WO2022169642A1

WO2022169642A1 - Okn-007 as a therapeutic agent for neurodegenerative diseases

Info

Publication number: WO2022169642A1
Application number: PCT/US2022/013793
Authority: WO
Inventors: Rheal TOWNER
Original assignee: Oklahoma Medical Research Foundation
Priority date: 2021-02-08
Filing date: 2022-01-26
Publication date: 2022-08-11
Also published as: WO2022169642A9

Abstract

The present invention includes compositions and methods for treating a neurodegenerative disease comprising: identifying that the patient is in need for treatment for the neurodegenerative disease having Aβ plaque formation; and providing an amount of 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4-disulfonyl α-phenyl tertiary butyl nitrone, OKN-007 or a pharmaceutically acceptable salt thereof, effective to prevent or reduce the neurodegenerative disease by preventing or decreasing Aβ plaques or Aβ plaque formation.

Description

OKN-007 AS A THERAPEUTIC AGENT FOR NEURODEGENERATIVE DISEASES

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to U.S. Provisional Application Serial No. 62/970,043, filed February 8, 2021, the entire contents of which is incorporated herein by reference.

TECHNICAL FIELD OF THE INVENTION

[0002] The present invention relates in general to the field of a therapeutic agent for the treatment of neurodegenerative diseases.

STATEMENT OF FEDERALLY FUNDED RESEARCH

[0003] Not Applicable.

BACKGROUND OF THE INVENTION

[0004] Without limiting the scope of the invention, its background is described in connection with neurodegenerative diseases.

[0005] One such treatment is taught in U.S. Patent No. 10,851,109, issued to de Roulet, et al., entitled “Compositions and methods of using the same for treatment of neurodegenerative and mitochondrial disease”, which is said to teach compounds and pharmaceutically acceptable salts thereof for the treatment and/or prevention of neurodegenerative and/or mitochondrial diseases, such as Parkinson's disease and Leigh's disease. The compounds and pharmaceutically acceptable salts thereof are of the class of nitrogenous bases, for example, pyrimidines, purines, pteridines, and pharmaceutically acceptable salts thereof.

[0006] However, a need remains for novel agents that can be used more broadly to treat neurodegenerative diseases.

SUMMARY OF THE INVENTION

[0007] In one embodiment, the present invention includes a method for treating a neurodegenerative disease in a patient that comprises administering to the patient a therapeutically effective amount of 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4-disulfonyl a- phenyl tertiary butyl nitrone (OKN-007) or a pharmaceutically acceptable salt thereof. [0008] In one embodiment, the present invention includes a method for treating a neurodegenerative disease in a patient with A [3 plaques comprising administering to the patient a therapeutically effective amount of 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4- disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof to increase the expression of transthyretin (TTR) and reduce or eliminate A0 plaques by forming A0/TTR complexes that reduce A[3 plaques or A[3 plaque formation. In one aspect, the neurodegenerative disease is selected from diseases with A [3 plaque formation selected from at least one of Alzheimer’s Disease, sepsis, alcoholism, or Parkinson’s disease. In another aspect, the therapeutically effective amount of 2,4-Disulfonyl-N-Tert- Butylnitrone, 2,4-disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof increases the formation of A0/TTR complexes. In another aspect, the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4-disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof is administered orally, intravenously, or intraperitoneally. In another aspect, the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4- disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof are from about 40 to 1,200 mg/kg body weight/day, 100 to 450 mg/kg body weight/day, 200 to 400 mg/kg body weight/day, 300 to 800 mg/kg body weight/day, 350 to 1,000 mg/kg body weight/day, or 400 to 1,100 mg/kg body weight/day. In another aspect, the 2,4- Disulfonyl-N-Tert-Butylnitrone, 2,4-disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof is at least one of: (1) administered at least one of continuously, intermittently, systemically, or locally, (2) administered one or more times a day for as long as the patient is in need of treatment for the neurodegenerative disease; (3) sequentially or concomitantly, with another pharmaceutical agent in a newly diagnosed neurodegenerative disease patient; (4) as a single agent or in combination with another pharmaceutical agent in a newly diagnosed neurodegenerative disease patient; (5) as a single agent or in combination with another pharmaceutical agent in a newly diagnosed neurodegenerative disease pediatric patient; or (6) a pediatric neurodegenerative disease patient. In another aspect, the neurodegenerative disease is not ALS. In another aspect, the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4-disulfonyl a-phenyl tertiary butyl nitrone, or a pharmaceutically acceptable salt thereof, is provided in a sustained-release formulation.

