WO2022174351A1 - Purine compounds for treating disorders - Google Patents

Purine compounds for treating disorders Download PDF

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Publication number
WO2022174351A1
WO2022174351A1 PCT/CA2022/050238 CA2022050238W WO2022174351A1 WO 2022174351 A1 WO2022174351 A1 WO 2022174351A1 CA 2022050238 W CA2022050238 W CA 2022050238W WO 2022174351 A1 WO2022174351 A1 WO 2022174351A1
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formula
compound
pharmaceutically acceptable
compounds
administration
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PCT/CA2022/050238
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French (fr)
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Mark Williams
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Marvel Biotechnology
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D473/00—Heterocyclic compounds containing purine ring systems
    • C07D473/02—Heterocyclic compounds containing purine ring systems with oxygen, sulphur, or nitrogen atoms directly attached in positions 2 and 6
    • C07D473/04—Heterocyclic compounds containing purine ring systems with oxygen, sulphur, or nitrogen atoms directly attached in positions 2 and 6 two oxygen atoms
    • C07D473/06—Heterocyclic compounds containing purine ring systems with oxygen, sulphur, or nitrogen atoms directly attached in positions 2 and 6 two oxygen atoms with radicals containing only hydrogen and carbon atoms, attached in position 1 or 3
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
    • C07D487/12—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains three hetero rings
    • C07D487/14—Ortho-condensed systems

Definitions

  • This invention relates generally to substituted purine compounds and salts thereof acting as adenosine A2a receptor (A2aR) antagonists for cancer, depression, anxiety, multiple sclerosis, NASH, sclerodermascleroderma, ADHD, Alzheimer's and Parkinsons, pharmaceutical compositions comprising such compounds, and methods of their use in treating cancer, depression, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer's and Parkinsons.
  • A2aR adenosine A2a receptor
  • Adenosine realizes its biological actions through a class of membrane specific receptors that belong to the super family of receptors coupled with G proteins. At least four subtypes of adenosine receptors have been identified: Al, A2a, Alb, and A3.
  • A2aR has been shown to provide a regulatory role in the immune system.
  • One A2aR antagonist, istradefylline has been shown to reduce motor impairment and in turn improve function in neurodegenerative diseases such as Parkinson's disease and related movement disorders (e.g. Huntington's Disease).
  • WO2013058681A2 discloses using A2aR antagonists for treating diseases of the central nervous system, oncological diseases and viral and bacterial diseases.
  • A2aR antagonists that are suitable for treating neurological diseases, fibrosis related diseases (NASH and scleroderma) and cancer.
  • NECA 5'-N-Ethylcarboxamidoadenosine, of Formula (II), or a pharmaceutically acceptable salt thereof, is an A2aR antagonist that may be useful for treating cancer, depression, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer's and Parkinsons:
  • SCH 442416 2-(2-Furanyl)-7-[3-(4-methoxyphenyl)propyl]-7H-pyrazolo [4,3-e][l,2,4]triazolo[l,5-c]pyrimidin-5-amine, of Formula (III), or a pharmaceutically acceptable salt thereof, is an A2aR antagonist that may be useful for treating cancer, depression, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer's and Parkinsons:
  • Istradefylline, (E)-8-(3,4-Dimethoxystyryl)-l,3-diethyl-7-methylxanthine, of Formula (IV), or a pharmaceutically acceptable salt thereof is an A2aR antagonist that may be useful for treating cancer, depression, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer's: [0011]
  • the invention also includes a pharmaceutical composition comprising a therapeutically effective amount of compound of formula I, and pharmaceutically acceptable salts thereof, and a pharmaceutically acceptable excipient.
  • This invention further includes a method for treating depression using compound of formula I and pharmaceutically acceptable salts thereof are provided.
  • the method comprises administering the one or more compound to a subject in need of such treatment, thereby treating depression.
  • This invention further includes a method for treating cancer, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer's or Parkinson's using compound of formula I and pharmaceutically acceptable salts thereof are provided.
  • the method comprises administering the one or more compound to a subject in need of such treatment, thereby treating depression, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer's and Parkinsons.
  • the compound is administered by intravenous injection, by injection into tissue, intraperitoneally, orally, or nasally.
  • the composition have a form of a solution, dispersion, suspension, powder, capsule, tablet, pill, time release capsule, time release tablet, or time release pill.
  • the invention encompasses a method comprising providing at least one such compound, measuring inhibition of A2aR activity for the compound and determining if the inhibition is above the expected level.
  • FIG. 1 is a graph showing the GPCR screening result of the compound of formula (I), SD-007;
  • FIG. 2 is a graph showing the GPCR screening result of the compound of formula (IV), istradefyline;
  • FIG. 3 is a graph showing the GPCR screening result of the compound of formula (II), 5'-/V-Ethylcarboxamidoadenosine (NECA);
  • FIG. 4 is a graph showing the GPCR screening result of the compound of formula (III), of SCH 442416;
  • FIG. 5 is a graph showing the brain versus plasma concentration of Formula (I) over time.
  • FIG. 6 is a graph showing the plasma concentration of Formula (I) over time for varying dosages.
  • FIG. 7 is a graph showing the brain versus plasma concentration of Formula (IV) over time.
  • FIG. 8 is a graph showing the plasma concentration of Formula (IV) over time for varying dosages.
  • FIG. 9 is a graph showing the dose response latency to immobility time of Formula (I) and (II) in the forced swim test of depression.
  • FIG. 10 is a graph showing the dose response immobilization time of Formula (I) and (II) in the forced swim test of depression.
  • the terms “pharmaceutically effective amount”, “therapeutically effective amount” or “therapeutically effective dose,” “effective amount” refer to the amount of the subject compound that will elicit the biological or medical response of a tissue, system, animal or human that is being sought by the researcher, veterinarian, medical doctor or other clinician.
  • therapeutically effective amount includes that amount of a compound that, when administered, is sufficient to prevent development of, or alleviate to some extent, one or more of the symptoms of the condition or disorder being treated. The therapeutically effective amount will vary depending on the compound, the disorder or condition and its severity and the age, weight, etc., of the mammal to be treated.
  • salts of the compounds of this invention include salts of the compounds of this invention that are prepared with relatively nontoxic acids or bases, depending on the particular substituents found on the compounds described herein.
  • base addition salts may be obtained by contacting the neutral form of such compounds with a sufficient amount of the desired base, either neat or in a suitable inert solvent.
  • salts may be derived from pharmaceutically acceptable inorganic bases that include aluminum, ammonium, calcium, copper, ferric, ferrous, lithium, magnesium, manganic, manganous, potassium, sodium, zinc and the like.
  • salts may be derived from pharmaceutically acceptable organic bases that include salts of primary, secondary and tertiary amines, including substituted amines, cyclic amines, naturally-occurring amines and the like, such as arginine, betaine, caffeine, choline, N,N'-dibenzylethylenediamine, diethylamine, 2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine, ethylenediamine, N-ethylmorpholine, N-ethylpiperidine, glucamine, glucosamine, histidine, hydrabamine, isopropylamine, lysine, methylglucamine, morpholine, piperazine, piperidine, polyamine resins, procaine, purines, theobromine, triethylamine, trimethylamine, tripropylamine, tromethamine and the like.
  • arginine betaine
  • caffeine choline
  • acid addition salts may be obtained by contacting the neutral form of such compounds with a sufficient amount of the desired acid, either neat or in a suitable inert solvent.
  • pharmaceutically acceptable acid addition salts include those derived from inorganic acids like hydrochloric, hydrobromic, nitric, carbonic, monohydrogencarbonic, phosphoric, monohydrogenphosphoric, dihydrogenphosphoric, sulfuric, monohydrogensulfuric, hydriodic, or phosphorous acids and the like, as well as the salts derived from relatively nontoxic organic acids like acetic, propionic, isobutyric, malonic, benzoic, succinic, suberic, fumaric, mandelic, phthalic, benzenesulfonic, p-tolylsulfonic, citric, tartaric, methanesulfonic, and the like.
