Preventive efficacy of serotonin 4 receptor agonists against stress

Abstract

Provided is a method for prophylactically treating stress-induced affective disorder or stress-induced psychosis in a subject.Also provided is a method for inducing and / or enhancing stress resilience in a subject.In certain embodiments, an effective amount of an agonist of serotonin 4 receptor (5-HT4R) or its pharmaceutically acceptable salt or derivative is administered to the subject before a stressor.

Description

Technical Field

[0001] Cross - Reference to Related Applications This application claims priority to U.S. Provisional Patent Application No. 62 / 831,517, filed on April 9, 2019, U.S. Provisional Patent Application No. 62 / 857,075, filed on June 4, 2019, and U.S. Provisional Patent Application No. 62 / 910,859, filed on October 4, 2019, the entire disclosures of which are hereby incorporated by reference herein in their entireties.

[0002] Field of the Invention The present invention relates to serotonin 4 receptor (5 - hydroxytryptamine (serotonin) receptor 4, or 5 - HT 4 R) agonist compositions and their use in methods for treating or preventing stress - induced mood disorders such as post - traumatic stress disorder (PTSD). In certain embodiments, the compositions of the present invention can be administered prior to exposure to a stressor.

Background Art

[0003] Background of the Invention Exposure to stress is an important factor in the development of major depressive disorder (MDD) and post - traumatic stress disorder (PTSD). According to the National Comorbidity Survey, approximately 60% of men and 51% of women have been exposed to one or more traumatic events during their lifetime. 7.8% of the total population experiences PTSD at some point in their life, and women (10.4%) experience this disorder at a significantly higher rate than men [5.0%] [1]. Women are 2 - 3 times more likely to suffer from stress - related anxiety or depressive disorders than men

[61] . Traditionally, mood disorders have been treated from an approach of symptom suppression. Existing drugs are aimed at reducing the effects of these chronic diseases but do not cure or prevent the diseases themselves. However, if drugs that enhance stress resilience are developed, they could potentially be used to defend against stress - induced mental disorders in at - risk populations.

[0004] Anxiety disorder is one of the most common mental disorders, with a lifetime prevalence of over 25% (A1) and an annual financial burden of over $40 billion (A2). Benzodiazepines (BZDs) are effective in treating most anxiety disorders and have been the standard treatment for many years, with a response of over 80% in reducing acute anxiety in patients (A3). However, their long-term daily use has been associated with the risks of dependence and memory loss. As a result, they have often been replaced by chronic treatment with serotoninergic agents such as selective serotonin reuptake inhibitors (SSRIs) that focus on the role of serotonin (5-HT) in treating anxiety disorders. However, since SSRIs have a delayed onset of action for several weeks and a non-responder rate of 40% in anxiety patients (A4), the development of new rapid-acting anxiolytics is required.

[0005] The inventors and others have recently reported that (R,S)-ketamine acts as a resilience enhancer (e.g., a prophylactic agent) against stress when administered one week before stress in mice [2 - 6]. In addition, limited data in human patients have demonstrated the potential of (R,S)-ketamine in preventing mental disorders such as post-traumatic stress disorder (PTSD) [7] and possibly postpartum depression (PPD) [8, 9] in a dose-specific manner. The effectiveness of prophylactic agents has been limited to (R,S)-ketamine until Gould and co-workers recently reported that group II metabotropic glutamate receptor (mGlu 2 / 3 ) antagonists are also protective

[10] . The inventors have previously reported that the SSRI Flx is ineffective as a prophylactic agent, but whether other serotoninergic agents can be effective prophylactic agents and / or whether the serotoninergic system is involved in prophylactic efficacy remains undetermined.

[0006] 5-HT 4 5-HT1AR is a promising target for treating depression and anxiety disorders. 5-HT 4 5-HT1AR responds to 5-HT to activate G SA metabotropic G protein-coupled receptor that stimulates the cyclic adenosine monophosphate (cAMP) / protein kinase A (PKA) signaling pathway [11-15]. 5-HT 4 R is highly expressed in the periphery including the heart and adrenal glands, as well as in regions such as the amygdala (AMG), medial prefrontal cortex (mPFC), nucleus accumbens (NAc), and hippocampus (HPC) in the brain [16, 17]. 5-HT 4 R knockout mice exhibit increased anxiety-like and depressive-like behaviors, while 5-HT 4 Activation of R stimulates neurogenesis in the HPC and produces a rapid antidepressant-like effect [18-21]. However, 5-HT 4 Whether and how R is involved in stress resilience has not yet been determined.

[0007] There is an unmet need for effective preventive treatments to prevent the onset of stress-induced mood disorders.

Summary of the Invention

Problems to be Solved by the Invention

[0008] Summary of the Invention The present disclosure provides a method for preventing or delaying stress-induced mood disorders or stress-induced psychosis in a subject in need thereof. The method may comprise administering to the subject, prior to exposure to a stressor, an effective amount of an activator of the serotonin 4 receptor (5-HT 4 R) (e.g., an agonist of the serotonin receptor (5-HT 4 R)), or a pharmaceutical composition comprising a pharmaceutically acceptable salt, analog, derivative or metabolite thereof.

[0009] The present disclosure also provides a method for inducing and / or enhancing stress resilience in a subject in need thereof. The method may comprise administering to the subject, prior to exposure to a stressor, an activator of the serotonin 4 receptor (5-HT 4 R) (e.g., an agonist of the serotonin receptor (5-HT 4administering an effective amount of a pharmaceutical composition comprising an agonist of R), or a pharmaceutically acceptable salt, analog, derivative or metabolite thereof. **Means for Solving the Problems**

[0010] 5-HT 4 The agonist of R may include 1-(4-amino-5-chloro-2-methoxyphenyl)-3-[1(n-butyl)-4-piperidinyl]-1-propanone HCl (RS-67,333 or RS67333), 4-amino-5-chloro-2,3-dihydro-N-[1-3-methoxypropyl)-4-piperidinyl]-7-benzofurancarboxamide monohydrochloride (purcaropride), 4-[4-[4-tetrahydrofuran-3-yloxy)-benzod]isoxazol-3-yloxymethyl]-piperidin-1-ylmethyl]-tetrahydropyran-4-ol (PF-04995274), or a combination thereof.

[0011] The pharmaceutical composition may be administered to the subject about 48 hours to about 3 weeks before the stress. In certain embodiments, the pharmaceutical composition is administered to the subject about 72 hours to about 2 weeks before the stress. In certain embodiments, the pharmaceutical composition is administered to the subject about 1 week before the stress.

[0012] In certain embodiments, the pharmaceutical composition is administered to the subject once prior to the stress.

[0013] In certain embodiments, the pharmaceutical composition is administered to the subject orally, intravenously, intranasally, or via injection.

[0014] The stress-induced mood disorder may include major depressive disorder (MDD) and / or post-traumatic stress disorder (PTSD). In certain embodiments, the stress-induced mood disorder is selected from the group consisting of depressive-like behaviors and related mood disorders, anhedonic behaviors and related mood disorders, anxiety disorders and related mood disorders, cognitive impairments and deficiencies and related disorders, stress-induced phobias, and combinations thereof.

[0015] In additional embodiments, stress-induced mood disorder includes stress-induced psychosis. In certain embodiments, stress-induced psychosis includes depressive behavior and / or anxiety behavior.

[0016] The methods of the invention may prevent or delay stress-induced cognitive impairment and / or decline.

[0017] The methods of the invention may further comprise administering to the subject an effective amount of an antidepressant, an antianxiety agent, or a combination thereof.

[0018] The methods of the invention may further comprise administering to the subject an effective amount of a selective serotonin reuptake inhibitor (SSRI), or a pharmaceutically acceptable salt or derivative thereof.

[0019] The methods of the invention may further comprise administering to the subject an effective amount of fluoxetine, paroxetine, sertraline, lithium, riluzole, prazosin, lamotrigine, ifenprodil, or a combination thereof.

[0020] The subject may be a mammal. In certain embodiments, the subject is a human. The subject may be female or male.

[0021] In certain embodiments, the pharmaceutical composition is administered in a booster series. BRIEF DESCRIPTION OF THE DRAWINGS

[0022]

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Mode for Carrying Out the Invention

[0023] Detailed Description of the Invention The present disclosure provides a method for prophylactically treating stress-induced affective disorder or stress-induced psychosis in a subject. Also included in the present disclosure is a method for inducing and / or enhancing stress resilience in a subject. In certain embodiments, a serotonin 4 receptor (5-HT ) antagonist, such as RS-67333 (RS67333), prucalopride, PF-04995274, or a pharma- ceutical acceptable salt, analog, derivative, or metabolite thereof, is used. 4 R) activators (e.g., serotonin 4 receptor (5-HT 4 An effective amount of an agonist of R) is administered to the subject prior to the stressor.

[0024] The agent / composition of the present invention may be administered therapeutically to achieve therapeutic benefit or prophylactically to achieve prophylactic benefit. Therapeutic benefit refers to eradication or amelioration of the underlying stress-induced affective disorder being treated, and / or eradication or amelioration of one or more of the symptoms associated with the underlying disorder. Prophylactic benefit refers to prevention or delay of the onset of stress-induced affective disorder, and / or prevention or delay of the onset of one or more of the symptoms associated with stress-induced affective disorder. In certain embodiments, an effective amount of the agent / composition of the present invention administered prevents stress-related disorder from developing or worsening into a more severe state.