[0009] In another embodiment, the present invention includes a method of preventing or treating a patient with a neurodegenerative disease comprising: identifying that the patient is in need for treatment for the neurodegenerative disease having A0 plaque formation; and providing an amount of 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4- disulfonyl a-phenyl tertiary butyl nitrone, or a pharmaceutically acceptable salt thereof, effective to prevent or reduce the neurodegenerative disease by preventing or decreasing A0 plaques or A0 plaque formation. In one aspect, the neurodegenerative disease is selected from diseases with A0 plaque formation selected from at least one of Alzheimer’s Disease, sepsis, alcoholism, or Parkinson’s disease. In another aspect, the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4-disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof increases the formation of A0/TTR complexes. In another aspect, the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4-disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof is administered orally, intravenously, or intraperitoneally. In another aspect, the 2,4-Disulfonyl-N-Tert- Butylnitrone, 2,4-disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof are from about 40 to 1,200 mg/kg body weight/day, 100 to 450 mg/kg body weight/day, 200 to 400 mg/kg body weight/day, 300 to 800 mg/kg body weight/day, 350 to 1,000 mg/kg body weight/day, or 400 to 1,100 mg/kg body weight/day. In another aspect, the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4-disulfonyl a- phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof is at least one of: (1) administered at least one of continuously, intermittently, systemically, or locally, (2) administered one or more times a day for as long as the patient is in need of treatment for the neurodegenerative disease; (3) sequentially or concomitantly, with another pharmaceutical agent in a newly diagnosed neurodegenerative disease patient; (4) as a single agent or in combination with another pharmaceutical agent in a newly diagnosed neurodegenerative disease patient; (5) as a single agent or in combination with another pharmaceutical agent in a newly diagnosed neurodegenerative disease pediatric patient; or (6) a pediatric neurodegenerative disease patient. In another aspect, the 2,4-Disulfonyl- N-Tert-Butylnitrone, 2,4-disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof inhibits immune suppression against the neurodegenerative disease. In another aspect, the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4-disulfonyl a- phenyl tertiary butyl nitrone, or a pharmaceutically acceptable salt thereof, is provided in a sustained-release formulation. In another aspect, the 2,4-Disulfonyl-N-Tert- Butylnitrone, 2,4-disulfonyl a-phenyl tertiary butyl nitrone, or a pharmaceutically acceptable salt thereof, is formulated or administered together, separately or sequentially with a chemotherapy or an adjuvant therapy.

[0010] In another embodiment, the present invention includes a method for treating a patient with a neurodeg enerative disease comprising: determining if the patient has a neurodegenerative disease that is caused by A [3 plaques or A [3 plaque formation; and if the patient has the neurodegenerative disease administering a therapeutically effective amount of a therapeutically effective amount of 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4- disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof sufficient to overcome the neurodegenerative disease. In one aspect, the neurodegenerative disease is selected from diseases with A [3 plaque formation selected from at least one of Alzheimer’s Disease, sepsis, alcoholism, or Parkinson’s disease. In another aspect, the therapeutically effective amount of 2,4-Disulfonyl-N-Tert- Butylnitrone, 2,4-disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof increases the formation of A0-transthy retin (TTR) complexes. In another aspect, the therapeutically effective amount of 2,4-Disulfonyl-N-Tert- Butylnitrone, 2,4-disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof is administered orally, intravenously, or intraperitoneally. In another aspect, the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4-disulfonyl a-phenyl tertiary butyl nitrone, or a pharmaceutically acceptable salt thereof, is provided in a sustained- release formulation. In another aspect, the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4- disulfonyl a-phenyl tertiary butyl nitrone, or a pharmaceutically acceptable salt thereof, is provided at 40 to 1,200 mg/kg body weight/day, 100 to 450 mg/kg body weight/day, 200 to 400 mg/kg body weight/day, 300 to 800 mg/kg body weight/day, 350 to 1,000 mg/kg body weight/day, or 400 to 1,100 mg/kg body weight/day. In another aspect, the neurodegenerative disease is not ALS.

BRIEF DESCRIPTION OF THE DRAWINGS

[0011] For a more complete understanding of the features and advantages of the present invention, reference is now made to the detailed description of the invention along with the accompanying figure and in which:

[0012] Figure 1 shows Transthyretin (TTR) levels detected by immunofluorescence (IF) in mouse brains treated with either OKN-007 (150 mg/kg; intravenous (i.v.) for 1 hour) or saline. There was a significant increase in TTR levels in the OKN-007-treated mouse brains, compared to saline controls (*p<0.05). DETAILED DESCRIPTION OF THE INVENTION

[0013] While the making and using of various embodiments of the present invention are discussed in detail below, it should be appreciated that the present invention provides many applicable inventive concepts that can be embodied in a wide variety of specific contexts. The specific embodiments discussed herein are merely illustrative of specific ways to make and use the invention and do not delimit the scope of the invention.