  • salts of amino acids such as arginate and the like, and salts of organic acids like glucuronic or galactunoric acids and the like (see, e.g., Berge, S. M. et al., "Pharmaceutical Salts," Journal of Pharmaceutical Science, 66:1-19, 1977).
  • Certain specific compounds of the present invention contain both basic and acidic functionalities that allow the compounds to be converted into either base or acid addition salts.
  • the neutral forms of the compounds is regenerated by contacting the salt with a base or acid and isolating the parent compound in the conventional manner.
  • the parent form of the compound differs from the various salt forms in certain physical properties, such as solubility in polar solvents, but otherwise the salts are equivalent to the parent form of the compound for the purposes of the present invention.
  • the "subject” is defined herein to include animals such as mammals, including, but not limited to, primates (e.g., humans), cows, sheep, goats, horses, dogs, cats, rabbits, rats, mice and the like. In some embodiments, the subject is a human.
  • treat include partially or completely delaying, alleviating, mitigating or reducing the intensity, progression, or worsening of one or more attendant symptoms of a disorder or condition and/or alleviating, mitigating or impeding one or more causes of a disorder or condition.
  • Treatments according to the invention may be applied preventively, prophylactically, pallatively or remedially.
  • the compounds of the present invention may also contain unnatural proportions of atomic isotopes at one or more of the atoms that constitute such compounds.
  • the compounds may be radiolabeled with isotopes, such as for example deuterium ( 2 H), tritium ( 3 H), iodine-125 ( 125 I) or carbon-14 ( 14 C). All isotopic variations of the compounds of the present invention, whether radioactive or not, are intended to be encompassed within the scope of the present invention.
  • A2A receptor antagonists are drug candidates for the treatment of diseases of the central nervous system (CNS), in particular Parkinson's disease [Pinna A. Novel investigational adenosine A2A receptor antagonists for Parkinson's disease. Expert Opinion on Investigational Drugs, 2009, 18 (11), 1619-1631]; depression [El Yacoubi M, Costentin J, Vaugeois JM (December 2003) "Adenosine A2A receptors and depression”. Neurology 61 (11 Suppl 6): S82-7. PMID 14663017; Kaster MP, Rosa AO, Rosso MM, Goulart EC, Santos AR, Rodrigues AL (January 2004).
  • Adenosine binding to adenosine type A2A receptors located on the cell membrane of lymphocytes, transmits an adenosine signal into cells by activating the intracellular accumulation of cyclic adenosine monophosphate (cAMP), which, in turn, reduces the ability of lymphocytes to attack tumor tissue [Sitkovardsky MV, Kjaerga Lukashev D, Ohta A. Hypoxia-Adenosinergic Immunosuppression: Tumor Protection by T Regulatory Cells and Cancerous Tissue Hypoxia. Clin Cancer Res (2008) 14 (19), 5947-5952.].
  • cAMP cyclic adenosine monophosphate
  • the compound of formula I was subjected to GPCR screening using A2aR as the target.
  • the screening was done using the GPCR Screening and Profiling Services by Eurofins DiscoverX Corporation.
  • NECA and SCH 442416 were also screened. The results are shown in FIGs. 1-4 and the Experiments.
  • A2aR antagonists can be useful for treating Alzheimer's disease, Parkinson's disease, attention-deficit hyperactivity disorder, fragile X syndrome, depression, anxiety. Thus the compounds herein are useful for treating neurological diseases.
  • Cronstein B. “Adenosine receptors and fibrosis: a translational view", F1000 Biol Rep. 2011; 3: 21, which is incorporated herein by reference in its entirety, discloses certain A2aR antagonists could be useful in treating fibrosis, in particular hepatic fibrosis (e.g. NASH) and dermal fibrosis (e.g. scleroderma).
  • One or more of the compounds described herein is useful for treating cancer, depression, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer's or Parkinsons.
  • the compounds of the invention are useful in their pure forms.
  • the compounds of this invention are useful as pharmaceutical compositions prepared with a therapeutically effective amount of the compounds, as defined herein, and a pharmaceutically acceptable excipient, for example a carrier, or a diluent.
  • the compounds are systemically administered, e.g., orally, in combination with a pharmaceutically acceptable vehicle such as an inert diluent or an assimilable edible carrier; or by inhalation or insufflation. They may be enclosed in hard or soft shell gelatin capsules, may be compressed into tablets, or may be incorporated directly with the food of the patient's diet.
  • a pharmaceutically acceptable vehicle such as an inert diluent or an assimilable edible carrier
  • the compounds may be combined with one or more excipients and used in the form of ingestible tablets, buccal tablets, troches, capsules, elixirs, suspensions, syrups, wafers, and the like.
  • the compounds may be combined with a fine inert powdered carrier and inhaled by the subject or insufflated.
  • the percentage of the compositions and preparations may, of course, be varied and may be a suitable percentage of the weight of a given unit dosage form.
  • the amount of compounds in such therapeutically useful compositions is such that an effective dosage level may be obtained.
  • the tablets, troches, pills, capsules, or the like also contains the following: binders such as gum tragacanth, acacia, corn starch or gelatin; excipients such as dicalcium phosphate; a disintegrating agent such as corn starch, potato starch, alginic acid and the like; a lubricant such as magnesium stearate; and a sweetening agent such as sucrose, fructose, lactose or aspartame or a flavoring agent such as peppermint, oil of wintergreen, or cherry flavoring may be added.
  • a capsule may contain, in addition to materials of the above type, a liquid carrier, such as a vegetable oil or a polyethylene glycol.
  • any material used in preparing any unit dosage form should be pharmaceutically acceptable and substantially non-toxic in the amounts employed.
  • the compounds are incorporated into sustained-release preparations and devices.
  • the compounds may be incorporated into time release capsules, time release tablets, and time release pills.
  • the compounds are administered intravenously or intraperitoneally by infusion or injection.
  • Solutions of the compounds may be prepared in water, optionally mixed with a nontoxic surfactant.
  • dispersions are prepared in glycerol, liquid polyethylene glycols, triacetin, or mixtures thereof or in oils. In some embodiments, these preparations contain a preservative to prevent the growth of microorganisms under ordinary conditions of storage and use.
  • the pharmaceutical dosage forms for injection or infusion is sterile aqueous solutions or dispersions or sterile powders comprising the compounds which are adapted for the extemporaneous preparation of sterile injectable or infusible solutions or dispersions, optionally encapsulated in liposomes.
  • the ultimate dosage form should be sterile, fluid and stable under the conditions of manufacture and storage.
  • the liquid carrier or vehicle is a solvent or liquid dispersion medium comprising, for example, water, ethanol, a polyol (for example, glycerol, propylene glycol, liquid polyethylene glycols, and the like), vegetable oils, nontoxic glyceryl esters, and suitable mixtures thereof.
  • the proper fluidity is maintained by the formation of liposomes, by the maintenance of the required particle size in the case of dispersions or by the use of surfactants.
  • various antibacterial and antifungal agents for example, parabens, chlorobutanol, phenol, sorbic acid, thimerosal, and the like is used to prevent of the action of microorganisms.
  • isotonic agents for example, sugars, buffers or sodium chloride is included.
  • agents delaying absorption for example, aluminum monostearate or gelatin is used to prolong absorption of the injectable compositions.
  • sterile injectable solutions is prepared by incorporating the compounds in the required amount in the appropriate solvent, and optionally with some of the other ingredients enumerated above, as may be required, followed by filter sterilization.
  • the composition may be vacuum dried and/or freeze dried, which yield a powder of the active ingredient plus any additional desired ingredient present in the previously sterile-filtered solutions.
  • the compounds are applied in pure form.
  • the compounds are administered to the skin as compositions or formulations, in combination with a dermatologically acceptable carrier, which may be a solid or a liquid.
