WO2019046534A1 - Deuterated indoloquinoline compounds - Google Patents

Abstract

The invention is directed to novel quinone reductase 2 (QR2) inhibitors represented by Formula (I): and pharmaceutically acceptable salts thereof. The variables for Formula (I) are defined herein and the compound comprises at least one deuterium atom. The invention also provides compositions comprising a compound of this invention and the use of such compositions in methods of preventing and/or therapeutically treating diseases and conditions that are beneficially prevented and/or treated by administering a QR2 inhibitor.

Description

DEUTERATED INDOLOQ UINOLINE COMPOUNDS CROSS REFERENCE TO RELATED APPLICATION

This application claims the benefit of priority of U.S. Provisional Application No. 62/552,540, filed August 31, 2017, the disclosure of which is incorporated herein by reference in its entirety.

BACKGROUND OF THE INVENTION

Melatonin, synthesized by the pineal gland, is a neurohormone which circulates in the blood of humans at higher concentrations during the night in order to transmit circadian rhythm to peripheral tissue via 2 high-affinity melatonin receptor binding sites (MTi and MT₂). A third significantly lower- affinity melatonin binding site (MT₃) was properly identified in 1997 (Zhao, et al., Proc Nat Acad Sci 1997, 94: 1669-1674) as quinone reductase 2 (QR2) with the same protein sequence/structure as MT₃. QR2, originally described in 1961 (Liao, S, and Williams -Ashman, H,: Biochem Biophys Res Commun 1961 4: 208-213), is a cytosolic enzyme closely related to quinone reductase 1 (QR1) but whose physiological function has been difficult to definitively clarify. Fu, et al. (J Biol Chem 2008, 283(35): 23829-23835 ) describe it as a catecholamine reductase and thus, it might be concluded that QR2 also effects the levels of such neurotransmitters as dopamine and norepinephrine. Melatonin is a moderate-to-weak inhibitor of the functional activity of QR2 with an IC₅₀ typically in the range of 10-100 μΜ (e.g., Calamini, B, et al., Biochem J 2008, 413: 81-91 ("Calamini 2008"); Ferry, G, et al., Chem Biol Interact 2010, 186: 103-109; Mailliet, F., Biochem Pharmacol 2005, 71: 74-88; Pegan, S, Protein Sci 2011, 20: 1182- 1195).

QR2 is a homo log of QR1, but differs in that it uses N-ribosyl- and N- alkyldihydronicotinamides as co-substrates rather than NADH or NADPH. Although the function of QR2 is not as well defined, the function of QR1 seems unquestionably to be that of detoxification. On the other hand, "QR2 may actually transform certain quinone substrates into more highly reactive compounds/radicals capable of causing cellular damage" (Calamini 2008 at p 81). Furthermore, specific inhibitors of QR2 have been shown to prevent the formation of reactive oxygen species by QR2, in vitro (Reybier et. al., Free Radic Res 2011, 45 (10): 1184-1195 ("Reybier 2011")).

Animal studies targeting QR2 functions have strongly linked this enzyme to cognition and neurodegenerative diseases. QR2 knock-out rodent models have demonstrated enhanced learning abilities as compared with their wild-type equivalents (Benoit, C, et al, J NeuroSci 2010, 30(38): 12690-12700 ("Benoit")), and QR2 mRNA expression level was found to be notably higher in the hippocampus of aged, memory- imp aired rats than in aged, memory- unimpaired rats (Brouillette, J and Quirion, R, Neurobiol Aging 2008, 29: 1721-1732).

Treatment with QR2 inhibitors was shown to reverse cognitive deficits in scopolamine- treated rats (Benoit) and improved learning in a mouse model of vascular dementia

(Fitzpatrick, J, et al, Neurology 2014, 83 (10 supp. PI): 237).

Likewise, a number of human studies have also correlated QR2 to diseases with cognitive impairment. A genetic mutation in the D-allele of the promoter region of human QR2 leads to increased expression of QR2. A higher prevalence of this mutation has been detected in patients with certain neurodegenerative diseases, such as Parkinson's disease (Wang, W, et al, J Gerontol 2008, 63A (2): 127-134; Harada, S, et al, Biochem Biophys Res Commun 2001, 288 (4): 887-892), alcohol withdrawal symptoms (Okubo T, et al, Alcohol Clin Exp Res 2003, 27: 68-71) and schizophrenia (Harada, S, et al, Psychiatr Genet 2003, 13: 205-209). QR2 has been shown to be overexpressed in the cortex of Alzheimer's Disease (AD) patients (Rappaport, A, et al, J Neurosci 2015, 35 (47): 15568-15581) and the level of QR2 was found to be significantly higher in the hippocampus of AD patients than in control subjects (Hashimoto, T and Nakai, M, Neurosci Lett 2011, 502: 10-12). The administration of melatonin, a QR2 inhibitor, to AD patients improved cognition as compared with AD subjects receiving placebo (Asayama, K, et al, J Nippon Med Sch 2003, 70 (4): 334-341) although this effect may have been due to melatonin's effect at the MTi and MT₂ binding sites.

Ambocarb (also known as carbacetam), a compound studied in the Ukraine, was established to be highly neuroprotective in rodent models of traumatic shock and hypoxia (I. V. Komissarov, A. V. Titevsky, 1. 1. Abramets, V. I. Dulenko, Yu. A. Nikolyukin, A. V. Kibal'nyi, T. A. Groshevoi, G. F. Sukhovoi, RU Pat. 2064793 [in Russian]; V. I. Dulenko, I. V. Komissarov, Yu. A. Nikolyukin, A. V. Kibal'nyi, A. V. Titevsky, E. Y. Leshchinskaya, UA Pat. 24293 [in Russian]). Similarly, a recent series of publications also demonstrated that treatment with ambocarb (carbacetam) resulted in an improved outcome in a polytrauma model in the rat, specifically with regard to the heart, liver, lungs, immune system and serum parameters (Kozak, D, Clinical Surgery 2014, (1), 40-42 [in Ukrainian] ; Kozak, D, and Volkov, K, Theoretical Medicine (Uzhgorod University publication) 2014, 2 (50), 3-6 [in Ukrainian]; Kozak, D, Medicinal and Clinical Chemistry 2015, 17 (3), 38-41 [in Ukrainian] ; Kozak, D, Advances in Clinical and Experimental Medicine 2015, (2-3), 58-60 [in

Ukrainian]) .

In addition to the positive outcomes in shock models, ambocarb was reported to enhance learning in rats (Titievsky, A, et al, Pharmacol Biochem Behav 1994, 47 (3) 681- 688) and was shown to be effective in patients with cognitive impairment and attention- span deficit, including patients with residual deficits from traumatic brain injury (Kut'ko, I, and Sitchenko, N, Lik Sprava 1995, 9-12: 96-98 [in Russian] ; Komissarov, I, and Leshchinskaya, E, Zh Akad Medychn Nauk Ukrainy 1998, 4: 199-215). The therapeutic target(s) of ambocarb at the time of these studies was not clearly established and was reported to be possibly related to NMDA receptor-mediated mechanisms (Abramets, I, et al., Neurophysiol 1993, 25 (3) 179-184), AMPA/kainate receptor-mediated mechanisms (Abramets, I, et al., Neurophysiol 1994, 26 (5) 365-372) and/or GABA_A receptor-mediated mechanisms

(Kopanitsa, M, et al., Clin Exp Pharm Physiol 2000, 27: 46-54). However, it was recently reported that ambocarb is potent at displacing 125 I-melatonin from the melatonin receptor subtype MT₃, with an IC₅₀ ~5 nM ( Shcherbakova, I, Chem Heterocycl Compd 2013, 49 (1) 2-18 ("Shcherbakova 2013"))· These data suggest that one of the key mechanism of action of ambocarb responsible for the observed effects in animal and clinical studies may in fact be related to inhibition of QR2 enzyme activity.

