Synergistic combination therapy for the treatment of ALS

Abstract

A method for inhibiting the progression of amyotrophic lateral sclerosis (ALS) in humans is disclosed that utilizes a synergistic combination of 2,4-dihydro-3H-pyrazol-3-one and deuterated arachidonic acid or a prodrug thereof, such as 11,11-D2-linoleic acid or an ester thereof.

Description

[Technical field]

[0001] cross reference This application claims the benefit of U.S. Provisional Application No. 63 / 310541, filed February 15, 2022, U.S. Provisional Application No. 63 / 310544, filed February 15, 2022, and U.S. Provisional Application No. 63 / 313190, filed February 23, 2022, and is a continuation of and claims the benefit of U.S. Patent Application No. 17 / 684279, filed March 1, 2022, all of which are incorporated by reference in their entireties.

[0002] Incorporation by Reference All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference.

[0003] Technical Field A method for inhibiting the progression of amyotrophic lateral sclerosis (ALS) in humans is disclosed. The method uses a synergistic combination of 3-methyl-1-phenyl-2-pyrazolin-5-one (commercially available under the name "edaravone" from Millipore Sigma, St. Louis, Missouri, USA, for the treatment of ALS) and a prodrug of deuterated arachidonic acid. The combination of edaravone and the prodrug has been shown to significantly reduce the rate of disease progression. [Background technology]

[0004] ALS is a debilitating and fatal neurodegenerative human disease that, despite researchers' best efforts, remains incurable, so doctors try to slow the progression of the disease and maintain patients' quality of life for as long as possible.

[0005] ALS is a fatal, slow-onset, progressive neurological disease with corresponding pathological features such as progressive muscle weakness, muscle atrophy, and spasticity, all reflecting the degeneration and death of upper or lower motor neurons. Once diagnosed, most patients experience disease progression at an initially slow rate, followed by rapid progression, typically resulting in death within 3-4 years, although some patients die sooner.

[0006] The underlying features of the disease involve lipid peroxidation (LPO) of polyunsaturated fatty acids (PUFAs) in motor neurons. Central to this oxidative pathway is the presence of an unstable bis-allylic hydrogen found in arachidonic acid, the abundant PUFA in neurons. The structure of arachidonic acid, including identification of the bis-allylic moiety, is as follows: [ka]

[0007] Arachidonic acid accumulates in cell membranes, and oxidative processes involving reactive oxygen species (ROS) act as initiators, resulting in the autoxidation of these PUFAs by abstraction of bis-allylic hydrogen and formation of oxidative reactive species in the PUFAs. Once the initial oxidation occurs at the first bis-allylic site, sequential oxidation of additional PUFAs occurs in the cell or mitochondrial membrane. The oxidative process begins with hydrogen abstraction at the bis-allylic site on the first PUFA, then the next PUFA, and so on, in a sequential manner. At some point, the oxidative process impairs or destroys neuronal viability, promoting pathologies that cause the generation of excessive amounts of ROS.

[0008] Previously, the art has disclosed that edaravone, an antioxidant and free radical scavenger, reduces the rate of loss of function characteristic of ALS. Similarly, the art has disclosed that deuteration of arachidonic acid at one or more bis-allylic sites in neurons can inhibit the progression of ALS. The stability of deuterium-carbon bonds against such oxidative processes is significantly stronger (more stable) than that of hydrogen-carbon bonds, such as deuterated arachidonic acid with two deuteriums substituted at position 13 (each hydrogen replaced by deuterium). This means that the carbon-deuterium bond reduces the generation of oxidative species at the bis-allylic sites, inhibiting lipid peroxidation. As a result, inhibition of this pathway promotes neuronal survival and inhibits disease progression.

[0009] As shown in the Examples herein, unpublished data from clinical trials indicate that patients treated with a prodrug of 13,13-D2-arachidonic acid benefited compared to untreated patients during periods of rapid functional decline associated with ALS. This benefit was manifested by a reduction in functional loss compared to placebo. Although this reduction was significant, there remains a need for therapies for ALS, which experience even greater rates of functional loss. Summary of the Invention

[0010] A method is disclosed in which the synergistic combination of edaravone and deuterated arachidonic acid or a prodrug thereof significantly inhibits the progression of ALS. When used in this manner, patients treated with such a combination experience a substantially reduced rate of loss of function compared to either drug alone. In fact, patients treated with both drugs experience minimal loss of function during the treatment period.

[0011] Thus, in one embodiment, a method of reducing the rate of disease progression of ALS in a patient is provided, the method comprising administering to the patient an effective amount of edaravone or a pharma- ceutically acceptable salt thereof in combination with an effective amount of deuterated arachidonic acid or a prodrug thereof, such that when so administered, the combination results in a reduction in the rate of loss of function that is greater than the additive reduction achieved by each drug alone.

[0012] In one embodiment, a method is provided for reducing the rate of functional loss in an ALS patient being treated with an effective amount of deuterated arachidonic acid or a prodrug thereof, the method further comprising administering to said patient an effective amount of edaravone or a pharma- ceutically acceptable salt thereof, such that when so administered, the combination provides a reduction in the rate of functional loss that is greater than the additive reduction achieved by each drug alone.

[0013] In one embodiment, a method is provided for reducing the rate of functional loss in an ALS patient being treated with an effective amount of edaravone or a pharma- ceutically acceptable salt thereof, the method further comprising administering to the patient an effective amount of deuterated arachidonic acid or a prodrug thereof, When so administered, the combination provides a reduction in the rate of functional loss that is greater than the additive reduction achieved by each drug alone.

[0014] In one embodiment, edaravone or a pharma- ceutically acceptable salt thereof is administered according to the Radicava® (edaravone) product insert, which is incorporated herein by reference in its entirety.

[0015] In one embodiment, deuterated arachidonic acid or a prodrug thereof is administered using a dosing regimen including a loading dose and a maintenance dose as described herein.

[0016] In one embodiment, the prodrug of deuterated arachidonic acid is 11,11-D2-linoleic acid or its ester. In vivo, a portion of this prodrug is converted to the corresponding 13,13-D2-arachidonic acid, which is the active agent in motor neurons. In general, it takes several weeks to several months to achieve therapeutic concentrations of 13,13-D2-arachidonic acid in motor neurons using this prodrug. However, the use of a loading dose followed by a maintenance dose increases the uptake of 13,13-D2-arachidonic acid (D2-AA) into motor neurons, providing sustained protection against LPO in a shorter time after treatment initiation.

[0017] In one embodiment, the deuterated arachidonic acid prodrug is an ester of 7,7,10,10,13,13-D6-arachidonic acid, which is converted in vivo to 7,7,10,10,13,13-D6-arachidonic acid (D6-AA). Administration of this compound avoids the need for in vivo conversion as required by 11,11-D2-linoleic acid, providing more rapid uptake into motor neurons and associated increased protection against LPO.

[0018] In one embodiment, when the prodrug is 11,11-D2-linoleic acid ethyl ester, a dose of between about 5.5 grams per day and about 12 grams per day, or between about 7 and about 12 grams per day, e.g., about 9 grams / day (e.g., 8.64 grams / day) is administered daily during a loading period, followed by a dose of between 3 and 8 grams per day, e.g., about 5 grams / day (e.g., 4.80 grams / day) or about 6 grams / day (5.76 grams / day) administered daily during a maintenance period, where the maintenance dose is less than the loading dose.

[0019] In one embodiment, edaravone is provided in a formulation sold under the name Radicava® by Mitsubishi Tanabe Pharma, Osaka, Japan. In one embodiment, edaravone is provided in an orally deliverable form in an aqueous composition according to U.S. Pat. No. 1,098,7341, the entirety of which is incorporated herein by reference.

[0020] In one embodiment, edaravone is combined with a prodrug of deuterated arachidonic acid ethyl ester. Because 11,11-D2-linoleic acid ethyl ester is an oil at room temperature, edaravone is preferably added to the oil. The recommended dose of edaravone is 60mg per administration, therefore, in one embodiment, this dose is combined with a capsule of 11,11-D2-linoleic acid ethyl ester that the patient takes every day. In one embodiment, nine 1-gram tablets, containing a total of 8.64 grams of 11,11-D2-linoleic acid ethyl ester (D2-LA ethyl ester), are administered to the patient three times a day in three 3 pill increments.

[0021] In one embodiment, edaravone is added to the oil phase of D2-LA ethyl ester. For example, edaravone can be added to three tablets containing D2-LA ethyl ester, and these tablets can be appropriately labeled so that they are taken together. For example, the capsule containing edaravone can be colored differently from the capsule that does not contain edaravone. Alternatively, 60 mg of edaravone can be divided equally into all nine tablets (i.e., about 6.6 mg per tablet). In either case, the two drugs can be mixed to form a combination suitable for oral administration.