  • the solid carriers include finely divided solids such as talc, clay, microcrystalline cellulose, silica, alumina and the like. In some embodiments, the solid carriers include nontoxic polymeric nanoparticles or microparticles.
  • the liquid carriers include water, alcohols or glycols or water/alcohol/glycol blends, in which the compounds can be dissolved or dispersed at effective levels, optionally with the aid of non-toxic surfactants. In some embodiments, adjuvants such as fragrances and additional antimicrobial agents are added to optimize the properties for a given use.
  • the resultant liquid compositions may be applied from absorbent pads, used to impregnate bandages and other dressings, or sprayed onto the affected area using pump-type or aerosol sprayers.
  • thickeners such as synthetic polymers, fatty acids, fatty acid salts and esters, fatty alcohols, modified celluloses or modified mineral materials are employed with liquid carriers to form spreadable pastes, gels, ointments, soaps, and the like, for application directly to the skin of the user.
  • the compounds are formulated in lyophilized form for parenteral administration.
  • Lyophilized formulations is reconstituted by addition of water or other aqueous medium and then further diluted with a suitable diluent prior to use.
  • the liquid formulation is a buffered, isotonic, aqueous solution.
  • the diluents are isotonic saline solution, 5% dextrose in water, and buffered sodium or ammonium acetate solution.
  • Pharmaceutically acceptable solid or liquid excipients may be added to enhance or stabilize the composition, or to facilitate preparation of the composition.
  • the pharmaceutical composition may additionally contain one or more other pharmacologically active agents in addition to a compound described herein.
  • useful dosages of the compounds may be determined by comparing their in vitro activity, and in vivo activity in animal models. Methods for the extrapolation of effective dosages in mice, and other animals, to humans are known to the art; for example, see U.S. Pat. No. 4,938,949, which is hereby incorporated by reference.
  • the amount of the compounds required for use in treatment will vary not only with the particular salt selected but also with the route of administration, the nature of the condition being treated and the age and condition of the patient and will be ultimately at the discretion of the attendant physician or clinician.
  • effective dosages and routes of administration of agents of the invention are conventional.
  • the exact amount (effective dose) of the compounds may vary from subject to subject, depending on, for example, the species, age, weight and general or clinical condition of the subject, the severity or mechanism of any disorder being treated, the particular compound or vehicle used, the method and scheduling of administration, and the like.
  • the therapeutically effective dose is determined empirically, by conventional procedures known to those of skill in the art.
  • an effective dose is estimated initially either in cell culture assays or in suitable animal models.
  • the animal model is used to determine the appropriate concentration ranges and routes of administration.
  • such information is then used to determine useful doses and routes for administration in humans.
  • a therapeutic dose is selected by analogy to dosages for comparable therapeutic agents.
  • the dosage may be in the range from about 0.001 to about 100 mg/kg, e.g., from about 0.01 to about 100 mg/kg of body weight per day, such as above about 0.1 mg per kilogram, or in a range of from about 1 to about 10 mg per kilogram body weight of the recipient per day.
  • a suitable dose may be about 0.3 mg/kg, 0.7 mg/kg, 1 mg/kg, 10 mg/kg, or 50 mg/kg of body weight per day.
  • the compounds are administered in unit dosage form; for example, containing 0.05 to 10000 mg, 0.5 to 10000 mg, 5 to 1000 mg, or about 100 mg of active ingredient per unit dosage form.
  • the compounds are administered to achieve peak plasma concentrations of, for example, from about 0.5 to about 75 mM, about 1 to 50 pM, about 2 to about 30 pM, or about 5 to about 25 pM.
  • Exemplary desirable plasma concentrations include at least or no more than 0.25, 0.5, 1, 5, 10, 25, 50, 75, 100 or 200 pM.
  • plasma levels may be from about 1 to 100 micromolar or from about 10 to about 25 micromolar. In some embodiments, this is achieved by the intravenous injection of a 0.05 to 5% solution of the compounds, optionally in saline, or orally administered as a bolus containing about 1-100 mg of the compounds.
  • desirable blood levels is maintained by continuous infusion to provide about 0.00005-5 mg per kg body weight per hour, for example at least or no more than 0.00005, 0.0005, 0.005, 0.05, 0.5, or 5 mg/kg/hr.
  • such levels are obtained by intermittent infusions containing about 0.0002-20 mg per kg body weight, for example, at least or no more than 0.0002, 0.002, 0.02, 0.2, 2, 20, or 50 mg of the compounds per kg of body weight.
  • the amount of purine compound used is between 0.1 to 5 mg per kg of the subject per day. In a preferred embodiment, the amount of purine compound used is between 0.2 to 1.3 mg per kg of the subject per day. In a further preferred embodiment, the amount of purine compound used is about 0.3 mg to 0.7 per kg of the subject per day.
  • the compounds are presented in a single dose or as divided doses administered at appropriate intervals, for example, as two, three, four or more sub-doses per day.
  • the sub-dose itself is further divided, e.g., into a number of discrete loosely spaced administrations; such as multiple inhalations from an insufflator.
  • a pharmaceutical composition of the present invention is packaged in a container with a label, or instructions, or both, indicating use of the pharmaceutical composition in the treatment of the indicated disease.
  • Cell lines were expanded from freezer stocks according to standard procedures. Cells were seeded in a total volume of 20 pL into black-walled, clear-bottom, Poly-D-lysine coated 384-well microplates and incubated at 37°C for the appropriate time prior to testing.
  • Assays were performed in 1 x Dye Loading Buffer consisting of lx Dye, lx Additive A and 2.5 mM Probenecid in HBSS / 20 mM Hepes. Probenicid was prepared fresh. Cells were loaded with dye prior to testing. Media was aspirated from cells and replaced with 20 pL Dye Loading Buffer. Cells were incubated for 30-60 minutes at 37°C.
  • agonist determination cells were incubated with sample to induce response. After dye loading, cells were removed from the incubator and 10 pL HBSS / 20 mM Hepes was added. 3x vehicle was included in the buffer when performing agonist dose curves to define the EC80 for subsequent antagonist assays. Cells were incubated for 30 minutes at room temperature in the dark to equilibrate plate temperature. Intermediate dilution of sample stocks was performed to generate 4X sample in assay buffer. Compound agonist activity was measured on a FLIPR Tetra (MDS). Calcium mobilization was monitored for 2 minutes and 10 pL 4X sample in HBSS / 20 mM Hepes was added to the cells 5 seconds into the assay. Allosteric Modulation Format
  • % Activity 100% x (mean RFU of test sample - mean RFU of vehicle control) / (mean MAX RFU control ligand - mean RFU of vehicle control). [0073] For positive allosteric mode assays, percentage modulation was calculated using the following formula:
  • % Modulation 100% x ((mean RFU of test sample - mean RFU of EC20 control) / (mean RFU of MAX control ligand - mean RFU of EC20 control)).
  • % Inhibition 100% x (1 - (mean RFU of test sample - mean RFU of vehicle control) / (mean RFU of EC80 control - mean RFU of vehicle control)).
  • FIGs 1 to 4 present the results of G-protein-coupled receptors (GPCR) screening using A2aR as the target by the GPCR Screening and Profiling Services of Eurofins DiscoverX Corporation.
  • GPCR G-protein-coupled receptors
  • Figures 1 to 4 show percent reacted (Y) versus concentration in mM (X).
  • Compounds were tested in antagonist mode with the requested GPCR Biosensor Assay.
  • For the antagonist assay data was normalized to the maximal and minimal response observed in the presence of EC80 ligand and vehicle.
  • the following EC80 concentrations were used: ADORA2A Calcium Flux: 0.021 pM NECA.
  • FIG. 1 shows the result for the compound of formula I.
  • the ICso was 0.1438 pM.
  • a higher ICso level indicates better binding with A2aR.