It is well characterized that following brain injury (TBI) or ischemic stroke there is a release of glutamate from the neuronal cells of the brain and a cascade of effects such as an increase in free radical formation and an increase in intracellular calcium (Prins, M, et. al., Dis Model Mech 2013, 6 (6), 1307-1315). In addition to QR2 inhibition, Shcherbakova (2013) reported that ambocarb and structurally similar compounds are inhibitors of neuronal L-type calcium channels. It was recently demonstrated that inhibitors of neuronal L-type calcium channels protect against ischemic brain injury in gerbils and rats (Hicks CA et al., Eur J Pharm, 2000, 408, 241-248). As stated previously, QR2 inhibitors have been shown to reduce free radical formation (Reybier 2011). Therefore, the combination of anti-oxidant activity and neuronal L-type calcium channel inhibition may be responsible for the neuroprotective effects or improvement of the residual effects of TBI that were demonstrated by ambocarb (referenced above). In addition, it was recently concluded in a series of publications by Starodubska et al., and Ziablitsev et al. that ambocarb (carbacetam) improved the cognitive deficit caused by TBI in rats (Starodubska et al., Advances in Clinical and Experimental Medicine 2017, (1) 76-79 [in Ukrainian] ; Starodubska et al., Current Issues in Modern Medicine 2017, 17 (2, No 58),50-54 [in Ukrainian]; Ziablitsev et al, Pathology 2017, 14 (1 No. 39), 95-99 [in Ukrainian] ; Ziablitsev et al, Journal Education Health Sport 2017, 7(1) 525-533, Ziablitsev et al, Morphologia 2017, 11 (2), 12-18 [in Ukrainian] ;

Ziablitsev et al, Current Issues in Modern Medicine 2017, 17 (3 No. 59), 25-29 [in

Ukrainian]; Ziablitsev et al, Trauma 2017, 18(2) 53-58 [in Ukrainian]).

In addition to the numerous reports referenced above on the effect of QR2 inhibition on cognitive performance, there are also reports of QR2 involvement in other disease conditions. In one report it was concluded that high levels of QR2 resulting from a genetic polymorphism may also be the cause of early tamoxifen withdrawal of breast cancer patients receiving adjuvant therapy (Jamieson, D, et al, Pharmacogenet Genomics 2011, 21 (12), 808-819). Therefore, co-administration of QR2 inhibitors may be effective in allowing these patients to maintain their breast cancer adjuvant therapy. In other reports, it has been shown that anti-malarials, such as the quinoline class (eg. chloroquine) and the indolone class, interact with QR2 either as inhibitors or substrates suggesting that QR2 inhibition may impart anti-malarial activity (Graves, P, et al, Mol Pharmacol 2002, 62, 1364-1372; Leung, K, and Shilton, B, J Biol Chem 2013, 288 (16), 11242-11251).

There are currently no satisfactory therapies available for the cognitive deficits resulting from TBI or from dementia in an ever-aging population. The studies summarized above suggest that novel QR2 inhibitors with an improved pharmacokinetic profile might be beneficial for the health of patients who suffer from attention-span deficit and/or decreased cognition, or from other diseases in which QR2 may be involved. There is a continuing need for new compounds to treat the aforementioned diseases and conditions.

SUMMARY OF THE INVENTION

This invention relates to novel quinone reductase 2 (QR2) inhibitors, and

pharmaceutically acceptable salts thereof. This invention also provides compositions comprising a compound of this invention and the use of such compositions in methods of preventing and/or therapeutically treating diseases and conditions that are beneficially prevented and/or treated by administering a QR2 inhibitor.

DETAILED DESCRIPTION OF THE INVENTION

The present disclosure features novel inhibitors of the QR2 enzyme for treating, preventing and/or delaying the onset and/or development of diseases and disorders of the central nervous system. "Inhibitors of the QR2 enzyme" refer to compounds able to inhibit the functional activity of the QR2 enzyme. The ability of a compound to "inhibit the QR2 enzyme" means that the compound causes a decrease in one or more of the enzyme activities evoked by the QR2 enzyme. For example, melatonin, an endogenous neurohormone, is a weak inhibitor of the QR2 enzyme with an IC₅₀ in the mid-μΜ range, depending on assay conditions. The compounds disclosed herein, as well as such compounds as iodo-melatonin and resveratrol have been shown to have IC₅₀ values in the low-μΜ to high-nM range, depending on the assay conditions.

The use of inhibitors of the QR2 enzyme may achieve a beneficial effect in a subject, as described herein. More specifically, the present disclosure demonstrates the ability of the compounds, which are inhibitors of the QR2 enzyme, to achieve improvement in cognitive function.

The term "cognitive function" is related to any mental process or state that involves but is not limited to learning, creation of imaginary thinking, awareness, reasoning, spatial ability, speech and language skills, language acquisition and capacity for judgment attention. Cognitive function is formed in multiple areas of brain such as hippocampus, cortex and other brain structures. It is assumed that long term memories are stored at least in part in cortex, and it is known that sensory information is acquired, consolidated and retrieved by a specific cortical structure, the gustatory cortex, which resides within the insular cortex.

Another region that has been shown to be responsible for memory formation and long term storage is hippocampus.

In humans, cognitive function can be measured by any known method, for example and without limitation, by the Clinical Global Impression of Change Scale (CIBIC-plus scale), the Mini Mental State Exam (MMSE), the Neuropsychiatric Inventory (NPI), the Clinical Dimentia Rating Scale (CDR), the Cambridge Neuropsychological Test Automated Battery (CANTAB) or the Sandoz Clinical Assessment-Geriatric (SCAG). Cognitive function may also be measured indirectly using imaging techniques such as Positron Emission Tomography (PET), functional Magnetic Resonance Imaging (fMRI), Single Photon

Emission Computed Tomography (SPECT) or any other imaging technique that allows brain function measurements. Cognitive and mental function for mental disorders such as Attention Deficit Disorder/ Attention Deficit Hyperactivity Disorder (ADD/ ADHD) can be measured based upon the Diagnostic and Statistical Manual of Mental Disorders (DSM-5) criteria and other screening measures for ADD/ ADHD. An improvement of one or more processes affecting the cognitive function in a subject will signify an improvement of the cognitive function in said subject. In certain embodiments, improving the cognitive function comprises improving learning, plasticity and/or memory.

The term "learning" relates to acquiring or gaining new, or modifying and reinforcing existing knowledge, behaviors, skills, or preferences.

The term "plasticity" relates to synaptic plasticity, brain plasticity or neuroplasticity associated with the ability of the brain to change with learning, and to change the already acquired memory. One measurable parameter reflecting plasticity is memory extinction.

The term "memory" relates to the process in which information is encoded, stored, and retrieved. In this disclosure, the term "memory" refers to all categories of human memory including sensory memory, short-term (or working) memory and long-term memory. The term "long-term memory" includes explicit (or conscious/declarative) memory and implicit (or subconscious/procedural) memory.

The term "subject" refers to either a human or a non-human animal.

Also described herein are techniques which may be used to obtain additional compounds substituted with deuterium as inhibitors of the QR2 enzyme.

The term "isotopic enrichment factor" at a particular position normally occupied by hydrogen means that the ratio between the abundance of deuterium at the position and the natural abundance of hydrogen at that position. By way of example, an isotopic enrichment factor of 3500 means that the amount of deuterium at the particular position is 3500 fold greater than natural abundance, or that 52.5% of the compounds have deuterium at the particular position (i.e., 52.5% deuterium incorporation at the given position).

When a particular position in a compound of the invention is designated by name or structure as containing hydrogen or deuterium, it is to be understood that the position can contain hydrogen at its natural abundance or can be enriched in deuterium with an isotopic enrichment factor of, for example, at least 835 (13% deuterium incorporation), of at least 1670 (26% deuterium incorporation, of at least 3500 (52.5% deuterium incorporation), at least 4500 (67.5% deuterium incorporation), at least 5000 (75% deuterium), at least 5500 (82.5% deuterium incorporation), at least 6000 (90% deuterium incorporation), at least 6333.3 (95% deuterium incorporation), at least 6466.7 (97% deuterium incorporation), at least 6600 (99% deuterium incorporation), or at least 6633.3 (99.5% deuterium incorporation).

