WO2014011754A1

WO2014011754A1 - Novel 1, 4-benzoxazinone compounds

Info

Publication number: WO2014011754A1
Application number: PCT/US2013/049899
Authority: WO
Inventors: Amir Tamiz; Aaron H. HEIFETZ
Original assignee: Amir Tamiz; Heifetz Aaron H
Priority date: 2012-07-10
Filing date: 2013-07-10
Publication date: 2014-01-16

Abstract

The present invention provides novel benzoxazinone compounds, pharmaceutical compositions comprising such compounds, and methods of treating disease using such compositions. Pharmaceutical compositions of the invention are particularly useful in the treatment of neurodegenerative diseases.

Description

NOVEL 1, 4- BENZOXAZINONE COMPOUNDS

[0001] The present invention relates compounds and compositions comprising such compounds that may be used to treat a variety of neurodegenerative conditions such as, for example, Alzheimer's disease, Parkinson's disease, Amyotropic lateral sclerosis (ALS), or Huntington's disease, and conditions such as ischemic stroke and traumatic brain injury.

[0002] Neurodegenerative diseases are characterized by an aberrant loss of neurons.

The loss is associated with neurofibrillary tangles, neuritic plaques, and neuronal cell death. There are currently no effective strategies to prevent the neuronal cell death in these pathologies.

[0003] There remains a need in the art for compounds and compositions capable of reducing the rate of and/or preventing neuronal death such as that associated with neurodegenerative diseases. This need and others are met by the present invention.

SUMMARY OF THE INVENTION

[0004] The present invention provides materials and methods to reduce the rate of and/or to prevent neuronal cell death, for example, the neuronal death associated with neurodegenerative diseases. The present invention provides novel 1,4-benzoxazinone compounds that have the ability to reduce the rate of and/or prevent neuronal cell death. The present invention further comprises compositions comprising the novel compounds, for example, pharmaceutical compositions. Compositions of the invention may comprise one or more pharmaceutically acceptable excipients, for example, salts, buffers, binders, sugars and the like.

[0005] Compositions of the invention may be formulated for any type of delivery.

For example, compositions of the invention may be formulated for intestinal delivery, e.g., may be delayed release compositions. Compositions of the invention may be formulated for pulmonary delivery.

[0006] The present invention also provides method of preventing, ameliorating and/or treating neurodegenerative diseases comprising administering a composition of the invention to a subject in need thereof. As used herein, a "subject" may be any mammal, for example, a human. Neurodegenerative diseases that may be treated according to the present invention include, but are not limited to, Alzheimer's disease, Parkinson's disease, Huntington's disease, and amyotropic lateral sclerosis. The present invention also provides methods of preventing, ameliorating and/or treating conditions that may result in neuronal cell death comprising administering a composition of the invention to a subject in need thereof. Such conditions include, but are not limited to, ischemic stroke and traumatic brain injury.

BRIEF DESCRIPTION OF THE DRAWINGS

[0007] Figures 1A, IB and 1C provide the structure of some compounds discussed in the present application.

[0008] Figure 2 provides micrographs of HT-22 cells treated with homocysteic acid and various compounds described herein.

[0009] Figure 3 is a bar graph showing % survival of cells treated with homocysteic acid (HCA) in the presence of various compounds of the invention.

[0010] Figure 4 is a bar graph showing % survival of cells treated with homocysteic acid (HCA) in the presence of various compounds of the invention.

DETAILED DESCRIPTION

[0011] Compounds of the invention are modified benzoxazinones as exemplified below.

[0012] In some embodiments, com ounds of the invention have the structure I

[0013] Where Rl is selected from the group consisting of: hydrogen, C1-C6 alkyl which may be straight chained, branched or cyclic; substituted C1-C6 alkyl which may be straight chained, branched or cyclic; C1-C6 alkenyl which may be straight chained, branched or cyclic; or substituted C1-C6 alkenyl which may be straight chained, branched or cyclic. Suitable substitutions include, but are not limited to, one or more halogens, one or more hydroxyl groups, one or more amino groups and combinations thereof. Preferred groups include methyl , ethyl, propyl, isopropyl and cyclopropyl.

[0014] R2 and R3 are independently selected from the group consisting of: hydrogen;

C1-C6 alkyl which may be straight chained, branched or cyclic; substituted C1-C6 alkyl which may be straight chained, branched or cyclic; C1-C6 alkenyl which may be straight chained, branched or cyclic; or substituted C1-C6 alkenyl which may be straight chained, branched or cyclic. R2 and R3 taken together may form a ring containing from 3 to 5 carbon atoms and optionally one or more heteroatoms.