  • FIG. 2 shows the result for the compound of formula IV, istradefyline.
  • the ICso was 0.0667 pM.
  • FIG. 3 shows the result for NECA.
  • the half maximal effective concentration (ECso) was 0.0076 mM.
  • FIG. 4 shows the result for SCH 442416.
  • the ICso was 0.0113 pM.
  • the forced swim test also known as the behavioral despair test, is used to test for depression-like behavior.
  • the test includes placing a rat or mouse inside a cylinder filled with water.
  • 'Floating behavior' (where the animal remains almost immobile and with its head above water) is used as a parameter to analyze 'hopelessness' and thus depression-like behavior.
  • Rodents given antidepressants swim longer than controls. Immobility time is decreased by various antidepressants.
  • vehicle 0.3% Tween 80
  • test article 2.5-10 mg/kg of formula I or Istradefylline (aka KW-6002, 2.5 mg/kg) one hour prior to testing.

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Abstract

Novel and known substituted purine compounds and salts thereof act as adenosine A2a receptor (A2aR) antagonists for cancer immunotherapy, depression, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer's and Parkinsons. Pharmaceutical compositions comprising such compounds, and methods of their use in treating depression are also taught.

Description

Purine Compounds for Treating Disorders
FIELD OF THE INVENTION
[0001] This invention relates generally to substituted purine compounds and salts thereof acting as adenosine A2a receptor (A2aR) antagonists for cancer, depression, anxiety, multiple sclerosis, NASH, sclerodermascleroderma, ADHD, Alzheimer's and Parkinsons, pharmaceutical compositions comprising such compounds, and methods of their use in treating cancer, depression, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer's and Parkinsons.
BACKGROUND OF THE INVENTION
[0002] Adenosine realizes its biological actions through a class of membrane specific receptors that belong to the super family of receptors coupled with G proteins. At least four subtypes of adenosine receptors have been identified: Al, A2a, Alb, and A3.
[0003] A2aR has been shown to provide a regulatory role in the immune system. One A2aR antagonist, istradefylline, has been shown to reduce motor impairment and in turn improve function in neurodegenerative diseases such as Parkinson's disease and related movement disorders (e.g. Huntington's Disease).
[0004] WO2013058681A2 discloses using A2aR antagonists for treating diseases of the central nervous system, oncological diseases and viral and bacterial diseases.
[0005] There is a need for A2aR antagonists that are suitable for treating neurological diseases, fibrosis related diseases (NASH and scleroderma) and cancer.
SUMMARY OF THE INVENTION
[0006] The inventors have found a purine compound useful as an A2aR antagonist for treating diseases including depression. [0007] In one aspect, 7FMI, a compound of Formula (I) or a pharmaceutically acceptable salt thereof is provided:
Figure imgf000003_0001
(E)-8-(3,4-dimethoxystyryl)-1 ,3-diethyl-7-(fluoromethyl)-3,7-dihydro-1 /-/-purine-2, 6-dione
[0008] In one aspect, it is found that NECA, 5'-N-Ethylcarboxamidoadenosine, of Formula (II), or a pharmaceutically acceptable salt thereof, is an A2aR antagonist that may be useful for treating cancer, depression, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer's and Parkinsons:
Figure imgf000003_0002
[0009] In one aspect, it is found that SCH 442416, 2-(2-Furanyl)-7-[3-(4-methoxyphenyl)propyl]-7H-pyrazolo [4,3-e][l,2,4]triazolo[l,5-c]pyrimidin-5-amine, of Formula (III), or a pharmaceutically acceptable salt thereof, is an A2aR antagonist that may be useful for treating cancer, depression, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer's and Parkinsons:
Figure imgf000004_0001
(III).
[0010] In one aspect, it is found that Istradefylline, (E)-8-(3,4-Dimethoxystyryl)-l,3-diethyl-7-methylxanthine, of Formula (IV), or a pharmaceutically acceptable salt thereof, is an A2aR antagonist that may be useful for treating cancer, depression, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer's:
Figure imgf000004_0002
[0011] The invention also includes a pharmaceutical composition comprising a therapeutically effective amount of compound of formula I, and pharmaceutically acceptable salts thereof, and a pharmaceutically acceptable excipient.
[0012] This invention further includes a method for treating depression using compound of formula I and pharmaceutically acceptable salts thereof are provided. The method comprises administering the one or more compound to a subject in need of such treatment, thereby treating depression.
[0013] This invention further includes a method for treating cancer, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer's or Parkinson's using compound of formula I and pharmaceutically acceptable salts thereof are provided. The method comprises administering the one or more compound to a subject in need of such treatment, thereby treating depression, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer's and Parkinsons.
[0014] In some embodiments, the compound is administered by intravenous injection, by injection into tissue, intraperitoneally, orally, or nasally. In some embodiments, the composition have a form of a solution, dispersion, suspension, powder, capsule, tablet, pill, time release capsule, time release tablet, or time release pill.
[0015] Method for synthesizing the compound for formula I are provided.
[0016] The invention encompasses a method comprising providing at least one such compound, measuring inhibition of A2aR activity for the compound and determining if the inhibition is above the expected level.
BRIEF DESCRIPTION OF THE DRAWINGS
[0017] FIG. 1 is a graph showing the GPCR screening result of the compound of formula (I), SD-007;
[0018] FIG. 2 is a graph showing the GPCR screening result of the compound of formula (IV), istradefyline;
[0019] FIG. 3 is a graph showing the GPCR screening result of the compound of formula (II), 5'-/V-Ethylcarboxamidoadenosine (NECA); [0020] FIG. 4 is a graph showing the GPCR screening result of the compound of formula (III), of SCH 442416;
[0021] FIG. 5 is a graph showing the brain versus plasma concentration of Formula (I) over time. [0022] FIG. 6 is a graph showing the plasma concentration of Formula (I) over time for varying dosages.
[0023] FIG. 7 is a graph showing the brain versus plasma concentration of Formula (IV) over time.
[0024] FIG. 8 is a graph showing the plasma concentration of Formula (IV) over time for varying dosages.
[0025] FIG. 9 is a graph showing the dose response latency to immobility time of Formula (I) and (II) in the forced swim test of depression.
[0026] FIG. 10 is a graph showing the dose response immobilization time of Formula (I) and (II) in the forced swim test of depression. DETAILED DESCRIPTION
[0027] Embodiments of the invention are discussed in detail below. In describing embodiments, specific terminology is employed for the sake of clarity. However, the disclosure is not intended to be limited to the specific terminology so selected. A person skilled in the relevant art will recognize that other equivalent parts can be employed and other methods developed without parting from the spirit and scope of the disclosure. All references cited herein are incorporated by reference as if each had been individually incorporated. [0028] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs.
[0029] As used in the specification and claims, the singular forms "a", "an" and "the" include plural references unless the context clearly dictates otherwise.
[0030] The term "comprising" as used herein will be understood to mean that the list following is non-exhaustive and may or may not include any other additional suitable items, for example one or more further feature(s), component(s) and/or ingredient(s) as appropriate.
[0031] The terms "pharmaceutically effective amount", "therapeutically effective amount" or "therapeutically effective dose," "effective amount" refer to the amount of the subject compound that will elicit the biological or medical response of a tissue, system, animal or human that is being sought by the researcher, veterinarian, medical doctor or other clinician. The term "therapeutically effective amount" includes that amount of a compound that, when administered, is sufficient to prevent development of, or alleviate to some extent, one or more of the symptoms of the condition or disorder being treated. The therapeutically effective amount will vary depending on the compound, the disorder or condition and its severity and the age, weight, etc., of the mammal to be treated.
