical composition and the second pharmaceutical composition together to treat a subject suffering from ALS.

[0074] 19. The package of item 18, wherein said amount of said second compound and said amount of pridopidine are prepared to be administered simultaneously or synchronously.

[0075] 20. A pharmaceutical composition comprising an amount of pridopidine and an amount of a second compound, wherein the second compound is riluzole, edaravone, or dextromethorphan / quinidine. BRIEF DESCRIPTION OF THE DRAWINGS

[0076] Figure 1a to Figure 1c .Axonal transport assay. Figure 1a . Experimental procedure for axonal transport assay. Figure 1b . Schematic representation of the experimental system used for axonal transport tracing in motor neurons (MNs). Figure 1c .Time-lapse images of axonal transport of Qdot-BDNF (bright orange).

[0077] Figure 2a to Figure 2b . Graph showing the effect of Pridopidine on the instantaneous velocity values ​​and particle stop counts of Qdot-BDNF along axons (LM (littermates) = WT (wild type)). Figure 2a Effect of Pridopidine on the instantaneous velocity values ​​(μm / sec) of Qdot-BDNF particles in WT (blue), SOD1G93A (red), or Sigma-1 receptor knockout (S1R- / -) (grey) MNs. Figure 2b Effect of Pridopidine on particle stall counts (number of stalls of Qdot-BDNF counted per second).

[0078] Figure 3 Schematic representation of the experimental procedure for the neuromuscular co-culture assay used to measure muscle innervation and neuromuscular junction function (NMJ). Spinal explants were cultured in the proximal compartment and primary muscle cells were cultured in the distal compartment.

[0079] Figure 4. Graph showing the effect of pridopidine on axonal growth of WT and SOD1G93A, measuring the number of troughs with axonal crossing into the myogenic compartment.

[0080] Figure 5a and Figure 5b . Results of co-culture experiments in microfluidic chamber (MFC). Figure 5a Upper panel: Phase image of a myocyte in the distal compartment connected by an axon (green arrow). Lower panel: High magnification image of the myocyte:MN contact point. Figure 5b .Muscle contraction trajectories extracted from measurements of the strength of the muscle contraction over time.

[0081] Figure 6 . Graph showing the effect of pridopidine on innervation rate in WT and SOD1G93A myocytes.

[0082] Figure 7a to Figure 7b . Graph showing the levels of contractile myocytes in MN-myocyte co-cultures. Figure 7a . Percentage of contracting myoblasts in co-cultures of ventral spinal cord (WT or SOD1G93A) slices and primary myocytes (SOD1G93A or LM control). Figure 7b Effects of pridopidine on neuromuscular junctions (NMJs) in muscle tissue from WT or SOD1G93A and in motor neurons from WT or S1R- / - mice.

[0083] Figure 8a to Figure 8b .Effect of Pridopidine on ERK levels. Figure 8a Effects of Pridopidine on total ERK (tERK) and phosphorylated ERK (pERK) levels from culture extracts of WT, SOD1G93A and S1R- / - MNs shown in immunoblots. Figure 8b Quantification of the effect of pridopidine on ERK activation as measured by tERK / pERK.

[0084] Figures 9a to 9c .Effect of Pridopidine on SOD1 aggregation. Figure 9a Visualization and quantification of fluorescently labeled spinal cords using NC500 to label mutant SOD1 aggregates in WT and SOD1G93A spinal cords treated or not with pridopidine. Figure 9b . Quantitative analysis of the number of aggregates per unit area identified in gray matter. Fig.9c . Quantitative analysis of the number of aggregates per unit area identified in white matter.

[0085] Figure 10a to Figure 10b .Effects of Pridopidine on muscle tissue. Fig.10a. Representative images of H&E-stained cross sections of gastrocnemius muscles from WT or SOD1G93A mice treated or not with pridopidine. Fig.10b .Evaluation of the effects of pridopidine on muscle fiber loss: Quantitative analysis of the effects of pridopidine on muscle fiber diameter.

[0086] Figure 11a to Figure 11b : Effects of pridopidine on NMJ. Fig.11a Effects of Pridopidine on NMJ retention in vivo. Fig.11b Quantitative analysis of the effect of pridopidine on the percentage of innervated NMJs. DETAILED DESCRIPTION OF THE INVENTION

[0087] The present invention provides a method for treating a subject suffering from amyotrophic lateral sclerosis (ALS), the method comprising periodically administering to the subject an amount of pridopidine effective to treat the human subject.

[0088] In an embodiment of the invention, the ALS is sporadic ALS.

[0089] In an embodiment of the invention, the ALS is familial ALS (FALS). In some embodiments, the ALS is juvenile ALS (JALS).

[0090] In some embodiments, the ALS is not FALS. In some embodiments, the ALS is not juvenile ALS (JALS).

[0091] In an embodiment of the present invention, the type of ALS is classic ALS, bulbar ALS, flail arm ALS, flail leg ALS, pyramidal ALS and respiratory ALS, progressive muscular atrophy, primary lateral sclerosis or progressive bulbar palsy.

[0092] In an embodiment of the invention, the subject carries a mutated form of a gene that causes or contributes to the pathogenesis of ALS. In some embodiments, the mutated form of the gene is selected from the group consisting of superoxide dismutase 1 (SOD1), TAR DNA binding protein encoding TDP-43 (TARDBP), fusion in sarcoma (FUS), p62 (SQSTM1), ubiliquin-2 (UBQLN2), TANK binding kinase 1 (TBK1), inhibitory protein 1 (PFN1), VCP or p97 (VCP), angiopoietin (ANG), optic nerve protein (OPTN), C9orf72, Sigma-1 receptor (S1R), tubulin alpha-4A (TUBA4A), dynamin activator protein (DCTN1), hnRNPA1 (HNRNPA1), matrix protein 3 (MATR3), Coiled-coil-helix-coiled-coil-helix domain containing 10 (CHCHD10) gene and any combination thereof.

[0093] In an embodiment of the invention, the amount of pridopidine is effective to inhibit or reduce the progression of ALS symptoms in said subject.

[0094] In an embodiment of the invention, the symptoms of ALS are clinical symptoms of ALS.

[0095] In an embodiment of the present invention, the symptoms of ALS are muscle weakness and insufficient growth, fasciculations and muscle cramps, spastic hypertonus, hyperreflexia, dysarthria, dysphagia and respiratory failure, behavioral disorders, dysexecutive disorders and frontotemporal dementia.

[0096] In an embodiment of the invention, the symptoms of ALS are neuropathological symptoms.

[0097] In some embodiments, the symptom is bulbar palsy or pseudobulbar effect (PBA).

[0098] In an embodiment of the invention, the symptoms of ALS are muscle atrophy, motor neuron loss, anterior horn cell loss, lateral column sclerosis or gliosis.

[0099] In one embodiment, the symptom of ALS is the rate of decline in (a) lung function, (b) sexual disability, or (c) lower extremity performance scores. In an embodiment of the invention, the amount of pridopidine is effective to cause survival of a subject or to cause neuroprotection in a subject.

[0100] In some embodiments of the invention, treating the subject with pridopidine results in a decrease or improvement in the subject's impairment in one or more of the following domains:

[0101] 1) speech, 2) salivation, 3) swallowing, 4) handwriting, 5) cutting food and handling utensils (with or without a gastrostomy), 6) dressing and hygiene habits, 7) turning and adjusting bedding, 8) walking, 9) climbing stairs, 10) breathing, 11) dyspnea, 12) orthopnea, and 13) insufficiency.

[0102] In some embodiments, a rating scale (e.g., the Amyotrophic Lateral Sclerosis Functional Rating Scale (ALSFRS) or the Revised ALSFRS (ALSFRS-R)) is used to monitor changes in the above domains in patients, and functional changes in patients are monitored over time.

[0103] In some embodiments, pseudobulbar effect (PBA) is monitored in a patient. In some embodiments, the severity and / or frequency of emotional outbursts in a subject experiencing PBA is reduced by treatment with pridopidine.

[0104] In an embodiment of the present invention, pridopidine is administered daily.

[0105] In an embodiment of the invention, pridopidine is administered more than once daily.

[0106] In an embodiment of the present invention, pridopidine is administered twice daily. In an embodiment of the present invention, pridopidine is administered three times daily.

[0107] In an embodiment of the invention, pridopidine is administered less than once daily, for example, once every other day, three times a week, twice a week, or once a week.

[0108] In an embodiment of the present invention, pridopidine is administered orally.

[0109] In some embodiments, a unit dose of the pharmaceutical composition contains 10-250 mg of pridopidine. In some embodiments, the composition comprises 45 mg, 67.5 mg, 90 mg or 112.5 mg of pridopidine.

[0110] In an embodiment, between 10-225 mg of pridopidine is administered to the patient daily. In another embodiment, between 45-180 mg of pridopidine is administered to the patient daily. In another embodiment, 10 mg, 22.5 mg, 45 mg, 67.5 mg, 90 mg, 100 mg, 112.5 mg, 125 mg, 135 mg, 150 mg, or 180 mg of pridopidine is administered to the patient daily.