Suitable substitutions include, but are not limited to, one or more halogens, one or more hydroxyl groups, one or more amino groups and combinations thereof. In some embodiments R2 and/or R3 may be methyl, hydroxymethyl, ethyl, hydroxyethyl, propyl, hydroxypropyl, isopropyl, hydroxyisopropyl, cyclopropyl, or

hydroxycyclopropyl.

[0015] Aryl is selected from the group consisting of phenyl and substituted phenyl.

Suitable substitutions include, but are not limited to, one or more halogens, one or more hydroxyl groups, one or more amino groups and combinations thereof. Aryl comprises mono, di and tri substituted phenyl groups. In some embodiments, aryl may be a monosubstituted phenyl group that may be substituted at the 2, 3, or 4 position. Some preferred embodiments include, but are not limited to, halo substituted phenyl, for example, chlorophenyl, fluorophenyl, and bromophenyl such as 4-halo substituted phenyl, for example, 4-chlorophenyl, 4-fiuorophenyl, and 4- bromophenyl. In other embodiments, aryl includes but is not limited to, alkyl substituted phenyl, for example, methylphenyl and ethylphenyl, for example, 4- methylphenyl or 4-ethylphenyl. In some embodiments, aryl includes haloalkyl substituted phenyl, for example, halomethyl and haloethyl. Examples include, but are not limited to, fluoromethyl, difluoromethyl, trifuoromethyl , chloromethyl, dichloromethyl and trichloromethyl. Preferred examples include, but are not limited to, 4-trichlormethyl and 4-trifluoromethyl substituted phenyl.

[0016] In some embodiments, aryl may be a disubstituted phenyl group that may be substituted at two of the 2, 3, 4, 5 or 6 positions. Some preferred embodiments include, but are not limited to, dihalo substituted phenyl, for example, dichlorophenyl, difluorophenyl, and dibromophenyl such as 3,4- or 3,5-dihalo substituted phenyl, for example, 3,4- or 3,5-dichlorophenyl, 3,4- or 3,5-difluorophenyl, and 3,4- or 3,5- dibromophenyl. In other embodiments, aryl includes but is not limited to, dialkyl substituted phenyl, for example, dimethylphenyl and diethylphenyl, for example, 3,4- or 3,5-dimethylphenyl or 3,4- or 3,5-diethylphenyl. In some embodiments, aryl includes dihaloalkyl substituted phenyl, for example, dihalomethyl and dihaloethyl. Examples include, but are not limited to,di-fluoromethyl,di- difluoromethyl,di- trifuoromethyl , di-chloromethyl,di- dichloromethyl and di-trichloromethyl. Preferred examples include, but are not limited to, 3,4- or 3,5-trichlormethyl and 3,4- or 3,5- trifluoromethyl substituted phenyl.

[0017] In one embodiment, l is hydrogen, R2 is hydrogen, R3 is hydrogen, and aryl is 3, 5, dibromo-phenyl.

[0018] In one embodiment, Rl is hydrogen, R2 is hydrogen, R3 is hydrogen, and aryl is 4-chloro-phenyl.

[0019] In one embodiment, Rl is hydrogen, R2 is hydrogen, R3 is hydrogen, and aryl is 4-triflouromethyl-phenyl.

[0020] In one embodiment, Rl is hydrogen, R2 is hydrogen, R3 is acetyl, and aryl is 4-chlorophenyl.

[0021] In one embodiment, Rl is hydrogen, R2 is methyl, R3 is methyl, and aryl is 4- chlorophenyl.

[0022] In one embodiment, Rl is hydrogen, R2 and R3 together form a piperazine ring, and aryl is 4-chloro.

[0023] In one embodiment, Rl is hydrogen, R2 and R3 are both hydroxyethyl, and aryl is 4-chloro-phenyl.

[0024] In one embodiment, Rl is methyl, R2 and R3 are both hydrogen, and aryl is 4- chlorophenyl. This compound is termed Compound F, Figure 1A..

[0025] In one embodiment, Rl is cyclopropyl, R2 and R3 are both hydrogen, and aryl is 4-chlorophenyl. SYNTHESIS OF COMPOUNDS OF THE INVENTION

[0027] The following scheme can be used to synthesize benzoxazinone compounds of the invention with various aryl groups.

[0028] Scheme I

Chemistry:

[0029] This scheme can be used to introduce modifications in the aryl group. 6- Nitro-2H-l,4-benzoxazin-3(4H)-one (Aldrich catalog number 662313) can be condensed with the desired benzaldehyde and then the nitro group reduced to yield compounds of the invention.