[0025] In certain embodiments, for prophylactic administration, the agent / composition of the present invention may be administered to a patient at risk of developing stress-induced affective disorder, even if the diagnosis of stress-induced affective disorder has not yet been made, or to a patient who has reported one or more of the physiological symptoms of stress-induced affective disorder. In certain embodiments, prophylactic administration is applied to avoid the onset of physiological symptoms of the underlying disorder before the symptoms are clinically manifested. In this latter embodiment, the treatment is preventative with respect to the associated physiological symptoms instead of the underlying indication. In certain embodiments, the agent / composition of the present invention is administered prior to the recurrence of the stressor. In certain embodiments, the agent / composition of the present invention is administered prior to the onset of the characteristic symptoms.

[0026] In a further embodiment, the present invention provides the use of the agent of the present invention, or a pharmaceutically acceptable salt or solvate thereof, a physiologically functional derivative or analog thereof, or a metabolite thereof, in the preparation of a medicament for the treatment of stress-induced mood disorders.

[0027] "Treating" or "treatment" of a condition, disorder or state includes (1) preventing or delaying the appearance of clinical symptoms of a condition, disorder or state in a person who may be troubled by, or be predisposed to, the condition, disorder or state but who has not yet experienced or presented the clinical symptoms of the condition, disorder or state; or (2) inhibiting the condition, disorder or state, i.e., arresting the onset or recurrence (in the case of maintenance treatment) of the disease, or at least halting, reducing or delaying one or more clinical symptoms, signs or tests thereof; or (3) alleviating the disease, i.e., causing at least a regression of the condition, disorder or state, or one or more clinical or subclinical symptoms or signs.

[0028] The benefit to the subject being treated is either statistically significant or at least perceptible to the patient or physician.

[0029] Examples of the agent of the present invention include 5-HT 4 R agonists such as RS-67,333 (RS67333), prucalopride, PF-04995274, pharmaceutically acceptable salts or solvates thereof, analogs thereof, derivatives thereof (e.g., physiologically functional derivatives or analogs thereof), or metabolites thereof, and combinations thereof.

[0030] "Prophylactically effective amount" refers to an amount effective to achieve the desired prophylactic result, the required dosage and duration. In certain embodiments, since the prophylactic dosage is used in a subject prior to or at an early stage of the disorder, the prophylactically effective amount is less than the therapeutically effective amount. In certain embodiments, the prophylactically effective amount is similar to, equivalent to, or greater than the therapeutically effective amount.

[0031] A therapeutically effective amount, or effective amount, of a drug is an amount effective to demonstrate the desired activity of the drug. The "therapeutically effective amount" will vary depending on the compound, the disorder and its severity, and the age, weight, physical condition and responsiveness of the subject being treated. In certain embodiments, 5-HT 4 An effective amount of an R agonist, or a pharmaceutically acceptable salt or solvate thereof, or a physiologically functional derivative or analog thereof, or a metabolite thereof, is an amount effective to prevent or delay the onset of stress-induced mood disorder and / or to reduce one or more of the symptoms of stress-induced mood disorder.

[0032] The present disclosure provides a method for preventing or delaying stress-induced mood disorder or stress-induced psychosis in a subject in need thereof. The method may comprise administering to the subject, prior to exposure to a stressor, an effective amount of a pharmaceutical composition comprising an activator of serotonin 4 receptor (5-HT 4 R) (e.g., an agonist of serotonin 4 receptor (5-HT 4 R)) or a pharmaceutically acceptable salt, analog, derivative or metabolite thereof.

[0033] The present disclosure also provides a method for inducing and / or enhancing stress resilience in a subject in need thereof. The method may comprise administering to the subject, prior to exposure to a stressor, an effective amount of a pharmaceutical composition comprising an activator of serotonin 4 receptor (5-HT 4 R) (e.g., an agonist of serotonin 4 receptor (5-HT 4 R)) or a pharmaceutically acceptable salt, analog, derivative or metabolite thereof.

[0034] The compositions of the present invention may be administered by any method known to those skilled in the art, including, but not limited to, intranasal, oral, transdermal, intraocular, intraperitoneal, inhalation, intravenous, intracerebroventricular (ICV), intracisternal injection or infusion, subcutaneous, implant, intravaginal, sublingual, urethral (e.g., urethral suppository), subcutaneous, intramuscular, intravenous, rectal, sublingual, mucosal, ophthalmic, spinal, intrathecal, intra-articular, intra-arterial, subarachnoid, bronchial, and lymphatic administration. Topical formulations may be in the form of gels, ointments, creams, aerosols, etc.; intranasal formulations may be delivered as sprays or droplets; transdermal formulations may be administered via transdermal patches or iontophoresis; inhalation formulations may be delivered using nebulizers or similar devices. The compositions may also take the form of tablets, pills, capsules, semi-solids, powders, sustained-release formulations, solutions, suspensions, elixirs, aerosols, or any other suitable composition.

[0035] In certain embodiments, the subject is treated with the agent / composition of the present invention via intravenous, oral, transdermal, or intranasal administration. In certain embodiments, the subject is injected with the agent / composition of the present invention.

[0036] In certain embodiments, the subject is treated with a single dose of an effective amount of the agent / composition of the present invention prior to, during, and / or after a stressor. In some aspects, the subject is treated with multiple doses of an effective amount of the agent / composition of the present invention prior to, during, and / or after a stressor.

[0037] In certain embodiments, 5-HT 4 R agonists (RS-67,333 (RS67333), prucalopride, and PF-04995274), or a pharmaceutically acceptable salt or solvate thereof, an analog thereof, a derivative thereof, or a metabolite thereof, are administered in a composition comprising a pharmaceutically acceptable carrier, excipient, or diluent. Also provided herein is the use of 5-HT 4An R agonist (RS-67,333 (RS67333), prucalopride, and PF-04995274), or a pharmaceutically acceptable salt or solvate thereof, an analog thereof, a derivative thereof, or a metabolite thereof, together with a pharmaceutically acceptable carrier, excipient or diluent, is a pharmaceutical composition.

[0038] "Patient" or "subject" refers to a mammal and includes human and veterinary subjects. In certain embodiments, the subject is a mammal.

[0039] The agent of the present invention (e.g., 5-HT 4 R activator) can, without limitation, activate / increase 5-HT 4 R activity, activate / increase 5-HT 4 R levels, and / or activate / increase 5-HT 4 R gene expression, and activate 5-HT 4 R through any mechanism including this. The terms "5-HT 4 R activator", "5-HT4 receptor activator", "5-HT4 receptor activator", and "5-HT 4 R activator" are used interchangeably herein.

[0040] "Activation", "upregulation" or "increase" means any positive effect exerted on the conditions being tested, which may be whole or partial. Thus, when the level or activity of a protein (e.g., 5-HT4 receptor or 5-HT 4 R) is detected, the agent / composition of the present invention can activate, upregulate or increase the level or activity of the protein (e.g., 5-HTR receptor or 5-HT 4 R). The activation or upregulation of the level or activity of the protein achieved by the agent of the present invention is the protein (e.g., 5-HT4 receptor or 5-HT 4At least 10%, such as at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or more, compared to the level or activity of (R).

[0041] The half-maximal effective concentration (EC50) refers to the concentration of a drug that induces a response that is half the difference between the basal value and the maximum value after a specified exposure time. pEC50 is defined as the negative logarithm of EC50: pEC50 = -log 10 (EC50).

[0042] In certain embodiments, the agents of the invention have a pEC50 in the range of about 3 to about 13, about 4 to about 12, about 5 to about 11, about 6 to about 10, about 6 to about 9, about 6 to about 8, about 6 to about 7, about 7 to about 10, about 7 to about 9, about 7 to about 8, about 8 to about 10, about 8 to about 9, about 9 to about 10, about 5 to about 10, about 6, about 6.5, about 7, about 7.5, about 8, about 8.5, about 9, about 9.5, or about 10 when activating 5-HT4 receptor activity.

[0043] Ki represents the affinity of a drug (an activator such as an agonist) for a receptor. When measured using a radioligand competition binding assay, it is the molar concentration of the competing ligand that occupies 50% of the receptor in the absence of the radioligand. pKi is the negative logarithm of the Ki value.

[0044] In certain embodiments, the agents of the invention have a pKi for the 5-HT4 receptor in the range of about 3 to about 13, about 4 to about 12, about 5 to about 11, about 6 to about 10, about 6 to about 9, about 6 to about 8, about 6 to about 7, about 7 to about 10, about 7 to about 9, about 7 to about 8, about 8 to about 10, about 8 to about 9, about 9 to about 10, about 5 to about 10, about 6, about 6.5, about 7, about 7.5, about 8, about 8.5, about 9, about 9.5, or about 10.

[0045] 5-HT 4 R agonist 5-HT 4R is a G protein-coupled receptor (GPCR) that activates the G protein Gs and stimulates the cAMP / PKA signaling pathway, resulting in phosphorylation of the cAMP response element-binding protein (CREB), and consequently, expression of a number of genes involved in neuroplasticity (A10). 5-HT 4 Most of R is expressed in the brains of primates and rodents, specifically in the medium spiny neurons of the striatum, the Cornu Ammonis (CA1 and CA3) of the hippocampus, the granule cells of the dentate gyrus, as well as the glutamatergic neurons of the cortex and amygdala (A11). In addition, 5-HT 4 R is also found in the hypothalamus, ventral pallidum, olfactory bulb, septal area, and substantia nigra. 5-HT 4 Mice lacking R exhibit anhedonic behavior and context-dependent anxiety-like behavior (A12), and various 5-HT 4 R agonists can exert antidepressant and anti-anxiety-like activities (A6).