[0014] To facilitate the understanding of this invention, a number of terms are defined below. Terms defined herein have meanings as commonly understood by a person of ordinary skill in the areas relevant to the present invention. Terms such as “a”, “an” and “the” are not intended to refer to only a singular entity, but include the general class of which a specific example may be used for illustration. The terminology herein is used to describe specific embodiments of the invention, but their usage does not limit the invention, except as outlined in the claims.

[0015] The present invention includes various methods of treating patients with compound 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4-disulfonyl a-phenyl tertiary butyl nitrone, also known as: OKN-007, NXY-059, 2,4-DSPBN, 2,4-disulfonyl phenyl tertbutyl nitrone, 2,4-ds-PBN and Cerovive. It has been found that OKN-007 is a small molecule that can easily cross the blood brain barrier and has been demonstrated to have no adverse effects on normal cells, unlike other agents that have systemic toxicity. OKN- 007 can be administered either orally in pill form (patient compliant), or intravenously.

[0016] Further, it was found that OKN-007 is a neuroprotective agent. There are currently no other compounds or agents that protect motor neurons. All other therapeutic approaches do not prevent late-stage disease progression.

[0017] The therapeutic methods of the present invention (which include prophylactic treatment) in general include administration of a therapeutically effective amount of the compositions described herein to a subject in need thereof, including a mammal, particularly a human. Such treatment will be suitably administered to subjects, particularly humans, suffering from, having, susceptible to, or at risk for a neurodegenerative disease, or having a symptom thereof. Determination of those subjects “at risk” can be made by any objective or subjective determination by a diagnostic test or opinion of a subject or health care provider (e.g., genetic test, enzyme or protein marker, marker (as defined herein), family history, and the like). [0018] As used herein, a “pharmaceutically acceptable” component is one that is suitable for use with humans and/or animals without undue adverse side effects (such as toxicity, irritation, and allergic response) commensurate with a reasonable benefit/risk ratio.

[0019] As used herein, the term “safe and effective amount” refers to the quantity of a component that is sufficient to yield a desired therapeutic response without undue adverse side effects (such as toxicity, irritation, or allergic response) commensurate with a reasonable benefit/risk ratio when used in the manner of this invention.

[0020] As used herein, the term “therapeutically effective amount” is meant an amount of a compound of the present invention effective to yield the desired therapeutic response. For example, an amount effective to delay, inhibit or reverse a neurogenerative disease. The specific safe and effective amount or therapeutically effective amount will vary with such factors as the particular condition being treated, the physical condition of the patient, the type of mammal being treated, the duration of the treatment, the nature of concurrent therapy (if any), and the specific formulations employed and the structure of the compounds or its derivatives.

[0021] As used herein, a “pharmaceutical salt” is salt for making an acid or base salts of the compound. Examples of pharmaceutically acceptable salts include, but are not limited to, mineral or organic acid salts of basic residues such as amines; alkali or organic salts of acidic residues such as phenols. Preferably the salts are made using an organic or inorganic acid. These preferred acid salts are chlorides, bromides, sulfates, nitrates, phosphates, sulfonates, formates, tartrates, maleates, malates, citrates, benzoates, salicylates, ascorbates, and the like. The preferred phenolate salts are the alkaline earth metal salts, sodium, potassium or lithium.

[0022] As used herein, a “pharmaceutical carrier” is a pharmaceutically acceptable solvent, suspending agent or vehicle, for delivering the OKN-007 compound to the animal or human. The carrier may be liquid or solid and is selected with the planned manner of administration in mind. Liposomes are also a pharmaceutical carrier.

[0023] OKN-007 is typically administered in admixture with suitable pharmaceutical salts, buffers, diluents, extenders, excipients and/or carriers (collectively referred to herein as a pharmaceutically acceptable carrier or carrier materials) selected based on the intended form of administration and as consistent with conventional pharmaceutical practices. Depending on the best location for administration, the OKN-007 may be formulated to provide, e.g., maximum and/or consistent dosing for the particular form for oral, rectal, topical, intravenous injection or parenteral administration. While the OKN- 007 may be administered alone, it will generally be provided in a stable salt form mixed with a pharmaceutically acceptable carrier. The carrier may be solid or liquid, depending on the type and/or location of administration selected.

