Compositions and methods for treating neurological conditions

Abstract

Compositions and methods for treating neurological conditions are provided. The present invention provides a pharmaceutical composition comprising a dopamine reuptake inhibitor or dopamine agonist, an adrenergic receptor antagonist, and a pharmaceutically acceptable carrier, and a method for treating a neurological disorder by administering the composition.

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of U.S. Provisional Application No. 63 / 426,165, filed November 17, 2022, which is incorporated herein by reference in its entirety. STATEMENT OF FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT This invention was made with U.S. government support under grant / contract numbers 1R43NS095422-01 and 1R43NS103696-01A1 awarded by the National Institutes of Health. The government has certain rights in this invention. The present invention relates to compositions and methods for treating neurological and neurodevelopmental disorders. [Background technology]

[0002] Effective medications for the treatment and amelioration of neurological disease states represent an unmet medical need. Many therapies have been developed to treat neurological and neurodevelopmental disorders, but their effectiveness has been limited. As such, widespread problems caused by these disorders remain, and hopes for effective and durable treatments are low. This unmet medical need is addressed by the present invention, exemplary compositions and methods of which are described in more detail below. Summary of the Invention

[0003] In one aspect, the present invention reflects the discovery that these neurological disorders involve an excess of adrenergic activity and a deficiency of dopaminergic activity. In one aspect, the present disclosure relates to a pharmaceutical composition comprising a dopamine reuptake inhibitor, an adrenergic receptor antagonist, and a pharmaceutically acceptable carrier. In another aspect, the present disclosure relates to a pharmaceutical composition comprising a dopamine agonist, an adrenergic receptor antagonist, and a pharmaceutically acceptable carrier. In one aspect, the present disclosure relates to methods of treating neurological and neurodevelopmental disorders by administering these pharmaceutical compositions to a subject in need thereof. Accordingly, one embodiment of the present invention provides pharmaceutical compositions comprising a dopamine reuptake inhibitor or dopamine agonist and an adrenergic receptor ("adrenergic receptor") antagonist for the treatment of a neurological disease or condition. In another aspect, the method (i.e., use of the composition in treatment) also includes measuring biomarker levels in a subject in need of treatment. This measuring step can occur before, during, and / or after treatment. In yet another aspect, the present disclosure relates to a pharmaceutical kit comprising a dopamine reuptake inhibitor or dopamine agonist, an adrenergic receptor antagonist, and reagents or devices for assessing and measuring at least one or more biomarkers.

[0004] The biomarkers used in the present method or kit may be glucose metabolism and metabolite profiles, and / or lipid metabolism and related metabolite profiles, and / or hormone profiles, and / or cytokine profiles, and / or cardiac function. Biomarkers of glucose metabolism may be, for example, glucose, lactate, pyruvate, the lactate-to-glucose ratio, or glucose disposal rate. Lipid metabolism and related metabolite profiles may be composed of β-hydroxybutyrate or acetoacetate, and / or high-density lipoproteins, cholesterol, triglycerides, or free fatty acids, or the disposal rates of β-hydroxybutyrate or acetoacetate. Biomarkers of hormone profiles may be, for example, cortisol or leptin. Biomarkers of cytokine profiles may be, for example, IL-10, IL-4, IL-17, IL-23, or their corresponding ratios. Biomarkers of cardiac function may be, for example, heart rate or heart rate variability. [Brief explanation of the drawings]