[0111] In an embodiment, the pharmaceutical composition is administered twice daily. In another embodiment, an equal amount of the pharmaceutical composition is administered at each administration. In an embodiment, the two doses are administered at least 6 hours, at least 7 hours, at least 8 hours, at least 9 hours, at least 10 hours, at least 11 hours apart. In some embodiments, the pharmaceutical composition is administered for at least 12 weeks, at least 20 weeks, at least 24 weeks, at least 26 weeks, at least 52 weeks, or at least 78 weeks.

[0112] In an embodiment of the present invention, the pridopidine is pridopidine hydrochloride.

[0113] In an embodiment of the invention, the subject is a human subject.

[0114] The present invention also provides pridopidine for use in treating a human subject suffering from ALS.

[0115] The present invention also provides a pharmaceutical composition for treating a human subject suffering from ALS, the pharmaceutical composition comprising an effective amount of pridopidine.

[0116] The present invention further provides a method for treating ALS, comprising periodically administering to a subject in need thereof an amount of pridopidine effective for treating ALS.

[0117] In an embodiment, the pharmaceutical composition includes an amount of pridopidine and an amount of a second compound, for example, a compound useful for treating a patient having ALS.

[0118] In some embodiments, the second compound is riluzole, edaravone, a combination of dextromethorphan and quinidine, or laquinimod.

[0119] In an embodiment, a pharmaceutical composition for treating a subject having ALS includes pridopidine and a second compound prepared to be administered simultaneously or synchronously.

[0120] In an embodiment, the pharmaceutical composition is in unit dosage form, which is useful for treating a subject suffering from ALS, the pharmaceutical composition comprising:

[0121] a) a certain amount of pridopidine;

[0122] b) an amount of a second compound,

[0123] wherein, upon concomitant administration of one or more of the unit dosage forms of the composition to the subject, the corresponding amounts of the second compound and the pridopidine in the composition are effective for treating the subject. In some embodiments, the second compound is riluzole, edaravone, or dextromethorphan / quinidine.

[0124] In an embodiment, the pharmaceutical composition comprises an amount of pridopidine for treating a subject with ALS as an add-on therapy to a second compound, the second compound being riluzole. In another embodiment, the pharmaceutical composition comprises an amount of pridopidine for treating a subject with ALS as an add-on therapy to a second compound, the second compound being edaravone. In another embodiment, the pharmaceutical composition comprises an amount of pridopidine for treating a subject with ALS as an add-on therapy to a second compound, the second compound being dextromethorphan / quinidine.

[0125] In an embodiment, the pharmaceutical composition includes an amount of pridopidine for treating a subject with ALS simultaneously or concurrently with a second compound, the second compound being riluzole. In another embodiment, the pharmaceutical composition includes an amount of pridopidine for treating a subject with ALS simultaneously or concurrently with a second compound, the second compound being edaravone. In another embodiment, the pharmaceutical composition includes an amount of pridopidine for treating a subject with ALS simultaneously or concurrently with a second compound, the second compound being dextromethorphan / quinidine. In another embodiment, the pharmaceutical composition includes an amount of pridopidine for treating a subject with ALS simultaneously or concurrently with a second compound, the second compound being laquinimod. In one embodiment, the pharmaceutical composition includes an amount of a compound for treating a subject with ALS, the compound being riluzole as an add-on therapy to pridopidine. In another embodiment, the pharmaceutical composition includes an amount of a compound for treating a subject with ALS, the compound being edaravone as an add-on therapy to pridopidine. In another embodiment, the pharmaceutical composition comprises an amount of a compound for treating a subject with ALS that is dextromethorphan / quinidine as an add-on therapy to pridopidine. In another embodiment, the pharmaceutical composition comprises an amount of a compound for treating a subject with ALS that is laquinimod as an add-on therapy to pridopidine.

[0126] In one embodiment, the pharmaceutical composition comprises an amount of a compound that is riluzole for treating a subject with ALS simultaneously or concurrently with pridopidine. In another embodiment, the pharmaceutical composition comprises an amount of a compound that is edaravone for treating a subject with ALS simultaneously or concurrently with pridopidine. In another embodiment, the pharmaceutical composition comprises an amount of a compound that is dextromethorphan / quinidine for treating a subject with ALS simultaneously or concurrently with pridopidine.

[0127] The present invention also provides a compound which is riluzole for use as an add-on therapy to pridopidine in treating a subject suffering from ALS.

[0128] The present invention also provides a compound which is edaravone for use as an add-on therapy to pridopidine in treating a subject suffering from ALS.

[0129] The present invention also provides a compound which is dextromethorphan / quinidine for use as an add-on therapy to pridopidine in treating a subject suffering from ALS.

[0130] The present invention also provides pridopidine for use as an add-on therapy to a compound that is riluzole in treating a subject suffering from ALS.

[0131] The present invention also provides pridopidine for use as an add-on therapy to a compound that is edaravone in treating a subject suffering from ALS.

[0132] The present invention also provides pridopidine for use as an add-on therapy to a compound that is dextromethorphan / quinidine in treating a subject suffering from ALS.

[0133] The present invention also provides pridopidine for use as an add-on therapy to a compound that is laquinimod in treating a subject suffering from ALS.

[0134] In one embodiment, the add-on therapy is for treating, preventing, or alleviating a symptom of ALS.

[0135] The present invention also provides a combination of a compound, which is riluzole, and pridopidine, for use in treating, preventing or alleviating the symptoms of ALS.

[0136] The present invention also provides a combination of a compound, which is edaravone, and pridopidine, for use in treating, preventing or alleviating the symptoms of ALS.

[0137] The present invention also provides a combination of a compound which is dextromethorphan / quinidine and pridopidine, the combination being useful for treating, preventing or alleviating the symptoms of ALS.

[0138] The present invention also provides use of pridopidine in manufacturing a medicament for treating ALS.

[0139] The methods, uses and combinations further comprise reducing the rate of neurological exacerbations in the subject.

[0140] In one embodiment, the methods of the present invention further comprise administering to the subject a therapeutically effective amount of a second compound, the second compound being riluzole or edaravone. In one embodiment, the methods of the present invention further comprise administering to the subject a therapeutically effective amount of a second compound, the second compound being dextromethorphan / quinidine. In one embodiment, the second compound is riluzole. In another embodiment, the second compound is edaravone. In another embodiment, the second compound is dextromethorphan / quinidine. In another embodiment, the second compound is laquinimod.

[0141] In an embodiment of the invention, pridopidine and said second compound are administered in one unit. In another embodiment, pridopidine and said second compound are administered in more than one unit.

[0142] In one embodiment, an amount of pridopidine and an amount of a second compound are administered simultaneously. In an embodiment, an amount of pridopidine and an amount of a second compound are administered concurrently.

[0143] In another embodiment, the administration of said second compound precedes the administration of said pridopidine. In another embodiment, the administration of said pridopidine precedes the administration of said second compound.

[0144] In one embodiment, the subject is receiving edaravone therapy prior to starting pridopidine therapy. In another embodiment, the subject is receiving riluzole therapy prior to starting pridopidine therapy. In another embodiment, the subject is receiving laquinimod therapy prior to starting pridopidine therapy. In another embodiment, the subject is receiving dextromethorphan / quinidine therapy prior to starting pridopidine therapy.

[0145] In another embodiment, the subject received edaravone therapy for at least 24 weeks, 28 weeks, 48 ​​weeks, or 52 weeks prior to starting pridopidine therapy. In another embodiment, the subject received riluzole therapy for at least 24 weeks, 28 weeks, 48 ​​weeks, or 52 weeks prior to starting pridopidine therapy. In another embodiment, the subject received dextromethorphan / quinidine therapy for at least 1 week, 2 weeks, 4 weeks, or 6 weeks prior to starting pridopidine therapy.

[0146] In one embodiment, the subject is receiving pridopidine therapy prior to starting edaravone therapy. In another embodiment, the subject is receiving pridopidine therapy for at least 24 weeks, 28 weeks, 48 ​​weeks, or 52 weeks prior to starting edaravone therapy.

[0147] In one embodiment, the subject received pridopidine therapy prior to initiating riluzole therapy. In another embodiment, the subject received pridopidine therapy for at least 24 weeks, 28 weeks, 48 ​​weeks, or 52 weeks prior to initiating riluzole therapy.

[0148] In one embodiment, the subject received pridopidine therapy prior to starting laquinimod therapy. In another embodiment, the subject received pridopidine therapy for at least 24 weeks, 28 weeks, 48 ​​weeks, or 52 weeks prior to starting laquinimod therapy.

[0149] In one embodiment, the subject is receiving pridopidine therapy prior to starting dextromethorphan / quinidine therapy. In another embodiment, the subject is receiving pridopidine therapy for at least 24 weeks, 28 weeks, 48 ​​weeks, or 52 weeks prior to starting dextromethorphan / quinidine therapy.