[0046] Regardless of whether in humans or rodents, the expression of the serotoninergic type 4 receptor (5-HT 4 ) is found in the limbic brain regions ( m PFC, HPC and NAc). In addition, the basal ganglia, namely the caudate nucleus and lenticular nucleus (putamen and globus pallidus), substantia nigra, and amygdala also express the 5-HT 4 receptor. 5-HT 4 receptor is expressed at the somatodendritic level and at the axon terminal level of the centrifugal spinal GABAergic neurons of the striatum, the Cornu Ammonis (CA1 and CA3) of the hippocampus, the granule cells of the dentate gyrus, as well as the glutamatergic neurons of the cortex, hippocampus and amygdala.

[0047] 5-HT 4 receptor is also found at the peripheral level, particularly at the cardiac level where activation exerts a positive inotropic effect, at the gastrointestinal level involved in intestinal motility, at the adrenal level playing a role in the secretion of corticosteroids, and at the bladder level causing smooth muscle contraction.

[0048] 5-HT 4The receptor is a receptor having seven transmembrane domains. The N-terminal region faces the extracellular environment, while the C-terminal domain conjugated to the Gs protein faces the cytoplasm. For example, activation of the 5-HT 4 receptor by an agonist can lead to the recruitment of the Gs protein that stimulates adenylate cyclase (AC) responsible for cAMP production. Protein kinase A (PKA) activated by cAMP modulates different ion currents, particularly potassium currents, and its inhibition leads to neuronal hyperexcitation. PKA can also phosphorylate the protein that binds to the response element for cAMP (CREB - cAMP response element binding protein), which results in an increase in the transcription of the neurotrophic brain factor (BDNF, brain-derived neurotrophic factor) involved in cognition, mood, and cell survival.

[0049] The term "agonist" may refer to a substance, agent, or compound that can bind to and activate one or more receptors such as 5-HT 4 R. The term "agonist" may refer to a compound that has the ability to initiate or enhance the biological function of a target protein (e.g., one or more receptors such as 5-HT 4 R), regardless of whether it enhances or initiates the activity or expression of the target protein. A 5-HT 4 R agonist may be a compound that activates the action of the 5-HT4 receptor. The term "agonist" may be defined in the context of the biological role of the target protein. In one embodiment, an agonist is an agent that binds to a receptor (e.g., 5-HT 4 R), activates the receptor, and produces a biological response. The agonists provided herein can specifically interact (e.g., bind) with the target protein, but compounds that initiate or enhance the biological activity of the target protein by interacting with other members of the signal transduction pathway of which the target protein is a member are also specifically included in this definition. A 5-HT 4 R agonist may be a compound or agent that activates the action of 5-HT 4 R. A 5-HT 4An R agonist is any agent that acts directly or indirectly through 5-HT 4 R, or acts on 5-HT 4 R, to produce a pharmacological effect. The terms "5-HT 4 R agonist", "5-HT4 receptor agonist", "5-HT4 receptor agonist" and "5-HT 4 R agonist" are used interchangeably herein.

[0050] A 5-HT 4 R agonist may be selective for the 5-HT4 receptor, or it may be non-selective and exhibit agonist or antagonist activity at other serotonin receptors. In one embodiment, the 5-HT 4 R agonist is selective for the 5-HT4 receptor.

[0051] A 5-HT 4 R agonist may include full agonists, partial agonists, 5-HT 4 R agonist / antagonist mixtures, etc.

[0052] "Full agonist" may refer to an agent that binds to and activates a receptor with the maximum response that the agonist can elicit at the receptor. Depending on the relative number of receptors and differences in receptor coupling, the agent may act as a full agonist in some tissues and as a partial agonist in other tissues.

[0053] A "partial agonist" can bind to and activate a given receptor but have only partial efficacy at the receptor compared to a "full agonist" or complete agonist. A partial agonist can act as an antagonist when it competes with a full agonist for receptor occupancy and causes a net decrease in receptor activation compared to the effects or activation observed with the full agonist alone. A partial agonist may refer to an agonist / antagonist mixture, which differentially affects receptor function within different dosage ranges. For example, a partial agonist may serve as an agonist at low doses and as an antagonist at high doses. A partial agonist may be a compound having reduced efficacy (typically 40 - 80%) for inducing a conformational change in the receptor compared to a full agonist, which induces an agonist effect at low doses but may induce an antagonist effect at high doses.

[0054] 5-HT 4 The R agonist may be an indole, benzamide, benzoate, aryl ketone, or benzamide.

[0055] 5-HT 4 Non-limiting examples of the R agonist include 1-(4-amino-5-chloro-2-methoxyphenyl)-3-[1(n-butyl)-4-piperidinyl]-1-propanone HCl (RS-67,333 or RS67333), 4-amino-5-chloro-2,3-dihydro-N-[1-3-methoxypropyl)-4-piperidinyl]-7-benzofurancarboxamide monohydrochloride (purcaropride), 4-[4-[4-tetrahydrofuran-3-yloxy)-benzo[d]isoxazol-3-yloxymethyl]-piperidin-1-ylmethyl]-tetrahydropyran-4-ol (PF-04995274), and combinations thereof. Also 5-HT 4Non-limiting examples of R agonists include 2-[1-(4-piperonyl)piperazinyl]benzothiazole (PPB), 5-methoxytryptamine, PRX-03140, cisapride, ((±)-cis-4-amino-5-chloro-N-[1-[3-(4-fluorophenoxy)propyl]-3-methoxy-4-piperidinyl]-2-methoxybenzamide monohydrate), BIMU-8 (2,3-dihydro-N-((3-exo)-8-methyl-8-azabicyclo[3.2.1]oct-3-yl]-3-(1-methylethyl)-2-oxo-1H-benzimidazole-1-carboxamide), RS67506 ((methylsulfonylamino)ethyl-4-piperidinyl]-1-propanone hydrochloride), mosapride (4-amino-5-chloro-2-ethoxy-N-[[4-((4-fluorophenyl)methyl]-2-morpholinyl]methyl]benzamide citrate), tegaserod (2-[(5-methoxy-1H-indol-3-yl)methylene]―N-pentyl-hydrazinecarboximide amide maleate), ML10302 (4-amino-5-chloro-2-methoxybenzoic acid 2-(1-piperidinyl)ethyl ester hydrochloride), velusetrag (TD-5108) (N-[(1R,3R,5S)-8-[(2R)-2-hydroxy-3-(N-methylmethanesulfonamide)propyl]-8-azabicyclo[3.2.1]octane-3-yl]-2-oxo-1-(propan-2-yl)-1,2-dihydroquinoline-3-carboxamide), naropride (ATI-7505) ([(3R)-1-azabicyclo[2.2.2]octane-3-yl]6-[(3S,4R)-4-[(4-amino-5-chloro-2-methoxybenzoyl)amino]-3-methoxypiperidin-1-yl]hexanoate, cinetapride (4-amino-N-[1-(cyclohex-3-en-1-ylmethyl)piperidin-4-yl]-2-ethoxy-5-nitrobenzamide), metoclopramide (4-amino-5-chloro-N-(2-(diethylamino)ethyl)-2-methoxybenzamide), lenzapride (ATL-1251, BRL24924, (±)-endo-4-amino-5-chloro-2-methoxy-N-(1-azabicyclo[3.3.1] Non-4-yl)benzamide), RQ-00000010 (4-{[4-({[4-(2,2,2-trifluoroethoxy)-1,2-benzisoxazol-3-yl]oxy}methyl)piperidin-1-yl]methyl}tetrahydro-2H-pyran-4-carboxylic acid), SUVN-D4010 (1-isopropyl-3-{5-[1-(3-methoxypropyl)piperidin-4-yl]-[1,3,4]oxadiazol-2-yl}-1H-indazole), TD-8954 (4-{ (4-[(2-isopropyl-1H-benzimidazole-4-carbonyl)amino]methyl}-piperidin-1-ylmethyl)piperidine-1-carboxylic acid methyl ester), SC53116 (4-amino-5-chloro-N-[[(1S,7aS)-hexahydro-1H-pyrrolidin-1-yl]methyl]-2-methoxy-benzamide), BIMU-1 (3-ethyl-2,3-dihydro-N-(8-methyl-8-azabicyclo[3.2.1]oct-3-yl)-2-oxo-1H-benzimidazole-1-carboxamide hydrochloride), Donepezil (MR31147, 1-(4-amino-5-chloro-2-methoxyphenyl)-3-[1-(cyclohexylmethyl)-4-piperidinyl]propan-1-one); LS650155 (Caeserod, 5-(8-amino-7-chloro-2,3-dihydrobenz[b][1,4]dioxin-5-yl)-3-(1-phenethylpiperidin-4-yl)-1,3,4-oxadiazol-2(3H)-one hydrochloride); PF-00885706: N-[2-[(1R,8S)-4-[[4-(cyclobutylamino)-5-(trifluoromethyl)pyrimidin-2-yl]amino]-11-azatricyclo[6.2.1.02,7]undeca-2(7),3,5-triene-11-yl]-2-oxoethyl]acetamide, and combinations thereof are included.