[0022] In one embodiment, the prodrug of deuterated arachidonic acid is 7,7,10,10,13,13-D6-arachidonic acid ethyl ester. This compound does not require in vivo conversion like D2-LA, and since D2-LA is only converted to D2-AA at about 10%, the amount of D6-AA ethyl ester required for treatment is about 10% of D2-LA. Furthermore, as shown in the examples, D6-AA has been shown to be about twice as active as D2-AA, although in an inflammation model. However, this is considered to be a good indicator for improving activity in this specification. Therefore, the amount of D6-AA required for treatment is about 0.05 to about 2 grams per day. In this case, one, two, or three capsules can be used to administer D6-AA. In one embodiment, this dose of D6-AA can be administered simultaneously by also including a 60 mg dose of edaravone in one, two, or three D6-AA capsules, or, when such capsules are used, by dividing the 60 mg dose of edaravone evenly among all capsules.

[0023] In one embodiment, a composition is provided comprising edaravone and deuterated arachidonic acid or a prodrug thereof, the composition being designed for oral, intravenous, or infusion administration.

[0024] In one embodiment, the deuterated arachidonic acid prodrug is 11,11-D2-linoleic acid or an ester thereof.

[0025] In one embodiment, the deuterated arachidonic acid prodrug is 7,7,10,10,13,13-D6-arachidonic acid ester.

[0026] In one embodiment, the ester is an ethyl ester.

[0027] In one embodiment, the disclosure provides a kit of parts comprising at least a daily dose of 2,4-dihydro-3H-pyrazol-3-one or a pharma- ceutically acceptable salt thereof and at least a daily dose of deuterated arachidonic acid or a prodrug thereof.

[0028] In one embodiment, the 7,7,10,10,13,13-D6-arachidonic acid or ester thereof is administered daily at a dose of about 0.05 to about 5 grams / day during a loading period, followed by daily administration at a dose of about 30% to about 70% of the loading dose during a maintenance period.

[0029] In one embodiment, the 7,7,10,10,13,13-D6-arachidonic acid or ester thereof is administered daily during a loading period at a dose of about 0.5 to about 5 grams per day.

[0030] In one embodiment, the 2,4-dihydro-3H-pyrazol-3-one or a pharma- ceutically acceptable salt thereof is administered as an infusion, an injectable composition, or an oral composition.

[0031] In one embodiment, the 2,4-dihydro-3H-pyrazol-3-one or a pharma- ceutically acceptable salt thereof is administered as an injectable or oral composition.

[0032] In one embodiment, the 2,4-dihydro-3H-pyrazol-3-one or a pharma- ceutically acceptable salt thereof is administered as an oral composition in combination with deuterated arachidonic acid or a prodrug thereof.

[0033] In one embodiment, the methods described herein further comprise limiting the patient's excess dietary polyunsaturated fatty acid intake during administration of said deuterated arachidonic acid or a prodrug thereof. [Brief description of the drawings]

[0034] The novel features of the present disclosure are set forth with particularity in the appended claims. A better understanding of the features and advantages of the present disclosure will be obtained by reference to the following detailed description that sets forth preferred embodiments, in which the principles of the disclosure are utilized, and the accompanying drawings, in which:

[0035] [Figure 1] Figure 1 compares the natural history of ALS with the corresponding loss of function to that of edaravone-treated patients after 6 months of treatment. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0036] In the following detailed description, reference is made to the accompanying drawings, which form a part hereof. In the drawings, like symbols typically refer to like elements, unless context dictates otherwise. The preferred embodiments described in the detailed description, drawings, and claims are not meant to be limiting. Other embodiments may be utilized, and other changes may be made, without departing from the scope of the subject matter presented herein. It will be readily understood that the aspects of the present disclosure, as generally described herein and illustrated in the drawings, can be arranged, substituted, combined, separated, and designed in a wide variety of different configurations, all of which are expressly contemplated herein.

[0037] Although specific embodiments and examples are disclosed below, the subject matter of the invention extends beyond the specifically disclosed embodiments to other alternative embodiments and / or uses, as well as modifications and equivalents thereof. Thus, the claims appended hereto are not limited by any of the specific embodiments described below. For example, in any method or process disclosed herein, the acts or operations of the method or process may be performed in any suitable order, and are not necessarily limited to the particular order disclosed. Although various operations may be described in sequence as multiple separate operations in a manner that will be helpful in understanding certain embodiments, the order of description should not be interpreted to imply that these operations are order-dependent. Furthermore, the structures, systems, and / or devices described herein may be embodied as integrated or separate components.

[0038] For purposes of comparing various embodiments, certain aspects and advantages of these embodiments are described. Not all such aspects or advantages are achieved by any particular embodiment. Thus, for example, various embodiments may be implemented in a manner that achieves or optimizes one or a group of advantages taught herein without necessarily achieving other aspects or advantages that may be taught or suggested herein.

[0039] A method for inhibiting the progression of amyotrophic lateral sclerosis (ALS) in humans is disclosed that utilizes a synergistic combination of 2,4-dihydro-3H-pyrazol-3-one (marketed under the trademark Radicava® and sold for the treatment of ALS) and a deuterated prodrug of arachidonic acid.

[0040] Before discussing the invention in more detail, the following terms will first be defined. Terms not defined will be given a definition in the context or a medically accepted definition.

[0041] The terms used in this specification are used only for the purpose of describing particular embodiments, and are not intended to limit the scope of the present invention. In this specification, the singular forms "a", "an", and "the" are intended to include the plural forms unless otherwise clear from the context.

[0042] As used herein, the terms "optional" or "optionally" mean that the subsequently described event or circumstance may or may not occur, and that both the occurrence and non-occurrence of the event or circumstance are included in the description.

[0043] As used herein, the term "about" when used before numerical designations such as temperature, time, amounts, and concentrations (including ranges thereof) indicates approximations that may vary by (+) or (-) 15%, 10%, 5%, or 1%, or any subrange or subvalue therebetween. Preferably, the term "about" when referring to a quantity or other characteristic such as dosage, means that the quantity may vary by ±10%.

[0044] As used herein, the term "comprising" or "comprises" is intended to mean that the compositions and methods include the recited elements, but do not exclude other elements.

[0045] As used herein, the term "consisting essentially of," when used to define compositions and methods, is meant to exclude any other elements that are essential to the combination for the purpose described. Thus, a composition consisting essentially of the elements defined herein does not exclude other materials or steps that do not materially affect the basic and novel characteristics of the claimed invention.

[0046] As used herein, the term "consisting of" means excluding other ingredients and substantial method steps that are more than trace elements. Embodiments defined by each of these transition terms are within the scope of this invention.

[0047] The term "edaravone" as used herein refers to 3-methyl-1-phenyl-2-pyrazolin-5-one, which can be represented by the following formula, or a salt thereof. [ka]

[0048] The term "deuterated arachidonic acid" as used herein refers to arachidonic acid, in which at least both hydrogen atoms at position 13 are replaced with two deuterium atoms, and its pharma- ceutically acceptable salts, unless the context requires otherwise. Such deuterated arachidonic acids include, but are not limited to, 13,13-D2-arachidonic acid, 10,10-13,13-D4-arachidonic acid, and 7,7,10,10,13,13-D6-arachidonic acid, and salts thereof. With respect to 7,7,10,10,13,13-D6-arachidonic acid, such compounds may further have moderate deuteration at the mono-allylic sites (i.e., positions 4 and 16) resulting from catalytic synthesis of the deuterated compound, as described in U.S. Pat. No. 1,073,0821, which is incorporated herein by reference in its entirety. Generally, up to about 40% of the total number of hydrogen atoms at these mono-allylic positions are replaced with deuterium, with deuterium typically being more prevalent at the 16 position than at the 4 position.

[0049] The term "prodrug" as used herein in connection with deuterated arachidonic acid ("drug") refers to a compound that provides the drug in vivo, unless the context requires otherwise. In one embodiment, the prodrug is an ester of deuterated arachidonic acid. Such deuterated arachidonic acids include 13,13-D2-arachidonic acid, 10,10-13,13-D4-arachidonic acid, and 7,7,10,10,13,13-D6-arachidonic acid.

[0050] In one embodiment, the prodrug may comprise 11,11-D2-linoleic acid or an ester thereof. In vivo, the ester group is removed to provide 11,11-D2-linoleic acid, and a portion of the compound is enzymatically converted to 13,13-D2-arachidonic acid. The unconverted portion of the compound is absorbed by the body and subsequently utilized as an energy source.