When a particular position in a compound of the invention is designated specifically by name or structure as "H" or "hydrogen", the position is understood to have hydrogen at its natural abundance isotopic composition.

When a particular position in a compound of the invention is designated specifically by name or structure as "D" or "deuterium", the position is understood to have deuterium at an abundance that is at least 3340 times greater than the natural abundance of deuterium, which is 0.015% (i.e., at least 50.1% incorporation of deuterium), at least 3500 times greater than the natural abundance of deuterium (52.5% deuterium incorporation), at least 4500 times greater than the natural abundance of deuterium (67.5% deuterium incorporation), at least 5000 (75% deuterium), at least 5500 times greater than the natural abundance of deuterium (82.5% deuterium incorporation), at least 6000 times greater than the natural abundance of deuterium (90% deuterium incorporation), at least 6333.3 times greater than the natural abundance of deuterium (95% deuterium incorporation), at least 6466.7 times greater than the natural abundance of deuterium (97% deuterium incorporation), at least 6600 times greater than the natural abundance of deuterium (99% deuterium incorporation), or at least 6633.3 times greater than the natural abundance of deuterium (99.5% deuterium incorporation).

When a chemical name or structure is silent as to whether a particular position in a compound normally occupied by hydrogen is isotopically enriched, it is intended that the particular position is occupied by hydrogen at its natural abundance. By way of example, the

term "phenyl" or

further designation as to isotopic enrichment indicates that all hydrogen atoms are present at natural abundance.

The term "compound," when referring to a compound of this disclosure, refers to a collection of molecules having an identical chemical structure, except that there may be isotopic variation among the constituent hydrogen atoms of the molecules. The relative amount of isotopic variation in a compound of this invention will depend upon a number of factors including the isotopic purity of deuterated reagents used to make the compound and the efficiency of incorporation of deuterium in the various synthesis steps used to prepare the compound. "D" and "d" both refer to deuterium. "H" refers to hydrogen.

"Substituted with deuterium" refers to the replacement of one or more hydrogen atoms with a corresponding number of deuterium atoms.

The invention is directed to a deuterated inhibitor of QR2 enzyme, containing a 2,3,4,7-tetrahydroindolo[2,3-c]quinoline core, and are provided by the chemical formula depicted in Structure I and the accompanying description:

or a pharmaceutically acceptable salt thereof.

X¹, X², X³, X⁴ andX⁵ are each independently one of: H or D;

X⁶, X⁷, X⁸and X⁹ are each independently H, D or a halogen;

R¹ and R² are each independently CH₃, CD₃, CHD₂ or C¾D; and

R³ is CH3, CD3, CHD₂ or C¾D, provided that the compound comprises at least one deuterium atom.

In a first embodiment, the deuterated inhibitor of QR2 enzyme is a compound represented by Formula (I) , or a pharmaceutically acceptable salt thereof, wherein X , X , X andX⁹ are each H, and the remaining variables are as described for Formula (I).

In a second embodiment, the deuterated inhibitor of QR2 enzyme is a compound represented by Formula (I) , or a pharmaceutically acceptable salt thereof, wherein X⁶, X⁷, X⁸ and X⁹ are each D, and the remaining variables are as described for Formula (I).

In a third embodiment, the deuterated inhibitor of QR2 enzyme is a compound represented by Formula (I) , or a pharmaceutically acceptable salt thereof, wherein R is C¾, and the remaining variables are as described for Formula (I) or in the first or second embodiment.

In a fourth embodiment, the deuterated inhibitor of QR2 enzyme is a compound represented by Formula (I) , or a pharmaceutically acceptable salt thereof, wherein R is CD₃, and the remaining variables are as described for Formula (I) or in the first or second embodiment.

In a fifth embodiment, the deuterated inhibitor of QR2 enzyme is a compound represented by Formula (I) , or a pharmaceutically acceptable salt thereof, wherein X¹, X², X³ and X⁴ are each H, and the remaining variables are as described for Formula (I) or in the first, second, third or fourth embodiment.

In a sixth embodiment, the deuterated inhibitor of QR2 enzyme is a compound represented by Formula (I) , or a pharmaceutically acceptable salt thereof, wherein X¹, X², X³ and X⁴ are each D, and the remaining variables are as described for Formula (I) or in the first, second, third or fourth embodiment.

In a seventh embodiment, the deuterated inhibitor of QR2 enzyme is a compound represented by Formula (I) , or a pharmaceutically acceptable salt thereof, wherein R¹ and R² are each CH₃, and the remaining variables are as described for Formula (I) or in the first, second, third, fourth, fifth or sixth embodiment.

In an eighth embodiment, the deuterated inhibitor of QR2 enzyme is a compound represented by Formula (I) , or a pharmaceutically acceptable salt thereof, wherein R¹ and R² are each CD₃, and the remaining variables are as described for Formula (I) or in the first, second, third, fourth, fifth or sixth embodiment.

In a ninth embodiment, the deuterated inhibitor of QR2 enzyme is a compound represented by Formula (I) , or a pharmaceutically acceptable salt thereof, wherein X^s is H, and the remaining variables are as described for Formula (I) or in the first, second, third, fourth, fifth, sixth, seventh or eighth embodiment.

In a ninth embodiment, the deuterated inhibitor of QR2 enzyme is a compound represented by Formula (I) , or a pharmaceutically acceptable salt thereof, wherein X^s is D, and the remaining variables are as described for Formula (I) or in the first, second, third, fourth, fifth, sixth, seventh or eighth embodiment.

Specific examples of deuterated inhibitors of QR2 enzyme of the invention include a compound selected from:

3,3,6-trimethyl-2,3,4,7-tetrahydroindolo-8,9, 10, 1 l-d₄-[2,3-c]quinolin- 1-one;

3,3-dimethyl-6-methyl-d3-2,3,4,7-tetrahydroindolo[2,3-c]qumolin-l-one;

3,3-dimethyl-d₆-6-methyl-2,3,4,7-tetrahydroindolo[2,3-c]qumolin- 1-one; and

3,3-dimethyl-d₆-6-methyl-d₃-2,3,4,7-tetrahydroindolo-8,9, 10, 1 l-d₄-[2,3-c]quinolin- 1- one; or a pharmaceutically acceptable salt of any of the foregoing. In certain embodiments, the compounds may be basic and form pharmaceutically acceptable salts with organic and inorganic acids.

Examples of suitable acids for such acid addition salt formation are hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, acetic acid, citric acid, oxalic acid, malonic acid, salicylic acid, p-aminosalycilic acid, malic acid, fumaric acid, succinic acid, ascorbic acid, maleic acid, sulfonic acid, phosphonic acid, perchloric acid, nitric acid, propionic acid, gluconic acid, lactic acid, tartaric acid, hydroxymaleic acid, pyruvic acid, pnenylacetic acid, benzoic acid, p-aminobenzoic acid, p-hydroxybenzoic acid, methanesulfonic acid, ethanesulfonic acid, nitrous acid, hydroxyethanesulfonic acid, ethylenesulfonic acid, p- toluenesulfonic acid, naphthylsulfonic acid, sulfanilic acid, camphersulfonic acid, mandelic acid, o-methylmandelic acid, hydrogen-benzenesulfonic acid, picric acid, adipic acid, D-o- tolyltartaric acid, tartronic acid, oc-toluic acid (o, m, p), naphthylamine sulfonic acid, and other mineral or carboxylic acids well known to those skilled in the art. The salts may be prepared by contacting the free base form with a sufficient amount of the desired acid to produce a salt in the conventional manner.

The free base forms may be regenerated by treating the salt with a suitable dilute aqueous base solution, such as dilute aqueous sodium hydroxide, potassium carbonate, ammonia and sodium bicarbonate. The free base forms may differ from their corresponding salt forms in certain physical properties, such as solubility in polar solvents. The free base forms may differ from their corresponding salt forms in certain pharmacokinetic parameters, such as bioavailability, resulting in different pharmacological effects.

The present disclosure includes the pharmaceutically active free base forms of the compounds and pharmaceutically active salts of these compounds, all stereoisomeric forms and regioisomeric forms of these compounds or prodrugs thereof.