[0150] In one embodiment, between 0.5 mg and 1.5 mg of laquinimod is administered to the patient daily. In another embodiment, 0.5 mg or 1.0 mg of laquinimod is administered to the patient daily. In one embodiment, laquinimod is administered orally.

[0151] In one embodiment, between 10-200 mg of riluzole is administered to the patient daily. In another embodiment, 50 mg, 100 mg, or 200 mg of riluzole is administered to the patient daily.

[0152] In one embodiment, riluzole is administered orally. In one embodiment, dextromethorphan / quinidine is administered orally.

[0153] In one embodiment, between 5-60 mg of edaravone is administered to the patient daily. In another embodiment, 30 mg or 60 mg of edaravone is administered to the patient daily.

[0154] In one embodiment, edaravone is administered by intravenous infusion. In another embodiment, edaravone is administered once daily for 10 days, followed by a 14-day drug-free period. In another embodiment, edaravone is administered once daily for 14 days, followed by a 14-day drug-free period.

[0155] In one embodiment, the amount of the second compound when used alone and the amount of pridopidine when used alone are respectively effective for treating the subject. In another embodiment, the amount of the second compound when used alone or the amount of pridopidine when used alone is less effective for treating the subject. In another embodiment, the amount of the second compound when used alone or the amount of pridopidine when used alone is not effective for treating the subject.

[0156] In one embodiment, pridopidine is used to assist in the administration of a second compound. In another embodiment, the second compound is used to assist in the administration of pridopidine.

[0157] In one embodiment, a loading dose of an amount different than the intended dose is administered at a time period beginning with regular administration.

[0158] In one embodiment, a method of enhancing axonal transport of BDNF in motor neurons in a subject having ALS is provided, the method comprising administering to the subject an amount of pridopidine effective to enhance axonal transport of BDNF in motor neurons in the subject.

[0159] In one embodiment, a method of enhancing ERK activation in motor neurons in a subject having ALS is provided, the method comprising administering to the subject an amount of pridopidine effective to enhance ERK activation in motor neurons in the subject.

[0160] In one embodiment, a method of preserving neuromuscular junction (NMJ) structure in muscle cells of a subject suffering from ALS is provided, the method comprising administering to the subject an amount of pridopidine effective to preserve neuromuscular junction structure in the subject's muscle.

[0161] Further provided is a method of improving muscle contraction in a subject suffering from ALS, the method comprising administering to the subject an amount of pridopidine effective to improve muscle contractile function in the subject.

[0162] Further provided is a method of improving innervation of muscle tissue in a subject having ALS, the method comprising administering to the subject an amount of pridopidine effective to improve innervation in the subject.

[0163] In one embodiment, a method of enhancing motor neuron axon growth in a subject having ALS is provided, the method comprising administering to the subject an amount of pridopidine effective to enhance motor neuron axon growth in the subject.

[0164] In one embodiment, a method of enhancing muscle cell survival in a subject having ALS is provided, the method comprising administering to the subject an amount of pridopidine effective to enhance muscle cell survival in the subject.

[0165] In one embodiment, a method of reducing the progression of muscle fiber wasting in a subject having ALS is provided, the method comprising administering to the subject an amount of pridopidine effective to reduce the progression of muscle fiber wasting in the subject.

[0166] In one embodiment, a method of reducing axonal degeneration in a subject having ALS is provided, the method comprising administering to the subject an amount of pridopidine effective to reduce axonal degeneration in the subject.

[0167] In one embodiment, a method of preserving NMJ formation in a subject having ALS is provided, the method comprising administering to the subject an amount of pridopidine effective to preserve NMJ formation in the subject.

[0168] In one embodiment, a method of preserving NMJ structure and function in a subject having ALS is provided, the method comprising administering to the subject an amount of pridopidine effective to preserve NMJ structure and function in the subject.

[0169] In one embodiment, a method of reducing protein aggregation in a subject having ALS is provided, the method comprising administering to the subject an amount of pridopidine effective to reduce protein aggregation in the subject.

[0170] In one embodiment, a method of reducing the progression of pseudobulbar disease in a subject having ALS is provided, the method comprising administering to the subject an amount of pridopidine effective to reduce the progression of pseudobulbar disease in the subject.

[0171] The present invention also provides a package, comprising:

[0172] A pharmaceutical composition comprising an amount of pridopidine; and

[0173] Optionally, instructions for using the pharmaceutical composition to treat a subject suffering from ALS.

[0174] The present invention also provides a therapeutic package for administering a drug to a subject or for use in administering a drug to a subject, the therapeutic package comprising:

[0175] one or more unit doses, each such unit dose comprising an amount of pridopidine therein, wherein upon administration to said subject, the amount of pridopidine in said unit dose is effective for treating ALS in said subject, and

[0176] A pharmaceutical container is provided for this purpose, the container containing one or more of the unit doses, the container optionally further containing or including a label directing the use of the package in treating a subject suffering from ALS. Such a unit dose contains 10-250 mg of pridopidine, or, for example, 10 mg, 22.5 mg, 45 mg, 67.5 mg, 90 mg or 112.5 mg of pridopidine.

[0177] The present invention also provides a package, comprising:

[0178] A first pharmaceutical composition comprising a certain amount of pridopidine and a pharmaceutically acceptable carrier;

[0179] a second pharmaceutical composition comprising an amount of a second compound and a pharmaceutically acceptable carrier, wherein the second compound is riluzole, edaravone or dextromethorphan / quinidine; and

[0180] Optionally, instructions for using the first pharmaceutical composition and the second pharmaceutical composition together to treat a subject having ALS.

[0181] In one embodiment, an amount of the second compound and an amount of pridopidine are prepared to be administered simultaneously or synchronously.

[0182] The present invention also provides a therapeutic package for administering to a subject suffering from ALS or for use in administering to a subject suffering from ALS, the therapeutic package comprising:

[0183] One or more unit doses, each such unit dose comprising:

[0184] A certain amount of Pridopidine and

[0185] an amount of a second compound, wherein the second compound is riluzole or edaravone or dextromethorphan / quinidine,

[0186] wherein the respective amounts of said pridopidine and said second compound in said unit dose are effective for treating said subject upon concomitant administration to said subject, and

[0187] A pharmaceutical container is thus completed, the container containing one or more of the unit doses, the container optionally further containing or including a label directing the use of the package in treating the subject.

[0188] The present invention further provides a method of combination therapy for treating ALS, comprising administering to a subject in need thereof a therapeutically effective amount of pridopidine and a therapeutically effective amount of riluzole, edaravone, or dextromethorphan / quinidine.

[0189] For the foregoing embodiments, each embodiment disclosed herein is contemplated to be applicable to each other disclosed embodiment. For example, elements described in the method embodiment can be used in the pharmaceutical composition, packaging and use embodiments described herein, and vice versa.

[0190] All combinations, sub-combinations and permutations of the various elements of the methods and uses described herein are contemplated and are within the scope of the invention.

[0191] Pharmaceutically acceptable salts

[0192] As used herein, "pridopidine" refers to pridopidine base or its pharmaceutically acceptable salts and derivatives, such as pridopidine and salts of deuterium-enriched type. Examples of deuterium-enriched pridopidine and salts and methods for preparing the same can be found in U.S. Application Publication Nos. 2013-0197031, 2016-0166559 and 2016-0095847, each of which is incorporated herein by reference in its entirety. In certain embodiments, pridopidine is a pharmaceutically acceptable salt, such as, HCl salt or tartrate. Preferably, in any embodiment of the present invention described herein, pridopidine is in the form of its hydrochloride.

[0193] Examples of pharmaceutically acceptable addition salts include, but are not limited to, nontoxic inorganic and organic acid addition salts such as hydrochloride, hydrobromide, nitrate, perchlorate, phosphate, sulfate, format, acetate, aconate, ascorbate, benzenesulfonate, benzoate, cinnamate, citrate, succinate, heptanoate, fumarate, glutamate, glycolate, lactate, maleate, malonate, mandelate, methanesulfonate, naphthalene-2-sulfonate, phthalate, salicylate, sorbate, stearate, succinate, tartrate, p-toluenesulfonate, etc. These salts can be formed by procedures well known and described in the art.

[0194] "Deuterium-enriched" means that the abundance of deuterium at any relevant site of a compound is greater than the abundance of deuterium naturally occurring at that site in a certain amount of the compound. The naturally occurring distribution of deuterium is about 0.0156%. Thus, in a "deuterium-enriched" compound, the abundance of deuterium at any relevant site thereof exceeds 0.0156%, and may range from more than 0.0156% to 100%. Deuterium-enriched compounds may be obtained by exchanging hydrogen for deuterium or synthesizing the compound with deuterium-enriched starting materials.

[0195] Pharmaceutical composition

[0196] Although the pridopidine used according to the invention can be administered in the form of the raw compound, a preferred administration of pridopidine (optionally in the form of a physiologically acceptable salt) is in a pharmaceutical composition together with one or more adjuvants, excipients, carriers, buffers, diluents and / or other conventional pharmaceutical auxiliaries.