[0051] In one embodiment, the prodrug may comprise 8,8,11,11-D4-linoleic acid or an ester thereof. In vivo, the ester group is removed to provide 8,8,11,11-D4-linoleic acid, and a portion of the compound is enzymatically converted to 10,10,13,13-D4-arachidonic acid. The unconverted portion of the compound is absorbed into the body and subsequently utilized as an energy source.

[0052] In one embodiment, the prodrug is an ester of deuterated arachidonic acid. Such esters include deuterated arachidonic acid, such as esters of 13,13-D2-arachidonic acid, 10,10-13,13-D4-arachidonic acid, and 7,7,10,10,13,13-D6-arachidonic acid. With respect to 7,7,10,10,13,13-D6-arachidonic acid, such compounds may further have moderate deuteration at mono-allylic sites (i.e., positions 4 and 16) resulting from catalytic synthesis of this compound, as described above.

[0053] As used herein, the term "linoleic acid" refers to the compound having the formula provided below and having deuterium at natural abundance at each hydrogen atom (i.e., about 0.0156% natural deuterium), and pharma- ceutically acceptable salts thereof. [ka]

[0054] Esters of linoleic or arachidonic acid are formed by replacing the -OH group with an -OR group by methods well known in the art, such esters being as defined herein below.

[0055] As used herein, unless the context requires otherwise, the term "deuterated linoleic acid or esters thereof" refers to 11,11-D2-linoleic acid or C1-6 alkyl esters, glycerol esters (including monoglycerides, diglycerides, and triglycerides), sucrose esters, phosphate esters (e.g., phospholipids), and the like. The particular ester group used is not critical, so long as the ester group is pharma- ceutically acceptable (non-toxic and biocompatible).

[0056] As used herein, unless the context requires otherwise, the term "deuterated arachidonic acid ester" refers to such deuterated arachidonic acid having alkyl esters of 1 to 6 carbon atoms, glycerol esters (including monoglycerides, diglycerides, and triglycerides), sucrose esters, phosphate esters (e.g., phospholipids), and the like. The particular ester group used is not critical, so long as the ester group is pharma- ceutically acceptable (non-toxic and biocompatible).

[0057] As used herein, the term "phospholipid" refers to any phospholipid that is a component of cell membranes. This term includes phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, and sphingomyelin. In motor neurons, cell membranes are rich in phospholipids that contain arachidonic acid.

[0058] As used herein, the term "disease pathology" refers to the causes, onset, structural / functional changes, and natural history associated with the disease. The term "natural history" refers to the progression of the disease in the absence of treatment with the methods described herein.

[0059] As used herein, the term "reduction in disease progression rate" refers to a reduction in the rate of disease progression after initiation of treatment compared to the patient's natural history. In one example, the rate of reduction in disease progression using the methods described herein is a reduction of at least 35% or at least 50% at a time point after initiation of treatment, such as 3 months to 24 months, such as 6 months or 1 year, compared to the patient's natural history. In ALS, disease progression rate is assessed using serial measurements of the Revised ALS Functional Rating Scale (ALSFRS-R) at www.mdcalc.com / revised-amyotrophic-lateral-sclerosis-functional-rating-scale-alsfrs-r, which is incorporated herein by reference in its entirety. This rating scale assesses 12 different components on a scale of 0 (worse) to 4 (best), including speech, salivation, swallowing, writing, walking, food handling, dress and hygiene, turning in bed, walking, climbing stairs, dyspnea, orthopnea, and respiratory failure.

[0060] A reduction in the rate of disease progression is confirmed by a decrease in the slope of decline (flattening of the curve) in the ALSFRS-R of the patient's relative functionality during treatment when compared to the slope of decline seen during the patient's natural history. Typically, the difference between the slope of decline measured before treatment and the slope measured at least 90 days after the start of treatment has a flattening level of at least about 30%, which corresponds to a change of 7.5 degrees (e.g., a decrease from a 25 degree slope of decline during the natural history to a 17.5 degree slope of decline, which represents a 40% decrease in slope). In any case, a decrease in the slope of decline indicates that the treatment is reducing the rate of disease progression in the patient.

[0061] As used herein, the term "patient" refers to a human patient, or a cohort of human patients, afflicted with a neurodegenerative disease treatable by administration of deuterated arachidonic acid or a prodrug thereof.

[0062] As used herein, the term "loading or priming dose" refers to an amount of deuterated linoleic acid or an ester thereof sufficient to result in a reduction in the rate of disease progression within at least about 45 days, preferably within about 30 days, after administration begins.

[0063] In one embodiment, 11,11-D2-linoleic acid or its ester has been found to have a wide therapeutic window and good patient tolerance.When a loading dose is used, the steady-state concentration of 13,13-D2-arachidonic acid in red blood cells reaches at least about 12%, preferably at least about 15%, more preferably at least about 20%, based on the total amount of arachidonic acid in red blood cells, including deuterated arachidonic acid, more quickly.See allowed U.S. patent application Ser. No. 17 / 169,271, which is incorporated herein by reference in its entirety.Other deuterated arachidonic acid prodrugs are also well tolerated in vivo.

[0064] As used herein, the term "maintenance dose" refers to a dose of 11,11-D2-linoleic acid less than the primer dose, which is sufficient to maintain therapeutic concentrations of 13,13-D2-arachidonic acid in the cell membranes of red blood cells and, therefore, motor neurons, and to maintain a stable rate of disease progression.

[0065] As used herein, the term "regular administration" refers to a dosing regimen that is substantially compatible with the administration described herein. In other words, regular administration includes patients who are at least 75% compliant, preferably at least 80% compliant, over a 30-day period. In some embodiments, the dosing regimen includes scheduled dosing breaks. For example, a dosing regimen that provides dosing 6 days per week is one form of regular dosing. In another example, a patient can take a break from dosing for about 3 or 7 days or more, such as for personal reasons, provided that the patient is at least 75% compliant.

[0066] The term "cohort" refers to a group of at least two patients for which results are averaged.

[0067] As used herein, the term "pharmaceutically acceptable salt" of a compound disclosed herein includes acid or base addition salts that retain the desired pharmacological activity and are not biologically undesirable (e.g., the salts are not overly toxic, allergenic, or irritating and are bioavailable) within the scope of the methods described herein. When a compound has a basic group, such as an amino group, a pharmaceutically acceptable salt can be formed with inorganic acids (such as hydrochloric acid, hydroboric acid, nitric acid, sulfuric acid, and phosphoric acid), organic acids (such as alginic acid, formic acid, acetic acid, benzoic acid, gluconic acid, fumaric acid, oxalic acid, tartaric acid, lactic acid, maleic acid, citric acid, succinic acid, malic acid, methanesulfonic acid, benzenesulfonic acid, naphthalenesulfonic acid, and p-toluenesulfonic acid), or acidic amino acids (such as aspartic acid and glutamic acid). When a compound has an acidic group, such as a carboxylic acid group, it can be formed with metals, such as alkali metals and alkaline earth metals (Na + , Li + , K + , Ca 2+ , Mg 2+ , Zn 2+ etc.); ammonia or organic amines (dicyclohexylamine, trimethylamine, trimethylamine, pyridine, picoline, ethanolamine, diethanolamine, triethanolamine, etc.); or basic amino acids (arginine, lysine, ornithine, etc.); such salts can be prepared in situ during the isolation and purification of the compounds, or by reacting the purified compounds, in their free base or acid form, respectively, with a suitable acid or base and isolating the salt thereby formed.

[0068] "Excessive PUFAs", "excessive PUFA intake", and the like, refer to a total PUFA intake (e.g., total PUFA intake per day) that reduces the conversion of 11,11-D2-linoleic acid to 13,13-D2-arachidonic acid compared to a diet with a lower total PUFA intake. In some embodiments, the patient is on a diet that restricts the intake of linoleic acid, arachidonic acid, and / or other PUFA compounds. The amount of PUFAs that a patient can consume depends on many factors, such as the patient's health, weight, age, other medications being taken, liver function, metabolism, and the like.

[0069] Typically, patients on a 2000 calorie diet consume up to about 22 grams of polyunsaturated fatty acids per day (news.christianacare.org / 2013 / 04 / nutrition-numbers-revealed-fat-intake / ), of which about 14 grams are linoleic acid for both men and women (www.ncbi.nlm.nih.gov / pmc / articles / PMC3650500 / ). Furthermore, only about 10% of the average intake of linoleic acid is converted to arachidonic acid in the liver. That means that on average, about 1.4 grams of arachidonic acid are produced per day. When patients consume too much PUFA, such as linoleic acid, the excess dilutes the available concentration of 11,11-D2-linoleic acid. This, in turn, affects the amount of 13,13-D2-arachidonic acid produced in the liver, all other factors held constant.