By way of example, the compounds of Structure I wherein R 1 , R2 and R 3 are CH₃ or CD₃, X⁵ is H, and X¹, X², X³, X⁴, X⁶, X⁷, X⁸ and X⁹ are H or D, can be prepared according to Scheme I.

More particularly, Scheme I involves a method of preparing an appropriate compound of Structure I using the techniques described in US 2011/0136844 and WO 2011/068990, and the appropriate isotopically enriched reagents and solvents with incorporated deuterium. Scheme I

The chemical synthesis involves a method of making the substituted 2-(2-acyl-lH- indol-3-yl)-5,5-dimethyl-cyclohexane-l,3-dione intermediate of Structure II (shown below) by reacting l-oxo-2,3,4,7-tetrahydro-lH-5-oxonia-7-azabenzo[c]fluorene tetrafluoroborates of Structure III without isolation with water.

The 2-(lH-indol-3-yl)cyclohexane-l,3-diones of Structure IV can be prepared according to Scheme I involving a method of reacting an appropriate 2-(l-acetyl-lH-indol-3- yl)cyclohexane-l,3-dione with sodium hydroxide.

The 2-(l-acetyl-lH-indol-3-yl)cyclohexane-l,3-dione can be prepared according to Scheme I involving a method of reacting an appropriate 1 -acetyl-1 ,2-dihydroindol-3-one with the appropriate cyclohexane-l,3-dione and triethylamine in acetic acid.

Structures II, III and IV are generally:

Structure II Structure III Structure IV

wherein:

X¹, X², X³, X⁴, X⁶, X⁷, X⁸ and X⁹ are independently one of: H or D;

R'. ^ and R³ are independently one of: CH₃ or CD₃;

The l-acetyl-4,5,6,7-d₄-l,2-dihydroindol-3-one can be prepared according to Scheme II using standard techniques (see, for example, Leung et al., J. Med. Chem., 55, 1844-1857 (2012)) involving a method of reacting N-(2-(2-chloroacetyl)-3,4,5,6-d₄-phenyl)acetamide with sodium hydride.

The N-(2-(2-chloroacetyl)-3,4,5,6-d₄-phenyl)acetamide can be prepared by reacting 1- (2-amino-3,4,5,6-d₄-phenyl)2-chloroethanone with acetic anhydride.

The l-(2-amino-3,4,5,6-d₄-phenyl)2-chloroethanone can be prepared by reacting aniline-d₇ with 2-chloroacetonitrile and boron trichloride.

Scheme Π

The 5,5-ciimethyl-d₆-cyclohexane-l,3-dione can be prepared according to Scheme ΙΠ using the standard techniques (see, for example, Young Ho Seo et al., Angewandte Chemie, International Ed. 50, 1342-1345 (2011)) by reacting diethyl malonate and mesityl-dio oxide in the presence of sodium ethylate.

Scheme ΠΙ

In order to use a compound of Structure I or a pharmaceutically acceptable salt or complex thereof for the treatment of humans and/or other animals, it is normally formulated in accordance with standard pharmaceutical practice as a pharmaceutical composition.

The compounds encompassed by Structure I may be administered by different routes including, but not limited to, intravenous, intraperitoneal, subcutaneous, intramuscular, oral, topical (transdermal), or transmucosal administration. For systemic administration, oral administration is preferred. For oral administration, for example, the compounds encompassed by Structure I may be formulated into conventional oral dosage forms such as capsules, tablets, and liquid preparations such as syrups, elixirs, and concentrated drops. These formulations may be designed for immediate release of the drug or they may be designed for a controlled release of the drug.

Compositions of Structure I and their pharmaceutically acceptable salts and/or complexes, which are active when given orally, may be formulated as syrups or other liquid compositions, tablets, capsules, and lozenges. A syrup or liquid formulation will generally consist of a suspension or solution of the compound or salt in a liquid carrier such as, for example, ethanol, peanut oil, olive oil, glycerin or water with or without a flavoring or coloring agent. Where the composition is in the form of a tablet, any pharmaceutical carrier routinely used for preparing solid formulations may be used. Examples of such carriers include magnesium stearate, terra alba, talc, gelatin, acacia, stearic acid, starch, lactose and sucrose. Where the composition is in the form of a capsule, any routine encapsulation is suitable, for example using the aforementioned carriers in a hard gelatin capsule shell. Where the composition is in the form of a soft gelatin shell capsule, any pharmaceutical carrier routinely used for preparing dispersions or suspensions may be utilized. For example, aqueous gums, celluloses, silicates or oils may be used to form a soft gelatin capsule shell.

Alternatively, injection (parenteral administration) may be used, e.g., intramuscular, intravenous, intraperitoneal, and subcutaneous. For injection, the compounds encompassed by Structure I may be formulated in liquid solutions, preferably, in physiologically compatible buffers or solutions, such as saline solution, Hank's solution, or Ringer's solution. Typical parenteral compositions consist of a solution or suspension of a compound or salt in a sterile aqueous or non-aqueous carrier optionally containing parenterally acceptable oil, for example polyethylene glycol, polyvinylpyrrolidone, lecithin, arachis oil or sesame oil. In addition, the compounds encompassed by Structure I may be formulated in solid form and redissolved or suspended immediately prior to use. Lyophilized forms can also be produced.

Typical compositions for inhalation are in the form of a powder, solution, suspension or emulsion that may be administered as a dry powder or in the form of an aerosol using a conventional propellant such as dichlorodifluoromethane or trichlorofluoromethane.

Systemic administration can also be achieved by transmucosal or transdermal methods. For transmucosal or transdermal administration, penetrants appropriate to the barrier to be permeated may also be used in the formulation. Such penetrants are generally known in the art, and include, for example, for transmucosal administration, bile salts and fusidic acid derivatives. In addition, detergents may be used to facilitate permeation.

Transmucosal administration, for example, may be through nasal sprays, rectal suppositories, or vaginal suppositories. A typical suppository formulation comprises a compound of Structure I or a pharmaceutically acceptable salt or complex thereof which is active when administered in this way, with a binding and/or lubricating agent, for example polymeric glycols, gelatins, cocoa-butter or other low-melting vegetable waxes or fats or their synthetic analogs.

For topical administration, the compounds encompassed by Structure I may be formulated into ointments, salves, gels, or creams, as is generally known in the art. Typical dermal and transdermal formulations comprise a conventional aqueous or non-aqueous vehicle, for example a cream, ointment, lotion or paste or are in the form of a medicated plaster, patch or membrane.

For ocular administration, the compounds encompassed by Structure I may be formulated into drops, sprays, or ointments, as is generally known in the art.

The amounts of various compounds encompassed by Structure I to be administered can be determined by standard procedures taking into account factors such as the compound IC₅₀, EC₅₀, the biological half-life of the compound, the age, size and weight of the subject, and the disease or disorder associated with the subject. The importance of these and other factors to be considered are known to those of ordinary skill in the art.

Amounts administered also depend on the routes of administration and the degree of oral bioavailability. For example, for compounds with low oral bioavailability, relatively higher doses may have to be administered.

The composition may be in unit dosage form. For oral application, for example, a tablet or capsule may be administered; for nasal application, a metered aerosol dose may be administered; for transdermal application, a topical formulation or patch may be

administered; and for transmucosal delivery, a buccal patch may be administered. In each case, dosing is such that the subject may administer a unit dose irrespective of weight or for a general weight range, rather than on a per kilogram body weight basis.

Each dosage unit for oral administration may contain from 0.01 to 500 mg, and in certain embodiments, from 1 to 100 mg, of a compound of Structure I or a pharmaceutically acceptable salt or complex thereof, calculated as the free base. The daily dosage for parenteral, nasal, oral inhalation, transmucosal or transdermal routes may contain from 0.01 mg to 100 mg of a compound of Structure I. A topical formulation may contain 0.01 to 5.0% of a compound of Structure I. The active ingredient may be administered as a single dose or in multiple doses, for example, from 2 to 6 times per day, sufficient to exhibit the desired activity, as is readily apparent to one skilled in the art.