[0056] RS-67,333 is a high-affinity 5-HT 4An agonist of the R part

[22] . This drug is effective in improving behavioral deficits, reducing the number of amyloid plaques and the level of amyloid-beta (Aβ) species, and reducing hippocampal astrogliosis and microgliosis in a 5×FAD mouse model of Alzheimer's disease (AD)

[23] . RS67333 is an aryl ketone. Incorporating an n-butyl group onto the piperidine increased agonist activity with great potency, optimal selectivity, and excellent bioavailability. Its high hydrophobicity aids passage through the blood-brain barrier and enables penetration into the brain (Eglen et al., Pharmacological characterization of two novel and potent 5-HT4 receptor agonists, RS 67333 and RS 67506, in vitro and in vivo. Br. J. Pharmacol. 1995;115(8):1387-92).

[0057] Prucalopride is a selective high-affinity 5-HT 4 R agonist

[24] . Prucalopride is a derivative of the benzofuran family that exhibits high selectivity for the 5-HT 4 receptor but no affinity for the hERG (human ether-à-go-go-related gene) channel. In 2018, it received FDA approval for chronic constipation and is currently being tested for chronic pseudo-obstruction. Prucalopride has also been tested in two separate clinical trials to investigate its effect on emotional processing after acute (e.g., single-dose) or chronic (e.g., 1-week) administration in healthy volunteers [25, 26].

[0058] PF-04995274 is a potent partial 5-HT 4It is an R agonist

[27] . Clinical trials were conducted to evaluate PF-04995274 alone or in combination with donepezil for scopolamine-induced deficits in psychomotor and cognitive function in healthy adults, but this trial was terminated not due to safety concerns

[28] . Currently, clinical trials are underway to test whether adjunctive administration of PF-04995274 has a positive effect on emotional processing and neuroactivity in treatment-resistant depression (TRD) patients mediated compared to placebo

[29] .

[0059] Tegaserod is a partial agonist of the 5-HT 4 receptor and has moderate affinity for the 5-HT 1 (agonist) and 5-HT 2A-C (antagonist) receptors.

[0060] Cisapride is a parasympathomimetic that increases acetylcholine released in the enteric nervous system by activating the 5-HT 4 receptor.

[0061] Sinitapride is a benzamide that acts as an agonist of the 5-HT 1A and 5-HT 4 receptors and as an antagonist of the 5-HT 2A receptor.

[0062] Mosapride is a selective 5-HT 4 receptor agonist, and its major active metabolite acts as a 5-HT 3 receptor antagonist.

[0063] Metoclopramide is an agonist of the 5-HT 4 and 5-HT 3A receptors. It is a D2 receptor antagonist. It is also an M1 muscarinic receptor agonist and an acetylcholinesterase inhibitor.

[0064] SUVN-D4010 is a potent, selective, and effective 5-HT4 receptor partial agonist and has good bioavailability via the oral route.

[0065] Examples of 5-HTR agonist / antagonist mixtures include, but are not limited to, buspirone, mianserin, trazodone, and mirtazapine.

[0066] The terms "serotonin," "5-hydroxytryptamine," and "5-HT" refer to the phenolic amine neurotransmitter produced from tryptophan by hydroxylation and decarboxylation in serotonergic neurons of the central nervous system and enterochromaffin cells of the gastrointestinal tract. Serotonin is a precursor of melatonin.

[0067] The term "pharmaceutically acceptable derivative" refers to any pharmaceutically acceptable salt, solvate, prodrug, such as an ester, or other precursor of a compound that can (directly or indirectly) provide the active compound or its active metabolite or residue upon administration to a recipient. Such salts include pharmaceutically acceptable base or acid addition salts, as well as pharmaceutically acceptable metal salts, ammonium salts, and alkylated ammonium salts. Such derivatives are recognizable to those skilled in the art without undue experimentation. Derivatives are described, for example, in Burger’s Medicinal Chemistry and Drug Discovery, 5th Edition, Vol 1: Principles and Practice, which is incorporated herein by reference. In certain embodiments, pharmaceutically acceptable derivatives include salts, solvates, esters, carbamates, and phosphate esters.

[0068] The agent / composition of the present invention may be administered by various routes including oral, intravenous (i.v. or IV), intranasal (i.n. or IN), intramuscular (i.m. or IM), caudal, intrathecal, and subcutaneous (s.c.) routes.

[0069] Pharmaceutical compound Agents used in the methods of the present invention include all hydrates, solvates, and complexes of the compounds described herein. When chiral centers or other forms of isomeric centers are present in the compounds of the present invention, all forms of such isomers or multiple isomers, including enantiomers and diastereomers, are intended to be included herein. Compounds containing chiral centers may be used as racemic mixtures, mixtures enriched in enantiomers, or the racemic mixtures may be separated using well-known techniques, and the individual enantiomers may be used alone. Compounds described in the present disclosure may exist in racemic form or as individual enantiomers. Enantiomers can be separated using known techniques such as those described in Pure and Applied Chemistry 69, 1469-1474, (1997) IUPAC. When the compound has an unsaturated carbon-carbon double bond, both cis (Z) and trans (E) isomers are within the scope of the present disclosure. When the compound can exist in tautomeric forms such as keto-enol tautomers, each tautomeric form is intended to be included in the present disclosure whether present equally or with one form predominating.

[0070] When the structure of the compounds used in the present disclosure contains an asymmetric carbon atom, such compounds can exist as racemates, racemic mixtures, and isolated single enantiomers. All such isomeric forms of these compounds are expressly included in the present disclosure. Each stereogenic carbon may be in the R or S configuration. Thus, it should be understood that isomers arising from such asymmetric forms (e.g., all enantiomers and diastereomers) are included within the scope of the present disclosure unless otherwise indicated. Such isomers can be obtained in substantially pure form by classical separation techniques such as those described in “Enantiomers, Racemates and Resolutions” by J. Jacques, A. Collet and S. Wilen, Pub. John Wiley & Sons, NY, 1981, and by stereochemically controlled synthesis. For example, resolution may be carried out by preparative chromatography on a chiral column.

[0071] The present disclosure also contemplates the use of all isotopes of the atoms present in the compounds disclosed herein. Isotopes include those atoms having the same atomic number but different mass numbers. Isotopically labeled compounds can be prepared by conventional techniques generally known to those of ordinary skill in the art or by processes similar to those described herein using appropriate isotopically labeled reagents in place of the unlabeled reagents employed.

[0072] The compounds of the present disclosure may be in salt form. As used herein, "salt" refers to an acid or base of a compound of the present invention, a salt of a compound of the present invention modified by making a salt. In the case of a compound used for treating mammals, the salt is pharmaceutically acceptable. Examples of pharmaceutically acceptable salts include, but are not limited to, salts of inorganic or organic acids with basic residues such as amines; alkalis or organic salts of acidic residues such as phenols. Salts can be prepared using organic or inorganic acids. Such acid salts include chlorides, bromides, sulfates, nitrates, phosphates, sulfonates, formates, tartrates, maleates, malates, citrates, benzoates, salicylates, ascorbates, etc. Phenolate salts are alkaline earth metal salts, sodium, potassium or lithium. The term "pharmaceutically acceptable salt" in this regard refers to addition salts of relatively non-toxic inorganic and organic acids or bases of the compounds of the present invention. These salts can be prepared in situ during the final isolation and purification of the compounds of the present invention, or by separately treating the purified compounds of the present invention in free base or free acid form with a suitable organic or inorganic acid or base, and isolating the thus formed salt. Representative salts include hydrobromide, hydrochloride, sulfate, bisulfate, phosphate, nitrate, acetate, valerate, oleate, palmitate, stearate, laurate, benzoate, lactate, phosphate, tosylate, citrate, maleate, fumarate, succinate, tartrate, napsylate, mesylate, glucoheptonate, lactobionate, and laurylphosphonate, etc. (see, for example, Berge et al. (1977) ”Pharmaceutical Salts”, J. Pharm. Sci. 66:1-19).

[0073] The method of the present invention also encompasses administering a physiologically functional derivative of a compound of the present invention. As used herein, the term "physiologically functional derivative" refers to a compound of the present invention, or its active metabolite, or a compound that is converted in vivo to produce a pharmaceutically acceptable salt, hydrate or solvate of this compound (e.g., a drug salt form). The conversion can occur by various mechanisms such as hydrolysis in the blood (e.g., by metabolism or chemical processes). Prodrugs are such derivatives, and discussions of the use of prodrugs are provided in T. Higuchi and W. Stella, “Pro-drugs as Novel Delivery Systems,” Vol.14 of the A.C.S. Symposium Series, and Bioreversible Carriers in Drug Design, ed. Edward b. Roche, American Pharmaceutical Association and Pergamon Press, 1987.