[0070] A patient is considered to have an excess intake of linoleic acid if their total PUFA intake is such that the amount of 13,13-D2-arachidonic acid enzymatically produced per day is less than about 70% of the amount that would be produced with an average PUFA intake.

[0071] pathology The discovery that some aldehydes react readily with sulfhydryl groups and disrupt vital metabolic processes has implicated the peroxidation of polyunsaturated fatty acids as a component of the pathology of many neurodegenerative diseases (Schauenstein, E.; Esterbauer, H. Formation and properties of reactive aldehydes. Ciba Found. Symp. (67):225-244; 1978). Such lipid peroxidation, whether as the primary cause of the disease or as a secondary consequence, is due to oxidative stress, which leads to neuronal death and contributes to the progression of ALS.

[0072] The oxidative stress that causes such peroxidation is due to the daily production of reactive oxygen species ("ROS") and an imbalance in detoxification, which leads to oxidative attack on the lipid membranes of cells. The lipid membranes of motor neurons as well as the endoplasmic reticulum and mitochondria are highly enriched in arachidonic acid, a 20-carbon polyunsaturated fatty acid ("PUFA") with four cis-unsaturations. Each of these four sites is separated by three bis-arylmethylene groups. These groups are particularly susceptible to oxidative damage by ROS, as well as to the effects of enzymes such as cyclooxygenase, cytochrome, and lipoxygenase, compared to arylmethylene and methylene groups.

[0073] Furthermore, once the bis-arylmethylene group in one arachidonic acid is oxidized by ROS, a cascade of further oxidation of other arachidonic acid groups in lipid membranes can occur because a single ROS can cause the oxidation of one arachidonic acid moiety by a free radical mechanism, which can then oxidize an adjacent arachidonic acid by the same free radical mechanism, which can then oxidize yet another adjacent arachidonic acid, a process called lipid chain autoxidation. The resulting damage leads to a high content of oxidized arachidonic acid moieties in cell membranes.

[0074] Oxidized arachidonic acid adversely affects the fluidity and permeability of the cell membrane of motor neurons. Furthermore, it can cause the oxidation of membrane proteins and can be converted into a number of highly reactive carbonyl compounds. The latter include reactive species such as acrolein, malonic dialdehyde, glyoxal, and methylglyoxal (Negre-Salvayre A, et al. Brit. J. Pharmacol. 2008; 153:6-20). However, the most prominent products of arachidonic acid oxidation are α,β-unsaturated aldehydes such as 4-hydroxynon-2-enal (4-HNE; generated from n-6 PUFAs such as LA or AA) and the corresponding ketoaldehydes (Esterfbauer H, et al. Free Rad. Biol. Med. 1991; 11:81-128). As mentioned above, these reactive carbonyls cross-link (biomolecules) through Michael addition or Schiff base formation pathways, perpetuating the underlying pathology of the disease.

[0075] Disease progression Once a patient is diagnosed with ALS, clinicians assess the rate of disease progression for that patient by assessing the patient's loss of function in the absence of treatment, as described herein. This rate is called the "natural history" of the disease, and is typically measured by standardized tests that measure the degree of functionality of the patient over a set period of time. For example, in the case of ALS, there is a standard test called the ALSFRS-R that measures the loss of physical functionality over time, which is used to measure the progression of the disease. The test has 12 items, each measured on a scale of 0 (worse) to 4 (best). The efficacy of a drug is evidenced by its ability to reduce the rate of disease progression. Even a small decrease in the rate of functional loss is considered meaningful.

[0076] Previously, treatment of ALS with deuterated 11,11-D2-linoleic acid or esters thereof (including in lipid bilayer form) has been shown to stabilize polyunsaturated fatty acids against ROS. Examples of such treatments are described in WO 2011 / 053870, WO 2012 / 148946, and WO 2020 / 102596, each of which is incorporated herein by reference in its entirety.

[0077] Each of these documents discloses that a portion of 11,11-D2-linoleic acid is converted in vivo to 13,13-D2-arachidonic acid, which is then taken up by motor neurons to stabilize these neurons from oxidative damage. This accumulation of 13,13-D2-arachidonic acid in the body occurs over a period of months until therapeutic concentrations are achieved. Once therapeutic concentrations of 13,13-D2-arachidonic acid are achieved, continued administration of 11,11-D2-linoleic acid or its esters is required to maintain such therapeutic concentrations.

[0078] Moreover, the dosing regimen employed must address the patient's need for rapid onset of therapeutic benefit, especially considering that loss of function typically occurs quite rapidly and frequently well before the end stage of the disease (typically in the range of about 2-5 years after diagnosis). Thus, any therapy for treating such neurodegenerative diseases must provide a significant therapeutic benefit within one month of initiating treatment, thereby preserving patient function as much as possible, and providing a substantial reduction in the rate of disease progression.

[0079] Preparation of compounds 11,11-D2-linoleic acid is known in the art and is commercially available.Furthermore, 11,11-D2-linoleic acid and its esters are described, for example, in US Patent No. 10052299 (incorporated herein in its entirety by reference).Deuterated arachidonic acid esters are known in the art, as provided in US Patent No. 10577304 and US Patent No. 10730821 (incorporated herein in its entirety by reference).

[0080] Methodology - 11,11-D2-Linoleic Acid or its Esters In one method described herein, a patient suffering from ALS is treated with 11,11-D2-linoleic acid or an ester thereof, which is a prodrug of 13,13-D2-arachidonic acid, which is converted to 13,13-D2-arachidonic acid in vivo after administration.

[0081] In one embodiment, 11,11-D2-linoleic acid or its ester is administered to a patient in an amount sufficient to produce a synergistic concentration in vivo when combined with edaravone. In one embodiment, such synergistic concentration is at least 3% as 13,13-D2-arachidonic acid in red blood cells or plasma based on the total concentration of arachidonic acid, including any deuterated arachidonic acid. In a preferred embodiment, a patient is administered sufficient 11,11-D2-linoleic acid or its ester to reach a steady-state concentration of 13,13-D2-arachidonic acid in red blood cells or plasma of at least about 12%, preferably at least about 15%, more preferably at least about 20%, based on the total amount of arachidonic acid, including deuterated arachidonic acid, found therein.

[0082] In one embodiment, the patient is administered a sufficient amount of deuterated LA or ester thereof such that the concentration of deuterated AA in red blood cells is at least about 3%, preferably at least 6%, more preferably at least 10%, and most preferably at least 15%, based on the total amount of arachidonic acid, including deuterated arachidonic acid, found therein. At any of these concentrations, the attending physician can correlate the concentration found to a therapeutic concentration of deuterated AA in neurons. The ratio of deuterated AA to the total amount of arachidonic acid, including deuterated AA, in red blood cells may be between about 0.5% and about 60%. In one embodiment, the ratio of deuterated AA to the total arachidonic acid in red blood cells may be between about 5% and about 50%, between about 10% and about 40%, between about 15% and about 30%, and between about 15% and about 25%. In some embodiments, the dose of deuterated LA is altered (e.g., increased) if the ratio of deuterated AA to the total amount of arachidonic acid in red blood cells, including deuterated AA, is less than a target amount, e.g., less than about 3%, less than about 6%, less than about 10%, less than about 15%, less than about 20%, less than about 30%, less than about 40%, less than about 50%, or less than about 60%.

[0083] In one embodiment, such administration involves the use of a dosing regimen comprising two dosing components: a first dosing component comprises a primer amount of 11,11-D2-linoleic acid or an ester thereof, and a second dosing component comprises a maintenance amount of 11,11-D2-linoleic acid or an ester thereof, the amount of 11,11-D2-linoleic acid or an ester thereof in the second dosing component being less than the amount in the first dosing component.

[0084] The amount of primer is designed to provide a rapid onset of therapeutic effect with the amount of 11,11-D2-linoleic acid or its ester used. Such therapeutic effect is measured by suppressing the disease progression of neurodegenerative disease, as described below. In one embodiment, the amount of primer takes into account various complex factors, such as the amount of PUFA that a patient ingests on a given day, the conversion rate of 11,11-D2-linoleic acid to 13,13-D2-arachidonic acid in vivo, and the general turnover rate (half-life) of lipids in the patient's neurons.

[0085] Regarding this last point, the lipid components of neurons are not static, but rather are replaced over time and have a finite half-life in the body. In general, only a small portion of the lipid components in cells are replaced each day. In the case of neurons, these cells are rich in arachidonic acid. The turnover of arachidonic acid in these membranes occurs from a stable pool of lipids containing arachidonic acid in the cerebrospinal fluid. This stable pool is then replaced and replenished over time by arachidonic acid in lipids newly ingested by the patient as part of the patient's diet, and by the biosynthesis of arachidonic acid from linoleic acid in the liver. In some embodiments, the maintenance dose of 11,11-D2-linoleic acid is set so that the conversion to 13,13-D2-arachidonic acid at least matches the rate of secretion from the body.