As used herein, "treatment" of a disease or disorder includes both therapeutic and prophylactic treatment. Therapeutic treatment refers to alleviating at least one symptom of the disease or disorder. Prophylactic treatment refers to delaying or suppressing the onset and/or development of the disease or disorder.

Diseases and disorders which might be therapeutically treated or prevented, based upon the affected cells, include central nervous system diseases or disorders such as neurodegenerative diseases, and neurological disorders and diseases. As discussed above, alterations in the QR2 enzyme activity have been identified in certain neurodegenerative diseases, and neurological diseases and disorders.

The term "neurodegenerative diseases" includes but is not limited to Mild Cognitive Impairment (MCI) or age-related dementia, vascular dementia, neurocognitive decline resulting from alcohol consumption, Alzheimer's disease, Parkinson's disease, Huntington's disease, Down syndrome, Guillain-Barre syndrome, amyotrophic-lateral sclerosis, AIDS- related dementia, fragile X-associated tremor/ataxia syndrome (FXTAS), progressive supranuclear palsy (PSP), and striatonigral degeneration (SND), which is included with olivopontocerebellear degeneration (OPCD) and Shy Drager syndrome (SDS) in a syndrome known as multiple syndrome atrophy (MSA), regenerative (recovery) treatment of CNS disorders such as spinal cord injury, acute neuronal injury (stroke, traumatic brain injury), guam-parkinsonism-dementia complex, corticobasal neurodegeneration, frontotemporal dementia.

In one embodiment, the term "limited neurodegenerative diseases" includes Mild Cognitive Impairment (MCI) or age-related dementia, vascular dementia, neurocognitive decline resulting from alcohol consumption, Alzheimer's disease, Parkinson's disease, Huntington's disease, Down syndrome, Pick's disease, brain injury, stroke, traumatic brain injury, corticobasal neurodegeneration, frontotemporal dementia. The term "neurological disorders and diseases" includes but is not limited to attention deficit hyperactivity disorder, adjustment disorders, mood disorders, delirium, dementia, amnestic and cognitive disorders, disorders usually first diagnosed in infancy, childhood, or adolescence, dissociative disorders (e.g. dissociative amnesia, depersonalization disorder, dissociative fugue and dissociative identity disorder), eating disorders, factitious disorders, impulse-control disorders, mental disorders due to general medical condition, mood disorders, other conditions that may be a focus of clinical attention, personality disorders, seizures, epilepsy, acute and chronic pain, schizophrenia and other psychotic disorders, sexual and gender identity disorders, sleep disorders, somatoform disorders, substance-related disorders, generalized anxiety disorder (e.g. acute stress disorder), panic disorder, phobia, agoraphobia, obsessive-compulsive disorder, stress, post-traumatic stress disorder, acute stress disorder, anxiety neurosis, nervousness, phobia, abuse, manic depressive psychosis, specific phobias, social phobia, adjustment disorder with anxious features.

In one embodiment, the term "limited neurological disorders and diseases" includes attention deficit hyperactivity disorder, adjustment disorders, anxiety disorders, delirium, amnestic disorders, dissociative disorders (e.g. dissociative amnesia, depersonalization disorder, dissociative fugue and dissociative identity disorder), eating disorders, factitious disorders, impulse-control disorders, personality disorders, other psychotic disorders, sexual and gender identity disorders, sleep disorders, somatoform disorders, phobia, agoraphobia, specific phobias, social phobia, and adjustment disorder with anxious features.

Examples

The following specific examples are included for illustrative purposes only and are not to be considered as limiting to this disclosure. The reagents and intermediates used in the following examples are either commercially available or can be prepared according to standard literature procedures by those skilled in the art of organic synthesis.

General Procedures

General Procedure A: Preparation of the compounds of Structure III

A compound of Structure IV (0.582 mol, 1 equiv) was added to a mixture of acetic acid (1.5 L) and acetic anhydride (147 mL, 1.56 mol, 2.68 equiv) under stirring at room temperature. Boron trifluoride diethyl etherate (150 mL, 1.22 mol, 2.1 equiv) was added dropwise. In approximately 15 minutes, a very thick yellow slurry was formed. The mixture was stirred at room temperature for 2 hours and then cooled at 5°C. Without isolation of the compound of Structure ΙΠ, the reaction mixture was treated with water as described in General Procedure B.

General Procedure B: Preparation of the compounds of Structure Π

The reaction mixture of General Procedure A was treated with water (pre-cooled with ice), and the resultant suspension was stirred at room temperature overnight. The precipitate was collected, washed with water and air-dried to give 85-92% of the corresponding compound of Structure II. The product was used in the next step without purification. General Procedure C: Preparation of the compounds of Structure I

The compound of Structure II (0.581 mol) and ammonium acetate (269 g, 3.48 mol, 6 equiv) were dissolved in a mixture of ethanol (1.8 L) and water (160 mL). The mixture was refiuxed for 1 hour and cooled to room temperature. The reaction mixture was concentrated under reduced pressure. The residue was diluted with water (4 L), and 33% aqueous ammonia solution (50 mL) was added to bring the solution to pH 8-9. The resulting solid was filtered off, washed with water (1 L) and dried in vacuum overnight at 40°C to afford a compound of Structure I with purity >99% in 58-90% yield.

Example 1

Preparation of 3.3.6-trimemvl-2.3.4.7-tetrahvdroindolo-8.9.10.1 l-d₄-r2.3-clquinol.n-l -one (Compound 225)

a) l-(2-Amino-3,4,5,6-d₄-phenyl)2-chloroethanone

To a stirred solution of 1M boron trichloride in hexane (2.97 L, 2.96 mol, 1.1 equiv), a solution of aniline-d7 (270 g, 2.7 mol, 1 equiv) in dry toluene (3.4 L) was added dropwise at a rate to keep the temperature below 0°C. To the resulting heterogeneous mixture containing aniline-d7 boron trichloride complex, 2-chloroacetonitrile (205 mL, 3.24 mol, 1.2 equiv) and anhydrous aluminum trichloride (395 g, 2.965 mol, 1.1 equiv) were added successively at 0°C. After warming to room tempwrature, the mixture was connected to a sodium hydroxide scrubber and refluxed for 5 hours, resukting in a two-layer solution. After cooling to 5°C, the mixture was poured into ice-cold (5°C) 2N D-hydrochloric acid in deuterium oxide (12 L) under fast stirring, and a yellow precipitate was formed. To hydrolyze the intermediate ketimine, the mixture was heated at 80°C under stirring until the precipitate was dissolved (1 hour). The mixture was cooled to room temperature and to a separatory funnel. The organic layer was separated and discarded. The aqueous layer was poured into water (20 L) and extracted with ethyl acetate (2 x 10 L). The combined organic layers were washed with saturated brine (10 L) and concentrated to dryness under reduced pressure. The crude product was purified over silica gel (3 kg), eluting with a gradient of 0 to 100% ethyl acetate in heptane to afford l-(2-amino-3,4,5,6-d₄-phenyl)2-chloroethanone as a yellow solid (230 g, 49% yield) which slowly darkened upon storage.

b) N-(2-(2-Chloroacetyl)-3 ,4,5 ,6-d₄-phenyl)acetamide

A mixture of l-(2-amino-3,4,5,6-d₄-phenyl)2-chloroethanone of Example la (230 g, 1.33 mol, 1 equiv) and acetic anhydride (1.6 L, 17.2 mol, 13 equiv) was heated under argon at 90°C for 1 hour. The mixture was cooled to room temperature, the solvent was evaporated under reduced pressure to dryness, and the residue was purified by column chromatography on silica gel (3 kg) eluting with a gradient of 0-100% ethyl acetate in heptanes to afford pure N-(2-(2-chloroacetyl)-3,4,5,6-d₄-phenyl)acetamide ( 174 g, 61% yield) as a yellow solid which slowly darkened upon storage. c) 1 -Acetyl-4,5,6,7-d₄-l ,2-dihydroindol-3-one