[0005] [Figure 1A]Comparison of tyrosine hydroxylase expression in different postmortem human brain sections (pons / putamen; Figure 1C). Compared to brain sections from individuals without a neurodegenerative condition, brain sections from frontotemporal dementia (FTD) showed increased levels of tyrosine hydroxylase expression in the pons and decreased expression of tyrosine hydroxylase in the putamen. [Figure 1B] Comparison of tyrosine hydroxylase expression in different postmortem human brain sections (pons / putamen; Figure 1C). Compared to brain sections from individuals without a neurodegenerative condition, brain sections from frontotemporal dementia (FTD) showed increased levels of tyrosine hydroxylase expression in the pons and decreased expression of tyrosine hydroxylase in the putamen. [Figure 2A] Figure 1 shows a comparison of lipid and glucose metabolic activity between healthy controls and patients with frontotemporal dementia. These analyses suggest that FTD conditions are associated with metabolic dysregulation, namely, increased lipid catabolism (β-hydroxybutyrate is a product of lipid β-oxidation) and altered glucose metabolism. [Figure 2B] Figure 1 shows a comparison of lipid and glucose metabolic activity between healthy controls and patients with frontotemporal dementia. These analyses suggest that FTD conditions are associated with metabolic dysregulation, namely, increased lipid catabolism (β-hydroxybutyrate is a product of lipid β-oxidation) and altered glucose metabolism. [Figure 2C] Figure 1 shows a comparison of lipid and glucose metabolic activity between healthy controls and patients with frontotemporal dementia. These analyses suggest that FTD conditions are associated with metabolic dysregulation, namely, increased lipid catabolism (β-hydroxybutyrate is a product of lipid β-oxidation) and altered glucose metabolism. [Figure 2D] Figure 1 shows a comparison of lipid and glucose metabolic activity between healthy controls and patients with frontotemporal dementia. These analyses suggest that FTD conditions are associated with metabolic dysregulation, namely, increased lipid catabolism (β-hydroxybutyrate is a product of lipid β-oxidation) and altered glucose metabolism. [Figure 3A]FIG. 1 shows a comparison of the behavioral and lipid metabolic patterns of drug-treated and untreated (vehicle) P301L mice with those of Swiss Webster (SW) mice (the background strain of transgenic P301L mice, used as a healthy control). [Figure 3B] FIG. 1 shows a comparison of the behavioral and lipid metabolic patterns of drug-treated and untreated (vehicle) P301L mice with those of Swiss Webster (SW) mice (the background strain of transgenic P301L mice, used as a healthy control). [Figure 3C] FIG. 1 shows a comparison of the behavioral and lipid metabolic patterns of drug-treated and untreated (vehicle) P301L mice with those of Swiss Webster (SW) mice (the background strain of transgenic P301L mice, used as a healthy control). [Figure 3D] FIG. 1 shows a comparison of the behavioral and lipid metabolic patterns of drug-treated and untreated (vehicle) P301L mice with those of Swiss Webster (SW) mice (the background strain of transgenic P301L mice, used as a healthy control). [Figure 3E] FIG. 1 shows a comparison of the behavioral and lipid metabolic patterns of drug-treated and untreated (vehicle) P301L mice with those of Swiss Webster (SW) mice (the background strain of transgenic P301L mice, used as a healthy control). [Figure 3F] FIG. 1 shows a comparison of the behavioral and lipid metabolic patterns of drug-treated and untreated (vehicle) P301L mice with those of Swiss Webster (SW) mice (the background strain of transgenic P301L mice, used as a healthy control). [Figure 4A]FIG. 1 shows a comparison of learning and memory behaviors between different drug-treated and untreated (vehicle) FmrlKO mice and FVB mice (the background strain of the transgenic FmrlKO mice, used as a healthy control). [Figure 4B] FIG. 1 shows a comparison of learning and memory behaviors between different drug-treated and untreated (vehicle) FmrlKO mice and FVB mice (the background strain of the transgenic FmrlKO mice, used as a healthy control). [Figure 4C] FIG. 1 shows a comparison of learning and memory behaviors between different drug-treated and untreated (vehicle) FmrlKO mice and FVB mice (the background strain of the transgenic FmrlKO mice, used as a healthy control). [Figure 4D] FIG. 1 shows a comparison of learning and memory behaviors between different drug-treated and untreated (vehicle) FmrlKO mice and FVB mice (the background strain of the transgenic FmrlKO mice, used as a healthy control). DETAILED DESCRIPTION OF THE INVENTION

[0006] Those skilled in the art will appreciate that the present invention can be embodied in many different forms, and should not be construed as limited to the embodiments set forth in this application. Rather, these embodiments are provided so that this disclosure will fully convey the invention to those skilled in the art. Many modifications and other embodiments of the invention will come to mind for those skilled in the art to which this invention pertains, having the benefit of the teachings presented herein. In particular, the neurological and neurodevelopmental disorders may be fragile X syndrome (FXS), neurofibromatosis I (NF1), tuberous sclerosis (TS), Down syndrome (DS), or autism spectrum disorder (ASD), amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), progressive supranuclear palsy (PSP), multiple system atrophy, Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease, and other related neurological diseases or conditions. These disorders also include, but are not limited to, neuropsychiatric conditions such as major depressive disorder (MDD), post-traumatic stress disorder (PTSD), and chemical dependency / substance abuse, such as cocaine use-related disorder. Collectively, these debilitating neurological conditions negatively impact the quality of life of patients and simultaneously place a burden on affected families and society as a whole.

[0007] The present invention is based on surprising results from studies using human postmortem brains and / or animal models of human neurological disease states. These results suggested that a catecholamine imbalance (e.g., but not limited to, excess adrenergic activity and deficiency of dopaminergic activity) is a component of disease pathogenesis. The present invention is also based on the finding that treatment of animal models of the human condition with a combination of drugs—one that enhances dopaminergic activity and another that attenuates adrenergic activity—can normalize the catecholamine imbalance and thereby alleviate disease symptoms. The present invention is also based on the finding that the same brain catecholamine imbalance affects many physiological functions. Physiological functions can include metabolism, such as changes in glucose disposal rate (including increases or decreases in glucose disposal rate), changes in glucose metabolic pathways (gluconeogenesis, glycolysis, and / or oxidative phosphorylation), and dyslipidemia (including increases in lipolysis or catabolism). Functions can include changes in cardiovascular function, such as changes in heart rate (both tachycardia and bradycardia) and / or heart rate variability (HRV). The same neurochemical functions can also include immunological states of pro- and anti-inflammatory activity (e.g., plasma IL-10 levels or the ratio of IL-10 to IL-17).