[0030] The following scheme can be used to synthesize compounds of the invention having various modifications to the benzoxazinone structure.

[0031] Scheme 2

[0032] This scheme can be used to introduce various modifications into the

benzoxazine ring structure. The appropriate 2-amino-phenol (e.g., 2-amino-4- nitrophenol Aldrich catalog number A70402) can be condensed with methyl bromoacetate (Aldrich catalog number 157910) and the product condensed with the appropriate benzaldehyde and treated as above. [0033] This scheme can be used to introduce modifications to the ring nitrogen of the benzoxazinone of the invention.

[0034] Scheme 3

[0035] Compositions

[0036] Typically, compositions, such as pharmaceutical compositions, comprising a compound of the invention, comprise a pharmaceutically effective amount of the compound. The pharmaceutically effective amount of compound (e.g., a

benzoxazinone of the invention) employed in any given composition may vary according to factors such as the disease state, age, sex, and weight of the individual. Dosage regimens may be adjusted to provide the optimum therapeutic response. For example, a single bolus may be administered, several divided doses may be administered over time or the dose may be proportionally reduced or increased as indicated by the exigencies of the therapeutic situation. Generally, the amount of compound used for preventing, ameliorating and/or treating a disease in a subject will be in the range of about 1.0 μg to 1 g, preferably about 1 mg to about 1000 mg, or from about 10 mg to about 100 mg, or from about 10 mg to about 50 mg, or from about 10 mg to about 25 mg of compound.

[0037] Compositions of the invention may comprise one or more benzoxazinone compounds at a level of from about 0.1 wt% to about 20 wt%, from about 0.1 wt% to about 18 wt%, from about 0.1 wt% to about 16 wt%, from about 0.1 wt% to about 14 wt%, from about 0.1 wt% to about 12 wt%, from about 0.1 wt% to about 10 wt%, from about 0.1 wt% to about 8 wt%, from about 0.1 wt% to about 6 wt%, from about 0.1 wt% to about 4 wt%, from about 0.1 wt% to about 2 wt%, from about 0.1 wt% to about 1 wt%, from about 0.1 wt% to about 0.9 wt%, from about 0.1 wt% to about 0.8 wt%, from about 0.1 wt% to about 0.7 wt%, from about 0.1 wt% to about 0.6 wt%, from about 0.1 wt% to about 0.5 wt%, from about 0.1 wt% to about 0.4 wt%, from about 0.1 wt% to about 0.3 wt%, or from about 0.1 wt% to about 0.2 wt% of the total weight of the composition. Compositions of the invention may comprise one or more benzoxazinone compounds at a level of about 0.1 wt%, about 0.2 wt%, about 0.3 wt%, about 0.4 wt%, about 0.5 wt%, about 0.6 wt%, about 0.7 wt%, about 0.8 wt%, or about 0.9 wt% based on the total weight of the composition.

[0038] Compositions of the invention may comprise one or more benzoxazinone compounds at a level of from about 1 wt% to about 20 wt%, from about 1 wt% to about 18 wt%, from about 1 wt% to about 16 wt%, from about 1 wt% to about 14 wt%, from about 1 wt% to about 12 wt%, from about 1 wt% to about 10 wt%, from about 1 wt% to about 9 wt%, from about 1 wt% to about 8 wt%, from about 1 wt% to about 7 wt%, from about 1 wt% to about 6 wt%, from about 1 wt% to about 5 wt%, from about 1 wt% to about 4 wt%, from about 1 wt% to about 3 wt%, or from about 1 wt% to about 2 wt% of the total weight of the composition

[0039] Compositions of the invention may formulated for pulmonary delivery (e.g., may be pulmonary dosage forms). Typically such compositions may be provided as pharmaceutical aerosols, e.g., solution aerosols or powder aerosols. Those of skill in the art are aware of many different methods and devices for the formation of pharmaceutical aerosols, for example, those disclosed by Sciarra and Sciarra, Aerosols, in Remington: The Science and Practice of Pharmacy, 20th Ed., Chapter 50, Gennaro et al. Eds., Lippincott, Williams and Wilkins Publishing Co., (2000).

[0040] The compositions of the invention may be formulated for enteric delivery, for example, may comprise one or more coatings, for example, delayed release coating containing one or more enteric agents. A delayed release coating is typically substantially stable in gastric fluid and substantially unstable (e.g., dissolves rapidly or is physically unstable) in intestinal fluid, thus providing for substantial release of the benzoxazinone compound from the composition in the duodenum or the jejunum.