To a 60% dispersion of sodium hydride in mineral oil (34.9 g, 872 mmol, 1.1 equiv) in dimethyiacetamide (1.5 L) was added a solution of N-(2-(2-chloroacetyl)-3,4,5,6-d₄- phenyl)acetamide of Example lb (171 g, 0.793 mol, 1 equiv) in dimethyiacetamide (1.5 L) while keeping the temperature below 0°C. The mixture was stirred at 0°C for 2 hours, pouredinto water (30 L) and acidified with IN hydrochloric acid (750 mL). and the mixture was extracted with ethyl acetate (2 X 12 L). The organic layers were combined, washed with saturated brine (10 L), dried over sodium sulfate, filtered and concentrated to dryness under reduced pressure. The crude product was purified by column chromatography on silica gel (3 kg) eluting with a gradient of 0-100% ethyl acetate in heptanes to afford 1- acetyl-4,5,6,7-d₄-l,2-dihydroindol-3-one as a light tan solid (83 g, 58% yield).

d) 2-( 1 -Acetyl-4,5 ,6,7-d₄- 1 H-indol-3-yl)cyclohexane- 1 ,3-dione

l-Acetyl-4,5,6,7-d₄-l^-dihydroindol-3-one of Example lc (188.5 g, 1.05 mol, 1 equiv) and 5,5-dimethyl-cyclohexane-l,3-dione (147 g, 1.05 mol, 1 equiv) were dissolved in glacial acetic acid (1 L), and triethylamine (148 mL, 1.05 mol, 1 equiv) was added at room temperature with stirring. The reaction mixture was refluxed for 6 hours, cooled to room temperature and concentrated under reduced pressure to dryness. Water (4 L) was added, and the resulting suspension was stirred at room temperature overnight. The precipitate was filtered, triturated with methyl terf-butyl ether (1 L), and dried under vacuum overnight at 40°C to afford 2-(l-acetyl-4,5,6,7-d₄-lH-indol-3-yl)cyclohexane-l,3-dione (243.4 g, 77% yield) as a light tan solid. e) 2-(4,5,6,7-d₄-lH-indol-3-yl)cyclohexane-l,3-dione

To a suspension of the 2-(l-acetyl-4,5,6,7-d₄-lH-indol-3-yl)cyclohexane-l,3-dione of Example Id (321.7 g. 1.07 mol, 1 equiv) in methanol (1.3 L), 2N sodium hydroxide aqueous solution (1.28 L, 3.2 mol, 3 equiv) was added at room temperature with stirring. The reaction mixture was heated at 60°C for 2 hours with stirring. The mixture was cooled to room temperature, poured into 2N HC1 (3 L) and extracted with ethyl acetate (6 L). The organic layer was separated, and the aqueous layer was extracted with ethyl acetate (4 L). The combined organic layers were washed with saturated brine (4 L), dried over sodium sulfate and concentrated under reduced pressure to afford 2-(4,5,6,7-d₄-lH-indol-3-yl)cyclohexane- 1,3-dione as a pale yellow solid (234 g, 85% yield) as a tan solid. f) 3,3,6-Trimethyl-l^xo-2,3,4,7-tetrahydro-lH-5-oxonia-8,9,10,l l-d₄-7- azabenzo[c]fluorene tetrafluoroborate

The title compound was prepared utilizing the procedures described in General Procedure A. Without isolation of the 3,3,6-trimethyl-l-oxo-2,3,4,7-tetrahydro-lH-5-oxonia- 8,9,10,1 l-d₄-7-azabenzo[c]fiuorene tetrafluoroborate, the next step involved formation of 2- (2-acetyl-l//-indol-3-yl)-5,5-dimethyl-cyclohexane-l,3-dione directly, as described in Example lg. g) 2-(2-AcetyM,5,6,7-d₄-1H-mdol-3-yl)-5,5-dimethyl-cyclohexane-l,3-dione

Utilizing the procedures described in General Procedure B, the title compound was prepared in 86% yield as a grey solid.

h) 3,3,6-Trimethyl-2,3,4 ,7-tetrahydroindolo-8,9,10,l l-d₄-[2,3-c]quinolin -1-one

Utilizing the procedures described in General Procedure C, the title compound was prepared as a light tan solid (148.3 g, 90% yield, >99% purity). 1H NMR (400 MHz, CDC1₃): δ 1.17 (s, 6H), 2.70 (s, 2H), 2.84 (s, 3H), 3.22 (s, 2H), 8.41 (s, 1H).

Example 2

Preparation of 3 ,3-dimethyl-6-methyl-d3-23.4,7-tetrahvdromdolo[2,3-c ]qumolin-1-)ne (Compound 254)

Utilizing the procedures described in Examples 1 d-h except substituting in Example Id: commercially available l-acetyl-l,2-dihydroindol-3-one for l-acetyl-4,5,6,7-d₄- l,2-dihydroindol-3-one; Example If: acetic anhydride-d₆ for acetic anhydride, acetic acid-d₄ for acetic acid, and deuterium oxide for water, Example 1 g: deuterium oxide for water; and Example lh: ammonium acetate-dj for ammonium acetate, EtOD for ethanol, and deuterium oxide for water, the title compound was prepared as a yellow solid (87 mg, 84% yield). 1H NMR (400 MHz, CDC1₃): δ 1.17 (s, 6H), 2.71 (s, 2H), 3.20 (s, 2H), 7.32-7.34 (m, 1H), 7.50- 7.51 (m, lH), 7.56-7.60 (m, 1 H), 8.45 (s, 1H), 9.35 (d, 1H).

Example 3

Preparation of 33-Dimethyl-d₆-6-methvl-2,3,4,7-tetrahydroindolo[2,3-clqumolin-l-one (Compound 255)

a) 5,5-Dimethyl-d₆-cyclohexane- 1 ,3-dione

Sodium hydride (93 mg, 3.9 mmol) was carefully added to dry ethanol (2.2 mL) at room temperature under argon to form sodium ethylate. After complete dissolution of sodium hydride, diethyl malonate (650 mg, 4 mmol) followed by mesityl-dio oxide (375 mg, 3.5 mmol) were added at room temperature. The reaction mixture was refluxed at 100°C for 3 hours and cooled to room temperature. A solution of potassium hydroxide (465 mg) in water (2.2 mL) was added at room temperature; the reaction mixture was heated at 110°C for 6 hours, and then cooled at room temperature. The mixture was acidified to pH 2 and heated at 50°C for 1 hour. The reaction mixture was cooled and extracted with ether three times. The organic layers were combined, washed with brine, dried over sodium sulfate, filtered and concentrated to dryness. The crude product was purified by column chromatography on silica gel (heptane-ethyl acetate 1 :1 + 1% acetic acid to 100% ethyl acetate + 1% acetic acid) to obtain 5,5-Dimethyl-d₆-cyclohexane-l,3-dione as a white solid (350 mg, 69% yield).

b) 3,3-dimethyl-d₆-6-methyl-2,3,4,7-tetrahydroindolo[2,3-c]qumolin-l-one

Utilizing the procedures described in Examples 1 d-h except substituting in Example Id commercially available 1 -acetyl- l,2-dihydroindol-3-one for l-acetyl-4,5,6,7-d₄-l,2- dihydroindol-3-one, and 5,5-dimethyl-d₆-cyclohexane-l,3-dione for 5,5-dimethyl- cyclohexane-l,3-dione, the title compound was prepared as a yellow solid (50 mg, 68% yield). 1HNMR (400 MHz, CDCI₃): δ 2.70 (s, 2H), 2.87 (s, 3H), 3.20 (s, 2H), 7.30-7.34 (m, lH), 7.50-7.52 (m, lH), 7.57-7.61 (m, 1 H), 8.45 (s, 1H), 9.35 (d, 1H).

Example 4

Preparation of 33-dimethyl-d₆-6-methyl-d₃-23.4J-tetrahvdromdolo-8.9.10.11-d₄- i2.3-c1quinolin-l-one (Compound 240)

Utilizing the procedures described in Examples 1 d-h except substituting in Example Id: 5,5-dimethyl-d₆-cyclohexane-l,3-dione for 5,5-dimethyl-cyclohexane-l,3-dione;

Example If: acetic anhydride-d₆ for acetic anhydride, acetic acid-d₄ for acetic acid, and deuterium oxide for water; Example lg: deuterium oxide for water, and Example lh:

ammonium acetate-d₄ for ammonium acetate, EtOD for ethanol, and deuterium oxide for water, the title compound was prepared as a yellow solid (37 mg, 58% yield). 1H NMR (400 MHz, CDCI₃): δ 2.67 (s, 2H), 3.19 (s, 2H), 8.50 (s, lH).