[0008] The present invention is also based on the findings that combined treatment of animal models of the human condition with multiple drugs (e.g., a drug with pharmacological activity that enhances dopaminergic activity and a drug that attenuates adrenergic activity) normalized physiological activity (e.g., lipid metabolism or glucose metabolism). Thus, specific physiological activities, such as metabolism, cardiac function, and / or immune status, can be used as objective biomarkers to assess brain catecholaminergic balance and treatment efficacy. The overall profile of tyrosine hydroxylase expression in different brain regions of FTD patients suggests the pathophysiology of catecholamine imbalance. As shown in Figures 1A and 1B, postmortem studies of neurodegenerative conditions typified by frontotemporal dementia (a type of condition commonly referred to as a tauopathy) found increased levels of tyrosine hydroxylase expression in the pons, which includes the locus coeruleus. Tyrosine hydroxylase is the rate-limiting enzyme in the synthesis of catecholamines (dopamine, norepinephrine, and epinephrine). The locus coeruleus is the primary brain region regulating the adrenergic state of the brain. Increased levels of tyrosine hydroxylase expression in the pons indicate increased adrenergic activity in the brain. These same studies also found decreased expression of the same enzyme in the putamen, which is part of the dorsal striatum and a key brain region regulating dopaminergic activity. A decrease in tyrosine hydroxylase levels in this brain region suggests a loss of dopamine synthesis capacity and dopaminergic cells, or a general loss of dopaminergic neurons, and therefore a deficiency in dopaminergic activity.

[0009] As shown in Figures 2A-2D, analysis of metabolomic data from patients with frontotemporal dementia compared to healthy individuals suggested metabolic abnormalities, such as increased lipid catabolism (β-hydroxybutyrate is a product of lipid β-oxidation) and dysregulated glucose metabolism. These metabolic changes suggest alterations in catecholamine balance, which is consistent with the differential expression of tyrosine hydroxylase mentioned above. P301L mice, which express frontotemporal dementia-associated mutations in the microtubule-associated protein tau (MAPT), are a mouse model of human neurological disorders such as frontotemporal dementia, Alzheimer's disease, or conditions collectively known as tauopathies. As shown in Figures 3A–3F, the behavioral and lipid metabolic patterns of drug-treated and untreated P301L mice were compared with those of SW mice (SW = Swiss Webster mice, the background strain of the transgenic P301L mice). Co-treatment of P301L mice with propranolol (a drug that attenuates β-adrenergic activity) and 3-(phenylpropyl)-sydnonimine-N-phenylcarbamoyl (a drug that enhances dopaminergic activity; see Chen et al., U.S. Patent No. 8,415,385) normalized the behavioral and lipid metabolic patterns of these P301L mice, modeling the human FTD condition.

[0010] As shown in Figures 4A-4D, brain studies of Fmr1-KO mice, a rodent model of human fragile X syndrome and neurodevelopmental disorders, demonstrated increased expression of dopamine reuptake proteins in the mouse striatum and decreased expression of norepinephrine reuptake proteins in the mouse brainstem (the locus coeruleus is a brainstem nucleus). Reuptake proteins regulate the extracellular concentrations of various catecholamines. The distinct expression patterns of these proteins suggested excessive adrenergic activity and a deficiency of dopaminergic activity (Figure 4B). The same knockout animals also exhibited increased glycolytic activity (increased plasma lactate levels), increased lipid catabolism (increased β-hydroxybutyrate), and dyslipidemia (low HDL and low cholesterol), which are similar to metabolic abnormalities in humans with fragile X syndrome. The altered metabolic activity suggested sympathetic dysregulation (i.e., catecholaminergic imbalance). Dopaminergic activity regulates sympathetic activation. Treatment of Fmr1KO mice with ACT01, which specifically activates dopaminergic activity, normalized glucose and lipid metabolism (Figure 4C).

[0011] The learning and memory behaviors of drug-treated and untreated (vehicle) Fmr1KO mice were compared with those of FVB mice (the background strain of transgenic Fmr1KO mice). Drug combinations, such as 3-(phenylpropyl)-sydnonimine-N-phenylcarbamoyl (also known as ACT01 or ACT) (25 mg / kg) and propranolol (Pro, 3 mg / kg), ACT01 (25 mg / kg), ACT01 (25 mg / kg) and prazosin (Praz, 2 mg / kg), and ACT01 (25 mg / kg) and carvedilol (Carv, 2 mg / kg), significantly (p<0.05) improved the learning and memory abilities (NOR index = novel object recognition index) of Fmr1KO mice. The effects of drug combinations were superior to those of single drugs. Specifically, co-treatment of Fmr1-KO mice with prazosin, carvedilol, or propranolol (drugs that attenuate adrenergic activity) and ACT01 (drug that enhances dopaminergic activity; see Chen et al., U.S. Pat. No. 8,415,385) restored learning and memory function and normalized lipid and glucose metabolic patterns in the animal model.