[0024] Techniques and compositions for making useful dosage forms using the present invention are described in one or more of the following references: Anderson, Philip O.; Knoben, James E.; Troutman, William G, eds., HANDBOOK OF CLINICAL DRUG DATA, Tenth Edition, McGraw-Hill, 2002; Pratt and Taylor, eds., PRINCIPLES OF DRUG ACTION, Third Edition, Churchill Livingston, New York, 1990; Katzung, ed., BASIC AND CLINICAL PHARMACOLOGY, Ninth Edition, McGraw Hill, 2007; Goodman and Gilman, eds., THE PHARMACOLOGICAL BASIS OF THERAPEUTICS, Tenth Edition, McGraw Hill, 2001 ; REMINGTON’ S PHARMACEUTICAL SCIENCES, 20th Ed., Lippincott Williams & Wilkins., 2000, and updates thereto; Martindale, THE EXTRA PHARMACOPOEIA, Thirty-Second Edition (The Pharmaceutical Press, London, 1999); all of which are incorporated by reference, and the like, relevant portions incorporated herein by reference.

[0025] For example, the OKN-007 may be included in a tablet. Tablets may contain, e.g., suitable binders, lubricants, disintegrating agents, coloring agents, flavoring agents, flow-inducing agents and/or melting agents. For example, oral administration may be in a dosage unit form of a tablet, gel cap, caplet or capsule, the active drug component being combined with a non-toxic, pharmaceutically acceptable, inert carrier such as lactose, gelatin, agar, starch, sucrose, glucose, methyl cellulose, magnesium stearate, dicalcium phosphate, calcium sulfate, mannitol, sorbitol, mixtures thereof, and the like. Suitable binders for use with the present invention include: starch, gelatin, natural sugars (e.g., glucose or beta-lactose), corn sweeteners, natural and synthetic gums (e.g., acacia, tragacanth or sodium alginate), carboxymethylcellulose, polyethylene glycol, waxes, and the like. Lubricants for use with the invention may include: sodium oleate, sodium stearate, magnesium stearate, sodium benzoate, sodium acetate, sodium chloride, mixtures thereof, and the like. Disintegrators may include: starch, methyl cellulose, agar, bentonite, xanthan gum, mixtures thereof, and the like.

[0026] OKN-007 may be administered in the form of liposome delivery systems, e.g., small unilamellar vesicles, large unilamellar vesicles, and multilamellar vesicles, whether charged or uncharged. Liposomes may include one or more: phospholipids (e.g., cholesterol), stearylamine and/or phosphatidylcholines, mixtures thereof, and the like. [0027] OKN-007 may also be coupled to one or more soluble, biodegradable, bioacceptable polymers as drug carriers or as a prodrug. Such polymers may include: polyvinylpyrrolidone, pyran copolymer, polyhydroxylpropylmethacrylamide-phenol, polyhydroxyethylasparta-midephenol, or polyethyleneoxide-polylysine substituted with palmitoyl residues, mixtures thereof, and the like. Furthermore, the OKN-007 may be coupled to one or more biodegradable polymers to achieve controlled release of the OKN-007, biodegradable polymers for use with the present invention include: polylactic acid, polyglycolic acid, copolymers of polylactic and polyglycolic acid, polyepsilon caprolactone, polyhydroxy butyric acid, polyorthoesters, polyacetals, polydihydropyrans, polycyanoacylates, and crosslinked or amphipathic block copolymers of hydrogels, mixtures thereof, and the like.

[0028] In one embodiment, gelatin capsules (gelcaps) may include the OKN-007 and powdered carriers, such as lactose, starch, cellulose derivatives, magnesium stearate, stearic acid, and the like. Like diluents may be used to make compressed tablets. Both tablets and capsules may be manufactured as immediate-release, mixed-release or sustained-release formulations to provide for a range of release of medication over a period of minutes to hours. Compressed tablets may be sugar coated or film coated to mask any unpleasant taste and protect the tablet from the atmosphere. An enteric coating may be used to provide selective disintegration in, e.g., the gastrointestinal tract.

[0029] For oral administration of OKN-007 in a liquid dosage form, the oral drug components may be combined with any oral, non-toxic, pharmaceutically acceptable inert carrier such as ethanol, glycerol, water, and the like. Examples of suitable liquid dosage forms include solutions or suspensions in water, pharmaceutically acceptable fats and oils, alcohols or other organic solvents, including esters, emulsions, syrups or elixirs, suspensions, solutions and/or suspensions reconstituted from non-effervescent granules and effervescent preparations reconstituted from effervescent granules. Such liquid dosage forms may contain, for example, suitable solvents, preservatives, emulsifying agents, suspending agents, diluents, sweeteners, thickeners, and melting agents, mixtures thereof, and the like.