[0041] The terms "stable in gastric fluid" or "stable in acidic environments" refers to a composition that releases 30% or less by weight of the total benzoxazinone compound in the composition in gastric fluid with a pH of 5 or less, or simulated gastric fluid with a pH of 5 or less, in approximately sixty minutes. Examples of simulated gastric fluid and simulated intestinal fluid include, but are not limited to, those disclosed in the 2005 Pharmacopeia 23NF/28USP in Test Solutions at page 2858 and/or other simulated gastric fluids and simulated intestinal fluids known to those of skill in the art, for example, simulated gastric fluid and/or intestinal fluid prepared without enzymes.

[0042] Compositions of the of the invention may release from about 0% to about 30%, from about 0% to about 25%, from about 0% to about 20%, from about 0% to about 15%, from about 0% to about 10%, 5% to about 30%, from about 5% to about 25%, from about 5% to about 20%, from about 5% to about 15%, from about 5% to about 10% by weight of the total benzoxazinone compound in the composition in gastric fluid with a pH of 5, or less or simulated gastric fluid with a pH of 5 or less, in approximately sixty minutes. As use herein, "about" used to modify a numerical value means within 10% of the value. Compositions of the invention may release about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, or about 10% by weight of the total benzoxazinone compound in the composition in gastric fluid with a pH of 5 or less, or simulated gastric fluid with a pH of 5 or less, in approximately sixty minutes.

[0043] The term "unstable in intestinal fluid" refers to a composition that releases 70% or more by weight of the total benzoxazinone compound in the composition in intestinal fluid or simulated intestinal fluid in approximately sixty minutes. The term "unstable in near neutral to alkaline environments" refers to a composition that releases 70% or more by weight of the total amount of benzoxazinone compound in the composition in intestinal fluid with a pH of 5 or greater, or simulated intestinal fluid with a pH of 5 or greater, in approximately ninety minutes. For example, a composition that is unstable in near neutral or alkaline environments may release 70% or more by weight of a compound of the invention in a fluid having a pH greater than about 5 (e.g., a fluid having a pH of from about 5 to about 14, from about 6 to about 14, from about 7 to about 14, from about 8 to about 14, from about 9 to about 14, from about 10 to about 14, or from about 11 to about 14) in from about 5 minutes to about 90 minutes, or from about 10 minutes to about 90 minutes, or from about 15 minutes to about 90 minutes, or from about 20 minutes to about 90 minutes, or from about 25 minutes to about 90 minutes, or from about 30 minutes to about 90 minutes, or from about 5 minutes to about 60 minutes, or from about 10 minutes to about 60 minutes, or from about 15 minutes to about 60 minutes, or from about 20 minutes to about 60 minutes, or from about 25 minutes to about 90 minutes, or from about 30 minutes to about 60 minutes.

[0044] In addition to benzoxazinone compounds, compound, compositions of the invention may further comprise one or more therapeutic agents. Therapeutic agents include, but are not limited to, steroids and other anti-inflammatory compounds. Suitable therapeutic agents may include one or more of aminosalicylates, corticosteroids, immunomodulators, antibiotics, cytokines, chemokines and biologic therapies.

[0045] Compositions of the invention may also comprise one or more

pharmaceutically acceptable excipients. Suitable excipients include, but are not limited to, buffers, buffer salts, bulking agents, salts, surface active agents, acids, bases, sugars, and binders.

[0046] Methods of use

[0047] The compositions of the invention can be used for preventing, slowing the onset of, ameliorating and/or treating any disease associated with neurodegeneration or neuronal cell death. Specific examples of disease of this type include, but are not limited to, Alzheimer's disease, Parkinson's disease, Amyotropic lateral sclerosis (ALS), or Huntington's disease, and conditions such as ischemic stroke and traumatic brain injury.

[0048] It will be readily apparent to one of ordinary skill in the relevant arts that other suitable modifications and adaptations to the methods and applications described herein are obvious and may be made without departing from the scope of the invention or any embodiment thereof. Having now described the present invention in detail, the same will be more clearly understood by reference to the following examples, which are included herewith for purposes of illustration only and are not intended to be limiting of the invention. EXAMPLES

EXAMPLE 1

[0049] Homocysteic Acid (HCA) Assay

[0050] Treatment of the mouse neuroblastoma HT-22 cell line with homocysteic acid (HCA) induces apoptosis through glutathione depletion and oxidative stress. The mouse HT-22 neuroblastoma cell line was purchased from ATCC (Manassas, VA, USA) and cultured in Dulbecco's modified Eagle's medium (DMEM) with 4.5 g/L glucose (without sodium pyruvate) supplemented with 10% FBS, 100 units/ml penicillin and 100 μg/ml streptomycin. Cultures were plated at ~ 30% confluence for HCA treatment. HCA was made as 150 mM stock solution adjusted to pH 7.5 and used at a final concentration of 1.5 mM.