[0012] In a further embodiment, the dopamine reuptake inhibitor is 3-(phenylpropyl)-sydnonimine-N-phenylcarbamoyl, 3-(p-methylbenzyl)-sydnonimine-N-phenylcarbamoyl, 3-(p-carboxybenzyl)-sydnonimine-N-phenylcarbamoyl, 3-phenethyl-sydnonimine-N-phenylcarbamoyl, 3-phenethyl-sydnonimine-N-(3',4'-dichloro-phenyl)-carbamoyl, 3-(p-nitrophenyl)- 3-(3',5'-difluorobenzyl)-sydnonimine-N-(3',4'-dinitrophenyl)-carbamoyl, 3-(p-fluorobenzyl)-sydnonimine-N-phenylcarbamoyl, 3-benzyl-sydnonimine-N-phenylcarbamoyl, 3-phenethyl-sydnonimine-N-(p-chlorophenyl)-carbamoyl, 3-phenethyl-sydnonimine-N-(m-trifluoromethyl)-phenylcarbamoyl, 3-(3',5'-difluorobenzyl)-sydnonimine-N -phenylcarbamoyl, 3-(m-fluorobenzyl)-sydnonimine-N-phenylcarbamoyl, 3-(p-trifluoromethyl-benzyl)-sydnonimine-N-phenylcarbamoyl, 3-(p-tert-butylbenzyl)-sydnonimine-N-phenylcarbamoyl, 3-(p-methylbenzyl)-sydnonimine-N-(p'-trifluoromethyl-phenyl)carbamoyl, and 3-(p-methylbenzyl)-sydnonimine Selected from, but not limited to, nonimine-N-(p-dimethylamino-phenyl)carbamoyl, mesocarb, altropane, amfonelic acid, amineptine, BTCP, 3C-PEP, DBL-583, difluoropine, GBR-12783, GBR-12935, GBR-13069, GBR-13098, GYKI-52895, iometopan, modafinil, armodafinil, RTI-229, and vanoxerine.

[0013] One embodiment of the present invention provides a pharmaceutical composition comprising a dopaminergic agent and an adrenoceptor antagonist for the treatment of a neurological disease or condition. In further embodiments, the dopamine agonist is selected from, but is not limited to, apomorphine, bromocriptine, cabergoline, ciladopa, dihydrexidine, dinapsoline, doxantrin, epicriptine, L-dopa, lisuride, pergolide, piribedil, pramipexole, propylnorapomorphine, quinagolide, ropinirole, rotigotine, loxindol, sumanirole.

[0014] In further embodiments, the adrenoceptor antagonist is selected from, but is not limited to, propranolol, bucindolol, carteolol, carvedilol, labetalol, nadolol, oxprenolol, penbutolol, pindolol, sotalol, timolol, acebutolol, atenolol, betaxolol, bisoprolol, celiprolol, metoprolol, nebivolol, esmolol, butoxamine, prazosin, terazosin, doxazosin, silodosin, alfuzosin, and tamsulosin.

[0015] In another embodiment, the dopamine reuptake inhibitor is selected from 3-(phenylpropyl)-sydnonimine-N-phenylcarbamoyl (or mesocarb, or modafinil) and the adrenoceptor antagonist is propranolol, or carvedilol, or prazosin, or doxazosin, or tamsulosin. One embodiment of the present invention encompasses the co-administration of adrenergic blocking agents, such as β-adrenergic antagonists (e.g., propranolol) and / or β-α-adrenergic antagonists (e.g., carvedilol or labetalol), with agents that enhance dopaminergic activity, such as 3-(phenylpropyl)-sydnonimine-N-phenylcarbamoyl (Chen et al., U.S. Pat. No. 8,415,385), or agents with similar pharmacological properties, such as modafinil, to control neurological disease states caused by catecholamine imbalance. In a further embodiment of the invention, dopamine activity is affected by 3-(phenylpropyl)-sydnonimine-N-phenylcarbamoyl and adrenergic activity is controlled by the concomitant use of propranolol, prazosin, or carvedilol. In a further embodiment of the invention, the exact composition (and / or dosage) of the dopaminergic potentiator and adrenergic attenuator is determined by the particular biomarker profile of an individual patient and / or the patient's stage of disease progression and is prescribed accordingly.

[0016] The following examples are intended to illustrate certain aspects of the present disclosure and should not be construed as limiting the scope of the claims. The contents of all references, pending patent applications, and published patents cited throughout this application are expressly incorporated herein by reference. In particular, the description and related drawings herein provide exemplary information to demonstrate the therapeutic effect of the combination of 3-(phenylpropyl)-sydnonimine-N-phenylcarbamoyl and propranolol on animal behavior, but do not limit the scope of the present invention. The above methods can also include measuring biomarker levels in a subject in need of treatment. Also contemplated are pharmaceutical kits comprising a dopamine reuptake inhibitor or dopamine agonist, an adrenergic receptor antagonist, and reagents or devices for assessing and measuring at least one or more biomarkers.