[0030] Liquid dosage forms for oral administration of the OKN-007 may also include coloring and flavoring agents that increase patient acceptance and therefore compliance with a dosing regimen. In general, water, a suitable oil, saline, aqueous dextrose (e.g., glucose, lactose and related sugar solutions) and glycols (e.g., propylene glycol or polyethylene glycols) may be used as suitable carriers for parenteral solutions. Solutions for parenteral administration include generally, a water-soluble salt of the active ingredient, suitable stabilizing agents, and if necessary, buffering salts. Antioxidizing agents such as sodium bisulfite, sodium sulfite and/or ascorbic acid, either alone or in combination, are suitable stabilizing agents. Citric acid and its salts and sodium EDTA may also be included to increase stability. In addition, parenteral solutions may include pharmaceutically acceptable preservatives, e.g., benzalkonium chloride, methyl- or propyl-paraben, and/or chlorobutanol. Suitable pharmaceutical carriers are described in REMINGTON'S PHARMACEUTICAL SCIENCES, Mack Publishing Company, a standard reference text in this field, relevant portions incorporated herein by reference.

[0031] Capsules. Capsules with OKN-007 may be prepared by filling standard two-piece hard gelatin capsules each with 10 to 500 milligrams of powdered active ingredient, 5 to 150 milligrams of lactose, 5 to 50 milligrams of cellulose and 6 milligrams magnesium stearate.

[0032] Soft Gelatin Capsules. A mixture of OKN-007 is dissolved in a digestible oil such as soybean oil, cottonseed oil or olive oil. The active ingredient is prepared and injected by using a positive displacement pump into gelatin to form soft gelatin capsules containing, e.g., 100-500 milligrams of the active ingredient. The capsules are washed and dried.

[0033] Tablets. A large number of tablets are prepared by conventional procedures so that the dosage unit of OKN-007, e.g., 1-500 milligrams of OKN-007, 0.2 milligrams of colloidal silicon dioxide, 5 milligrams of magnesium stearate, 50-275 milligrams of microcrystalline cellulose, 11 milligrams of starch and 98.8 milligrams of lactose. Appropriate coatings may be applied to increase palatability or delay absorption.

[0034] To provide an effervescent tablet appropriate amounts of, e.g., monosodium citrate and sodium bicarbonate, are blended together and then roller compacted, in the absence of water, to form flakes that are then crushed to give granulates. The granulates are then combined with the active ingredient, drug and/or salt thereof, conventional beading or filling agents and, optionally, sweeteners, flavors and lubricants.

[0035] Injectable solution. A parenteral composition suitable for administration by injection is prepared by stirring 1.5% by weight of active ingredient in deionized water and mixed with, e.g., up to 10% by volume propylene glycol and water. The solution is made isotonic with sodium chloride and sterilized using, e.g., ultrafiltration. [0036] Suspension. An aqueous suspension is prepared for oral administration so that each 5 ml contain 100 mg of finely divided active ingredient, 200 mg of sodium carboxymethyl cellulose, 5 mg of sodium benzoate, 1.0 g of sorbitol solution, U.S.P., and 0.025 ml of vanillin.

[0037] OKN-007 as a Therapeutic agent for Neurodegenerative Diseases. From rat brain microarray data comparing normal rats either untreated or treated with OKN-007 (via drinking water for 1 week; 18 mg/kg/day), it was established that transthyretin (TTR; transports thyroxine and retinol; originally called prealbumin) gene expression was altered. TTR, a transport protein in the serum and cerebrospinal fluid (CSF), is mainly synthesized by the liver and the choroid plexus of the brain (Vieira and Saraiva, 2014).

[0038] Synthesis of TTR by epithelial cells of the choroid plexus is the main source of CSF TTR (Vieira and Saraiva, 2014; Aleshire et al., 1983), but TTR mRNA has also been found in the cortex, hippocampus and cerebellum in murine or human brains (Vieira and Saraiva, 2014; Carro et al., 2002; Stein and Johnson, 2002; Buxbaum et al., 2008; Li etal., 2011).

[0039] It was previously found that production of A0 (P-amyloid), its precursor, or its related peptides induces neuronal TTR transcription and the synthesis and the presence of Ap/TTR complexes in vivo, suggesting that increased TTR production coupled with the interaction between TTR and Ap and/or its related peptides may play a role in natural resistance to human Alzheimer’s disease (AD) (Li et al., 2011).