[0074] Dosage In certain embodiments, an effective amount of the agent of the present invention is a dosage of about 0.01 to about 3 mg / kilogram of subject body weight (mg / kg), i.e., about 0.01 mg / kg to about 3 mg / kg body weight. In certain embodiments, an effective amount of the compound of the present invention is from 0.001 to approximately 3 mg / kg body weight, from 0.001 to approximately 2 mg / kg body weight, from about 0.01 mg / kg to about 3 mg / kg body weight, from about 0.01 to about 2 mg / kg body weight, from about 0.01 to about 1.5 mg / kg body weight, from about 0.05 to about 1.4 mg / kg body weight, from about 0.05 to about 1.3 mg / kg body weight, from about 0.05 to about 1.2 mg / kg body weight, from about 0.05 to about 1.1 mg / kg body weight, from about 0.01 to about 1 mg / kg body weight, or from about 0.05 to about 0.7 mg / kg body weight. In some aspects, the dosage is from about 0.05 to about 0.5 mg / kg. In some aspects, the dosage is less than about 0.5 mg / kg body weight, less than about 0.4 mg / kg body weight, or less than about 0.3 mg / kg body weight. In some aspects, an effective dosage of the compound of the present invention is a dosage in the range of about 0.01 mg / kg to about 1.5 mg / kg body weight. In some aspects, an effective dosage of the compound of the present invention is a dosage in the range of about 0.01 mg / kg to about 1 mg / kg body weight. In some aspects, an effective amount of the compound of the present invention is a dosage in the range of about 0.01 mg / kg to about 0.75 mg / kg body weight. In some aspects, an effective amount of the compound of the present invention is a dosage in the range of about 0.75 mg / kg to about 1.5 mg / kg body weight. In some aspects, an effective amount of the compound of the present invention is a dosage in the range of about 0.5 mg / kg to about 1.2 mg / kg body weight. In some aspects, an effective amount of the compound of the present invention is a dosage in the range of about 0.05 mg / kg to about 0.5 mg / kg. In some aspects, an effective amount of the compound of the present invention is a dosage of about 0.2 mg / kg or about 0.4 mg / kg body weight.In some embodiments, the dosage of the compounds of the present invention is about 0.01 to about 1 mg / kg, about 0.1 to about 0.5 mg / kg, about 0.8 to about 1.2 mg / kg, about 0.7 to about 1.1 mg / kg, about 0.05 to about 0.7 mg / kg, about 0.01 mg / kg, about 0.05 mg / kg, about 0.1 mg / kg, about 0.2 mg / kg, about 0.3 mg / kg, about 0.4 mg / kg, about 0.5 mg / kg, about 0.6 mg / kg, about 0.7 mg / kg, about 0.8 mg / kg, about 0.9 mg / kg, about 1.0 mg / kg, about 1.1 mg / kg, about 1.2 mg / kg, about 1.3 mg / kg, about 1.4 mg / kg, about 1.5 mg / kg, about 1.6 mg / kg, about 1.7 mg / kg, about 1.8 mg / kg, about 1.9 mg / kg, about 2.0 mg / kg, or about 3 mg / kg body weight.

[0075] In certain embodiments, the dosage of the compounds of the present invention per administration is about 1 to about 250 mg, about 10 mg to about 300 mg, about 10 mg to about 250 mg, about 10 to about 200 mg, about 15 to about 175 mg, about 20 to about 175 mg, about 8 mg to about 32 mg, about 50 mg to about 75 mg, about 25 to about 150 mg, about 25 to about 125 mg, about 25 to about 100 mg, about 50 to about 100 mg, about 50 mg to about 75 mg, about 75 mg to about 100 mg, or about 75 mg to about 200 mg, 1 mg, 2 mg, 4 mg, 5 mg, 10 mg, 15 mg, 20 mg, 25 mg, 30 mg, 35 mg, 40 mg, 45 mg, 50 mg, 55 mg, 60 mg, 65 mg, 70 mg, 75 mg, 80 mg, 85 mg, 90 mg, 95 mg, 100 mg, 110 mg, 120 mg, 130 mg, 140 mg, 150 mg, 160 mg, 170 mg, 180 mg, 190 mg, 200 mg, 210 mg, 220 mg, 230 mg, 240 mg, and 250 mg. In some embodiments, the dosage of the compounds of the present invention is about 50 mg. In some embodiments, the dosage of the compounds of the present invention is about 75 mg. In some embodiments, the total dosage of the compounds of the present invention is about 100 mg.

[0076] In certain embodiments, the therapeutically effective amount of the agent of the present invention is less than the level that causes one or more side effects of the agent.

[0077] In some embodiments, the (therapeutically) effective amount of agent d of the present invention is from about 0.01 mg to about 1000 mg, from about 0.01 mg to about 500 mg, from about 0.1 mg to about 250 mg, or any amount or range therein. In another embodiment, the (therapeutically) effective amount of the agent of the present invention is, for example, 0.01 mg, 0.025 mg, 0.05 mg, 0.1 mg, 0.5 mg, 1 mg, 5 mg, 10 mg, 25 mg, 50 mg, 55 mg, 60 mg, 65 mg, 70 mg, 75 mg, 80 mg, 90 mg, 95 mg, 100 mg, 150 mg, 200 mg, 250 mg, or 500 mg.

[0078] In certain embodiments, the therapeutically effective amount of the agent of the present invention may be adjusted according to the condition of the disease / disorder being treated or prophylactically treated, the age, weight, general health, gender, and diet of the subject, the dosing interval of the drug, the route of administration, the excretion rate, and the combination.

[0079] The initial dose of the agent of the present invention is larger, and then one or more smaller maintenance doses may follow. Other ranges are possible depending on the response of the subject to the treatment. The initial dose may be the same as, lower than, or higher than the next dosing dose.

[0080] The agent / composition of the present invention may be administered once a day, once a week, once every two weeks, several times a day, twice a week, every other day, once every two weeks, four times a year, several times a week, twice a week, once a month, etc. The duration and frequency of treatment may depend on the response of the subject to the treatment.

[0081] In certain embodiments, the subject may be administered 1 dose, 2 doses, 3 doses, 4 doses, 5 doses, 6 doses, or more of the agent / composition of the present invention. In certain embodiments, a single dose of the agent / composition of the present invention is administered by the method of the present invention. In certain embodiments, multiple doses (e.g., 2 doses, 3 doses, 4 doses, 5 doses, 6 doses, 7 doses, 8 doses, 9 doses, 10 doses or more) of the agent / composition of the present invention are administered by the method of the present invention.

[0082] In certain embodiments, where there is more than one dose of the agent / composition of the invention administered to a subject, the second dose is lower than the first dose. In certain embodiments, the second dose is an amount that is at most half, one quarter, or one tenth the amount of the first dose.

[0083] The number and frequency of doses may be determined based on the subject's response to the administration of the composition, such as whether one or more of the patient's symptoms improve, and / or whether the subject tolerates the administration of the composition without experiencing an adverse reaction.

[0084] In certain embodiments, the agent / composition of the invention is administered at least once a day, at least twice a day, at least three times a day, or more frequently. In certain embodiments, the agent / composition of the invention is administered at least once a week, at least twice a week, at least three times a week, or more frequently. In certain embodiments, the agent / composition of the invention is administered at least once a month or at least twice a month.

[0085] Treatment using the methods of the invention can be continued as long as necessary.

[0086] Time frame of administration In certain embodiments, the agent / composition of the invention is administered to the subject prior to a stressor. In certain embodiments, the agent / composition of the invention is administered to the subject both prior to and after a stressor. In certain embodiments, the agent / composition of the invention is administered to the subject after a stressor. In certain embodiments, the agent / composition of the invention is administered to the subject prior to a stressor and again prior to the recurrence of the stressor or a different stressor.

[0087] In certain embodiments, the agent / composition of the present invention is administered to a subject about 12 hours to about 4 weeks, about 18 hours to about 4 weeks, about 1 day to about 3.5 weeks, about 2 days to about 3 weeks, about 3 days to about 3 weeks, about 4 days to about 3 weeks, about 5 days to about 3 weeks, about 6 days to about 3 weeks, about 2 days to about 2.5 weeks, about 3 days to about 2.5 weeks, about 4 days to about 2.5 weeks, about 5 days to about 2.5 weeks, about 6 days to about 2.5 weeks, about 1 week to about 2.5 weeks, about 1 week to about 2.5 weeks, about 1 week to about 2 weeks, about 5 minutes to about 3 days, about 10 minutes to about 2 days, about 15 minutes to about 24 hours, about 20 minutes to about 12 hours, about 30 minutes to about 8 hours, about 45 minutes to about 5 hours, about 1 hour to about 12 hours, about 2 hours to about 5 hours, about 5 minutes, about 10 minutes, about 15 minutes, about 20 minutes, about 30 minutes, about 45 minutes, about 1 hour, about 2 hours, about 3 hours, about 4 hours, about 5 hours, about 6 hours, about 8 hours, about 10 hours, about 12 hours, about 15 hours, about 1 day, about 1.5 days, about 2 days, about 3 days, about 4 days, about 5 days, about 6 days, about 1 week, about 8 days, about 9 days, about 10 days, about 11 days, about 12 days, about 13 days, about 2 weeks, about 2.5 weeks, about 3 weeks, about 3.5 weeks, or about 4 weeks before and / or after.

[0088] In certain embodiments, the administration of the agent / composition of the present invention is continued for up to 2 days, up to 3 days, up to 4 days, up to 5 days, up to 6 days, up to 1 week, up to 2 weeks, up to 3 weeks, up to 4 weeks, about 2 days, about 3 days, about 4 days, about 5 days, about 6 days, about 1 week, about 2 weeks, about 3 weeks, about 4 weeks, about 5 weeks, about 6 weeks, about 7 weeks, about 8 weeks, about 9 weeks, about 10 weeks, or a longer period.

[0089] In certain embodiments, the agent / composition of the present invention is administered 1 time, 2 times, at least 2 times, at least 3 times, at least 4 times, at least 5 times, at least 6 times, at least 7 times, at least 8 times, at least 9 times, or more per treatment.