[0032] The term "pharmaceutically acceptable salts" in this disclosure includes salts of the compounds of this invention that are prepared with relatively nontoxic acids or bases, depending on the particular substituents found on the compounds described herein. When compounds of the present invention contain relatively acidic functionalities, base addition salts may be obtained by contacting the neutral form of such compounds with a sufficient amount of the desired base, either neat or in a suitable inert solvent. For example, salts may be derived from pharmaceutically acceptable inorganic bases that include aluminum, ammonium, calcium, copper, ferric, ferrous, lithium, magnesium, manganic, manganous, potassium, sodium, zinc and the like. For example, salts may be derived from pharmaceutically acceptable organic bases that include salts of primary, secondary and tertiary amines, including substituted amines, cyclic amines, naturally-occurring amines and the like, such as arginine, betaine, caffeine, choline, N,N'-dibenzylethylenediamine, diethylamine, 2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine, ethylenediamine, N-ethylmorpholine, N-ethylpiperidine, glucamine, glucosamine, histidine, hydrabamine, isopropylamine, lysine, methylglucamine, morpholine, piperazine, piperidine, polyamine resins, procaine, purines, theobromine, triethylamine, trimethylamine, tripropylamine, tromethamine and the like. When compounds of the present invention contain relatively basic functionalities, acid addition salts may be obtained by contacting the neutral form of such compounds with a sufficient amount of the desired acid, either neat or in a suitable inert solvent. Examples of pharmaceutically acceptable acid addition salts include those derived from inorganic acids like hydrochloric, hydrobromic, nitric, carbonic, monohydrogencarbonic, phosphoric, monohydrogenphosphoric, dihydrogenphosphoric, sulfuric, monohydrogensulfuric, hydriodic, or phosphorous acids and the like, as well as the salts derived from relatively nontoxic organic acids like acetic, propionic, isobutyric, malonic, benzoic, succinic, suberic, fumaric, mandelic, phthalic, benzenesulfonic, p-tolylsulfonic, citric, tartaric, methanesulfonic, and the like. Also included are salts of amino acids such as arginate and the like, and salts of organic acids like glucuronic or galactunoric acids and the like (see, e.g., Berge, S. M. et al., "Pharmaceutical Salts," Journal of Pharmaceutical Science, 66:1-19, 1977). Certain specific compounds of the present invention contain both basic and acidic functionalities that allow the compounds to be converted into either base or acid addition salts.
[0033] In some embodiments, the neutral forms of the compounds is regenerated by contacting the salt with a base or acid and isolating the parent compound in the conventional manner. The parent form of the compound differs from the various salt forms in certain physical properties, such as solubility in polar solvents, but otherwise the salts are equivalent to the parent form of the compound for the purposes of the present invention.
[0034] The "subject" is defined herein to include animals such as mammals, including, but not limited to, primates (e.g., humans), cows, sheep, goats, horses, dogs, cats, rabbits, rats, mice and the like. In some embodiments, the subject is a human.
[0035] The terms "treat", "treating", "treatment" and grammatical variations thereof as used in this disclosure, include partially or completely delaying, alleviating, mitigating or reducing the intensity, progression, or worsening of one or more attendant symptoms of a disorder or condition and/or alleviating, mitigating or impeding one or more causes of a disorder or condition. Treatments according to the invention may be applied preventively, prophylactically, pallatively or remedially.
[0036] The compounds of the present invention may also contain unnatural proportions of atomic isotopes at one or more of the atoms that constitute such compounds. For example, the compounds may be radiolabeled with isotopes, such as for example deuterium (2H), tritium (3H), iodine-125 (125I) or carbon-14 (14C). All isotopic variations of the compounds of the present invention, whether radioactive or not, are intended to be encompassed within the scope of the present invention.
[0037] A2A receptor antagonists are drug candidates for the treatment of diseases of the central nervous system (CNS), in particular Parkinson's disease [Pinna A. Novel investigational adenosine A2A receptor antagonists for Parkinson's disease. Expert Opinion on Investigational Drugs, 2009, 18 (11), 1619-1631]; depression [El Yacoubi M, Costentin J, Vaugeois JM (December 2003) "Adenosine A2A receptors and depression". Neurology 61 (11 Suppl 6): S82-7. PMID 14663017; Kaster MP, Rosa AO, Rosso MM, Goulart EC, Santos AR, Rodrigues AL (January 2004). "Adenosine administration produces an antidepressant-like effect in mice: evidence for the involvement of A1 and A2A receptors". Neuroscience Letters 355 (1-2): 21-4. doi: 10.1016 / j.neulet.2003.10.040. PMID 14729225; Labato KR, Binfare RW, Budni J, Rosa AO, Santos AR, Rodrigues AL (May 2008). "Involvement of the adenosine A1 and A2A receptors in the antidepressant-like effect of zinc in the forced swimming test". Progress in Neuro-psychopharmacology & Biological Psychiatry 32 (4): 994-9. doi: 10.1016 / j.pnpbp.2008.01.012. PMID 18289757.], cognitive disorders [Takahashi RN, Pamplona FA, Prediger RD (2008). "Adenosine receptor antagonists for cognitive dysfunction: a review of animal studies". Frontiers in Bioscience: a Journal and Virtual Library 13 (13): 2614-32. doi: 10.2741 / 2870. PMID 17981738.], and inflammatory processes [Sullivan GW (November 2003) "Adenosine A2A receptor agonists as anti-inflammatory agents". Current Opinion in Investigational Drugs (London, England: 2000) 4 (11): 1313-9. PMID 14758770; Lappas CM, Sullivan GW, Linden J (July 2005). "Adenosine A2A agonists in development for the treatment of inflammation". Expert Opinion on Investigational Drugs 14 (7): 797-806. doi: 10.1517 / 13543784.14.7.797. PMID 16022569.].
[0038] More recently, it has been shown that adenosine, which accumulates near tumor tissue [Blay J., White T.D., Oskin D.W. The Extracellular Fluid of Solid Carcinomas Contains immunosuppressive Concentrations of Adenosine. Cancer Research, (1997) 57, 2602-2605.], Significantly inhibits the ability of activated T-lymphocytes and natural killer cells to bind and kill tumor cells [Hoskin D. W, Reynolds T., Blay J. 2-Chloroadenosine inhibits the MHC unrestricted cytolytic activity of anti-CD3-activated killer cells: evidence for the involvement of a non-Al / A2 cell-surface adenosine receptor. Cell Immunol. (1994) 159:85-93. Hoskin D.W., Reynolds T., Blay, J. Adenosine as a possible inhibitor of killer T-cell activation in the microenvironment of solid tumors. Int. J. Cancer, (1994) 59: 854-855.]. It has been shown that hypoxia, which develops in solid tumors, initiates the formation of increased concentrations of adenosine near the tumor tissue. Adenosine, binding to adenosine type A2A receptors located on the cell membrane of lymphocytes, transmits an adenosine signal into cells by activating the intracellular accumulation of cyclic adenosine monophosphate (cAMP), which, in turn, reduces the ability of lymphocytes to attack tumor tissue [Sitkovardsky MV, Kjaerga Lukashev D, Ohta A. Hypoxia-Adenosinergic Immunosuppression: Tumor Protection by T Regulatory Cells and Cancerous Tissue Hypoxia. Clin Cancer Res (2008) 14 (19), 5947-5952.]. Based on these observations [Ohta A., Gorelik E., Simon J., Prasad SJ, Ronchese F., Lukashev D., Wong MKK, Huang X., Caldwell S., Liu K., Patrick Smith P., Chen J .-F., Jackson E. K, Apasov S., Abrams S., Sitkovsky M. A2A adenosine receptor protects tumors from antitumor T cells. Proc Natl Acad Sci, USA (2006) 103,13132-13137. Lukashev D., Sitkovsky M., Ohta A .. From "Hellstrom Paradox" to anti-adenosinergic cancer immunotherapy. Purinergic Signaling (2007) 3, 129-134] proposed an innovative approach to enhancing the efficacy of anticancer vaccines using an adjuvant that inhibits the A2A adenosine receptor [Ohta A., Sitkovsky M. Methods And Compositions For Improving Immune Responses, WO 2008/14748]. [0039] The inventors developed the compounds of formulae I and developed efficient methods for preparing the compounds for formulae I that differs from the synthesis of Istradefylline in the final step :
[0040]
Figure imgf000011_0001
Scheme 1
[0041] To ascertain whether the compounds are suitable A2aR antagonists, the compound of formula I was subjected to GPCR screening using A2aR as the target. The screening was done using the GPCR Screening and Profiling Services by Eurofins DiscoverX Corporation. For comparison, NECA and SCH 442416 were also screened. The results are shown in FIGs. 1-4 and the Experiments.