[0017] The biomarkers used in the method and the kit may be any combination of glucose metabolism (including metabolite profiles), lipid metabolism (including metabolite profiles), hormone profiles, cytokine profiles, or cardiac function. Biomarkers of glucose metabolism may be, for example, glucose, lactate, pyruvate, the lactate-to-glucose ratio, or glucose disposal rate. Biomarkers of lipid metabolism and profile may be, for example, β-hydroxybutyrate, acetoacetate, high-density lipoprotein, or β-hydroxybutyrate or acetoacetate disposal rate. Biomarkers of hormone profiles may be, for example, insulin, cortisol (basal and evening cortisol levels), and / or leptin (basal and postprandial leptin levels). Biomarkers of cytokine profiles may be, for example, IL-10, IL-4, IL-17, IL-23, or their corresponding ratios. Biomarkers of cardiac function may be, for example, heart rate or heart rate variability. [Example]

[0018] Example 1: Postmortem study of frontotemporal dementia (FTD) Postmortem studies of neurodegenerative conditions, such as frontotemporal dementia (a type of condition commonly referred to as a tauopathy), have shown that individuals with FTD have increased levels of tyrosine hydroxylase expression in the pons and decreased levels in the putamen, compared with individuals without neurodegenerative diseases. The overall profile of tyrosine hydroxylase expression in different brain regions suggested the pathophysiology of catecholamine imbalance.

[0019] Postmortem human brain, pons, and putamen (part of the dorsal striatum) were stored at -80°C and used for protein analysis. Tissue sections were homogenized (approximately 100 μg tissue / 1 μL lysis buffer). Protein concentration was determined using a BCA assay (Pierce). The total protein concentration of denatured lysates from each animal was adjusted to the same total protein concentration (e.g., 1 μg / μL) so that each lane of the precast gel was loaded with the same amount of total protein (e.g., ≥5 μg / lane). Blots were developed using standard techniques, and images were captured and analyzed using ImageJ. Data were presented as relative optical density (OD) after normalization to the mean expression level of healthy controls. As shown in Figure 1A, the expression of tyrosine hydroxylase in the pons of FTD patients tended to be significantly increased compared to controls (p = 0.0526). In contrast, the expression level of this enzyme in the putamen of FTD patients was significantly decreased (p = 0.0258) (Figure 1B). The locus coeruleus, a nucleus in the pons, controls the brain's norepinephrine status, while the putamen, part of the dorsal striatum, contributes to the central dopamine status. The overall results of the protein analysis suggested an alteration in the central catecholamine status, namely, increased adrenergic activity and decreased dopaminergic activity.

[0020] (Example 2) Metabolomic data of FTD patients Analysis of metabolomic data from patients with frontotemporal dementia suggests metabolic abnormalities, such as increased lipid catabolism (β-hydroxybutyrate is a product of lipid β-oxidation) and increased glycolytic activity. Metabolic changes are associated with alterations in catecholamine balance, i.e., excessive sympathetic activity due to adrenergic activation and dopaminergic deficiency. Medical records containing metabolomic data from FTD patients, along with age-matched controls, were extracted from the UK Biobank (UKBB). Patients were divided into six age cohorts: <60 years, 60-64 years, 65-69 years, 70-74 years, 75-79 years, and ≥80 years. Metabolomic data were used to establish age-related patterns of lipid catabolic activity and to compare lipid catabolic activity under the influence of FTD.

[0021] As shown in Figure 2A, analysis of plasma β-hydroxybutyrate levels obtained from controls suggested a strong correlation between age and plasma β-hydroxybutyrate levels (R 2 =0.9924, p<0.0001, Y=0.0004622 * X + 0.02688). The analysis showed that "normal" metabolic aging involves a predictable course of increased lipid catabolism. Meanwhile, the same analysis using FTD patient data also suggested a correlation between age (or disease progression) and β-hydroxybutyrate levels in the FTD cohort (R 2 =0.5909, p<0.0094, Y=0.002753 * X-0.1113). Compared to controls, Figure 2B (slope: F = 4.484, DFn = 1, DFd = 7, P = 0.0720) 1 ), FTD showed an age- or progression-related inflection of accelerated lipid catabolism, suggesting excessive sympathetic (adrenergic excess and dopaminergic deficit) activation under the FTD condition. As shown in Figure 2C, analysis of plasma glucose and lactate levels obtained from controls suggested age-dependent changes in plasma lactate and glucose levels (R 2 =0.8453, p=0.0095, Y=-0.4567 * X+6.004). This analysis demonstrated that "normal" metabolic aging involves a predictable process in the decline of glycolytic activity.

[0022] Unlike healthy subjects (controls), the same analysis using data from FTD patients in Figure 2D showed no correlation, but did show trends suggesting increased glycolytic activity or dysregulation of glucose metabolism associated with age or disease progression. Example 3: Study of P301L mice (vs. SW) and testing of treatment effects In this experiment, P301L mice (Taconic Tau-Model2508, JNPL3(P301L)) were used to model human FTD, tauopathy, and Alzheimer's disease (Samaey, C., et al., Early Cognitive and Behavioral Deficits in Mouse Models for Tauopathy and Alzheimer's Disease. Front Aging Neurosci, 2019. 11: p. 335. Li, MZ, et al., Intracellular accumulation of tau inhibits autophagosome formation by activating TIA1-amino acid-mTORC1 signaling. Mil Med Res, 2022. 9(1): p. 38. Wenger, K., et al., Common mouse models of tauopathy reflect early but not late human disease. Mol Neurodegener, 2023. 18(1): p. 10.). An experimental outline is shown in Figure 3A. The mouse model expressed a transgene for the human P301L mutation of the microtubule-associated protein tau (MAPT) on a mixed background (Swiss Webster and C57 / B6). Between 14 and 15 weeks of age, metabolic profiles and behavioral patterns appeared different from control animals. The timing of these observed differences is similar to the timing of previously reported behavioral changes (Samaey, et al., 2019).