[0090] In certain embodiments, the agent / composition of the present invention is administered at least once a day, at least twice a day, at least three times a day, at least once a week, at least twice a week, at least three times a week, at least once a month, at least twice a month, or more frequently. The treatment can be continued as long as necessary. The agent / composition of the present invention can be administered once a day, once a week, once every two weeks, several times a day, twice a week, every other day, once every two weeks, four times a year, several times a week, twice a week, once a month, etc. The duration and frequency of treatment may depend on the response of the subject to the treatment.

[0091] Stressor A stressor is a stimulus that causes stress. It can be an event or other factor that disrupts the homeostasis of the body's temperature, blood pressure, and / or other functions. In certain embodiments, a stressor is a traumatic or stressful event. Since humans have refined their brains and thought processes, anticipating a breakdown can also be a stressor. In certain embodiments, a stressor is injury, trauma, combat, war, surgery, accident, illegal injury, child abuse, natural or man-made disaster, collision, grief, hunger, heat, cold, chemical exposure, autoimmune disease, infectious disease, viral infection, cancer, fatigue, physical pain, neuropathy, hyperalgesia, allodynia, emotional pain, or depression. A traumatic event can be an event or something that threatens the life of an individual or a person close to them, or something that has been witnessed. U.S. Patent Application Publication No. 20140018339.

[0092] A stressor can be acute or chronic.

[0093] There are numerous physiological processes that are modified by the response to stress. Among these, the levels of cortisol, corticotropin, catecholamines, and serotonin are modified. These levels return to basal values after acute stress is removed (McEwen N Eng J Med 1998 338(3):171-179). These biochemical stress markers sequentially lead to pathological health states and psychosocial disorders. As a result, stress plays an important role in physical and mental health. Stress can affect the onset or susceptibility to disease. It can also affect the progression or course of a disease, even if there is another pathophysiological action underlying the disease. Recovery from existing diseases can also be delayed by stress. For example, stress is a contributing factor in hypertension, heart disease, headache, colitis, irritable bowel syndrome, temporomandibular joint disorder, cancer, gastric ulcer, insomnia, skin disorders, and asthma. Stress can also exacerbate other conditions such as multiple sclerosis, diabetes, herpes, mental illness, substance abuse, and mental disorders characterized by the presence of violence or aggressive tendencies. In particular, stress contributes to functional somatic disorders, mood disorders, and major depressive disorder (MDD). These include disorders such as chronic fatigue syndrome (CFS), fibromyalgia (FMS), Gulf War syndrome, anxiety disorders, and post-traumatic stress disorder (PTSD). Stressors that disrupt normal exercise or sleep patterns.

[0094] Additional examples of use include administration prior to military deployment to protect military personnel (frontline combat soldiers, battlefield surgeons, etc.) and military working dogs from stress. Potential non-military examples of use include, but are not limited to, police officers, firefighters, first responders, emergency medical technicians (EMTs), emergency room (ER) physicians, correctional officers (and inmates), humanitarian aid workers, and refugees.

[0095] In certain embodiments, the subject may be administered the agent or composition of the invention prior to a situation where the subject (such as an initial responder or military personnel) may be exposed to traumatic stress immediately following exposure to traumatic stress and / or when the subject feels that PTSD symptoms may appear.

[0096] Resilience to stress Resilience to stress refers to the ability of a subject to adapt or change well and / or maintain physiological, neurological, or mental homeostasis when faced with a stressor (e.g., adversity). As used herein, the term "enhancing resilience" refers to increasing the ability of a subject to experience a stressor (e.g., a traumatic event) without suffering from stress-induced mood disorders and / or with fewer post-event symptoms or homeostatic breakdowns and / or normal activities of daily life. In certain embodiments, improving resilience may prevent stress-induced mood disorders. In certain embodiments, improving resilience may reduce at least one of the signs, symptoms, or symptom clusters of stress-induced mood disorders. In certain embodiments, the methods of the invention enhance the subject's resilience to stress, assist in the defense against stressor-related psychiatric morbidity, reduce the functional consequences (e.g., PTSD, etc.) of stressor-induced disorders, and reduce morbidity and mortality in medicine.

[0097] The Connor-Davidson Resilience Scale (CD-RISC) is a 25-item self-report scale, each of which is rated on a 5-point scale (0 - 4), with higher scores reflecting greater resilience (Connor K M & Davidson, J R T. Development of a new resilience scale: the Connor-Davidson Resilience Scale (CD-RISC). Depression and Anxiety, 2003: 18: 71-82).

[0098] Resilience, mental growth, and life satisfaction may be measured by the CD-RISC, Purpose in Life Scale, abbreviated as MOS, Social Support Survey, PTGI, and Q-LES-Q.

[0099] Combined therapy The agent or composition of the present invention may be administered to a subject alone or in combination with one or more other treatments / agents.

[0100] In certain embodiments, the second agent is an antidepressant, an antianxiety agent, or a combination thereof. In certain embodiments, the second agent is a serotonin reuptake inhibitor (SRI) or a selective serotonin reuptake inhibitor (SSRI). In certain embodiments, the second agent is fluoxetine, paroxetine, sertraline, lithium, riluzole, prazosin, lamotrigine, ifenprodil, or a combination thereof. In certain embodiments, the second agent is a dual serotonin norepinephrine reuptake inhibitor compound (DRI). In certain embodiments, the second agent is venlafaxine, duloxetine, milnacipran, or a combination thereof. In certain embodiments, the second agent is a non-tricyclic triple reuptake inhibitor (TRI).

[0101] In certain embodiments, the agent or composition of the present invention is administered to a subject in combination with one or more treatments / agents such as antidepressants, analgesics, muscle relaxants, anorectics, stimulants, antiepileptic drugs, and sedatives / hypnotics. Non-limiting examples of compounds that may be administered in combination with the compound or composition of the present invention include Neurontin, pregabalin, pramipexole, L-DOPA, amphetamine, tizanidine, clonidine, tramadol, morphine, tricyclic antidepressants, codeine, carbamazepine, sibutramine, amphetamine, Valium, trazodone, and combinations thereof.

[0102] In certain embodiments, combination therapy means simultaneous administration of the agents in the same dosage form, simultaneous administration in separate dosage forms, or separate administration of the agents.

[0103] In certain embodiments, the second agent / treatment is used as adjuvant therapy to the agent or composition of the invention. In certain embodiments, the treatment includes a phase in which treatment with the second agent / treatment is carried out after treatment with the agent or composition of the invention has ended. In certain embodiments, the treatment includes a phase in which treatment with the agent or composition of the invention overlaps with treatment with the second agent / treatment.

[0104] Combination therapy can be sequential or administered simultaneously. In either case, these drugs and / or treatments are said to be "co-administered." It must be understood that "co-administered" does not necessarily mean that the drugs and / or treatments are administered in a combined form (i.e., they may be administered separately (e.g., as separate compositions or formulations) at the same or different sites at the same or different times or together (e.g., in the same formulation or composition)).

[0105] In certain embodiments, a subject is treated simultaneously (or concomitantly) with the agent or composition of the invention and a second agent. In certain embodiments, a subject is first treated with the agent or composition of the invention, followed by the end of treatment with the compound or composition of the invention and the initiation of treatment with the second agent. In certain embodiments, the agent or composition of the invention is used as the first treatment, for example, by administration of 1, 2, or 3 doses, and the second agent is administered to extend the effect of the agent or composition of the invention or to amplify the effect of the agent or composition of the invention. One of ordinary skill in the art will recognize other variations of the presented regimen, such as starting treatment of a subject with the compound or composition of the invention, followed by treatment of the subject with the second agent as adjuvant therapy to the treatment with the compound or composition of the invention for a subsequent period, followed by discontinuation of treatment with the compound or composition of the invention.

[0106] The compounds of the present invention and other pharmaceutically active agent(s) may be administered together or separately, and when administered separately, this may be done simultaneously or sequentially in any order. The amounts of the agents of the present invention and other pharmaceutically active agent(s), as well as the relative timing of administration, will be selected to achieve the desired combination therapy effect.