[0042] The screening results showed this compounds to have substantial A2aR antagonist properties. [0043] Robert D. Leone, Ying-Chun Lo, and Jonathan D. Powell, "A2aR antagonists: Next generation checkpoint blockade for cancer immunotherapy", Comput Struct Biotechnol J. 2015; 13: 265-272, which is incorporated herein by reference in its entirety, discloses certain A2aR antagonists that can be useful for cancer immunotherapy. Thus, the compounds described herein are useful for treating cancer. Domenici MR eta/. "Adenosine A2A receptor as potential therapeutic target in neuropsychiatric disorders", Pharmacol Res 2019; 147: 104338, which is incorporated herein by reference in its entirety, discloses certain A2aR antagonists can be useful for treating Alzheimer's disease, Parkinson's disease, attention-deficit hyperactivity disorder, fragile X syndrome, depression, anxiety. Thus the compounds herein are useful for treating neurological diseases. Cronstein B. "Adenosine receptors and fibrosis: a translational view", F1000 Biol Rep. 2011; 3: 21, which is incorporated herein by reference in its entirety, discloses certain A2aR antagonists could be useful in treating fibrosis, in particular hepatic fibrosis (e.g. NASH) and dermal fibrosis (e.g. scleroderma).
[0044] One or more of the compounds described herein is useful for treating cancer, depression, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer's or Parkinsons.
[0045] In some embodiments, the compounds of the invention are useful in their pure forms. In some embodiments, the compounds of this invention are useful as pharmaceutical compositions prepared with a therapeutically effective amount of the compounds, as defined herein, and a pharmaceutically acceptable excipient, for example a carrier, or a diluent.
[0046] In some embodiments, the compounds are systemically administered, e.g., orally, in combination with a pharmaceutically acceptable vehicle such as an inert diluent or an assimilable edible carrier; or by inhalation or insufflation. They may be enclosed in hard or soft shell gelatin capsules, may be compressed into tablets, or may be incorporated directly with the food of the patient's diet. For oral therapeutic administration, the compounds may be combined with one or more excipients and used in the form of ingestible tablets, buccal tablets, troches, capsules, elixirs, suspensions, syrups, wafers, and the like. The compounds may be combined with a fine inert powdered carrier and inhaled by the subject or insufflated. The percentage of the compositions and preparations may, of course, be varied and may be a suitable percentage of the weight of a given unit dosage form. The amount of compounds in such therapeutically useful compositions is such that an effective dosage level may be obtained.
[0047] In some embodiments, the tablets, troches, pills, capsules, or the like also contains the following: binders such as gum tragacanth, acacia, corn starch or gelatin; excipients such as dicalcium phosphate; a disintegrating agent such as corn starch, potato starch, alginic acid and the like; a lubricant such as magnesium stearate; and a sweetening agent such as sucrose, fructose, lactose or aspartame or a flavoring agent such as peppermint, oil of wintergreen, or cherry flavoring may be added. It is appreciated that a capsule may contain, in addition to materials of the above type, a liquid carrier, such as a vegetable oil or a polyethylene glycol. Various other materials may be present as coatings or to otherwise modify the physical form of the solid unit dosage form. For instance, tablets, pills, or capsules may be coated with gelatin, wax, shellac or sugar and the like. A syrup or elixir may contain the active compound, sucrose or fructose as a sweetening agent, methyl and propylparabens as preservatives, a dye and flavoring such as cherry or orange flavor. It is understood that any material used in preparing any unit dosage form should be pharmaceutically acceptable and substantially non-toxic in the amounts employed. In some embodiments, the compounds are incorporated into sustained-release preparations and devices. For example, the compounds may be incorporated into time release capsules, time release tablets, and time release pills.
[0048] In some embodiments, the compounds are administered intravenously or intraperitoneally by infusion or injection. Solutions of the compounds may be prepared in water, optionally mixed with a nontoxic surfactant. In some embodiments, dispersions are prepared in glycerol, liquid polyethylene glycols, triacetin, or mixtures thereof or in oils. In some embodiments, these preparations contain a preservative to prevent the growth of microorganisms under ordinary conditions of storage and use.
[0049] In some embodiments, the pharmaceutical dosage forms for injection or infusion is sterile aqueous solutions or dispersions or sterile powders comprising the compounds which are adapted for the extemporaneous preparation of sterile injectable or infusible solutions or dispersions, optionally encapsulated in liposomes. In all cases, the ultimate dosage form should be sterile, fluid and stable under the conditions of manufacture and storage. In some embodiments, the liquid carrier or vehicle is a solvent or liquid dispersion medium comprising, for example, water, ethanol, a polyol (for example, glycerol, propylene glycol, liquid polyethylene glycols, and the like), vegetable oils, nontoxic glyceryl esters, and suitable mixtures thereof. In some embodiments, the proper fluidity is maintained by the formation of liposomes, by the maintenance of the required particle size in the case of dispersions or by the use of surfactants. In some embodiments, various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, sorbic acid, thimerosal, and the like is used to prevent of the action of microorganisms. In some embodiments, isotonic agents, for example, sugars, buffers or sodium chloride is included. In some embodiments, agents delaying absorption, for example, aluminum monostearate or gelatin is used to prolong absorption of the injectable compositions.
[0050] In some embodiments, sterile injectable solutions is prepared by incorporating the compounds in the required amount in the appropriate solvent, and optionally with some of the other ingredients enumerated above, as may be required, followed by filter sterilization. In the case of sterile powders for the preparation of sterile injectable solutions, the composition may be vacuum dried and/or freeze dried, which yield a powder of the active ingredient plus any additional desired ingredient present in the previously sterile-filtered solutions.
[0051] In some embodiments, for topical administration, the compounds are applied in pure form. In some embodiments, the compounds are administered to the skin as compositions or formulations, in combination with a dermatologically acceptable carrier, which may be a solid or a liquid.
[0052] In some embodiments, the solid carriers include finely divided solids such as talc, clay, microcrystalline cellulose, silica, alumina and the like. In some embodiments, the solid carriers include nontoxic polymeric nanoparticles or microparticles. In some embodiments, the liquid carriers include water, alcohols or glycols or water/alcohol/glycol blends, in which the compounds can be dissolved or dispersed at effective levels, optionally with the aid of non-toxic surfactants. In some embodiments, adjuvants such as fragrances and additional antimicrobial agents are added to optimize the properties for a given use. The resultant liquid compositions may be applied from absorbent pads, used to impregnate bandages and other dressings, or sprayed onto the affected area using pump-type or aerosol sprayers.
[0053] In some embodiments, thickeners such as synthetic polymers, fatty acids, fatty acid salts and esters, fatty alcohols, modified celluloses or modified mineral materials are employed with liquid carriers to form spreadable pastes, gels, ointments, soaps, and the like, for application directly to the skin of the user.
[0054] In some embodiments, the compounds are formulated in lyophilized form for parenteral administration. In some embodiments, Lyophilized formulations is reconstituted by addition of water or other aqueous medium and then further diluted with a suitable diluent prior to use. In some embodiments, the liquid formulation is a buffered, isotonic, aqueous solution. In some embodiments, the diluents are isotonic saline solution, 5% dextrose in water, and buffered sodium or ammonium acetate solution. Pharmaceutically acceptable solid or liquid excipients may be added to enhance or stabilize the composition, or to facilitate preparation of the composition.