[0086] The rate of synthesis of arachidonic acid in the body is normally limited to the maximum amount of arachidonic acid that the liver can produce in a given day. Therefore, only a small portion of ingested linoleic acid is converted to arachidonic acid, while the majority of linoleic acid remains intact. As a result of this rate limitation, synthesis of arachidonic acid from linoleic acid is delayed after administration of deuterated linoleic acid, and erythrocyte 13,13-D2-arachidonic acid concentrations continue to increase after the dosing regimen is switched from the primer to the maintenance dose.

[0087] Thus, the selection of the dosage of 11,11-D2-linoleic acid or its esters described herein addresses each of the above factors and establishes a dosage level that allows the accumulation of sufficient 11,11-D2-linoleic acid in the body and thus the production of therapeutic levels of 13,13-D2-arachidonic acid in red blood cells. When this is achieved, the data in the Examples show that there is a significant reduction in the rate of disease progression.

[0088] In some embodiments, the loading dose of the dosing regimen described herein includes a sufficient amount of 11,11-D2-linoleic acid or its ester to maximize the conversion of 11,11-D2-linoleic acid to 13,13-D2-arachidonic acid in vivo when the 11,11-D2-linoleic acid or its ester is absorbed by the patient. Once the conversion is maximized, the generated 13,13-D2-arachidonic acid accumulates in the body until it reaches a therapeutic concentration in the patient. During this process, 13,13-D2-arachidonic acid is absorbed systemically into cells in the body, including neurons, at a rate based on the exchange rate or turnover rate of lipids in the cell membranes of these motor neurons.

[0089] In one embodiment, the loading dose comprises about 7 to about 12 grams of 11,11-D2-linoleic acid or esters thereof, preferably about 8.64 grams of 11,11-D2-linoleic acid ethyl ester per day (3 capsules administered three times a day), and the maintenance dose is less than the loading dose, for example about 5.84 grams of 11,11-D2-linoleic acid (3 capsules administered two times a day, or 2 capsules administered three times a day). In some embodiments, the daily or periodic administration of 11,11-D2-linoleic acid or esters thereof in a primer amount ranges from about 5.5 to about 12 grams per day, such as about 5.5 g, about 6 g, about 6.5 g, about 7 g, about 7.5 g, about 8 g, about 8.5 g, about 8.64 g, about 9 g, about 9.5 g, about 10 g, about 10.5 g, about 11 g, about 11.5 g, and about 12 g. In some embodiments, the daily or periodic administration of 11,11-D2-linoleic acid or esters thereof in a primer amount ranges from about 5.5 to about 12 grams per day, such as about 5.5 g, about 6 g, about 6.5 g, about 7 g, about 7.5 g, about 8 g, about 8.5 g, about 8.64 g, about 9 g, about 9.5 g, about 10 g, about 10.5 g, about 11 g, about 11.5 g, and about 12 g.

[0090] In one embodiment, and as shown in the examples, synergistic results using edaravone and 11,11-D2-linoleic acid or its esters can be achieved by using only about 1.5 grams to about 12 grams of 11,11-D2-linoleic acid or its esters. However, the goal of achieving rapid onset of therapeutic effect is best achieved by using a loading dose and a maintenance dose as described above.

[0091] In one embodiment, the primer dose or loading dose is continued for at least about 24 days, or at least about 45 days, so that, for example, therapeutic concentration of 11,11-D2-linoleic acid in vivo is quickly achieved, thereby reducing the rate of disease progression.In one embodiment, after the administration of the primer dose is completed, a maintenance dose is administered periodically.In one embodiment, about 70% or less of the primer dose of 11,11-D2-linoleic acid or its ester is administered per day.Based on the safety profile of D2-LA, the primer dose or loading dose can be used throughout the treatment period at the discretion of the attending physician.

[0092] Methodology - 7,7,10,10,13,13-D6-arachidonic acid or its esters As mentioned above, the deuterated arachidonic acid or ester thereof administered to the patient includes 7,7,10,10,13,13-D6-arachidonic acid or ester thereof ("D6-AA"), as that term is defined herein. The three bis-allylic carbon atoms on these compounds have substantially all of their hydrogens replaced with deuterium, thus providing these compounds with superior reduction to reduce or eliminate lipid autoxidation. Thus, these compounds significantly reduce the rate of progression of ALS. In a preferred embodiment, the D-6 arachidonic acid or ester thereof is delivered in a stepwise manner comprising a first and a second administration component. The primer amount, which is the first administration component, follows the above protocol, except that about 0.05 to about 5 grams, or about 0.5 to about 5 grams of D6-AA or ester thereof is used in the primer, daily or periodically. The maintenance amount used is less than the loading amount, and is generally between about 30% to about 70% of the loading amount.

[0093] In one embodiment, the primer dose or loading dose is continued for at least about 24 days, or at least about 45 days, so that, for example, therapeutic concentrations of 7,7,10,10,13,13-D6-arachidonic acid are rapidly achieved in vivo, thereby reducing the rate of disease progression. In one embodiment, after the administration of the primer dose is completed, a maintenance dose is administered periodically. In one embodiment, about 70% or less of the primer dose of 7,7,10,10,13,13-D6-arachidonic acid or its ester is administered per day. Based on the safety profile of D6-AA, the primer dose or loading dose can be used throughout the treatment period at the discretion of the attending physician.

[0094] In the methods described herein, D6-AA or an ester thereof is administered to a patient to accumulate therapeutic concentrations of D6-AA for use in the methods described herein.

[0095] In one embodiment, a sufficient amount of D6-AA or an ester thereof is administered to the patient such that the concentration of D6-AA in red blood cells or plasma is at least about 3%, preferably at least 6%, more preferably at least 10%, and most preferably at least 15%, based on the total amount of arachidonic acid, including deuterated arachidonic acid, found therein. At any of these concentrations, the attending physician can correlate the concentration found to a therapeutic concentration of D6-AA in neurons. The ratio of D6-AA to the total amount of arachidonic acid, including deuterated AA, in red blood cells or plasma may be between about 0.5% and about 60%. In one embodiment, the ratio of D6-AA to the total arachidonic acid in red blood cells or plasma may be between about 5% and about 50%, between about 10% and about 40%, between about 15% and about 30%, and between about 15% and about 25%. In some embodiments, the dose of D6-AA is altered (e.g., increased) if the ratio of D6-AA to the total amount of arachidonic acid in red blood cells, including deuterated AA, is less than a target amount, e.g., less than about 3%, less than about 6%, less than about 10%, less than about 15%, less than about 20%, less than about 30%, less than about 40%, less than about 50%, or less than about 60%.

[0096] In one embodiment, such administration involves the use of a dosing regimen comprising two dosing components: a first dosing component comprises a primer amount of D6-AA or an ester thereof, and a second dosing component comprises a maintenance amount of D6-AA or an ester thereof, the amount of D6-AA or an ester thereof in the second dosing component being less than the amount in the first dosing component.

[0097] The amount of primer is designed to provide a rapid onset of therapeutic effect with the amount of D6-AA or its ester used. Such therapeutic effect is measured by suppressing the disease progression of neurodegenerative disease, as described below. In one embodiment, the amount of primer takes into account various complex factors, such as the amount of PUFA that patient takes in a given day and the general turnover rate (half-life) of lipids in patient's neurons.

[0098] Regarding this last point, the lipid components of neurons are not static, but rather are replaced over time and have a finite half-life in the body. In general, lipid components in lipids are only partially replaced each day. In the case of neurons, these cells are rich in arachidonic acid. The turnover of arachidonic acid in these membranes occurs from a stable pool of lipids containing arachidonic acid in the cerebrospinal fluid. This stable pool is then replaced and replenished over time by arachidonic acid in lipids newly ingested by the patient as part of the patient's diet, and by the biosynthesis of arachidonic acid from linoleic acid in the liver. In some embodiments, the maintenance dose of D6-AA or its ester is set so that the amount of D6-AA ester administered matches the secretion rate of D6-AA from the body.

[0099] The methods described herein are based in part on the discovery that when neuronal lipid membranes are stabilized against LPO, the progression of neurodegenerative diseases is substantially inhibited. This is believed to be because hydrogen atoms in arachidonic acid are replaced with deuterium atoms, rendering deuterated arachidonic acid significantly more stable against ROS than arachidonic acid containing only hydrogen atoms. As described above, this stability manifests itself in a weakened lipid autoxidation cascade, thereby inhibiting the rate of disease progression.