[0023] First (Figures 3B and 3C), to establish the relevance of the model to the human condition, i.e., dopaminergic deficiency and norepinephrine excess, we analyzed the expression levels of the norepinephrine transporter (NET) in the mouse brainstem (BS-NET) and the dopamine transporter (DAT) in the striatum (ST-DAT), Figures 3A and 3B. Mice were sacrificed at 15 weeks of age. Brain sections were isolated, frozen, and stored until analysis. The expression levels of DAT in the striatum and NET in the brainstem were analyzed using the same method as described above. Intracellular overexpression of tau leads to mTORC1 activation (Li, MZ, et al., Intracellular accumulation of tau inhibits autophagosome formation by activating TIA1-amino acid-mTORC1 signaling. Mil Med Res 2022. 9(1): p. 38). Such activation leads to upregulation of the dopamine transporter (DAT) and downregulation of the norepinephrine transporter (NET) (Bermingham and Blakely, 2016). Increased DAT expression reduces extracellular concentrations of dopamine, while decreased NET expression increases extracellular concentrations of norepinephrine. As shown in Figures 3B and 3C, P301L mice exhibited upregulated DAT expression in the striatum and decreased expression in the brainstem, thus creating a state of adrenergic excess and dopaminergic deficiency similar to the human condition of FTD. P301L mice were divided into two cohorts (n=5 in each cohort), one of which was treated with a combination of ACT01 (at 25 mg / kg) and propranolol (at 3 mg / kg), while the other P301L mice and age-matched SW mice were treated with vehicle as disease and healthy controls, respectively.

[0024] Treatment effects on animal behavior - locomotor activity (Figure 3D) Mice were placed in a pre-conditioned open field (box), and open field activity was recorded at a fixed pixel resolution. The distance traveled in the open field over several minutes (locomotor activity) was assessed using a freeware application (Kinovea) as the total number of pixel points traveled within 1 minute. Nest-building behavior (Figure 3E) Untreated mice (housed individually) are given 2x2 cotton nesting material (VWR, 10279-140) at the onset of the dark period, and the effect of treatment is assessed by measuring the weight of the "untreated" nesting material.

[0025] As shown in Figures 3D and 3E, simultaneous enhancement of dopaminergic activity by ACTO1 and modulation of adrenergic activity by propranolol normalized the behavioral patterns of the animals. Treatment effects on lipid metabolism in animals The resulting metabolic changes in male P301L mice were also characterized. In this study, serum β-hydroxybutyrate levels in drug-treated and untreated P301L mice were compared with SW mice (SW = Swiss Webster mice, the background strain of transgenic P301L mice) (Figure 3F). As shown, simultaneous mediation of various catecholaminergic activities by the dopamine reuptake inhibitor ACT01 and the β-blocker propranolol normalized lipid catabolic activity.

[0026] Example 4: Study of Fmr1KO mice (vs. FVB mice) and treatment efficacy test In this study, we compared the learning and memory behaviors of Fmr1KO mice treated with different drugs with those of untreated (vehicle) Fmr1KO mice with those of FVB mice (the background strain of transgenic Fmr1KO mice; "FVB" refers to the susceptibility of FVB mice to Friend leukemia virus B). <c-ch>Fmr1 <tm1cgr> / J (Stock No: 004624|FMR1 KO)) was used as an FXS / autism model, and age-matched male mice (FVB.129P2-Pde6b<+>Tyr <c-ch>(Bernardet, M. and W. E. Crusio, Fmr1 KO mice as a possible model of autistic features. Scientific World Journal, 2006. 6: pp. 1164-76. Hays, S. A., K. M. Huber, and J. R. Gibson, Altered neocortical rhythmic activity states in Fmr1 KO mice are due to enhanced mGluR5 signaling and involve changes in excitatory circuitry. J. Neurosci. 2011. 31(40): pp. 14223-34. Willemsen, R. and R. F. Kooy, Mouse models of fragile X-related disorders. Dis. Model. Mech. 2023. 16(2).) The experimental outline and results are shown in Figures 4A–4D.

[0027] Patients with fragile X syndrome develop macrocephaly, including enlargement of the putamen, at approximately 5 years of age (Shen, MD, et al., Subcortical Brain Development in Autism and Fragile X Syndrome: Evidence for Dynamic, Age- and Disorder-Specific Trajectories in Infancy. Am J Psychiatry, 2022.179(8): pp. 562-572; Williams, CA, A. Dagli, and A. Battaglia, Genetic disorders associated with macrocephaly. Am J Med Genet A, 2008.146A(15): pp. 2023-37; and Hazlett, HC, et al., Trajectories of early brain volume development in fragile X syndrome and autism. J Am Acad Child Adolesc Psychiatry, 2012.51(9): pp. 921-33). mTOR is an important mechanism for regulating growth. Excessive growth suggests excessive mTOR activity. TOR-related mutations cause macrocephaly in ASD (Yeung, KS, et al., Identification of mutations in the PI3K-AKT-mTOR signaling pathway in patients with macrocephaly and developmental delay and / or autism. Mol Autism, 2017. 8: p. 66.).