[0055] In some embodiments, the pharmaceutical composition may additionally contain one or more other pharmacologically active agents in addition to a compound described herein.
[0056] In some embodiments, useful dosages of the compounds may be determined by comparing their in vitro activity, and in vivo activity in animal models. Methods for the extrapolation of effective dosages in mice, and other animals, to humans are known to the art; for example, see U.S. Pat. No. 4,938,949, which is hereby incorporated by reference.
[0057] It is appreciated that the amount of the compounds required for use in treatment will vary not only with the particular salt selected but also with the route of administration, the nature of the condition being treated and the age and condition of the patient and will be ultimately at the discretion of the attendant physician or clinician. [0058] In some embodiments, effective dosages and routes of administration of agents of the invention are conventional. The exact amount (effective dose) of the compounds may vary from subject to subject, depending on, for example, the species, age, weight and general or clinical condition of the subject, the severity or mechanism of any disorder being treated, the particular compound or vehicle used, the method and scheduling of administration, and the like. In some embodiments, the therapeutically effective dose is determined empirically, by conventional procedures known to those of skill in the art. For example, persons skilled in the art may refer to The Pharmacological Basis of Therapeutics, Goodman and Gilman, eds., Macmillan Publishing Co., New York. In some embodiments, an effective dose is estimated initially either in cell culture assays or in suitable animal models. In some embodiments, the animal model is used to determine the appropriate concentration ranges and routes of administration. In some embodiments, such information is then used to determine useful doses and routes for administration in humans. In some embodiments, a therapeutic dose is selected by analogy to dosages for comparable therapeutic agents.
[0059] For example, the dosage may be in the range from about 0.001 to about 100 mg/kg, e.g., from about 0.01 to about 100 mg/kg of body weight per day, such as above about 0.1 mg per kilogram, or in a range of from about 1 to about 10 mg per kilogram body weight of the recipient per day. For example, a suitable dose may be about 0.3 mg/kg, 0.7 mg/kg, 1 mg/kg, 10 mg/kg, or 50 mg/kg of body weight per day.
[0060] In some embodiments, the compounds are administered in unit dosage form; for example, containing 0.05 to 10000 mg, 0.5 to 10000 mg, 5 to 1000 mg, or about 100 mg of active ingredient per unit dosage form.
[0061] In some embodiments, the compounds are administered to achieve peak plasma concentrations of, for example, from about 0.5 to about 75 mM, about 1 to 50 pM, about 2 to about 30 pM, or about 5 to about 25 pM. Exemplary desirable plasma concentrations include at least or no more than 0.25, 0.5, 1, 5, 10, 25, 50, 75, 100 or 200 pM. For example, plasma levels may be from about 1 to 100 micromolar or from about 10 to about 25 micromolar. In some embodiments, this is achieved by the intravenous injection of a 0.05 to 5% solution of the compounds, optionally in saline, or orally administered as a bolus containing about 1-100 mg of the compounds. In some embodiments, desirable blood levels is maintained by continuous infusion to provide about 0.00005-5 mg per kg body weight per hour, for example at least or no more than 0.00005, 0.0005, 0.005, 0.05, 0.5, or 5 mg/kg/hr. In some embodiments, such levels are obtained by intermittent infusions containing about 0.0002-20 mg per kg body weight, for example, at least or no more than 0.0002, 0.002, 0.02, 0.2, 2, 20, or 50 mg of the compounds per kg of body weight.
[0062] In an embodiment, the amount of purine compound used is between 0.1 to 5 mg per kg of the subject per day. In a preferred embodiment, the amount of purine compound used is between 0.2 to 1.3 mg per kg of the subject per day. In a further preferred embodiment, the amount of purine compound used is about 0.3 mg to 0.7 per kg of the subject per day.
[0063] In some embodiments, the compounds are presented in a single dose or as divided doses administered at appropriate intervals, for example, as two, three, four or more sub-doses per day. In some embodiments, the sub-dose itself is further divided, e.g., into a number of discrete loosely spaced administrations; such as multiple inhalations from an insufflator.
[0064] In some embodiments, a pharmaceutical composition of the present invention is packaged in a container with a label, or instructions, or both, indicating use of the pharmaceutical composition in the treatment of the indicated disease.
[0065] It is understood that one or more compounds of this invention may be used independently or in any combination thereof.
Example 1
[0066] The compounds of formulae I, II were screened for GPCR activity using A2aR as the target. For comparison, NECA and SCH 442416 were also screened. NECA is an A2aR agonist. SCH 442416 is an A2aR antagonist. Assay Design: Calcium Mobilization Cell Handling
[0067] Cell lines were expanded from freezer stocks according to standard procedures. Cells were seeded in a total volume of 20 pL into black-walled, clear-bottom, Poly-D-lysine coated 384-well microplates and incubated at 37°C for the appropriate time prior to testing.
Dye Loading
[0068] Assays were performed in 1 x Dye Loading Buffer consisting of lx Dye, lx Additive A and 2.5 mM Probenecid in HBSS / 20 mM Hepes. Probenicid was prepared fresh. Cells were loaded with dye prior to testing. Media was aspirated from cells and replaced with 20 pL Dye Loading Buffer. Cells were incubated for 30-60 minutes at 37°C.
Agonist Format
[0069] For agonist determination, cells were incubated with sample to induce response. After dye loading, cells were removed from the incubator and 10 pL HBSS / 20 mM Hepes was added. 3x vehicle was included in the buffer when performing agonist dose curves to define the EC80 for subsequent antagonist assays. Cells were incubated for 30 minutes at room temperature in the dark to equilibrate plate temperature. Intermediate dilution of sample stocks was performed to generate 4X sample in assay buffer. Compound agonist activity was measured on a FLIPR Tetra (MDS). Calcium mobilization was monitored for 2 minutes and 10 pL 4X sample in HBSS / 20 mM Hepes was added to the cells 5 seconds into the assay. Allosteric Modulation Format
[0070] For allosteric determination, cells were pre-incubated with sample followed by agonist induction at the EC20 concentration. Intermediate dilution of sample stocks was performed to generate 3X sample in assay buffer. After dye loading, cells were removed from the incubator and 10 pL 3x sample was added. Cells were incubated for 30 minutes at room temperature in the dark to equilibrate plate temperature. Vehicle concentration was 1%. Compound allosteric activity was measured on a FLIPRTetra (MDS). Calcium mobilization was monitored for 2 minutes and 10 pL of 4X EC20 agonist in HBSS / 20 mM Hepes was added to the cells 5 seconds into the assay.
Antagonist Format
[0071] For antagonist determination, cells were pre-incubated with sample followed by agonist challenge at the EC80 concentration. Intermediate dilution of sample stocks was performed to generate 3X sample in assay buffer. After dye loading, cells were removed from the incubator and 10 pL 3X sample was added. Cells were incubated for 30 minutes at room temperature in the dark to equilibrate plate temperature. Vehicle concentration was 1%. Compound antagonist activity was measured on a FLIPRTetra (MDS). Calcium mobilization was monitored for 2 minutes and 10 pL EC80 agonist in HBSS / 20 mM Hepes was added to the cells 5 seconds into the assay.
Data Analysis
[0072] Compound activity was analyzed using CBIS data analysis suite (Chemlnnovation, CA). For agonist mode assays, percentage activity is calculated using the following formula:
% Activity =100% x (mean RFU of test sample - mean RFU of vehicle control) / (mean MAX RFU control ligand - mean RFU of vehicle control). [0073] For positive allosteric mode assays, percentage modulation was calculated using the following formula:
% Modulation =100% x ((mean RFU of test sample - mean RFU of EC20 control) / (mean RFU of MAX control ligand - mean RFU of EC20 control)). [0074] For antagonist and negative allosteric modulation mode assays, percentage inhibition is calculated using the following formula:
% Inhibition =100% x (1 - (mean RFU of test sample - mean RFU of vehicle control) / (mean RFU of EC80 control - mean RFU of vehicle control)).