[0100] In one embodiment, the methods described herein address this challenge by employing a dosing regimen that delivers sufficient amounts of D6-AA to provide therapeutic concentrations of deuterated arachidonic acid in neurons, which, when administered in appropriate amounts, reduces the level of LPO, thereby effectively inhibiting the progression of ALS.

[0101] In one of the methods described herein, a patient suffering from ALS is treated with D6-AA ester, which is a prodrug of D6-AA. Alternatively, D6-AA (including its pharmaceutically acceptable salt) can be directly administered to the patient as its active agent. When administered, D6-AA ester is converted to D6-AA. In either case, this D6-arachidonic acid is easily absorbed by the body.

[0102] Methodology - 3-methyl-1-phenyl-2-pyrazolin-5-one (edaravone) or its pharma- ceutically acceptable salts Edaravone is commercially available from Millipore Sigma, and the method of administration of the drug is described in the product insert (incorporated herein by reference). In one embodiment, edaravone is administered as an infusion solution or the like as described in the product insert for Radicava® available from Mitsubishi Tanabe Pharma, Osaka, Japan. In another embodiment, edaravone is administered intravenously as described in https: / / medlineplus.gov / druginfo / meds / a617027.html (incorporated herein by reference in its entirety). In another embodiment, oral doses of edaravone can be administered as described in U.S. Pat. No. 1,098,7341 (incorporated herein by reference in its entirety). In yet another embodiment, edaravone can be formulated into a lipid-based nanosystem as described in Parikh, et al., Drug Del., 24(1):962-978 (2017).

[0103] combination The method of the present invention uses a synergistic combination of edaravone and deuterated arachidonic acid or an ester thereof, or a prodrug thereof, to significantly reduce the rate of functional decline in patients suffering from ALS compared to the use of each drug alone.

[0104] Edaravone may be administered simultaneously with, before, or after deuterated arachidonic acid or a prodrug or ester thereof. The deuterated arachidonic acid or a prodrug or ester thereof may be administered in the same formulation as edaravone or in a different formulation.

[0105] For example, deuterated arachidonic acid or a prodrug or ester thereof may be administered periodically, such as daily or every other day. The current recommended dose of edaravone is 60 mg administered as an intravenous infusion (30 mg / 100 mL) over 60 minutes. Edaravone may be administered orally, for example, at between 50 mg / day and 150 mg / day, such as 60 mg / day or 105 mg / day. In some embodiments, edaravone is administered in an initial treatment cycle of 14 days of daily doses followed by a 14-day drug-free period. In some embodiments, edaravone is administered in subsequent treatment cycles as follows: 10 days of daily doses followed by a 14-day drug-free period during the 14-day period.

[0106] In one embodiment, a method is provided for reducing the rate of functional loss in an ALS patient being treated with edaravone, the method further comprising treating the patient with an effective amount of deuterated arachidonic acid or a prodrug thereof, such that when so administered, the combination provides a reduction in the rate of functional loss that is greater than the additive reduction achieved by each drug alone.

[0107] In one embodiment, the mode of administration of edaravone is not critical and any mode that provides the desired treatment can be used, including infusion, intravenous administration, oral delivery, transdermal delivery, pulmonary delivery, and the like.

[0108] In one embodiment, deuterated arachidonic acid or an ester thereof, or a prodrug thereof, is administered as D2-LA or an ester thereof, having a deuteration at position 13. In vivo, a portion of the D2-LA is bioconverted to D2-AA, thereby providing the patient with active deuterated D2-AA.

[0109] In one embodiment, D2-LA ethyl ester is administered in a stepped dosing regimen, including a loading dose followed by a maintenance dose. The loading dose includes about 7 to about 12 grams of orally delivered D2-LA per day. The maintenance dose is less than the loading dose and is also administered orally. Typically, the maintenance dose is about 65% or less of the loading dose, preferably about 30% to about 65% of the loading dose. The loading dose is continued for a period sufficient to achieve a steady state concentration of D2-AA in vivo, which is typically at least about 30 days, or 45 days, or about 60 days from the start of treatment, based on the attending physician's assessment. The specific timing of the transition from the loading dose to the maintenance dose is at the discretion of the attending physician. See, for example, the above-mentioned allowed US patent application Ser. No. 17 / 169271. Periodic analysis of D2-AA concentration in red blood cells provides a basis for evaluating whether the absorption of D2-LA and the subsequent biotransformation of D2-LA to D2-AA are proceeding appropriately. See, e.g., U.S. Provisional Application No. 63 / 177794, International Application No. PCT / US22 / 15368, and International Application No. PCT / US22 / 15366, each of which is incorporated by reference in its entirety into this specification.

[0110] In one embodiment, the amount of D2-LA or its ester administered to a patient can be reduced to reflect the synergistic effect achieved by the combination described herein.Even though not preferred, synergistic effect is observed when combined with edaravone at about 3 grams per day or 5 grams per day.

[0111] As for D6-AA, it can be administered as an ester prodrug or as D6-arachidonic acid or its pharma- ceutically acceptable salt. The drug is administered orally as described above. Again, periodic blood tests can be used to assess proper absorption of the drug. As described above, a graded dosing regimen is preferred. However, although not preferred, a consistent amount of drug or prodrug, such as 0.6 grams per day, can also be administered.

[0112] In one embodiment, a method is provided for reducing the rate of functional loss in a patient suffering from ALS and being treated with an effective amount of deuterated arachidonic acid or a prodrug thereof, comprising administering to said patient an effective amount of edaravone. When so administered, the combination provides a reduction in the rate of functional loss that is greater than the additive reduction achieved by each drug alone. The above administration methods can be used.

[0113] In one embodiment, edaravone and deuterated arachidonic acid or its prodrug can be incorporated into a formulation administered to a patient to allow the drug to be systemically incorporated.For example, D2-LA ethyl ester and D6-AA ethyl ester are both oils that can be used as carriers for edaravone.For example, edaravone can be added to any of these oils, and the combination can be encapsulated or administered as oil.In the latter case, the combination can further include sweeteners, stabilizers, colorants, and the like.For example, edaravone (as free base) is added to D2-LA or D2-AA in an effective amount.The combination is stirred as necessary to provide a homogeneous oil solution.If necessary, a small amount of co-solvent such as AA can be mixed in to ensure the homogeneity of the solution, especially for D2-AA.

[0114] Thus, in one embodiment, a composition suitable for administration to a patient is provided, the composition comprising D2-LA ester or D6-AA ester, and the composition further comprising an effective amount of edaravone, which eliminates the need for separate administration of the two drugs in a sequential manner.

[0115] Disease progression In ALS, inhibition of disease progression can be readily calculated by comparing the rate of functional decline after initiation of drug therapy with the rate of functional decline before drug therapy (the natural history of functional decline) using the known and established rate of functional decline measured by the ALS Functional Rating Scale-Revised (ALSFRS-R). Because the rate of functional decline is not perceptible on a daily basis, functional decline is typically measured monthly and assessed over a period of time, such as every 1-24 months, such as every 3 months, every 6 months, or every year.

[0116] As described in the examples below, the rate of functional decline is premised on measuring the progression of an individual or cohort with respect to the natural history of the disease before treatment. The individual or cohort average of functional decline is then determined at a time point such as 3 months, 6 months, or 12 months after the start of treatment. The rate of decline based on the natural history of the cohort is set as the denominator. The numerator is set to the delta between the natural disease progression rate and the rate of functional decline after a set treatment period according to the present invention. The resulting fraction is multiplied by 100 to determine the rate of change. Below is an example of this analysis:

[0117] Cohort A has a natural mean annualized rate of functional decline over a one year period of 28. Six months after initiating treatment with the present invention, Cohort A had a mean annualized rate of functional decline of 14. This provides a delta of 14 degrees; that is, taking 14 as the numerator and 28 as the denominator and multiplying the result by 100 gives a 50 percent annualized reduction.

[0118] Generally, the methods of the present invention provide a mean rate of change in functional decline for a cohort of at least 30%, more preferably at least about 35%, or at least about 40%, or at least about 45%, or at least about 50%, or at least about 55%, or at least about 60%. In some embodiments, the change in functional decline is measured over a period of, for example, 3 months to 24 months, such as 3 months, 6 months, or annually. The rate of functional decline can be measured over any intermediate period between 3 months and 1 year.

[0119] Parts Kits The method described herein can be carried out using a kit of parts.For example, edaravone, either as injection or infusion, is designed to be administered once a day for 10 days, and then not administered for 18 days.When edaravone is orally administered as gel capsules, the kit of parts includes a first set of edaravone capsules and a second set of deuterated arachidonic acid or its prodrug.For D2-LA ester, the second set includes multiple capsules for administration on each day.