[0028] Activation of TOR activity has also been reported in the rodent model Fmr1KO (Huang, W.C., Y. Chen, and D.T. Page, Hyperconnectivity of prefrontal cortex to amygdala projections in a mouse model of macrocephaly / autism syndrome. Nat Commun, 2016. 7: p. 13421. Sharma, A., et al., Dysregulation of mTOR signaling in fragile X syndrome. J Neurosci, 2010. 30(2): p. 694-702). Although the relationship between TOR activity and the expression levels of various catecholamine transporters (or reuptake proteins) has been well documented (Bermingham, DP and RD Blakely, Kinase-dependent Regulation of Monoamine Neurotransmitter Transporters. Pharmacol Rev, 2016. 68(4): pp. 888–953), to examine catecholaminergic dysregulation, we analyzed the expression of NET in the cortex and DAT in the striatum (Figure 4B). Protein analysis (Western blot analysis using the method described above in Figures 1A and 1B) of brain sections (isolated according to the experimental procedure described above) from Fmr1KO mice (postnatal day 30) showed increased expression of DAT and decreased expression of NET (unlike neurodegenerative human and animal brains, protein expression levels are normalized to β-actin). Upregulation of DAT decreased extracellular dopamine, whereas downregulation of NET increased extracellular norepinephrine. This analysis established the fact that catecholamine status also changes in a similar manner as discussed above.

[0029] Treatment effects on metabolic activity in rodents: As shown in Figure 4C, these fragile X human model animals exhibited altered metabolic activity (e.g., lipids) similar to those observed in fragile X humans (Berry-Kravis, E., et al., Cholesterol levels in fragile X syndrome. Am J Med Genet A, 2015. 167A(2): pp. 379-84.), FTD patients, and P301L mice. These changes suggested excessive sympathetic activation. Dopaminergic enhancement by ACT01 appeared to attenuate sympathetic activation and normalize metabolic activity in the fragile X human model animals. Unexpected drug combination effects on rodent learning and memory: A novel object recognition test was used to assess the treatment effects of various catecholaminergic drugs (including drug combinations) on cognitive behaviors, namely learning and memory.

[0030] As shown in the experimental scheme (Figure 4A), mice were weaned on P21 (postnatal day 21) and immediately began daily treatment with the dopamine reuptake inhibitor 3-(phenylpropyl)-sydnonimine-N-phenylcarbamoyl (ACT01 or ACT), or the adrenergic antagonists propranolol (Pro), prazosin (Pra), carvedilol (Carv), or their corresponding combinations with ACT01. ACT01 was administered by oral gavage at 25 mg / kg, propranolol at 2 mg / kg, and prazosin at approximately 0.8 mg / kg; carvedilol at approximately 2 mg / kg. The control group received oral vehicle. After two days of treatment, animals followed the experimental protocol, receiving daily drug treatment 2 h before training / testing. Novel object recognition: On day 1 (habituation phase), each animal was introduced into the center of the opaque plastic arena and allowed to explore freely for 20 min. On the second day (sample phase), two identical objects were placed in opposite corners, and the mice were introduced into the center of the arena for a 5-minute training period. 24 hours later, the mice were returned to the arena for a 5-minute test period in which one of the objects was replaced with a novel one. Object recognition was measured using a preference index, which is the ratio of the time spent exploring the novel object to the total time spent exploring both objects.

[0031] The study showed (Figure 4D) that these drug combinations, such as 3-(phenylpropyl)-sydnonimine-N-phenylcarbamoyl (ACT01 or ACT) alone significantly enhanced cognition in animals (Fmr1-KO mice), whereas adrenergic blockers alone or individually showed no significant effect, although some trend was observed. The effect of the drug combination was significantly superior to any single drug (p<0.05 considered statistically significant).

Claims

1. a. dopamine reuptake inhibitors; b. Adrenergic receptor antagonists; and c. A pharmaceutically acceptable carrier A pharmaceutical composition comprising:

2. Dopamine reuptake inhibitors include 3-(phenylpropyl)-sydnonimine-N-phenylcarbamoyl, 3-(p-methylbenzyl)-sydnonimine-N-phenylcarbamoyl, 3-(p-carboxybenzyl)-sydnonimine-N-phenylcarbamoyl, 3-phenethyl-sydnonimine-N-phenylcarbamoyl, 3-phenethyl-sydnonimine-N-(3',4'-dichloro-phenyl)-carbamoyl, 3-(p-nitrophenethyl)-sydnonimine-N-phenylcarbamoyl, nonimine-N-(3',4'-dinitro-phenyl)-carbamoyl, 3-(p-fluorobenzyl)-sydnonimine-N-phenylcarbamoyl, 3-benzyl-sydnonimine-N-phenylcarbamoyl, 3-phenethyl-sydnonimine-N-(p-chlorophenyl)-carbamoyl, 3-phenethyl-sydnonimine-N-(m-trifluoromethyl)-phenylcarbamoyl, 3-(3',5'-difluorobenzyl)-sydnonimine-N-phenyl carbamoyl, 3-(m-fluorobenzyl)-sydnonimine-N-phenylcarbamoyl, 3-(p-trifluoromethyl-benzyl)-sydnonimine-N-phenylcarbamoyl, 3-(p-tert-butylbenzyl)-sydnonimine-N-phenylcarbamoyl, 3-(p-methylbenzyl)-sydnonimine-N-(p'-trifluoromethyl-phenyl)carbamoyl, and 3-(p-methylbenzyl)-sydnonimine 2. The pharmaceutical composition of claim 1, wherein the compound is amine-N-(p-dimethylamino-phenyl)carbamoyl, mesocarb, altropane, amfonelic acid, amineptine, BTCP, 3C-PEP, DBL-583, difluoropine, GBR-12783, GBR-12935, GBR-13069, GBR-13098, GYKI-52895, iometopan, modafinil, armodafinil, RTI-229, or vanoxerine.