[0040] Immunofluorescence (IF) data for TTR from mouse brains was obtained from mice that were either treated with OKN-007 or saline (i.v. for 1 hour; 150 mg/kg). OKN- 007 treatment was able to significantly increase TTR protein levels (see Fig. 1). By increasing TTR protein levels, OKN-007 acts as a therapeutic agent for the elimination of Ap. The reduction or elimination of Ap shows that OKN-007 can be used not only AD, but several neuroinflammatory diseases that form Ap, and then lead to neurodegeneration (e.g. endotoxemia from sepsis, alcoholism, Parkinson’s disease, etc.), by forming Ap/TTR complexes, and therefore preventing the formation of Ap plaques.

[0041] Figure 1 shows Transthyretin (TTR) levels detected by immunofluorescence (IF) in mouse brains treated with either OKN-007 (150 mg/kg; i.v. for 1 hour) or saline. There was a significant increase in TTR levels in the OKN-007-treated mouse brains, compared to saline controls (*p<0.05). Thus, the present invention can be used with immunohistochemistry (IHC) or IF to detect levels of TTR protein. [0042] Immunofluorescence and confocal microscopy. Immunofluorescence was conducted following the protocol described by Smith N, Saunders D, Lerner M, et al. In vivo and ex vivo assessment of bladder hyper-permeability and using molecular targeted magnetic resonance imaging to detect claudin-2 in a mouse model for interstitial cystitis. PLoS One 2020;15:e0239282, relevant portions incorporated herein by reference. Briefly, brain tissues were paraffin-embedded and sectioned at 6 pm. Following deparaffinization, antigen retrieval and blocking in 20% Aquablock buffer (Abeam, MA), sections were incubated with primary antibodies, rabbit anti-occludin (1 :200, Cat. #: 71-1500, RRID:AB_88065, Invitrogen, Carlsbad, CA) and mouse anti-transthyretin (1 :200, Cat. #: 856302, RRID: AB_2783455, Biolegend, San Diego, CA), overnight at 4°C. The secondary antibodies, goat anti-rabbit Alexa Fluor 594 and goat anti-mouse Alexa Fluor 488, were incubated for 1 hour at RT. Negative controls without primary antibodies were conducted following the same protocol. Fluorescent images were captured using Zeiss LSM 880 confocal microscope (Zeiss, Germany) with a 63X objective. 3 randomly selected regions with positive immunostaining were imaged for each animal. All the images were captured by using the same laser powers and offsets.

[0043] Image analysis. For transthyretin, image thresholds were automatically performed in Fiji/ImageJ. The percentage of transthyretin expression area was calculated by Analyze Particle in Fiji/ImagJ with default settings. For transthyretin, percentage of immunolabeled area were calculated and the raw data is averaged %area of each animal. The y-axis of the graph can also be described as a percentage of transthyretin expression area.

[0044] It is contemplated that any embodiment discussed in this specification can be implemented with respect to any method, kit, reagent, or composition of the invention, and vice versa. Furthermore, compositions of the invention can be used to achieve methods of the invention.

[0045] It will be understood that particular embodiments described herein are shown by way of illustration and not as limitations of the invention. The principal features of this invention can be employed in various embodiments without departing from the scope of the invention. Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, numerous equivalents to the specific procedures described herein. Such equivalents are considered to be within the scope of this invention and are covered by the claims. [0046] All publications and patent applications mentioned in the specification are indicative of the level of skill of those skilled in the art to which this invention pertains. All publications and patent applications are herein incorporated by reference to the same extent as if each individual publication or patent application was specifically and individually indicated to be incorporated by reference.

[0047] The use of the word “a” or “an” when used in conjunction with the term “comprising” in the claims and/or the specification may mean “one,” but it is also consistent with the meaning of “one or more,” “at least one,” and “one or more than one.” The use of the term “or” in the claims is used to mean “and/or” unless explicitly indicated to refer to alternatives only or the alternatives are mutually exclusive, although the disclosure supports a definition that refers to only alternatives and “and/or.” Throughout this application, the term “about” is used to indicate that a value includes the inherent variation of error for the device, the method being employed to determine the value, or the variation that exists among the study subjects.

[0048] As used in this specification and claim(s), the words “comprising” (and any form of comprising, such as “comprise” and “comprises”), “having” (and any form of having, such as “have” and “has”), “including” (and any form of including, such as “includes” and “include”) or “containing” (and any form of containing, such as “contains” and “contain”) are inclusive or open-ended and do not exclude additional, unrecited features, elements, components, groups, integers, and/or steps, but do not exclude the presence of other unstated features, elements, components, groups, integers and/or steps. In embodiments of any of the compositions and methods provided herein, “comprising” may be replaced with “consisting essentially of’ or “consisting of’. As used herein, the term “consisting” is used to indicate the presence of the recited integer (e.g., a feature, an element, a characteristic, a property, a method/process step or a limitation) or group of integers (e.g., feature(s), element(s), characteristic(s), property(ies), method/process steps or limitation(s)) only. As used herein, the phrase “consisting essentially of’ requires the specified features, elements, components, groups, integers, and/or steps, but do not exclude the presence of other unstated features, elements, components, groups, integers and/or steps as well as those that do not materially affect the basic and novel characteristic(s) and/or function of the claimed invention.