[0120] In one embodiment, the kit of parts can include a first row containing capsules with a dose of 60 mg of edaravone in one capsule or 30 mg in two capsules. A second row can contain capsules with 1 / 9 of the daily dose of D2-LA ester in nine capsules. These capsules can be divided into three groups of three capsules, each group being separate from the next group, to clarify that the dose is taken three times a day, in the morning, midday, and evening. In one embodiment, the edaravone capsules are colored to distinguish them from the D2-LA ester capsules. For example, the edaravone capsules can be colored red and the D2-LA ester capsules can be colored green. The kit of parts can include only one day's dose per container, or multiple days' doses, or multiple days' doses per container. Each of these containers can be labeled to indicate that the dose contained therein is taken prior to any administration of D2-LA ester alone ("first container"). Preferably, in any 28-day administration period, the maximum number of administrations of the combination in one container is within 10 days.This prevents edaravone from being administered continuously for more than 10 days.It is understood that when using a maintenance dose of D2-LA ester, the number of capsules required for administration may or may not be reduced.If the number of capsules is not reduced, the size of the capsule may be reduced or pharma- ceutically acceptable excipients may be added to the capsule.

[0121] The kit of parts can include additional or additional containers ("second containers"), each containing a daily dose or multiple days' doses of D2-LA ester. The second containers contain the remaining doses for the 28-day administration period. Preferably, the second containers are marked to indicate that they should not be used until the doses in the first container have been taken. The doses of D2-LA ester in the second containers can be arranged and colored in the same way as in the first container.

[0122] In the case of D6-AA ester, the amount of drug required for treatment is intended to be less than that of D2-LA ester described above. Thus, in this embodiment, the kit of parts is substantially the same as above, except for the fact that there are fewer capsules for D6-AA ester than for D2-LA ester.

[0123] In one embodiment, the capsule may contain a combination of edaravone mixed or solubilized in deuterated arachidonic acid or its prodrug. In such a case, the kit of parts may contain a container of a first set of capsules, which includes a daily dose of edaravone in a capsule containing D2-LA ester ("first set of capsules"). In one embodiment, the first set of capsules contains a dose of edaravone in one capsule or divided into the number of capsules required by the D2-LA ester. For example, if nine separate capsules are required for a dose of D2-LA, the total dose of edaravone may be divided into one capsule, three capsules, six capsules, or nine capsules. Any subset of capsules containing edaravone should be appropriately colored to distinguish them from capsules that do not. As mentioned above, the maximum number of doses in such a container is 10 doses to meet the current dosage requirements for edaravone. Alternatively, the first set of capsules for each day of the 10 day period can be provided in a single day container, a multi-day container (e.g., 5 days), or in one container for all 10 days. When a maintenance dose of D2-LA ester is used, the number of capsules is understood to be .

[0124] The kit of parts can include an additional or additional container for capsules to be used from day 11 to day 28 of the treatment regimen ("second set of capsules"). In one embodiment, the daily dose of the second set of capsules can be included in a single-day container or a multi-day container. Preferably, the container for the second set of capsules is labeled to indicate that the container is not to be used until after the first set of capsules is taken. The doses of D2-LA ester in these containers can be arranged in the same format (3 x 3) as described above. As above, if a maintenance dose is used, the number of capsules can be reduced, or the activity per capsule can be reduced to maintain the same number of capsules.

[0125] In the case of D6-AA ester, the amount of drug required for treatment is intended to be less than that of D2-LA ester described above. Thus, in this embodiment, the kit of parts is substantially the same as above, except for the fact that there are fewer capsules for D6-AA ester than for D2-LA ester.

[0126] When edaravone is administered as an injectable composition, the kit of parts can include both the daily dose of D2-LA ester or D6-AA ester described above, and a syringe containing an injectable edaravone composition or multiple daily doses. In one embodiment, the kit of parts can include a primary container that contains multiple syringes, up to 10, and a corresponding number of daily doses of D2-LA ester or D6-AA ester. As described above, the primary container can limit the number of syringes to 10, and place capsules containing either ester as described above. The secondary container can include the remaining 18 days of capsules containing either ester to ensure a full 28-day pharmaceutical administration regimen.

[0127] As will be appreciated, the details of the containers in each embodiment of the kit of parts can be varied, so long as a specific dose of edaravone sufficient to complete the dosing regimen for 10 days at 60 mg per day and either a loading or maintenance dose of D2-LA ester or D6-AA ester for the entire 28-day dosing regimen is provided. It is contemplated that sufficient containers can be provided to allow multiple 28-day dosing regimens to be shipped to a hospital, pharmacy, primary care physician, patient's caregiver, or patient.

[0128] The container used is not critical and may include, for example, a blister pack for capsules, a sealed box comprising paper, plastic, or metal, one or more bottles, and the like. The container will be labeled or contain appropriate labeling to ensure that a caregiver or patient is instructed on the proper administration of both medications. EXAMPLES

[0129] The methods and compositions described herein will be further understood by reference to the following examples. These examples are intended to be merely exemplary of the invention. These methods and compositions are not limited in scope by the preferred embodiments, which are intended to merely illustrate one aspect of the invention. Any functionally equivalent methods and compositions are considered equivalents. In addition to those described herein, various modifications of the invention will become apparent to those skilled in the art from the foregoing description and accompanying drawings. Such modifications are within the scope of the appended claims. In these examples, the following terms are used herein and have the following meanings. Unless defined, abbreviations have their conventional medical meaning. D2-AA = 13,13-D2-arachidonic acid D6-AA = as defined herein AA = arachidonic acid ALS = amyotrophic lateral sclerosis ALSFRS-R = ALS Functional Rating Scale-Revised D2-LA = 11,11-D2-linoleic acid LA = linoleic acid LPO = lipid peroxidation Radicava (registered trademark) = Edaravone infusion formulation sold by Mitsubishi Tanabe Pharma (Osaka, Japan) RBC = red blood cells

[0130] Example 1 – Demonstration of synergistic effects using combination therapy of D2-AA and Edaravone This example reports the results of an expanded access program in which 17 ALS patients were treated with D2-LA. Of these seven patients, three patients were treated with edaravone before and during treatment with D2-LA, and four patients were treated with D2-LA alone during the treatment protocol. Although the current dosing regimen is a loading dose of about 8.64 g / day of D2-LA for about one month, followed by a maintenance dose of about 4.80 g / day or about 5.76 g / day of D2-LA for the maintenance period, all patients in this study were treated with a daily dose of 2.88 g / day of LA for two weeks, followed by a dose of 5.76 g / day of D2-LA for the remainder of the study.

[0131] Before starting treatment, each patient's functionality was assessed using the ALSFRS-R scoring system and an initial score was recorded. At least 6 months after starting treatment, each patient's functionality was assessed again using the ALSFRS-R scoring system, and the results were then extrapolated to provide a common 6-month treatment window.

[0132] Patients were divided into two groups as follows: 1. Patients who received edaravone during treatment with D2-LA; 2. Patients who did not receive edaravone during treatment with D2-LA.

[0133] The ALSFRS-R scores immediately before the start of treatment were then compared between the two groups based on the adjusted results for the scores at 6 months of treatment. Table 1 below provides the mean ALSFRS-R scores after 6 months of treatment for patients treated with D2-AA alone (n=12) and for patients treated with D2-AA and Radicava® (n=5).

[0134] To evaluate the benefit associated with pretreatment with edaravone in the second group of patients, see Figure 1, which provides data from the manufacturer of edaravone (Tanabe Mitsubishi). According to the data, edaravone provides a benefit of 2.49 score units compared to the control group at 6 months. Since these patients were pretreated with edaravone before the start of treatment with D2-LA, the minimum value of the synergistic effect would be the result if it were assumed that pretreatment had not been reached 6 months before the start of D2-LA administration. In such an assumption, part of the benefit of edaravone treatment would have to be credited to the figures reported for the natural history. Nevertheless, even if we attribute the entire benefit of 2.49 score units provided by edaravone to the D2-LA treatment mentioned above, there is a benefit achieved by the combination of these two drugs, which still exceeds the additive effect by more than 2.5 score units. This difference is likely due to synergistic effects. Assuming that these patients had been pretreated with edaravone more than 6 months prior to treatment with D2-LA, any benefit achieved by edaravone would have been fully accounted for in the patients' natural history scores. Thus, the synergistic effect achieved by combining these drugs provided a 5-point advantage over the additive benefit of each drug alone.

[0135] Furthermore, because the administration of D2-LA in this study did not utilize the current administration regimen, it is possible that further improvements could be obtained that could lead to further synergistic effects.