[0051] HT-22 cells were treated with no additives (Un), 1.5 mM HCA, or 1.5 mM HCA plus various concentrations of the test compound. Appearance of the cultures at 24 hr after treatment was visualized by phase-contrast microscopy (Phase). Cell death was evaluated using DAPI staining. Viability of the cells is expressed as percentage of untreated cultures (control).

[0052] The viability status of neuronal cultures was evaluated by phase-contrast microscopy and quantified by staining cell nuclei with DAPI. Briefly, the cells were fixed in 4% paraformaldehydefor 20 min at 48^°C. After washing in phosphate- buffered saline, diamidino-2-phenylindole hydrochloride (DAPI; 1 μg/ml in phosphate-buffered saline) was added for 15 min at room temperature and viewed under ultraviolet light (260 nm).

[0053] Structures of various compounds are shown in Figure 1A and IB. The

micrographs used to analyze cell viability are shown in Figure 2. Figures 3 and 4 provide bar graphs quantifying the results of the HCA assay.

[0054] While the foregoing invention has been described in some detail for purposes of clarity and understanding, it will be appreciated by one skilled in the art from a reading of this disclosure that various changes in form and detail can be made without departing from the true scope of the invention and appended claims. All patents and publications cited herein are entirely incorporated herein by reference.

Claims

What is claimed is:

1. A compound of formula I, wherein

I

Rl is selected from the group consisting of: hydrogen, C1-C6 alkyl which may be straight chained, branched or cyclic; substituted C1-C6 alkyl which may be straight chained, branched or cyclic; C1-C6 alkenyl which may be straight chained, branched or cyclic; or substituted C1-C6 alkenyl which may be straight chained, branched or cyclic;

R2 and R3 are independently selected from the group consisting of: hydrogen; C1-C6 alkyl which may be straight chained, branched or cyclic; substituted C1-C6 alkyl which may be straight chained, branched or cyclic; C1-C6 alkenyl which may be straight chained, branched or cyclic; or substituted C1-C6 alkenyl which may be straight chained, branched or cyclic, R2 and R3 taken together may form a ring containing from 3 to 5 carbon atoms and optionally one or more heteroatoms; and

Aryl is selected from the group consisting of phenyl, mono, di, or tri substituted phenyl that may be substituted at the 2, 3, 4, and/or 5 position.

2. A compound according to claim 1, wherein Rl is hydrogen, R2 is hydrogen, R3 is hydrogen, and aryl is 2, 5, dibromo-phenyl.

3. A compound according to claim 1, wherein Rl is hydrogen, R2 is hydrogen, R3 is hydrogen, and aryl is 4-chloro-phenyl.

4. A compound according to claim 1, wherein Rl is hydrogen, R2 is hydrogen, R3 is hydrogen, and aryl is 4-triflouromethyl-phenyl.

5. A compound according to claim 1, wherein Rl is hydrogen, R2 is hydrogen, R3 is acetyl, and aryl is 4-chlorophenyl.

6. A compound according to claim 1, wherein Rl is hydrogen, R2 is methyl, R3 is methyl, and aryl is 4-chlorophenyl.

7. A compound according to claim 1, wherein Rl is hydrogen, R2 and R3 together form a piperazine ring, and aryl is 4-chloro.

8. A compound according to claim 1, wherein Rl is hydrogen, R2 and R3 are both hydroxyethyl, and aryl is 4-chloro-phenyl.

9. A compound according to claim 1, wherein Rl is methyl, R2 and R3 are both hydrogen, and aryl is 4-chlorophenyl.

10. A compound according to claim 1, wherein Rl is cyclopropyl, R2 and R3 are both hydrogen, and aryl is 4-chlorophenyl.

11. A composition comprising a compound according to any one of claims 1-10, and further comprising a pharmaceutically acceptable excipient.

12. A method of ameliorating, treating and/or preventing a neurodegenerative disease, comprising administering a composition according to claim 11 to a subject in need thereof.

13. A method of treating ischemic stroke and/or traumatic brain injury, comprising administering a composition according to claim 11 to a subject in need thereof.