[0075] Figures 1 to 4 present the results of G-protein-coupled receptors (GPCR) screening using A2aR as the target by the GPCR Screening and Profiling Services of Eurofins DiscoverX Corporation.
[0076] In the screening, the activation of the A2aR resulted in calcium mobilization, which was monitored using a calcium-sensitive dye. When the calcium was released, the fluorescence of the dye was increased and the increase was measured in real-time.
[0077] Figures 1 to 4 show percent reacted (Y) versus concentration in mM (X). Compounds were tested in antagonist mode with the requested GPCR Biosensor Assay. For the antagonist assay, data was normalized to the maximal and minimal response observed in the presence of EC80 ligand and vehicle. The following EC80 concentrations were used: ADORA2A Calcium Flux: 0.021 pM NECA.
[0078] FIG. 1 shows the result for the compound of formula I. The ICso was 0.1438 pM. A higher ICso level indicates better binding with A2aR.
[0079] FIG. 2 shows the result for the compound of formula IV, istradefyline. The ICso was 0.0667 pM. [0080] FIG. 3 shows the result for NECA. The half maximal effective concentration (ECso) was 0.0076 mM.
[0081] FIG. 4 shows the result for SCH 442416. The ICso was 0.0113 pM.
Example 2
[0082] The compound of formula I was tested in a study of depression using mice and the forced swim test. The results are shown in Figures 9 and 10.
[0083] The forced swim test, also known as the behavioral despair test, is used to test for depression-like behavior. The test includes placing a rat or mouse inside a cylinder filled with water. 'Floating behavior' (where the animal remains almost immobile and with its head above water) is used as a parameter to analyze 'hopelessness' and thus depression-like behavior. Rodents given antidepressants swim longer than controls. Immobility time is decreased by various antidepressants.
[0084] 32 male CD-I mice (N=8 mice per group) were treated orally with either vehicle (0.3% Tween 80) or test article (2.5-10 mg/kg) of formula I or Istradefylline (aka KW-6002, 2.5 mg/kg) one hour prior to testing. At t=0, mice were placed in a glass cylinder filled with water. After a period of vigorous activity, the mouse would adopt a characteristic immobile posture which is readily scored. Over a 6-minute test session, the latency to first immobility is recorded (in seconds). The duration of immobility (in seconds) during the last 4 minutes of the test is also measured.
[0085] The embodiments of the present invention described above are intended to be examples only. The present invention may be embodied in other specific forms. Alterations, modifications and variations to the invention may be made without departing from the intended scope of the present invention. While the system, devices and processes disclosed and shown herein may comprise a specific number of elements/components, the systems, devices and assemblies could be modified to include addition or fewer of such elements/components. For example, while any of the elements/components disclosed may be referenced as being singular, the embodiments disclosed herein could be modified to include a plurality of such elements/components. Selected features from one or more of the above-described embodiments may be combined to create alternative embodiments not explicitly described. All values and sub-ranges within disclosed ranges are also disclosed. The subject matter described herein intends to cover and embrace all suitable changes in technology. All references mentioned are hereby incorporated by reference in their entirety.

Claims

1. A compound of formula
Figure imgf000023_0001
(I) or a pharmaceutically acceptable salt thereof.
2. A method for treating depression, the method comprising administration of a compound of formula (I)
Figure imgf000023_0002
(I) and pharmaceutically acceptable salts thereof.
3. A method for treating cancer, the method comprising administration of a compound of formula
Figure imgf000024_0001
(I) and pharmaceutically acceptable salts thereof.
4. A method for treating one of more of depression, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer’s and Parkinsons, the method comprising administration of a compound of formula
Figure imgf000024_0002
(I) and pharmaceutically acceptable salts thereof.
5. The method of claims 4 to 6, wherein the administration is oral.
6. The method of claims 4 to 6, wherein the administration is injection.
7. The method of claims 4 to 8, wherein the administration is with a pharmaceutically acceptable excipient.
8. A pharmaceutical composition for treating depression, the composition comprising a compound of formula (I) and pharmaceutically acceptable salts thereof.
Figure imgf000025_0001
9. Use of compound of formula (I), or a pharmaceutically acceptable salt thereof for treating depression, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer's or Parkinsons. 10. A pharmaceutical composition for treating depression, anxiety, multiple sclerosis,
NASH, scleroderma, ADHD, Alzheimer’s or Parkinsons, the composition comprising
Figure imgf000025_0002
11. A method for treating depression, the method comprising administration of one or more compound of formula (II) or (IV)
s ra e y e,
(E)-8-(3,4-Dimethoxystyryl)-l,3-diethyl-7-methylxanthine, of Formula (IV) and pharmaceutically acceptable salts thereof for treating cancer, depression, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer’s.
12. A method for treating cancer, the method comprising administration of a compound of formula
Figure imgf000026_0001
SCH 442416, 2-(2-Furanyl)-7-[3-(4-methoxyphenyl)propyl]-7H-pyrazolo
[4,3-e][1 ,2,4]triazolo[1 ,5-c]pyrimidin-5-amine, of Formula (III), and pharmaceutically acceptable salts thereof.
13. A method for treating one of more of depression, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer’s and Parkinsons, the method comprising administration of one or more compound
Figure imgf000027_0001
NECA, 5'-N-Ethylcarboxamidoadenosine, of
Formula (II), and pharmaceutically acceptable salts thereof.
14. The method of claims 11 to 13, wherein the administration is oral.
15. The method of claims 11 to 13, wherein the administration is injection.
16. The method of claims 11 to 13, wherein the administration is with a pharmaceutically acceptable excipient.
17. A pharmaceutical composition for treating depression, the composition comprising one or more compound of formula (II) and (IV) and pharmaceutically acceptable salts thereof.
18. Use of compound of formula formula (II) NECA, 5'-N-Ethylcarboxamidoadenosine, of Formula (II), or a pharmaceutically acceptable salt thereof for treating depression, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer's or Parkinsons.
19. A pharmaceutical composition for treating depression, anxiety, multiple sclerosis, NASH, scleroderma, ADHD, Alzheimer’s or Parkinsons, the composition comprising
Figure imgf000028_0001
NECA, 5'-N-Ethylcarboxamidoadenosine, of
Formula (II).
20. The method, use or composition of claims 11 to 19, in a dosage of 1-400 mg/day, preferably 10-80 mg/day, more preferably 20-40 mg/day.
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Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
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CN110041331A (en) * 2019-05-17 2019-07-23 广东东阳光药业有限公司 Styryl xanthine derivative and application thereof
WO2020125779A1 (en) * 2018-12-21 2020-06-25 Sunshine Lake Pharma Co., Ltd. 8-substituted styryl xanthine derivatives and uses thereof
WO2020216152A1 (en) * 2019-04-24 2020-10-29 Dongguan Hec New Drug R&D Co., Ltd. 8-substituted aryl vinyl xanthine derivatives and uses thereof

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Publication number Priority date Publication date Assignee Title
WO2013058681A2 (en) * 2011-10-18 2013-04-25 Общество С Ограниченной Ответственностью "Ньювак" (Ооо "Ньювак") Substituted phenoxyacetic acids and esters and amides thereof, comprising a 2,6-dioxo-2,3,6,7-tetrahydro-1н-purine-8-yl fragment and constituting а2a adenosine receptor antagonists, and the use thereof
WO2020125779A1 (en) * 2018-12-21 2020-06-25 Sunshine Lake Pharma Co., Ltd. 8-substituted styryl xanthine derivatives and uses thereof
WO2020216152A1 (en) * 2019-04-24 2020-10-29 Dongguan Hec New Drug R&D Co., Ltd. 8-substituted aryl vinyl xanthine derivatives and uses thereof
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