Example 5

Preparation of 3 J.6-trimethvl-2.3.4.7-tetrahydroindolo-8.9.10.11 -d₄-[2.3-c1quinolin- 1 -one hydrochloride (Compound 225'HCI) To a solution of 3,3,6-trimethyl-2,3,4,7-tetrahyclromdolo-8,9,10,11-d4-[2,3-c]quinolin

-1-one of Example lh (100 g, 0.354 mol, 1 equiv) in 1,4-dioxane (1.2 L), 4N hydrochloride solution in 1,4-dioxane (89 mL, 0.354 mol, 1 equiv) was added under stirring at room temperature. The heterogeneous mixture was stirred for 4 hours, diluted with diethyl ether (3 L) and filtered off to afford the title compound as a yellow solid (108 g, 96% yield, >99% purity).

Example 6

Preparation of 3.3.6-trimethvl-2.3.4.7-tetrahvdroindolo-8.9.10.1 l-d₄-[2,3-c]quinolin- 1-one mesilate (Compound 225.MsOH)

To a solution of methanesulfonic acid (0.023 mL, 0.354 mmol, 1 equiv) in ethanok (3.5 mL), 3,3,6-trimethil-2,3,4,7-tetrahydroindolo-8,9,l(),ll-d4-[2,3-c]quinolin -1-one of Example lh (0.1 g, 0.354 mmol, 1 equiv) was added under stirring at room temperature resulting in dissolution before salt precipitation. The heterogeneous mixture was stirred overnight and concentrated under reduced pressure. The residue was triturated with diethyl ether (30 mL) and filtered off to afford a yellowish tan solid (0.1 g, 75% yield).

The specific approaches and compounds shown above are not intended to be limiting. The chemical structures in the Scheme I herein depict variables that are hereby defined commensurately with chemical group definitions (moieties, atoms, etc.) of the corresponding position in the compound structures herein, whether identified by the same variable name (i.e., X¹, X², X³, X⁴, X⁵, X⁶, X⁷, X⁸ and X⁹, andR¹, R² and R³) or not. The suitability of a chemical group in a compound structure for use in the synthesis of another compound is within the knowledge of one of ordinary skill in the art.

Additional methods of synthesizing compounds of Structure I and their synthetic precursors, including those within routes not explicitly shown in schemes herein, are within the means of chemists of ordinary skill in the art. Synthetic chemistry transformations and protecting group methodologies (protection and deprotection) useful in synthesizing the applicable compounds are known in the art.

Additional compounds of the invention are provided in Table 1 below.

Pharmaceutically acceptable salts of these compounds are also included in the invention.

Example 7

Evaluation of the Inhibition of Reductase Activity in Recombinant Human Ouinone

Reductase 2 Enzyme (OR2: NQO2)

There is no single, universally-accepted method to evaluate the activity of quinone reductase 2 (QR2; NQ02). The enzyme requires 2 co-substrates; one as a 2-electron acceptor and one as a 2-electron donor. The 2 substrates vary across the many published methods of QR2 activity, as do their method of measurement. In addition, the source and concentration of the enzyme vary. As noted by Pegan et al. (Protein Sci 2011, 20, 1182-1195 ("Pegan 2011")), QR2 inhibitors vary in their IC50 values between activity assay methods but the rank order of inhibitory potency is often similar. For example, the ICso of 2-iodomelatonin ranges from 1 to 16 μΜ under four different assay conditions (Mailliet et al., Biochem Pharmacol 2005, 71, 74-88; Calamini 2008; and Pegan 2011) and resveratrol ranges from 0.1 to 70 μΜ across six different assay conditions (Calamini 2008; Ferry et al., Chemico-Biol Interactions 2010, 186, 103-109; and St. John etal, Bioorg Med Chem 2013, 21 (19), 6022-6037).

Compounds of Structure I were tested using the following assay conditions:

Human quinone reductase 2 (hQR2) enzymatic activity was measured using 0.4 nM recombinant hQR2 enzyme (Reaction Biology Corp; NQ02, #NQO-l 1-312), 50 uM menadione as substrate and 25 uM of dihydrobenzylnicotamide (BNAH) as co-substrate. Reaction was performed in a buffer consisting of 50mM Tris/HCl, pH 8.5, 200 mM NaCl, 0.05% Brij30, ().2uM FAD and 0.25% DMSO. Compounds were serially diluted (3x) in DMSO and delivered to reaction wells using acoustic technology (ECHO, Labcyte). Final DMSO concentration in assay was 0.5%. Compounds were pre-incubated with hQR2 for 10 minutes and the reaction was initiated by the addition of menadione and BNAH. Activity was followed in real time at 460/25 nM with excitation at 340/60 nM using Envision plate reader (Perkin Elmer). Slopes were calculated using Excel from the linear portion of the reaction.

Activity in the presence of DMSO alone was considered as 100% and used to calculate % activity in the presence of variable compound concentrations. The inhibitory concentration 50% (IC50) was determined using GraphPad prism software by fitting data to sigmoidal dose response (variable slope) equation.

Table 2: Evaluation of the Inhibition of OR2 Enzyme

Example 8

Evaluation of Metabolic Stability in Pooled Human Liver Microsomes

Human liver microsomes (HLM) were acquired from Xenotech (Cat. HI 500) and nicotinamide adenine dinucleotide phosphate (NADPH; catalog #N1630) was obtained from Sigma-Aldrich.

The compounds of Structure I were first dissolved in DMSO to a concentration of 2 mM, followed by dilution in acetonitrile to a concentration of 200 μΜ, and then diluted further to 4 μΜ with 100 mM potassium phosphate, pH 7.4 and containing a 4 mM concentration of NADPH. Pooled human liver microsomes were diluted in 100 mM potassium phosphate buffer, pH 7.4 (warmed to 37°C) to a concentration of 1 mg/mL.

To start the reaction, equal volumes (195 μΙ_) of the solutions of test or control articles and HLM were mixed in polypropylene 96-well plates; the final concentrations in the assay incubations of the test or control articles was 2 μΜ and of HLM was 0.5 mg/mL.

Immediately after mixing, duplicate 50 μL aliquots were removed to represent the concentrations at 0 minutes (To) and then quenched with 300 μΐ of ACN containing the analytical internal standard (IS) to precipitate protein. The assay incubation plate was maintained at 37°C with gentle agitation. Aliquots were similarly removed and quenched after 10, 20, 30, 45 and 60 minutes.

The quenched samples were maintained at 4°C until they were centrifuged at 2000 x g (3100 rpm) for 10 minutes in a refrigerated (4°C) centrifuge. Aliquots (50 μL) of the supernatants were removed and diluted with 100 μL water to reduce the % organic content prior to bioanalysis using an ABI Sciex 5500 LC/MS/MS instrument.

Analysis of the concentration data was performed using Microsoft Excel 2010. The half-life (t½) values were calculated from the slopes of the linear regression of the log of the percentage of compound remaining at each time-point:

t½ = 0.693/k,

where k = - [slope of linear regression of % compound remaining (In) vs incubation time in min]

Table 3: Evaluation of Metabolic Stability in Pooled Human Liver Micrososmes

Example 9

Evaluation of the Pharmacokinetics in Rats of Compound 225 (Example lh) administered as Compound 225 HC1 (Example 5)

Male Sprague-Dawley rats with surgically-implanted jugular-vein catheters were given single intravenous bolus injections of 3 mg/kg Compound 225 HC1 (Example 5) dissolved in a 20% aqueous Captisol^® solution (Charles River Laboratories, Worcester, MA). Blood samples were collected at periodic intervals, centrifuged and the plasma analyzed by an LC/MS/MS method developed to quantify the concentration of Compound 225 (Example lh). Ambocarb was used as the internal standard for the analytical assay. The resulting analytical data were subjected to noncompartmental pharmacokinetic analysis.