[0136] Alternatively, due to the synergistic effects demonstrated above, the dose of D2-LA could be reduced from 8.64 grams per day to approximately 5.76 grams per day or less and still achieve significant synergistic effects. [Table 1]

[0137] Example 2 This example involves an assessment of the relative reduction of LPO in an inflammation model, and thereby the relative activity of D2-AA (derived by bioconversion of D2-LA) compared to D6-AA.

[0138] Specifically, it is well understood that inflammation plays an important role in many neurodegenerative diseases. In this example, administration of LPS is known to promote inflammation through various mechanisms, such as the secretion of inflammatory cytokines and eicosanoids, and the induction of ROS. In this example, LPS was used to confirm the degree of inflammation resulting from ROS-induced oxidation of H-AA, D2-AA (obtained by D2-LA administration), and D6-AA in mouse lungs. Specifically, four groups of mice were used. Group 1 was a control mouse treated with H-LA control mice. Group 2 mice were administered D2-LA for 6 weeks. It is understood that both H-LA and D2-LA are partially converted in vivo to provide AA and 13,13-D2-AA, respectively. Group 3 mice were administered H-AA for 6 weeks. Group 4 mice were administered D6-AA for 6 weeks.

[0139] All groups were then administered a single intranasal dose of LPS to induce acute lung inflammation. The degree of inflammatory response was based on the interalveolar septa distance, with larger septal distances indicating higher levels of inflammation. Animals were sacrificed and the interalveolar septal distances were measured. Table 2 provides the average magnitude of interalveolar septal spatial distances across all groups. [Table 2]

[0140] The above results show that mice treated with D2-AA (by administration of D2-LA prodrug) had a reduction in the spatial distance between alveolar septa by about 25% compared to mice treated with H-LA, whereas mice treated with D6-AA had a reduction in the spatial distance by about 60%, indicating that D6-AA is advantageous in treating inflammation.

[0141] The above data suggest that D6-AA provides at least a two-fold benefit over D2-AA in this model of inflammation, and we use these data to estimate the protection against LPO offered by D6-AA compared to D2-AA.

[0142] Using this data, we can predict that the amount of D6-AA needed to achieve similar results as D2-LA would be 1 / 10 of 8.64 grams / day (since bioconversion is no longer required) x 0.5 (since D6-AA is approximately twice as active as D2-AA), or 0.43 grams per day. Furthermore, considering that 5.76 grams of D2-LA produced enough D2-AA biologically to provide synergy with edaravone (5.76 grams per day for synergy / 8.64 grams per day for loading), we can estimate that D6-AA would require only 0.43 grams per day x (5.76 / 8.64) = 0.29 grams per day, or 0.05 grams per day to achieve synergy. ***

[0143] While preferred embodiments of the present disclosure have been shown and described herein, it will be apparent to those skilled in the art that such embodiments are provided by way of example only. The present disclosure is not intended to be limited by the specific examples provided herein. The present disclosure has been described with reference to the foregoing specification, but the descriptions and explanations of the embodiments herein are not intended to be construed in a limiting sense. Those skilled in the art will appreciate that numerous variations, changes, and substitutions are possible without departing from the present disclosure. Furthermore, it should be understood that all aspects of the present disclosure are not limited to the specific depictions, configurations, or relative proportions set forth herein, which depend upon a variety of conditions and variables. It should be understood that various alternatives to the embodiments of the disclosure described herein may also be employed in implementing the present disclosure. It is therefore intended that the present invention cover such alternatives, modifications, variations, or equivalents. It is intended that the following claims define the scope of the present disclosure, and that methods and structures within the scope of these claims, and their equivalents, are encompassed therein.

Claims

1. 1. A method of reducing the rate of disease progression of ALS in a patient, comprising administering to the patient a therapeutically effective amount of 2,4-dihydro-3H-pyrazol-3-one or a pharma- ceutically acceptable salt thereof in combination with a therapeutically effective amount of deuterated arachidonic acid or a prodrug thereof.

2. 1. A method for reducing the rate of functional loss in an ALS patient being treated with a therapeutically effective amount of deuterated arachidonic acid or a prodrug thereof, the method further comprising administering a therapeutically effective amount of 2,4-dihydro-3H-pyrazol-3-one or a pharma- ceutically acceptable salt thereof.

3. 1. A method for reducing the rate of functional loss in an ALS patient being treated with a therapeutically effective amount of 2,4-dihydro-3H-pyrazol-3-one or a pharma- ceutically acceptable salt thereof, further comprising administering a therapeutically effective amount of deuterated arachidonic acid or a prodrug thereof.

4. 4. The method of any one of claims 1, 2, or 3, wherein the rate of loss of function in the patient is less than the rate found in the aggregate achieved by either drug alone.

5. 5. The method of claim 4, wherein the deuterated arachidonic acid or a prodrug thereof is administered using a dosing regimen comprising a loading dose and a maintenance dose as described herein.

6. 2. The method of claim 1, wherein the deuterated arachidonic acid prodrug is 11,11-D2-linoleic acid or an ester thereof.

7. The method of claim 6, wherein the ester is an alkyl ester having 1 to 6 carbon atoms.

8. 2. The method of claim 1, wherein the deuterated arachidonic acid or a prodrug thereof is an ester of 7,7,10,10,13,13-D6-arachidonic acid.

9. The method of claim 8, wherein the ester is an alkyl ester having 1 to 6 carbon atoms.

10. 2. The method of claim 1, wherein the deuterated arachidonic acid is 7,7,10,10,13,13-D6-arachidonic acid or a pharma- ceutically acceptable salt thereof.

11. 7. The method of claim 6, wherein 11,11-D2-linoleic acid ethyl ester is administered daily at a dose of about 9 grams / day during a loading period of the dosing regimen, followed by daily administration at a dose of about 30% to about 65% of the loading dose during a maintenance period of the dosing regimen.

12. 12. The method of claim 11, wherein the 2,4-dihydro-3H-pyrazol-3-one or a pharma- ceutically acceptable salt thereof is administered as an infusion, an injectable composition, an oral composition, or in combination with the deuterated arachidonic acid or a prodrug thereof.

13. 13. The method of claim 12, wherein the deuterated arachidonic acid or a prodrug thereof is 7,7,10,10,13,13-D6-arachidonic acid, a pharma- ceutically acceptable salt thereof, or an ester thereof.

14. 14. The method of claim 13, wherein the deuterated arachidonic acid prodrug is 7,7,10,10,13,13-D6-arachidonic acid ethyl ester.

15. A composition suitable for systemic ingestion by a patient comprising a 2,4-dihydro-3H-pyrazol-3-one or a pharma- ceutically acceptable salt thereof and deuterated arachidonic acid or a prodrug thereof.

16. 16. The composition of claim 15, wherein the composition is suitable for oral, intravenous, or infusion administration.

17. 16. The composition of claim 15, wherein the deuterated arachidonic acid prodrug is 11,11-D2-linoleic acid or an ester thereof.

18. 17. The composition of claim 16, wherein the ester is an ethyl ester.

19. 16. The composition of claim 15, wherein the deuterated arachidonic acid prodrug is 7,7,10,10,13,13-D6-arachidonic acid ethyl ester.

20. 17. The composition of claim 16, wherein the ester is an ethyl ester.

21. A kit of parts comprising at least a daily dose of 2,4-dihydro-3H-pyrazol-3-one or a pharma- ceutically acceptable salt thereof, and at least a daily dose of deuterated arachidonic acid or a prodrug thereof.

22. 2. The method of claim 1, wherein 7,7,10,10,13,13-D6-arachidonic acid or an ester thereof is administered daily at a dose of about 0.05 to about 5 grams / day during a loading period, followed by daily administration at a dose of about 30% to about 70% of the loading dose during a maintenance period.

23. 23. The method of claim 22, wherein 7,7,10,10,13,13-D6-arachidonic acid or ester thereof is administered daily during the loading period at a dose of about 0.5 to about 5 grams per day.

24. 23. The method of claim 22, wherein the 2,4-dihydro-3H-pyrazol-3-one or a pharma- ceutically acceptable salt thereof is administered as an infusion, an injectable composition, or an oral composition.

25. 10. The method of claim 1, wherein the 2,4-dihydro-3H-pyrazol-3-one or a pharma- ceutically acceptable salt thereof is administered as an injectable or oral composition.

26. 10. The method of claim 1, wherein the 2,4-dihydro-3H-pyrazol-3-one or a pharma- ceutically acceptable salt thereof is administered in combination with deuterated arachidonic acid or a prodrug thereof as an oral composition.

27. 4. The method of claim 1, 2, or 3, further comprising restricting the patient's excess dietary intake of polyunsaturated fatty acids during administration of the deuterated arachidonic acid or a prodrug thereof.

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