3. 3. The pharmaceutical composition according to claim 2, wherein the dopamine reuptake inhibitor is 3-(phenylpropyl)-sydnonimine-N-phenylcarbamoyl, or modafinil, or mesocarb.

4. a. dopamine agonists; b. Adrenergic receptor antagonists; and c. A pharmaceutically acceptable carrier A pharmaceutical composition comprising:

5. 5. The pharmaceutical composition according to claim 4, wherein the dopamine agonist is apomorphine, cabergoline, dihydrexidine, dinapsoline, doxantrin, epicriptine (β-dihydroergocryptine), L-dopa (levodopa and L-3,4-dihydroxyphenylalanine), lisuride, piribedil, pramipexole, quinagolide, ropinirole, rotigotine, lokiindole, or sumanirole.

6. 6. The pharmaceutical composition according to claim 5, wherein the dopamine agonist is L-dopa, apomorphine, or rotigotine.

7. 7. The pharmaceutical composition of any one of claims 1 to 6, wherein the adrenoceptor antagonist is propranolol, bucindolol, carteolol, carvedilol, labetalol, nadolol, oxprenolol, penbutolol, pindolol, sotalol, timolol, acebutolol, atenolol, betaxolol, bisoprolol, celiprolol, metoprolol, nebivolol, esmolol, butoxamine, prazosin, terazosin, doxazosin, silodosin, alfuzosin, or tamsulosin.

8. 8. The pharmaceutical composition of claim 7, wherein the adrenergic receptor antagonist is propranolol, carvedilol, prazosin, doxazosin, or tamsulosin.

9. A method for treating a neurological disease, comprising administering the composition of any one of claims 1 to 8 to a subject in need thereof.

10. 10. The method of claim 9, wherein the neurological disease is Fragile X syndrome (FXS), Neurofibromatosis I (NF1), Tuberous Sclerosis (TS), Down's Syndrome (DS), or Autism Spectrum Disorder (ASD), Amyotrophic Lateral Sclerosis (ALS), Frontotemporal Dementia (FTD), Progressive Supranuclear Palsy (PSP), Alzheimer's Disease (AD), Parkinson's Disease (PD), Major Depressive Disorder (MDD), Post-Traumatic Stress Disorder (PTSD), or Cocaine Use Related Disorder.

11. 11. The method of claim 10, wherein the neurological disease is fragile X syndrome, autism spectrum disorder (ASD), amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), or Alzheimer's disease (AD).

12. further comprising measuring a level of a biomarker in the subject, wherein the biomarker is: a. glucose metabolism and metabolite profile (e.g., glucose, pyruvate, lactate), or b. Lipid metabolism and profile, or c. hormone profile, or d. cytokine profile, or e. Cardiac function 12. The method according to any one of claims 9 to 11, wherein

13. The method of claim 12, wherein the measuring step comprises measuring glucose metabolism and its profile, and / or lipid metabolism and its profile, and / or hormone profile (including dynamic profile), and / or cytokine profile (or ratios thereof), or biomarker levels of cardiac function.

14. 14. The method of claim 12 or 13, wherein the glucose metabolism biomarker is glucose, lactate, or pyruvate, or the lactate to glucose ratio, or glucose disposal rate.

15. 15. The method of any one of claims 12 to 14, wherein the biomarkers of lipid metabolism and profile are β-hydroxybutyrate, or acetoacetate, and / or high density lipoproteins, triglycerides, or the ratio of triglycerides to β-hydroxybutyrate, or the ratio of free fatty acids to β-hydroxybutyrate, or the disposal rate of β-hydroxybutyrate or acetoacetate.

16. 16. The method of any one of claims 12 to 15, wherein the metabolite profile consists of β-hydroxybutyrate, lactate, and glucose.

17. The method of any one of claims 12 to 16, wherein the biomarkers of the hormone profile are cortisol (basal and evening), leptin (basal and postprandial), or cytokine profile.

18. The method of any one of claims 12 to 17, wherein the biomarker of the hormone profile is basal leptin and / or postprandial leptin.

19. 17. The method of claim 16, wherein the biomarkers of the hormone profile are the biomarkers of the cytokine profile IL-10, IL-4, and / or IL-17.

20. The method of any one of claims 12 to 19, wherein the biomarker of cardiac function is heart rate or heart rate variability.

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