[0197] In one embodiment, the present invention provides a pharmaceutical composition comprising pridopidine or a pharmaceutically acceptable salt or derivative thereof, together with one or more pharmaceutically acceptable carriers, and optionally other therapeutic and / or prophylactic ingredients known and used in the art (including but not limited to riluzole, edaravone or (dextromethorphan / quinidine).

[0198] The carrier or carriers must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and suitable for administration to a human subject.

[0199] Combination therapy

[0200] When the present invention includes a combination of an active compound and one or more additional therapeutic and / or preventive ingredients, the combination of the present invention can be formulated together with at least one pharmaceutically acceptable carrier, additive, adjuvant or vehicle for simultaneous or synchronous administration thereof. This means that the combination of two active compounds can be administered as:

[0201] - as a combination which is part of the same pharmaceutical preparation, the two active compounds then being administered simultaneously, or

[0202] - as a combination of two units, each unit having one active substance which gives rise to the possibility of simultaneous or simultaneous administration.

[0203] The effects of any single drug are related to its absorption, distribution, and elimination. When two drugs are introduced into the body, each drug can affect the absorption, distribution, and elimination of the other drug, and thus change the effects of the other drug. For example, one drug can inhibit, activate, or induce the production of enzymes involved in the metabolic pathway that eliminates the other drug (Guidance for Industry, 1999).

[0204] The interaction between two drugs not only affects the expected therapeutic activity of each drug, but the interaction can increase the level of toxic metabolites (Guidance for Industry, 1999). The interaction can also strengthen or alleviate the side effects of each drug. Therefore, when two drugs are administered to treat a disease, it is impossible to predict what changes will occur in the negative properties of each drug.

[0205] In addition, it is difficult to predict exactly when the effects of an interaction between two drugs will become apparent. For example, a metabolic interaction between drugs may become apparent after the initial administration of the second drug, after both drugs reach steady-state concentrations, or after one drug is discontinued (Guidance for Industry, 1999).

[0206] In one example, experiments have shown that the combined administration of GA and interferon (IFN) abolished the clinical effectiveness of a therapy (Brod 2000). In another experiment, it was reported that the addition of pridopidine to a combined therapy with IFN-β antagonized its upregulatory effects. Therefore, when two drugs are administered to treat the same condition, it is unpredictable whether each drug will complement, influence, or interfere with the therapeutic activity of the other drug in human subjects.

[0207] Application

[0208] The pharmaceutical composition of the present invention can be used by any convenient approach suitable for the desired therapy. Preferred routes of administration include oral administration (particularly tablets, capsules, dragees, powders or liquid forms), intranasal administration, intradermal administration and parenteral administration (particularly skin, subcutaneous, intramuscular or intravenous injection). The pharmaceutical composition of the present invention can be manufactured by those skilled in the art by using standard methods and conventional techniques suitable for the desired preparation. When necessary, compositions adapted to sustained release of active ingredients can be used.

[0209] Tablet can contain suitable binding agent, lubricant, splitting agent (disintegrant), coloring agent, flavor enhancer, drainage agent and fluxing agent.For example, for the oral administration of the tablet or capsule of unit dosage form, active drug component can be combined with oral nontoxic pharmaceutically acceptable inert carrier (such as, lactose, gelatin, agar, starch, sucrose, glucose, methylcellulose, dicalcium phosphate, calcium sulfate, mannitol, sorbitol, microcrystalline cellulose etc.).Suitable binding agent includes starch, gelatin, natural sugar (such as, glucose or beta-lactose), corn starch, natural gum and synthetic gum (such as, gum arabic, tragacanth or sodium alginate), polyvidone, carboxymethyl cellulose, polyethylene glycol, wax etc.The lubricant used in these dosage forms includes sodium oleate, sodium stearate, sodium benzoate, sodium acetate, sodium chloride, stearic acid, sodium stearyl fumarate, talcum powder etc. Disintegrators (disintegrators) include, but are not limited to, starch, methyl cellulose, agar, bentonite, xanthan gum, cross-linked sodium carboxymethyl cellulose, sodium starch glycolate, and the like.

[0210] The general techniques and compositions for making dosage forms useful in the present invention are described in the following references: Modern Pharmaceutics, Chapters 9 and 10 (Banker & Rhodes, Editors, 1979); Pharmaceutical Dosage Forms: Tablets (Lieberman 1981); Ansel, Introduction to Pharmaceutical Dosage Forms 2nd Edition (1976); Remington's Pharmaceutical Sciences, 17th ed. (Mack Publishing Company, Easton, Pa., 1985); Advances in Pharmaceutical Sciences (David Ganderton, Trevor Jones, Eds., 1992); Advances in Pharmaceutical Sciences Vol 7. (David Ganderton, Trevor Jones, James McGinity, Eds., 1995); Aqueous Polymeric Coatings for Pharmaceutical Dosage Forms (Drugs and the Pharmaceutical Sciences, Series 36 (James McGinity, Ed., 1989); Pharmaceutical Particulate Carriers: Therapeutic Applications: Drugs and the Pharmaceutical Sciences, Vol 61 (Alain Rolland, Ed., 1993); Drug Delivery to the Gastrointestinal Tract (Ellis Horwood Books in the Biological Sciences. Series in Pharmaceutical Technology; J.G. Hardy, S.S. Davis, Clive G. Wilson, Eds); Modern Pharmaceutics Drugs and the Pharmaceutical Sciences, Vol. 40 (Gilbert S. Banker, Christopher T.Rhodes, Eds.) These references are incorporated herein by reference in their entirety.

[0211] the term

[0212] As used herein, and unless otherwise indicated, each of the following terms shall have the definitions shown below.

[0213] As used herein, "riluzole" refers to riluzole or a pharmaceutically acceptable salt and derivative thereof, for example, deuterium-enriched riluzole and salts. Riluzole is described in the Prescribers' Digital Reference, which is incorporated herein by reference (Riluzole PDR 2017).

[0214] As used herein, "edaravone" refers to edaravone or a pharmaceutically acceptable salt and derivative thereof, for example, deuterium-enriched edaravone and salts. Edaravone is described in the Prescribers' Digital Reference, which is incorporated herein by reference (Edaravone PDR 2017).

[0215] "A combination of dextromethorphan and quinidine" or "dextromethorphan / quinidine" or "dextromethorphan hydrobromide / quinidine sulfate" refers to a combination of dextromethorphan hydrobromide (20 mg) and quinidine sulfate (10 mg), such as, It is a drug currently on the market for the treatment of pseudobulbar effect (PBA), especially in ALS patients. Acts on sigma-1 and NMDA receptors in the brain. Recent data suggest that this combination has effects on bulbar function in ALS, but not other aspects of motor function (Smith 2017). Dextromethorphan hydrobromide / quinidine sulfate is described in the Prescribers' Digital Reference, which is incorporated herein by reference (Dextromethorphan hydrobromide / quinidine sulfate PDR 2017).

[0216] As used herein, the "amount" or "dose" of pridopidine measured in milligrams refers to the number of milligrams of underivatized pridopidine base present in the preparation, regardless of the form of the preparation. "A dose of 45 mg of pridopidine" means that the amount of pridopidine in the preparation is sufficient to provide 45 mg of underivatized pridopidine base with a naturally occurring isotopic distribution, regardless of the form of the preparation. Therefore, when in the form of a salt (e.g., pridopidine hydrochloride), the mass of the salt form necessary to provide a dose of 45 mg of underivatized pridopidine base will be greater than 45 mg due to the presence of additional salt ions. Similarly, when in the form of a deuterium-enriched derivative, the mass of the derivatized form necessary to provide a dose of 45 mg of underivatized pridopidine base with a naturally occurring isotopic distribution will be greater than 45 mg due to the presence of additional deuterium.

[0217] Any range disclosed herein means that all hundredths, tenths, and integer unit amounts within the range are specifically disclosed as part of the present invention. Thus, for example, 0.01 mg to 50 mg means that unit amounts of 0.02, 0.03...0.09; 0.1; 0.2...0.9; and 1, 2...49 mg are included as embodiments of the present invention. Any time range disclosed herein (i.e., week, month, or year) means that all time lengths of days and / or weeks within the range are specifically disclosed as part of the present invention. Thus, for example, 3-6 months means that 3 months and 1 day, 3 months and 1 week, and 4 months are included as embodiments of the present invention.

[0218] As used herein, "about" in the context of a value or range means ± 10% of the stated or claimed value or range.

[0219] As used herein, "monotherapy" refers to treatment with a single active agent, for example, treatment with pridopidine alone.

[0220] As used herein, "adjunctive" refers to treatment or administration with an additional compound together with the main compound, for example, to increase the efficacy or safety of the main compound or to enhance its activity.

[0221] As used herein, "regular administration" refers to repeated / repeated administration separated by time periods. The time periods between administrations are preferably sometimes consistent. Regular administration can include, for example, administration once a day, administration twice a day, administration three times a day, administration four times a day, administration once a week, administration twice a week, administration three times a week, administration four times a week, etc.