The mean (istandard deviation) estimates were as follows:

Clearance (CUs) = 51 (2) mlJmin/kg)

Volume of distribution (ν_Μ) = 1.4 (0.07) L/kg

Terminal elimination half-life (t_ia) = 1.41 (0.03) hr

Shcherbakova (US20110021776 Al) reports that the pharmacokinetics of ambocarb following 2 or 10 mg/kg intravenous doses to rats was dose-dependent, with a CI range of 71- 106 mlJmin/kg and a tm range of 0.37-0.765 hr.

These results are consistent with the increase in metabolic stability for Compound 225 (Example lh) shown for the human liver microsomes in Example 6.

Example 10

Evaluation of Brain Levels in Rats of Compound 225 (Example lh) administered as

Compound 225 HC1 (Example 5)

Male Wistar rats (N=20) were orally administered various doses (6-12 mg/kg) of Compound 225 HCL (Example 5) dissolved in a 20% aqueous Captisol^® solution (Charles River Laboratories, Finland). Blood and brain samples were collected from each rat at a specified time after dosing. Blood samples were centrifuged and plasma collected. Brain samples were flash frozen and later homogenized with water for analysis. Plasma and homogenized brain samples were analyzed by an LC/MS/MS method developed to quantify the concentration of Compound 225 (Example lh) (Charles River Laboratory, Worcester, MA). The mean (±SD) brain-to-plasma ratio was 3.54 (± 0.69). These data confirm that Compound 225 (Example lh) partitions readily from the plasma into the brain. In addition, these data are consistent with the estimated Vss value following iv dosing in Example 7 and confirm that Compound 225 (Example 5) reaches the therapeutic target for CNS indications.

The above description fully discloses the invention including certain embodiments thereof. Modifications and improvements of the embodiments specifically disclosed herein are within the scope of the following claims. Without further elaboration, it is believed that one skilled in the area can, using the preceding description, utilize the present invention to its fullest extent. The Examples are not intended to limit the present disclosure, although the specifics recited herein may include independently patentable subject matter.

Claims

1. A compound of Structure I:

or a pharmaceutically acceptable salt thereof, wherein:

X¹, X², X³, X⁴ andX⁵ arc each independently one of: H or D;

X⁶, X⁷, X⁸and X⁹ are each independently H, D or a halogen;

R¹ and R² are each independently CH₃, CD₃, CHD₂ or CH₂D; and

R³ is CH₃, CD₃, CHD₂ or CH₂D, provided that the compound comprises at least one deuterium atom.

2. The compound of claim 1, or a pharmaceutically acceptable salt thereof, wherein X⁶, X⁷, X⁸ and X⁹ are each H.

3. The compound of claim 1, or a pharmaceutically acceptable salt thereof, wherein X⁶, X⁷, X⁸ and X⁹ are each D.

4. The compound of any one of claims 1-3, or a pharmaceutically acceptable salt

thereof, wherein R is Cl¾.

5. The compound of any one of claims 1-3, or a pharmaceutically acceptable salt

thereof, wherein R³ is CD3.

6. The compound of any one of claims 1-5, or a pharmaceutically acceptable salt

thereof, wherein X¹, X², X³ and X⁴ are each H.

7. The compound of any one of claims 1-5, or a pharmaceutically acceptable salt

thereof, wherein X¹, X², X³ and X⁴ are each D.

8. The compound of any one of claims 1-7, or a pharmaceutically acceptable salt

thereof, wherein R and R are each C¾.

9. The compound of any one of claims 1-5, or a pharmaceutically acceptable salt

thereof, wherein R and R" are each CD3.

10. The compound of any one of claims 1-9, wherein X⁵ is H.

11. The compound of any one of claims 1 -9, wherein X⁵ is D.

12. A compound of claim 1 or a pharmaceutically acceptable salt thereof, selected from:

3,3,6-trimethyl-2,3,4,7-tetrahydroindolo-8,9,10,ll-d4-[2,3-c]qumolin-l-one;

3,3-dimethyl-6-methyl-d3-2,3,4,7 etrahydromdolo[2,3-c]quinolin-l-one;

3,3-dimethyl-d6-6-methyl-2,3,4,7-tetrahydroindolo[2,3-c]quinolin-l-one; and

3,3-dimethyl-d6-6-methyl-d3-2,3,4,7-tetrahydroindolo-8,9,10,ll-d4-[2,3-c]qumolm-l- one.

13. A compound of claim 1 selected from at least one of:

3,3,6-trimethyl-2,3,4,7-tetrahydroindolo-8,9,10,ll-d4-[2,3-c]qumolin-l-one;

3,3-dimethyl-6-methyl-d3-2,3,4,7-tetrahydroindolo[2,3-c]quinolin-l-one;

3,3-dimethyl-d6-6-methyl-2,3,4,7-tetrahydroindolo[2,3-c]qumolin-l-one;

3,3-dimethyl-d6-6-methyl-d3-2,3,4,7-tetrahydroindolo-8,9,10,ll-d4-[2,3-c]quinolin-l- one.

14. A composition comprising a compound according to any one of claims 1 -13 or a pharmaceutically acceptable salt thereof, wherein the composition is formulated for pharmaceutical use and the carrier is a pharmaceutically acceptable carrier.

15. A composition according to claim 14 additionally comprising a second therapeutic agent.

16. A composition according to claim 14 or 15, wherein the second therapeutic agent is a neuroactive compound, such as an NMDA receptor antagonist (e.g., memantine), a cholinesterase inhibitor (e.g., donepezil, galantamine, rivastigmine), a modulator of neurotransmitters (e.g., paroxetine, venlafaxine, escitalopram) or a natural product (e.g., melatonin).

17. A composition according to any one of claims 14 to 16, wherein the composition is used for the treatment or prevention of a disease or condition selected from: chronic neurodegenerative conditions (such as Alzheimer's disease, Parkinson's disease); psychiatric disorders (such as depression or major depressive disorder, schizophrenia, obsessive compulsive disorder, generalized anxiety, attention deficit

disorder/attention deficit hyperactivity disorder), neurocognitive decline caused by alcohol consumption; alcohol withdrawal symptoms; and other cognitive disorders, such as age-related dementia or those arising from traumatic brain injury (TBI)/ concussions or stroke.

18. A composition according to any one of claims 14 to 16, wherein the composition is used for cognitive enhancement, such as to treat dementia, including dementia arising from disease states such as Alzheimer's disease, Parkinson's disease and/or alcoholism.

19. A composition according to any one of claims 14 to 16, wherein the composition is used for cognitive enhancement, such as to treat cognitive disorders arising from traumatic brain injury (TBI)/ concussions.

20. A method of treating a subject suffering from, or susceptible to, a disease or condition that is beneficially treated by an agent that inhibits the enzyme quinone reductase 2 (QR2), comprising the step of administering to the subject in need thereof an effective amount of a compound of any one of claims 1-13 or a pharmaceutically acceptable salt thereof.

21. The method according to claim 20, wherein the disease or condition is selected from: chronic neurodegenerative conditions (such as Alzheimer's disease, Parkinson's disease); psychiatric disorders (such as depression or major depressive disorder, schizophrenia, obsessive compulsive disorder, generalized anxiety, attention deficit disorder/attention deficit hyperactivity disorder); neurocognitive decline caused by alcohol consumption; alcohol withdrawal symptoms; and other cognitive disorders, such as age-related dementia or those arising from traumatic brain injury (TBI)/ concussions or stroke.

22. The method according to claim 20, wherein the disease or condition is dementia, includ ng dementia arising from disease states such as Alzheimer's disease,

Parkinson's disease and/or alcoholism.

23. The method according to claim 20, wherein the disease or condition is cognitive impairment, including cognitive impairment arising from traumatic brain injury (TBI)/ concussions.

24. The method according to claim 20, wherein the disease is malaria.

25. The method according to claim 20, wherein the disease or condition is toxicity of tamoxifen in the treatment regimen of a cancer patient.