[0107] In various embodiments, the treatment (e.g., the compositions provided herein and a second agent in combination therapy) is administered at intervals of less than 5 minutes, less than 30 minutes, 1 hour, about 1 hour, about 1 to about 2 hours, about 2 to about 3 hours, about 3 to about 4 hours, about 4 to about 5 hours, about 5 to about 6 hours, about 6 to about 7 hours, about 7 to about 8 hours, about 8 to about 9 hours, about 9 to about 10 hours, about 10 to about 11 hours, about 11 to about 12 hours, about 12 to 18 hours, 18 to 24 hours, 24 to 36 hours, 36 to 48 hours, 48 to 52 hours, 52 to 60 hours, 60 to 72 hours, 72 to 84 hours, 84 to 96 hours, or 96 to 120 hours. In certain embodiments, the treatment is administered at intervals of 24 hours or less, or 48 hours or less. In certain embodiments, two or more treatments are administered within the same patient visit. In other embodiments, the compositions provided herein and the second agent are administered simultaneously. In other embodiments, the compositions provided herein and the second agent are administered at intervals of about 2 to 4 days, about 4 to 6 days, about 1 week, about 1 to 2 weeks, or more than 2 weeks. In certain embodiments, administration of the same agent may be repeated, and the administrations may be separated by at least 1 day, 2 days, 3 days, 5 days, 10 days, 15 days, 30 days, 45 days, 2 months, 75 days, 3 months, or 6 months. In other embodiments, administration of the same agent may be repeated, and the administrations may be separated by at least 1 day, 2 days, 3 days, 5 days, 10 days, 15 days, 30 days, 45 days, 2 months, 75 days, 3 months, or 6 months.In certain embodiments, the compositions and second agents provided herein are administered in a continuous and spaced-apart manner, such that the compositions provided herein act together with other agents to provide a greater benefit than when administered in other ways. For example, the second active agents can be administered sequentially at the same time or at different times in any order, but if not administered at the same time, they must be administered at times close enough to provide the desired therapeutic or prophylactic effect. In one embodiment, the compositions and second active agents provided herein exert their effects at overlapping times. Each second active agent can be administered separately, in any suitable form, and by any suitable route. In other embodiments, the compositions provided herein are administered before, simultaneously with, or after the administration of the second active agent. The term "about" refers to ±10% of the reference value. In other embodiments, multiple treatment procedures are performed on the patient simultaneously, i.e., individual doses of the second agent are administered separately but within an interval such that the compounds provided herein can act together with the second active agent. For example, one component can be administered once every two weeks or once every three weeks in combination with another component that can be administered once a week. In other words, the dosing regimens are carried out simultaneously even if the therapeutic agents are not administered at the same time or on the same day. The second agent can act additively or synergistically with the compounds provided herein. In one embodiment, the compositions provided herein are administered simultaneously with one or more second agents in the same pharmaceutical composition. In another embodiment, the compositions provided herein are administered simultaneously with one or more second agents in another pharmaceutical composition. In yet another embodiment, the compositions provided herein are administered prior to or subsequent to the administration of the second agent. Also contemplated is the administration of the compositions and second agents provided herein by the same or different routes of administration, e.g., orally and parenterally. In certain embodiments, when the compositions provided herein are administered simultaneously with a second agent that produces potentially harmful effects, including but not limited to toxicity, the second active agent can advantageously be administered at a dose that is below the threshold at which harmful side effects are induced.

[0108] Included by the disclosure of the present invention is a method for prophylactically treating a subject prior to a stressor. In certain embodiments, the agents / compositions and methods of the present invention prevent or delay stress-induced mood disorders or stress-induced psychoses in a subject. In certain embodiments, stress-induced mood disorders include major depressive disorder and post-traumatic stress disorder.

[0109] Stress-induced mood disorder There are numerous disorders caused or exacerbated by stress. The agents / compositions and methods of the present invention can prevent or delay stress-induced mood disorders or stress-induced psychoses. Stress-induced mood disorders or stress-induced psychoses that can be prevented or treated by the agents / compositions and methods of the present invention include addictive disorders such as substance abuse, anorexia nervosa, bulimia nervosa, obesity, smoking addiction, and weight addiction; anxiety disorders such as agoraphobia, generalized anxiety disorder, obsessive-compulsive disorder, panic attacks, performance anxiety, phobias, and post-traumatic stress disorder (PTSD); mental disorders such as stress-induced mental disorders; autoimmune diseases such as allergies, asthma, fibromyalgia, fibromytosis, lupus, multiple sclerosis, rheumatoid arthritis, Sjogren's syndrome, and vitiligo; cancers such as bone cancer, brain cancer, breast cancer, cervical cancer, colon cancer, Hodgkin's disease, leukemia, liver cancer, lung cancer, lymphoma, multiple myeloma, ovarian cancer, pancreatic cancer, and prostate cancer; cardiovascular disorders such as arrhythmia, arteriosclerosis, Buerger's disease, essential hypertension, arteriolosclerosis, mitral valve prolapse, palpitations, peripheral vascular disease, Raynaud's disease, stroke, tachycardia, and Wolff-Parkinson-White syndrome; and developmental disorders such as attention deficit disorder, concentration problems, conduct disorder, dyslexia, hyperactivity, speech and language disorders, and learning disorders, but are not limited thereto.

[0110] Anxiety disorder The agents / compositions and methods of the present invention can prevent or delay anxiety disorders. The five major types of anxiety disorders are panic disorder, obsessive-compulsive disorder, post-traumatic stress disorder, generalized anxiety disorder, and phobias (including social phobia, also known as social anxiety disorder). Each anxiety disorder has its own distinct features, but they are all grouped together under the common theme of excessive and irrational fear and dread. Anxiety disorders are commonly associated with depression, eating disorders, substance abuse, or another anxiety disorder.

[0111] Panic disorder is characterized by repeated episodes of intense fear that often strike without warning. Physical symptoms include chest pain, heart palpitations, shortness of breath, dizziness, abdominal pain, feelings of unreality, and fear of death. Obsessive-compulsive disorder is characterized by repeated unwanted thoughts or compulsive behaviors that seem uncontrollable or unstoppable. Generalized anxiety disorder is characterized by excessive worry and tension about daily events and activities that lasts for at least six months. It is often accompanied by physical symptoms such as fatigue, trembling, muscle tension, headache, or nausea, even when there is little or no reason to expect the worst. Phobias are characterized by two main types of fears, namely social phobia and specific phobia. People with social phobia have an overwhelming and disabling fear of scrutiny, embarrassment, or humiliation in social situations, which leads to the avoidance of many potentially enjoyable and meaningful activities. People with specific phobia are plagued by an extreme and disabling irrational fear of something that poses little or no actual danger, which leads to the avoidance of the object or situation and can unnecessarily limit people's lives.

[0112] Post-traumatic stress disorder (PTSD) Typically, subjects suffering from PTSD have been exposed to a traumatic event in which the person witnessed, experienced, or was confronted with an event or events involving actual or threatened death or serious injury, or a threat to the physical integrity of self or others, and intense fear, helplessness, or horror in which the person's response was involved.

[0113] Repeating the intrusion memory of trauma exposure is one of the core symptoms of PTSD. Patients with PTSD are known to exhibit learning and memory impairments during neuropsychological tests. Other core symptoms of PTSD include stress sensitivity (startle), increased tension and anxiety, memory impairment, and social withdrawal.

[0114] In certain embodiments, the method of the invention prevents or inhibits the development of post-traumatic stress disorder (PTSD) in a subject. In certain embodiments, the method of the invention prevents or inhibits the development of one or more PTSD-like symptoms. In certain embodiments, the subject may be administered the agent or composition of the invention prior to a situation in which the subject (such as a first responder or military personnel) may be exposed to traumatic stress, immediately following exposure to traumatic stress, and / or when the subject feels that there is a likelihood that PTSD symptoms may appear in the subject.

[0115] Typically, the traumatic event is persistently re-experienced in one or more of the following ways: recurrence and intrusive distressing recollections of the event, including images, thoughts, or perceptions; recurrent distressing dreams of the event; feelings of reliving the traumatic event as if it were recurring, including experiences, illusions, hallucinations, and dissociative flashback episodes that occur upon waking or when aroused; intense psychological distress at exposure to internal or external cues that symbolize or are similar to an aspect of the traumatic event; physiological reactivity at exposure to internal or external cues that symbolize or are similar to an aspect of the traumatic event. Individuals suffering from PTSD also have persistent avoidance of trauma-related stimuli and numbing of general responsiveness, which is indicated by three or more of the following: efforts to avoid trauma-related thoughts, feelings, or conversations; efforts to avoid activities, places, or people that arouse recollections of the trauma; inability to recall an important aspect of the trauma; markedly diminished interest in or participation in significant activities; feelings of detachment or estrangement from others; restricted range of affect (e.g., inability to feel loving); a sense of foreshortened future (e.g., not expecting to have a career, marriage, children, or a normal lifespan); persistent symptoms of increased arousal (not present before the trauma), as indicated by two or more of the following: difficulty falling or staying asleep; irritability or outbursts of anger; difficulty concentrating; hypervigilance; exaggerated startle response. The disturbance lasts for at least one month and causes clinically significant distress or impairment in social, occupational, or other important areas of functioning.

[0116] In certain embodiments, the compounds or compositions of the invention prevent, reduce, eliminate, or delay one or more of the symptoms, including, but not limited to, re-experiencing of traumatic experiences in the form of intrusive memories, nightmares, flashbacks; emotional and physical reactions caused by reminders of the trauma; distancing from others; decreased interest in activities or other people; emotional numbing; avoidance of traumatic reminders; symptoms of increased arousal, including sleep disturbances, hypersensitivity, hypervigilance, decreased concentration; increased startle reflex; and combinations thereof.

[0117] Regardless of what the root cause of the problem may be, some people with PTSD repeatedly relive the trauma in the form of nightmares and intrusive daytime recollections. They may also experience other sleep problems, feel alienated or numb, or be easily startled. They may lose interest in things they once enjoyed and have difficulty feeling affectionate. They may feel angrier, more aggressive, or violent than before. Things that remind them of the trauma are very painful and can lead them to avoid certain places or situations where they recall those memories.

[0118] This disorder may be accompanied by depression, substance abuse, or one or more other anxiety disorders. In severe cases, the person may have difficulty working or engaging in social activities.