[0222] As used herein, "combination" refers to a collection of agents used in therapy by simultaneous or synchronous administration. Simultaneous administration refers to the administration of an admixture (regardless of a true mixture, suspension, emulsion, or other physical combination) of pridopidine and a second compound (e.g., riluzole). In this case, the combination may be an admixture or separate container of pridopidine and a second compound combined just before administration. Synchronous administration or concomitant administration refers to the separate administration of pridopidine and a second compound (e.g., riluzole) at the same time, or separate administration together at a sufficiently close time to observe additive activity or preferably synergistic activity related to the separate activity of pridopidine or the second compound, or separate administration in a sufficiently close time difference (to allow the separate therapeutic effects of each agent to overlap).

[0223] As used herein, "add-on" or "add-on therapy" refers to a collection of agents used in a therapy, wherein a subject receiving therapy begins a first treatment regimen of one or more agents and then begins a second treatment regimen of one or more different agents in addition to the first treatment regimen, such that all agents used in the therapy are not started at the same time. For example, adding pridopidine therapy to a patient already receiving riluzole therapy.

[0224] As used herein, "effective" when used to refer to an amount of pridopidine means that the amount of pridopidine is sufficient to produce the desired therapeutic response. In preferred embodiments, the amount of pridopidine administered does not result in adverse side effects (such as toxicity, irritation, or allergic reactions).

[0225] "Administering to a subject" or "administering to a (human) patient" refers to administering, dispensing or applying a drug, medication or treatment to a subject / patient to relieve, cure or alleviate the symptoms associated with a disease, disorder or condition, such as a pathological state.

[0226] As used herein, "treating" encompasses inducing inhibition, regression, or arrest of a disease or condition, or attenuating, suppressing, inhibiting, lessening the severity of, eliminating or substantially eliminating, or ameliorating the symptoms of a disease or condition.

[0227] "Inhibition" of disease progression or disease complications in a subject refers to preventing or alleviating disease progression and / or disease complications in a subject.

[0228] "Symptoms" associated with a disease or condition include any clinical or laboratory manifestation associated with the disease or condition, and are not limited to what a subject can feel or observe.

[0229] As used herein, a "subject" suffering from a disease, disorder or condition refers to a subject that has been clinically diagnosed as having the disease, disorder or condition.

[0230] Glial cell line-derived neurotrophic factor (GDNF) is a protein encoded by the GDNF gene and is thought to promote the survival of many types of neurons.

[0231] Brain-derived neurotrophic factor (BDNF) is a protein produced by neurons and serves to maintain function and promote neuronal growth and neurogenesis.

[0232] The following numbered clauses define various aspects and features of the invention:

[0233] 1. A method for treating a subject suffering from amyotrophic lateral sclerosis (ALS), the method comprising: periodically administering to the subject an amount of pridopidine effective to treat the subject.

[0234] 2. The method of clause 1, wherein the amount of pridopidine is effective to inhibit or reduce the progression of symptoms of ALS in the subject.

[0235] 3. The method of item 2, wherein the symptom is muscle stiffness, muscle weakness, muscle wasting, muscle cramps, difficulty speaking, difficulty swallowing, difficulty breathing, difficulty chewing, difficulty walking, fasciculations and / or worsening of posture.

[0236] 4. The method of any one of clauses 1-3, wherein the ALS is sporadic ALS.

[0237] 5. The method of any one of clauses 1 to 4, wherein the amount of pridopidine is administered daily, or wherein the amount of pridopidine is administered more than once daily.

[0238] 6. The method of any one of clauses 1-4, wherein the amount of pridopidine is administered twice daily.

[0239] 7. The method of any one of clauses 1-4, wherein the amount of pridopidine is administered less than once daily.

[0240] 8. The method of any one of clauses 1-7, wherein the amount of pridopidine is administered orally.

[0241] 9. The method of any one of clauses 1 to 8, wherein the amount of pridopidine administered is 22.5 mg to 225 mg per day.

[0242] 10. The method of clause 9, wherein the amount of pridopidine administered is 45 mg per day to 180 mg per day.

[0243] 11. The method of clause 9, wherein the amount of pridopidine administered is 5 mg, 10 mg, 22.5 mg, 45 mg, 67.5 mg, 90 mg, 100 mg, 112.5 mg, 125 mg, 135 mg, 150 mg or 180 mg per day.

[0244] 12. The method of any one of clauses 1-11, wherein the periodic administration continues for at least 24 weeks.

[0245] 13. The method of any one of clauses 1 to 12, wherein the pridopidine is pridopidine hydrochloride.

[0246] 14. The method of any one of clauses 1-13, wherein the subject is a human subject.

[0247] 15. The method of any one of clauses 1-14, further comprising: administering to the subject a therapeutically effective amount of a second compound.

[0248] 16. The method of clause 15, wherein the second compound is riluzole, edaravone, or dextromethorphan / quinidine.

[0249] 17. The method of clauses 15-16, wherein the pridopidine and the second compound are administered in one unit.

[0250] 18. The method of clauses 15-16, wherein the pridopidine and the second compound are administered in more than one unit.

[0251] 19. The method of any of clauses 15-18, wherein the second compound is riluzole.

[0252] 20. The method of clause 19, wherein 10 mg-200 mg, or 50 mg, 100 mg, or 200 mg of riluzole is administered to the subject daily.

[0253] 21. The method of any one of clauses 15-20, wherein the riluzole is administered orally.

[0254] 22. The method of any of clauses 15-18, wherein the second compound is edaravone.

[0255] 23. The method of clause 22, wherein 5-60 mg, or 30 mg, or 60 mg of edaravone is administered to the subject daily.

[0256] 24. The method of any of clauses 15-18 and 22-23, wherein the edaravone is administered by intravenous infusion.

[0257] 25. The method of any one of clauses 15-18 and 22-24, wherein the edaravone is administered once daily for 14 days or 10 days, followed by a 14-day drug-free period.

[0258] 26. The method of any of clauses 15-18, wherein the second compound is dextromethorphan / quinidine.

[0259] 27. The method of clause 26, wherein 10, 20, or 40 mg of dextromethorphan is administered to the subject daily and 5, 10, or 20 mg of quinidine is administered to the subject daily.

[0260] 28. The method of any of clauses 26-27, wherein the dextromethorphan / quinidine is administered orally.

[0261] 29. The method of any one of clauses 15-28, wherein the amount of pridopidine and the amount of the second compound are administered simultaneously.

[0262] 30. The method of any of clauses 15-28, wherein administration of the second compound substantially precedes administration of pridopidine.

[0263] 31. The method of any of clauses 15-28, wherein administration of pridopidine substantially precedes administration of the second compound.

[0264] 32. The method of any of clauses 15-28, wherein the subject is receiving edaravone therapy, dextromethorphan / quinidine therapy, or riluzole therapy prior to initiating pridopidine therapy.

[0265] 33. The method of clause 32, wherein the subject has been receiving edaravone therapy, dextromethorphan / quinidine therapy, or riluzole therapy for at least 24 weeks, 28 weeks, 48 ​​weeks, or 52 weeks prior to initiating pridopidine therapy.

[0266] 34. The method of any of clauses 15-28, wherein the subject is receiving pridopidine therapy prior to initiating edaravone therapy, dextromethorphan / quinidine therapy, or riluzole therapy.

[0267] 35. The method of clause 34, wherein the subject has been receiving pridopidine therapy for at least 24 weeks, 28 weeks, 48 ​​weeks, or 52 weeks prior to initiating edaravone therapy, dextromethorphan / quinidine therapy, or riluzole therapy.

[0268] 36. The method of any of clauses 15-35, wherein each of said amount of said second compound when taken alone and said amount of pridopidine when taken alone are effective to treat said subject.

[0269] 37. The method of any of clauses 15-35, wherein the amount of the second compound when used alone, the amount of pridopidine when used alone, or each such amount when used alone is not effective to treat the subject.

[0270] 38. The method of any of clauses 15-35, wherein the amount of the second compound when used alone, the amount of pridopidine when used alone, or each such amount when used alone is less effective in treating the subject.

[0271] 39. The method of any one of clauses 15-38, wherein the pridopidine is administered adjunctively to the second compound.

[0272] 40. The method of any one of clauses 15-38, wherein the second compound is administered adjunctively to the pridopidine.

[0273] 41. The method of any one of clauses 1-40, wherein a loading dose of a different amount than the intended dose is administered at a time period beginning with the periodic administration.

[0274] 42. A method of enhancing axonal transport of BDNF in motor neurons in a subject suffering from ALS, the method comprising administering to the subject an amount of pridopidine effective to enhance axonal transport of BDNF in motor neurons of the subject.

[0275] 43. A method of improving NMJ formation and function in a subject suffering from ALS, the method comprising administering to the subject an amount of pridopidine effective to improve NMJ formation and muscle contractile function in the subject.