[0049] The term “or combinations thereof’ as used herein refers to all permutations and combinations of the listed items preceding the term. For example, “A, B, C, or combinations thereof’ is intended to include at least one of: A, B, C, AB, AC, BC, or ABC, and if order is important in a particular context, also BA, CA, CB, CBA, BCA, ACB, BAC, or CAB. Continuing with this example, expressly included are combinations that contain repeats of one or more item or term, such as BB, AAA, AB, BBC, AAABCCCC, CBBAAA, CABABB, and so forth. The skilled artisan will understand that typically there is no limit on the number of items or terms in any combination, unless otherwise apparent from the context.

[0050] As used herein, words of approximation such as, without limitation, “about”, “substantial” or “substantially” refers to a condition that when so modified is understood to not necessarily be absolute or perfect but would be considered close enough to those of ordinary skill in the art to warrant designating the condition as being present. The extent to which the description may vary will depend on how great a change can be instituted and still have one of ordinary skill in the art recognize the modified feature as still having the required characteristics and capabilities of the unmodified feature. In general, but subject to the preceding discussion, a numerical value herein that is modified by a word of approximation such as “about” may vary from the stated value by at least ±1, 2, 3, 4, 5, 6, 7, 10, 12 or 15%.

[0051] All of the compositions and/or methods disclosed and claimed herein can be made and executed without undue experimentation in light of the present disclosure. While the compositions and methods of this invention have been described in terms of preferred embodiments, it will be apparent to those of skill in the art that variations may be applied to the compositions and/or methods and in the steps or in the sequence of steps of the method described herein without departing from the concept, spirit and scope of the invention. All such similar substitutes and modifications apparent to those skilled in the art are deemed to be within the spirit, scope and concept of the invention as defined by the appended claims.

[0052] To aid the Patent Office, and any readers of any patent issued on this application in interpreting the claims appended hereto, applicants wish to note that they do not intend any of the appended claims to invoke paragraph 6 of 35 U.S.C. § 112, U.S.C. § 112 paragraph (f), or equivalent, as it exists on the date of filing hereof unless the words “means for” or “step for” are explicitly used in the particular claim. [0053] For each of the claims, each dependent claim can depend both from the independent claim and from each of the prior dependent claims for each and every claim so long as the prior claim provides a proper antecedent basis for a claim term or element.

REFERENCES

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Claims

WHAT IS CLAIMED IS:

1. A method for treating a neurodegenerative disease in a patient with A0 plaques comprising administering to the patient a therapeutically effective amount of 2,4- Disulfonyl-N-Tert-Butylnitrone, 2,4-disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof to increase the expression of transthyretin (TTR) and reduce or eliminate A0 plaques by forming A0/TTR complexes that reduce A0 plaques or A0 plaque formation.

2. The method of claim 1, wherein the neurodegenerative disease is selected from diseases with A0 plaque formation selected from at least one of Alzheimer’s Disease, sepsis, alcoholism, or Parkinson’s disease.

3. The method of claim 1 , wherein the therapeutically effective amount of 2,4- Disulfonyl-N-Tert-Butylnitrone, 2,4-disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof increases the formation of A0/TTR complexes.

4. The method of claim 1, wherein the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4- disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof is administered orally, intravenously, or intraperitoneally.

5. The method of claim 1, wherein the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4- disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof are from about 40 to 1,200 mg/kg body weight/day, 100 to 450 mg/kg body weight/day, 200 to 400 mg/kg body weight/day, 300 to 800 mg/kg body weight/day, 350 to 1,000 mg/kg body weight/day, or 400 to 1,100 mg/kg body weight/day.

6. The method of claim 1, wherein the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4- disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof is at least one of: (1) administered at least one of continuously, intermittently, systemically, or locally, (2) administered one or more times a day for as long as the patient is in need of treatment for the neurodegenerative disease; (3) sequentially or concomitantly, with another pharmaceutical agent in a newly diagnosed neurodegenerative disease patient; (4) as a single agent or in combination with another pharmaceutical agent in a newly diagnosed neurodegenerative disease patient; (5) as a single agent or in combination with another pharmaceutical agent in a newly diagnosed neurodegenerative disease pediatric patient; or (6) a pediatric neurodegenerative disease patient.