[0276] 44. A method of improving innervation of muscle tissue in a subject suffering from ALS, the method comprising administering to the subject an amount of pridopidine effective to improve innervation in the subject.

[0277] 45. A method of enhancing motor neuron axon growth in a subject suffering from ALS, the method comprising administering to the subject an amount of pridopidine effective to enhance motor neuron axon growth in the subject.

[0278] 46. ​​A method of increasing muscle contraction in a subject suffering from ALS, the method comprising administering to the subject an amount of pridopidine effective to increase muscle contraction in the subject.

[0279] 47. A method of restoring muscle contraction in a subject suffering from ALS, the method comprising administering to the subject an amount of pridopidine effective to improve muscle contraction in the subject.

[0280] 48. A pharmaceutical composition comprising an effective amount of pridopidine for use in treating a subject suffering from ALS.

[0281] 49. Use of an amount of pridopidine in the manufacture of a medicament for treating a subject suffering from ALS.

[0282] 50. A package, comprising:

[0283] a) a pharmaceutical composition comprising an amount of pridopidine; and optionally

[0284] b) instructions for using the pharmaceutical composition to treat a subject suffering from ALS.

[0285] 51. A therapeutic package for or use in administering a drug to a subject, the therapeutic package comprising:

[0286] a) one or more unit doses, each such unit dose comprising an amount of pridopidine therein, wherein upon administration to said subject, the amount of pridopidine in said unit dose is effective for treating ALS in said subject, and

[0287] b) a pharmaceutical container complete therefor, said container containing one or more said unit doses, said container further containing or comprising a label directing the use of said package in treating a subject suffering from ALS.

[0288] 52. A package, comprising:

[0289] a) a first pharmaceutical composition comprising a certain amount of pridopidine and a pharmaceutically acceptable carrier;

[0290] b) a second pharmaceutical composition comprising an amount of a second compound and a pharmaceutically acceptable carrier, wherein the second compound is riluzole, edaravone or dextromethorphan / quinidine; and optionally

[0291] c) instructions for using the first and second pharmaceutical compositions together to treat a subject suffering from ALS.

[0292] 53. The package of clause 52, wherein said amount of the second compound and said amount of pridopidine are prepared to be administered simultaneously or synchronously.

[0293] 54. A therapeutic package for or use in administering medication to a subject suffering from ALS, the therapeutic package comprising:

[0294] a) one or more unit doses, each such unit dose comprising:

[0295] i) a certain amount of Pridopidine and

[0296] ii) an amount of a second compound, wherein the second compound is riluzole, edaravone, or dextromethorphan / quinidine;

[0297] wherein the respective amounts of the pridopidine and the second compound in the unit dose are effective for treating the subject upon concomitant administration to the subject, and

[0298] b) a pharmaceutical container completed therefor, said container containing one or more of said unit doses, said container further containing or comprising a label directing use of said package in treating said subject.

[0299] 55. A pharmaceutical composition comprising an amount of pridopidine and an amount of a second compound, wherein the second compound is riluzole, edaravone, or dextromethorphan / quinidine.

[0300] 56. The pharmaceutical composition of clause 55, for use in treating a subject suffering from ALS, wherein the pridopidine and the second compound are prepared to be administered simultaneously, synchronously or concomitantly.

[0301] 57. A pharmaceutical composition in unit dosage form useful for treating a subject suffering from ALS, the pharmaceutical composition comprising:

[0302] a) a certain amount of pridopidine;

[0303] b) an amount of a second compound, wherein the second compound is riluzole, edaravone or dextromethorphan / quinidine,

[0304] wherein, upon concomitant administration of one or more of said unit dosage forms of said composition to said subject, the respective amounts of said second compound and said pridopidine in said composition are effective for treating said subject.

[0305] 58. A pharmaceutical composition comprising an amount of pridopidine for use as add-on therapy to a second compound, which is riluzole, edaravone, or dextromethorphan / quinidine, in treating a subject suffering from ALS.

[0306] 59. A pharmaceutical composition comprising an amount of pridopidine for simultaneous, concurrent or concomitant treatment of a subject suffering from ALS with a second compound, the second compound being riluzole, edaravone or dextromethorphan / quinidine.

[0307] 60. A pharmaceutical composition comprising an amount of a compound that is riluzole, edaravone, or dextromethorphan / quinidine for use as add-on therapy to pridopidine in treating a subject suffering from ALS.

[0308] 61. A pharmaceutical composition comprising an amount of a compound that is riluzole, edaravone, or dextromethorphan / quinidine for use in treating a subject suffering from ALS simultaneously, concurrently, or concomitantly with pridopidine.

[0309] 62. A compound which is riluzole, edaravone, or dextromethorphan / quinidine for use as an add-on therapy to pridopidine in treating a subject suffering from ALS.

[0310] 63. Pridopidine for use as an add-on therapy to a compound which is riluzole, edaravone, or dextromethorphan / quinidine in treating a subject suffering from ALS.

[0311] 64. The add-on therapy of clause 63, wherein the therapy is for treating, preventing or alleviating a symptom of ALS.

[0312] 65. A combination of pridopidine and a compound which is riluzole, edaravone or dextromethorphan / quinidine, for use in treating, preventing or alleviating the symptoms of ALS.

[0313] Throughout this application, certain publications and patent application disclosures are cited. The complete citations of these publications can be found immediately before the claims. The disclosures of these publications and patent application disclosures are hereby incorporated by reference to more fully describe the state of the art to which the present invention relates.

[0314] The present invention will be better understood by reference to the following "Experimental Details", but one skilled in the art will readily appreciate that the specific experiments detailed are merely illustrative of the invention as more fully described in the claims that follow.

[0315] Experimental details

[0316] Experiment 1: Axonal transport assay

[0317] Healthy motor neurons (MNs) extend axons over long distances and form synapses with muscles through a changing extracellular microenvironment. The ability of neurons to maintain this specialized morphology depends on cytoskeletal elements and the continuous transport of proteins and organelles to and from the cell body. Cytoskeletal alterations are a major pathway involved in the pathogenesis of ALS, affecting axonal transport, growth, and neuromuscular junction (NMJ) function (Eykens and Robberecht, 2015). Alterations in axonal transport are one of the first cellular processes to emerge in neurodegenerative diseases including ALS. Axonal transport was assessed using an in vitro compartment system of microfluidic chambers that separates the neuronal cell body from its axons and synapses, which enables the study of retrograde / anterograde transport ( Figure 1b ; Zahavi 2015; Ionescu 2016).

[0318] Quantum dot-labeled BDNF (Qdot BDNF) is retrogradely transported in axons of motor neuron explants grown in a microfluidic chamber (MFC). The MFC was used to analyze Qdot BDNF axonal transport. Axonal transport of BDNF has been studied in the SOD1 model of ALS (SOD1G93A) (Bilsland 2010; Perlson 2009; De Vos 2007). The effect of pridopidine on axonal transport of Qdot BDNF along motor neurons was evaluated in spinal cord explants from SOD1 and WT littermates (LM). Experimental flow chart of axonal transport assay (from left to right, Figure 1a ): SOD1G93A or wild-type (LM) spinal cord explants were plated in MFCs. About 5 days after plating, axons began to cross into the distal compartment. On the 6th day after plating, a certain amount of pridopidine was added to both compartments. On the 7th day, Qdot-BDNF was added to the distal compartment and axonal transport was imaged. Schematic diagram of the microfluidic chamber system ( Figure 1b ): Spinal cord (SC) explants from WT or SOD1G93A E12.5 mouse embryos were plated in the proximal compartment of a microfluidic chamber. MNs from the spinal cord sent their axons to the distal compartment. At this point, only Qdot-BDNF was applied to this compartment. Axonal transport of Qdot-BDNF was then visualized and recorded by a high-resolution spinning disk confocal imaging system.

[0319] QdotBDNF was tracked along axons of motor neuron explant cultures using spinning disk confocal microscopy. Time-lapse images of axonal transport of Qdot-BDNF acquired at 60X magnification ( Figure 1c). The yellow arrow points to a single Qdot-BDNF particle that is transported retrogradely toward the cell body (left). Scale bar: 10 μm. The figure below shows the motion of the complete Qdot-BDNF time-lapse movie. Scale bar: horizontal 10 μm; vertical 100 seconds ( Figure 1c )

[0320] On day 6 of the experiment, vehicle and pridopidine were added to both sides of the chamber at 2 concentrations (0.1 μM and 1 μM), and after overnight incubation with pridopidine, Qdot BDNF was added to the distal compartment ( Figure 1a and Figure 1b ). Six independent biological replicates from 6 different cultures were tested, so that from each culture and explants with neurons / glial cells, ~250 BDNF particles were tracked along axons in the trough. The velocity n refers to the movement of a single BDNF particle. The experiment was repeated with MNs from sigma1 receptor knockout mice (S1R KO or S1R- / -) (Langa, 2003). Ventral spinal cord slices from S1R KO mouse embryos were cultured and plated in MFCs as described above, and axonal transport of Qdot-BDNF was analyzed.

[0321] SOD1G93 and S1R KO explants with or without pridopidine were compared to littermate controls (LM).

[0322] Qdot-BDNF particle tracking was performed on Bitplane Imaris using the semi-automatic dot tracking feature. Inclusion criteria for particle analysis: tracking duration >10 frames; average velocity ≥0.2 μm / s; stop duration: velocity <0.1 μm / s for 3 frames. Data were then exported to MATLAB for further analysis of particle transport, including instantaneous velocity ( Figure 2a ); and stop counting ( Figure 2b ).

[0323] result:

[0324] Figure 2ashowed that in SOD1G93A motor neurons, pridopidine enhanced the instantaneous velocity of BDNF axonal transport. The instantaneous velocity of BDNF retrograde transport is normally reduced in SOD1G93A motor neurons. SOD1G93AMNs displayed slower velocities compared to WT MNs. Pridopidine treatment accelerated the instantaneous velocity in both WT MNs (0.1μM) and SOD1G93A MNs (0.1μM and 1μM). Application of 25μM or 100μM riluzole to SOD1G93A MNs did not affect the instantaneous velocity. S1R- / - MNs showed defects in BDNF axonal transport. Pridopidine at 0.1μM or 1μM did not restore these defects in S1R KO MNs ( Figure 2a ).

[0325] Particle stall counts (the number of stalls per second that Qdot-BDNF was counted as) show that pridopidine reduces the number of stalls during axonal transport in WT (blue) MNs (0.1 μM only) and SOD1G93A (red) MNs (0.1 μM and 1 μM). Axonal transport parameters of Qdot-BDNF in S1R- / - MNs show that they are unresponsive to pridopidine at any concentration tested ( Figure 2b ).

[0326] These results suggest that pridopidine enhances axonal transport of BDNF in SOD1G93A motor neurons and has the ability to correct ALS-related defects.

[0327] Experiment 2: Axon-muscle growth / degeneration assay

[0328] An early event in the pathogenesis of ALS is axonal degeneration. The compartment co-culture microfluidic chamber system was used to determine whether pridopidine modifies axonal degeneration ( Figure 3 ). Primary muscle cells from pre-symptomatic (P60) WT or SOD1G93A mice were cultured. On the 6th day, primary skeletal muscle myoblasts were cultured in the distal compartment of the MFC. About six days later (day 12), ventral spinal cord explants from WT or SOD1G93A E12.5 mouse embryos expressing HB9-GFP (specific motor neuron marker) were laid in the proximal compartment, and then pridopidine or vehicle was applied to both compartments. Pridopidine was updated every other day. Two days after the explants were laid (day 14), the growth and degeneration of motor axons were assessed using real-time imaging on a spinning disk confocal system. Axon growth was tracked by imaging every 10 minutes for 8 hours. The experiment was repeated three times.

[0329] result:

[0330] Data suggest that pridopidine increases axonal growth ( Figure 4). Compared to WT (LM) myocytes, myocytes carrying the SOD1G93A mutation have a reduced number of healthy axons that are able to cross into the distal compartment of the chamber of the microfluidic chamber. Treatment with 1 μM pridopidine (rightmost bar) shows an increase in the number of axons that cross into the distal compartment (the compartment with the muscle cells). (Y axis is the average number of slots with axons crossing into the muscle compartment).

[0331] These results suggest that pridopidine enhances axonal growth and has the ability to correct ALS-related defects.

[0332] Experiment 3: Measurement of neuromuscular junction (NMJ) formation and function

[0333] Synaptic disruption is the earliest cellular compartment to be destroyed in ALS. To test the ability of pridopidine to affect synaptic function in the ALS model, cultures from Experiment 2 were grown for approximately four additional days (Day 18) when axons projected into the distal compartment and made NMJ connections. In this co-culture, primary myocytes fully differentiated and formed connections with MN axons, and these connections were postsynaptic markers (AchR (Acetylcholine R)) colocalized with HB9:GFP neurons. Figure 5a : Top: Phase image of a myocyte in the distal compartment connected by an axon (green arrow). Scale bar: 20 μm. Bottom: High magnification image of myocyte:MN contact reveals NMJ formation as shown by colocalization of postsynaptic AChRs with HB9::GFP axons and 3-dimensional colocalization of pre- and postsynaptic markers (coloc). To assess NMJ function, 1000 frames of movies of muscle contraction were acquired at a frame rate of 30 frames per second ( Figure 5b ). Muscle contraction trajectories extracted from time-dependent intensity measurements of muscle contractions show flat trajectories for non-contracted, immobile myocytes (top), and trajectories for contracting myocytes exhibit multiple burst events (bottom).

[0334] To investigate the effects of pridopidine on MN and NMJ formation and function, 0.1 or 1 μM pridopidine or vehicle was added. Measurements of the percentage of innervation in myotubes and the percentage of innervation-induced contractions were assessed using live cell imaging as described previously (Ionescu 2015; Zahavi 2015). Briefly, the contractile activity of muscles in the distal compartment of the MFC overlapped by at least one axon was examined. Muscles were classified into two categories: “contracted” or “non-contracted”, depending on their motile activity during the movie. The motility of the muscles was verified by generating intensity plots over time for each muscle ( Figure 5b The number of contracted myofibers in each chamber was divided by the total number of myofibers analyzed in the same chamber to give the percentage of contracted myotubes as output for NMJ activity.

[0335] result:

[0336] Treatment of muscle cells with pridopidine induced innervation, which resulted in a higher probability of burst muscle patterning. Muscles carrying the SOD1 mutation had a lower innervation rate than WT (wild-type) muscles (20% innervation compared to ~40% in WT). 1 μM pridopidine restored the innervation rate of muscles carrying the SOD1 mutation to near WT levels ( Figure 6 ).

[0337] The percentage of contracted myofibers in the co-cultures showed that in the absence of MNs, only ~10% of the muscles were contracted, while in cultures that included MNs, this was 74%. Co-culture combinations that included at least one cell type expressing SOD1G93A showed a significantly lower percentage of contracted myocytes ( Figure 7a ). Treatment of SOD1G93A myocytes co-cultured with pridopidine (0.1 μM and 1 μM) increased the percentage of contracted myocytes and restored neuromuscular activity to WT levels. The combination of S1R- / -MNs and WT myocytes resulted in a low number of contracted myotubes compared to WT MNs. As indicated by the same concentration of pridopidine in co-culture with WT neurons, the application of 0.1 μM pridopidine to S1R- / - co-culture did not restore neuromuscular activity. The increase in the percentage of contracted myocytes observed after treatment with 1 μM pridopidine was significantly lower than the percentage of contracted myocytes in untreated and treated WT cultures. Data are shown as mean ± SEM. *p-value <0.05; **p-value <0.01, ***p-value <0.001, ****p-value <0.0001. (Student's t test).

[0338] Experiment 4: Activation of ERK in WT and SOD1G93A MNs

[0339] The ERK pathway promotes many cellular functions, including proliferation and differentiation. ERK phosphorylation (activation) in neurons is associated with neurotrophic signaling, such as stimulation of BDNF, which in turn promotes neuroprotection and neuronal survival (Bonni 1999). It has been previously determined that pridopidine enhances BDNF signaling in the mouse striatum through S1R, which in turn enhances ERK activation (Geva 2016). Primary MN cultures at 2DIV were starved overnight in neurotrophin-free and serum-free medium (PNB). The next day, cultures were treated with pridopidine or with BDNF as a positive control for 30 minutes.

[0340] result:

[0341] Immunoblotting analysis of phosphorylated ERK (pERK) showed significant activation of ERK by pridopidine (0.1 μM and 1 μM) as early as 30 min in culture in WT MNs (left panel) and SOD1G93A (middle panel), but not in S1R- / - MNs (right panel). Figure 8a ). Quantification of pERK revealed an increase of ∼3.5- and ∼4-fold in WT MNs after 0.1 μM and 1 μM pridopidine, respectively. SOD1G93A showed an increase of ∼2.9- and ∼8.5-fold in pERK after 0.1 μM and 1 μM pridopidine, respectively. Pridopidine did not induce ERK phosphorylation in S1R- / - MNs. Data are shown as mean pERK / ERK ratio ± SEM. *p-value < 0.05, ~p-value < 0.1 (Student's t-test) ( Figure 8b ).

[0342] Experiment 5: Effect of Pridopidine on mutant SOD1 aggregates in the spinal cord of SOD1G93A mice

[0343] Pridopidine induces neuroprotective properties by activating S1R, as shown by its effects on axonal transport, axonal degeneration, NMJ function, and ERK activation. S1R is located on the ER membrane in close proximity to the outer mitochondrial membrane, where mutant SOD1 protein tends to aggregate in the spinal cord of SOD1G93A mice (Millecamps and Julien 2013). Presymptomatic SOD1G93A mice (5 weeks of age) and wild-type (WT) controls were treated with saline, 3 or 30 mg / kg of pridopidine by daily sc (subcutaneous) administration for 11 weeks (until 16 weeks of age). At the end of the experiment, lumbar spinal cords (L1-L6) were extracted, fixed and embedded for cryosectioning. Next, 10 μM sections were prepared and stained with NSC500 dye to visualize SOD1 aggregates ( 2010). To evaluate the effect of in vivo pridopidine treatment on the number of mutant SOD1 aggregates in the gray and white matter of the spinal cord.