7. The method of claim 1, wherein the neurodegenerative disease is not ALS.

8. The method of claim 1, wherein the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4- disulfonyl a-phenyl tertiary butyl nitrone, or a pharmaceutically acceptable salt thereof, is provided in a sustained-release formulation.

9. A method of preventing or treating a patient with a neurodegenerative disease comprising: identifying that the patient is in need for treatment for the neurodegenerative disease having A0 plaque formation; and providing an amount of 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4-disulfonyl a- phenyl tertiary butyl nitrone, or a pharmaceutically acceptable salt thereof, effective to prevent or reduce the neurodegenerative disease by preventing or decreasing A0 plaques or A0 plaque formation.

10. The method of claim 9, wherein the neurodegenerative disease is selected from diseases with A0 plaque formation selected from at least one of Alzheimer’s Disease, sepsis, alcoholism, or Parkinson’s disease.

11. The method of claim 9, wherein the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4- disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof increases the formation of A0/TTR complexes.

12. The method of claim 9, wherein the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4- disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof is administered orally, intravenously, or intraperitoneally.

13. The method of claim 9, wherein the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4- disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof are from about 40 to 1,200 mg/kg body weight/day, 100 to 450 mg/kg body weight/day, 200 to 400 mg/kg body weight/day, 300 to 800 mg/kg body weight/day, 350 to 1,000 mg/kg body weight/day, or 400 to 1,100 mg/kg body weight/day.

14. The method of claim 9, wherein the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4- disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof is at least one of: (1) administered at least one of continuously, intermittently, systemically, or locally, (2) administered one or more times a day for as long as the patient is in need of treatment for the neurodegenerative disease; (3) sequentially or concomitantly, with another pharmaceutical agent in a newly diagnosed neurodegenerative disease patient; (4) as a single agent or in combination with another pharmaceutical agent in a newly diagnosed neurodegenerative disease patient; (5) as a single agent or in combination with another pharmaceutical agent in a newly diagnosed neurodegenerative disease pediatric patient; or (6) a pediatric neurodegenerative disease patient.

15. The method of claim 9, wherein the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4- disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof inhibits immune suppression against the neurodegenerative disease.

16. The method of claim 9, wherein the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4- disulfonyl a-phenyl tertiary butyl nitrone, or a pharmaceutically acceptable salt thereof, is provided in a sustained-release formulation.

17. The method of claim 9, wherein the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4- disulfonyl a-phenyl tertiary butyl nitrone, or a pharmaceutically acceptable salt thereof, is formulated or administered together, separately or sequentially with a chemotherapy or an adjuvant therapy.

18. A method for treating a patient with a neurodegenerative disease comprising: determining if the patient has a neurodegenerative disease that is caused by A0 plaques or A0 plaque formation; and if the patient has the neurodegenerative disease administering a therapeutically effective amount of a therapeutically effective amount of 2,4-Disulfonyl-N-Tert- Butylnitrone, 2,4-disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof sufficient to overcome the neurodegenerative disease.

19. The method of claim 18, wherein the neurodegenerative disease is selected from diseases with A0 plaque formation selected from at least one of Alzheimer’s Disease, sepsis, alcoholism, or Parkinson’s disease.

20. The method of claim 18, wherein the therapeutically effective amount of 2,4- Disulfonyl-N-Tert-Butylnitrone, 2,4-disulfonyl a-phenyl tertiary butyl nitrone or a pharmaceutically acceptable salt thereof increases the formation of AP-transthyretin (TTR) complexes.

21. The method of claim 18, wherein the therapeutically effective amount of 2,4- Disulfonyl-N-Tert-Butylnitrone, 2,4-disulfonyl a-phenyl tertiary butyl nitrone or a

17 pharmaceutically acceptable salt thereof is administered orally, intravenously, or intraperitoneally.

22. The method of claim 18, wherein the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4- disulfonyl a-phenyl tertiary butyl nitrone, or a pharmaceutically acceptable salt thereof, is provided in a sustained-release formulation.

23. The method of claim 18, wherein the 2,4-Disulfonyl-N-Tert-Butylnitrone, 2,4- disulfonyl a-phenyl tertiary butyl nitrone, or a pharmaceutically acceptable salt thereof, is provided at 40 to 1,200 mg/kg body weight/day, 100 to 450 mg/kg body weight/day, 200 to 400 mg/kg body weight/day, 300 to 800 mg/kg body weight/day, 350 to 1,000 mg/kg body weight/day, or 400 to 1,100 mg/kg body weight/day.

24. The method of claim 18, wherein the neurodegenerative disease is not ALS.

18