[0344] result:

[0345] Figure 9a-Left: Low magnification representative images of fluorescently labeled spinal cords from 4 groups of mice. Right: High magnification images of the area marked by a square in the left image. Scale bars: Left: 500 μm; Right: 50 μm. From top to bottom: WT vehicle, SOD1G93A vehicle, SOD1G93A 0.3 mg / kg pridopidine, SOD1G93A 30 mg / kg, all stained with NSC500 dye to mark mutant SOD1 protein aggregates. A significant increase in the number of mSOD1 aggregates was observed in both the gray and white matter of the spinal cord of SOD1G93A mice compared to WT mice. 30 mg / kg of pridopidine significantly reduced the number of aggregates in both the gray and white matter of the SOD1G93A spinal cord by ~50% ( Figures 9a to 9c ). Data are shown as mean ± SEM. *p-value < 0.05; **p-value < 0.01 (one-way ANOVA followed by Fisher's LSD post hoc comparison test). ( Figure 9b to Figure 9c The y-axis is the number of NSC500-positive SOD1 aggregates per square millimeter).

[0346] Experiment 6: In vivo assessment of myofiber atrophy and preservation of NMJ

[0347] NMJ destruction and subsequent skeletal muscle loss are two major pathologies of ALS. The effect of pridopidine on muscle fiber atrophy and NMJ retention was evaluated in vivo. Presymptomatic SOD1G93A mice and wild-type (WT) controls (5 weeks old) were treated with saline, 3 or 30 mg / kg of pridopidine by daily sc administration for 11 weeks. At 16 weeks of age, gastrocnemius muscles of mice treated with vehicle or pridopidine (30 mg / kg sc) were extracted from SOD1G93A and WT mice. Muscle cross sections were stained with hematoxylin and eosin (H&E), and the average muscle fiber diameter ( Fig.10a NMJ preservation was assessed by confocal imaging of colocalized presynaptic (NFH+synaptophysin-I (green)) and postsynaptic (AchR (BTX; red)) markers and counting the number of fully innervated NMJs in the gastrocnemius muscle ( Fig.11a ).

[0348] result:

[0349] Fig.10a: Representative images of H&E stained cross sections of gastrocnemius muscle from the following 4 groups of mice: WT-vehicle treated, WT-30mg / kg pridopidine treated, SOD1G93A-vehicle treated, and SOD1G93A-30mg / kg pridopidine treated mice. The muscle histology of SOD1G93A-vehicle mice was poor, and revealed that the diameter of its muscle fibers was smaller ( 1.34 × 10 ... Figure 10a to Figure 10b Pridopidine (30 mg / kg daily, sc) resulted in a significant increase in myofiber diameter of -4 μm in SOD1G93A and -5 μm in WT muscle.

[0350] NMJ preservation in muscles of SOD1G93A vehicle-treated mice showed the expected large loss of NMJs of ∼60% compared to WT mice, and morphological changes in the postsynaptic apparatus in muscles of SOD1G93A mice ( Figure 11a to Figure 11b ). Surprisingly, pridopidine treatment limited the loss of NMJs in SOD1G93A mice to ˜20%. Data are shown as mean±SEM. *p-value<0.05; **p-value<0.01; ***p-value<0.001 (double-blind Student's t-test).

[0351] Overall, these results show that pridopidine plays a neuroprotective role in ALS cell and animal models. In vitro, in SOD1G93A MN, pridopidine enhances BDNF axonal transport, upregulates ERK activation, enhances axonal growth, restores muscle innervation and improves NMJ formation and function. These neuroprotective effects are mediated by S1R. In vivo pridopidine treatment of SOD1G93A ALS mice reduces the aggregation of mutant SOD1 in the spinal cord (one of the hallmark disease phenotypes), increases the diameter of muscle fibers reduced by ALS, and retains the degenerated NMJ observed in diseased tissues. These data support the use of pridopidine as a neuroprotectant, and the use of S1R as a therapeutic target for treating ALS patients.

[0352] Additional ALS animal models useful for testing pridopidine have been described, for example, in McGoldrick (2013).

[0353] In the figures, the abbreviations are as follows: Geno. = genotype (ie, wild type (LM), mutant SOD1), Prido. = Pridopidine, mpk = milligrams per kilogram.

[0354] Experiment 7: Treatment of ALS in human subjects

[0355] Regular oral administration of pridopidine provides a clinically meaningful advantage in reducing symptoms of ALS in human subjects suffering from ALS. Pridopidine therapy provides efficacy in treating patients without undue adverse side effects and is effective in at least one of the following embodiments:

[0356] 1. The therapy is effective in improving the symptoms of ALS;

[0357] 2. The therapy is effective in enhancing BDNF axonal transport and / or enhancing ERK activation in motor neurons;

[0358] 3. The therapy is effective in improving NMJ formation and preservation, preserving NMJ structure, preserving NMJ function and / or improving the rate of innervation of muscle tissue;

[0359] 4. The therapy is effective to enhance motor neuron axon growth and / or reduce axonal degeneration including motor neuron axon degeneration;

[0360] 5. The therapy is effective in enhancing muscle cell survival, increasing muscle fiber diameter and function, reducing the progression of muscle fiber wasting, and / or improving muscle contraction; and / or

[0361] 6. The therapy is effective in reducing SOD1 aggregation and / or alleviating pseudobulbar disease progression.

[0362] In some patients, the attending physician administers pridopidine and a second compound, wherein the second compound is riluzole, edaravone, dextromethorphan / quinidine. In some embodiments, the second compound is laquinimod.

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[0395] U.S. Patent No. 7,923,459

[0396] U.S. Patent No. RE46117

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Claims

1. Use of a pridopidine salt in the preparation of a medicament for treating amyotrophic lateral sclerosis (ALS) in a subject.

2. The use according to claim 1, wherein the amyotrophic lateral sclerosis is sporadic amyotrophic lateral sclerosis.

3. The use of claim 1 or 2, wherein the amount of pridopidine salt is effective to inhibit or reduce the progression of amyotrophic lateral sclerosis symptoms in the subject.

4. The method of claim 3, wherein the symptoms are muscle stiffness, muscle weakness, muscle wasting, muscle cramps, difficulty speaking, difficulty swallowing, difficulty breathing, difficulty chewing, difficulty walking, fasciculations and / or worsening of posture.

5. The use of any one of claims 1, 2 and 4, wherein an amount of pridopidine salt is effective in the subject suffering from amyotrophic lateral sclerosis to: enhance axonal transport of brain-derived neurotrophic factor (BDNF) in motor neurons, enhance ERK activation in motor neurons, improve neuromuscular junction (NMJ) formation and preservation, increase the innervation rate of muscle tissue, enhance motor neuron axon growth, reduce axon degeneration, enhance muscle cell survival, enhance muscle fiber diameter and function, reduce superoxide dismutase 1 (SOD1) aggregation, reduce pseudobulbar disease progression, reduce muscle fiber loss progression and / or improve muscle contraction.

6. The use according to any one of claims 1, 2 and 4, wherein a certain amount of pridopidine salt is administered daily, or more than once a day.

7. The use according to any one of claims 1, 2 and 4, wherein the amount of pridopidine salt is administered twice daily.

8. The use according to any one of claims 1, 2 and 4, wherein the amount of pridopidine salt is administered orally.

9. The use according to any one of claims 1, 2 and 4, wherein the pridopidine salt is pridopidine hydrobromide, nitrate, perchlorate, phosphate, sulfate, formate, acetate, aconate, ascorbate, benzenesulfonate, benzoate, cinnamate, citrate, succinate, heptanoate, fumarate, glutamate, glycolate, lactate, maleate, malonate, mandelate, methanesulfonate, naphthalene-2-sulfonate, phthalate, salicylate, sorbate, stearate, succinate, tartrate or p-toluenesulfonate.

10. The use of any one of claims 1, 2 and 4, wherein when the medicament is used, a therapeutically effective amount of a second compound is also administered to the subject, wherein the second compound is riluzole, edaravone or dextromethorphan / quinidine.

11. The use according to claim 10, wherein the second compound is administered before the administration of the pridopidine salt.

12. The use of claim 10, wherein the pridopidine salt is administered prior to administration of the second compound.

13. The use of claim 10, wherein the pridopidine salt is administered adjunctively to the second compound.

14. The use of claim 10, wherein the second compound is administered adjunctively to the pridopidine salt.

15. The use of any one of claims 1, 2, and 4, wherein the amount of pridopidine salt is effective to preserve NMJ structure, preserve NMJ function, and / or reduce motor neuron axon degeneration in said subject suffering from amyotrophic lateral sclerosis.

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