[0119] Major depressive disorder Major depressive disorder refers to a classification of syndromes characterized by negative emotions and recurrent depressive episodes without any prior history of independent episodes of mania and hyperactivity that meet the criteria for bipolar disorder. Multiple subtypes of major depressive disorder are recognized, including those with atypical features, psychotic components, etc. The age of onset, severity, duration, and frequency of depressive episodes all vary widely. This disorder can begin at any age. The symptoms of major depressive disorder typically develop over several days to weeks. Prodromal symptoms include general anxiety disorder, panic attacks, phobias, or depressive symptoms, which can occur for several months before the episode. Individual episodes also typically last 3 to 12 months, but recurrence is not very common. Most patients are symptom-free between episodes, but a small number of patients, mainly the elderly, may develop persistent depression. Individual episodes of any severity are often suddenly triggered by stressful life events. Common symptoms of depressive episodes include reduced concentration and attention; reduced self-esteem and confidence; self-blame and guilt, thoughts or actions of self-harm or suicide; sleep disturbances; and loss of appetite. In certain embodiments, major depressive episodes occur following psychosocial stressors such as the death of a loved one, marital separation, childbirth, or the end of an important relationship.

[0120] The mood decline is mostly unchanged daily and is often unresponsive to the environment, but may show characteristic diurnal variation as the day progresses. Similar to bipolar episodes, clinical findings show significant individual variation, and atypical findings are particularly common in adolescence. In some cases, anxiety, distress, and motor agitation are more prominent when the patient is depressed, and mood changes may also be masked by additional features such as irritability, excessive alcohol consumption, histrionic behavior, and exacerbation of pre-existing phobic or obsessive-compulsive symptoms, or by hypochondriasis.

[0121] Psychiatric evaluation In certain embodiments, the effect or efficacy of treatment with the agents / compositions of the invention is evaluated by the subject and / or a medical professional, such as the subject's physician. In certain embodiments, the evaluation is performed within about 10 minutes, within about 15 minutes, within about 20 minutes, within about 25 minutes, within about 0.5 hour, within about 1 hour, within about 2 hours, within about 2.5 hours, within about 3 hours, within about 3.5 hours, within about 4 hours, within about 4.5 hours, within about 5 hours, within about 5.5 hours, within about 6 hours, within about 6.5 hours, within about 7 hours, within about 7.5 hours, within about 8 hours, within about 8.5 hours, within about 9 hours, within about 9.5 hours, within about 10 hours, within about 10.5 hours, within about 11 hours, within about 11.5 hours, within about 12 hours, within about 18 hours, within about 1 day, within about 2 days, within about 3 days, within about 4 days, within about 5 days, within about 6 days, within about 1 week, within about 2 weeks, within about 3 weeks, within about 4 weeks, within about 1 month, within about 2 months, within about 3 months, within about 4 months, within about 5 months, within about 6 months, within about 1 year, within about 2 years, or longer following stress and / or administration of the agents / compositions of the invention.

[0122] A psychiatric evaluation of a patient treated by the methods of the invention can be performed to determine whether the method is effective. In certain embodiments, the psychiatric evaluation may be performed before, during, and / or after treatment. If the psychiatric evaluation is performed both before and after (and / or during) treatment by the methods of the invention, the results of the pre-treatment evaluation can provide a baseline for comparison to the results of the evaluation during and / or after treatment. In certain embodiments, the psychiatric evaluation is performed only after treatment.

[0123] Psychophysiological stress tests can be performed to measure the amount of stress-induced anxiety present in various systems of the body (i.e., the muscular, cardiovascular, digestive, respiratory, and nervous systems). These stress tests are routinely used in the art. The test results are compared to both regional and national norms to determine whether the individual exhibits an excessive amount of physiological anxiety and whether it can recover from a standardized stressor over an appropriate period of time.

[0124] Psychiatric tests can be used to monitor a subject to determine the emotional and / or social etiology of stress disorders. These tests are known in the art and include health-related assessments, mental health assessments, personality tests, and personality type assessments.

[0125] In certain embodiments, clinician-administered evaluations and / or self-reporting means are used for the purpose of measuring baseline symptoms as well as (1) the overall severity of the disorder, (2) core symptoms, and (3) the drug effect on depressive mood.

[0126] Non-limiting examples of psychiatric evaluation tools and questionnaires include the following scales.

[0127] Diagnostic and Statistical Manual of Mental Disorders (DSM-5) includes revised diagnostic criteria for PTSD. See American Psychiatric Association: Diagnostic and Statistical Manual of Mental Disorders, Fifth Edition. Arlington, Va., American Psychiatric Association, 2013. See also ptsd.va.gov / professional / PTSD-verview / dsm5_criteria_ptsd.asp.

[0128] The Structured Clinical Interview for DSM-IV Axis I Disorders, Patient Edition (SCID-P) is a semi-structured interview that provides focused questions and follow-up questions asked by clinicians to assist in diagnosis. First et al., Structured Clinical Interview for DSM-IV TR Axis I Disorders, Research Version, Patient Edition (SCID-I / P). New York: New York State Psychiatric Institute, Biometrics Research; 2001. It includes an overview for obtaining information about demographic attributes, employment, chief complaint, current medical history, past history, treatment history, and current functioning. The body of the SCID-P contains nine modules designed to diagnose a total of 51 mental disorders.

[0129] The SCID-P for DSM-5 is the SCID-Patient version, a new edition of the SCID modified to incorporate the new DSM-5 criteria.

[0130] The Clinician-Administered PTSD Scale (CAPS) is a structured clinical interview designed to assess the essential features of PTSD as defined by the DSM-IV. Weathers et al., Clinician-administered PTSD scale: a review of the first ten years of research. Depress Anxiety. 2001; 13(3):132-156. The CAPS can be used to provide a categorical rating of the diagnostic status and a quantitative index of symptom severity. Both frequency scores and intensity scores are obtained for each individual symptom. The total score on the CAPS is based on an individual's responses to 17 items that assess the frequency and intensity of current PTSD symptoms. CAPS subscales are used to assess specific symptom clusters. The total score can range from 0 to 136.

[0131] The Clinician-Administered PTSD Scale for DSM-5 (CAPS-5) is a 30-item structured interview that can be used to make a current (past 1 month) diagnosis of PTSD, to make a lifetime diagnosis of PTSD, and to assess PTSD symptoms over the past week. The CAPS-5 is a 30-item questionnaire corresponding to the DSM-5 diagnosis for PTSD. The language of the CAPS-5 reflects both changes to existing symptoms and the addition of new symptoms in the DSM-5. Weathers, F. W., et al (2013). The Clinician-Administered PTSD Scale for DSM-5 (CAPS-5).

[0132] The Treatment Outcome PTSD Scale (TOP-8) is a brief interviewer-administered scale specifically designed for the assessment of signs and symptoms common to PTSD that change in response to treatment (Davidson, J. R., & Colket, J. T. (1997). The eight-item treatment-outcome post-traumatic stress disorder scale: A brief measure to assess treatment outcome in post-traumatic stress disorder. International Clinical Psychopharmacology, 12(1), 41-45). The TOP-8 consists of 8 items, each of which is measured on a scale of 0 to 4 and defined anchors are provided for each item. The items represent three core features of PTSD, and the maximum possible score is 32.

[0133] The Hamilton Psychiatric Rating Scale for Anxiety (HAM-A) is a widely used observational rating scale for the severity of anxiety disorders. This scale consists of 14 items. Each item is rated on a scale of 0 to 4. This scale is administered to assess the severity of anxiety disorders and improvement during the course of treatment. The HAM-A total score is the sum of the 14 items, and scores range from 0 to 56. Hamilton M. The Assessment of Anxiety-States by Rating. Br J Med Psychol. 1959; 32(1):50-55.

[0134] The Montgomery-Asberg Depression Rating Scale (MADRS) is a 10-item instrument used for the assessment of depressive symptoms in adults and for the rating of any changes in those symptoms. Montgomery S. A., et al., A new depression scale designed to be sensitive to change. Br J Psychiatry. 1979 April; 134:382-389. Each of the 10 items is rated on a scale of 0 - 6, with different descriptors for each item. These individual item scores are added together to form a total score, which can range from 0 - 60 points.

[0135] The Young Mania Rating Scale, item 1 (YMRS-1), is used to rate mood elevation on the day of infusion. Young R C, et al. Rating-Scale for Mania - Reliability, Validity and Sensitivity. Br J Psychiatry. 1978; 133(NOV):429-435.

[0136] The Brief Psychiatric Rating Scale (BPRS) is used to rate acute behavioral changes during infusion. Overall J E et al., The Brief Psychiatric Rating-Scale. Psychol. Rep. 1962; 10(3):799-812. Four important BPRS items for positive (+) symptoms of psychosis are used: disturbance of conceptual integration, behavior due to hallucinations, suspiciousness, and unusual thought content. Three items representing negative (-) symptoms of psychosis will also be used: flattening of affect, emotional withdrawal, and psychomotor retardation.

[0137] The Clinician-Administered Dissociative States Scale (CADSS) is used to measure dissociative effects during infusion. Bremner JD, et al., Measurement of Dissociative States with the Clinician-Administered Dissociative States Scale (CADSS). J Trauma Stress. 1998; 11(1):125-136. The scale includes 19 questions and 8 observational evaluations, scored from 0 (none) to 4 (extreme). The CADSS measures impairments in body perception, environmental perception, time perception, memory perception, and depersonalization.

[0138] The Patient Rating Inventory of Side Effects (PRISE) is a patient self-report used to qualify side effects by identifying and evaluating the tolerance of each symptom. Levine J, Schooler NR. SAFTEE: A technique for the systematic assessment of side effects in clinical trials. Psychopharmacol Bull.1986; 22(2):343-381.

[0139] The Clinical Global Impression (CGI) scale assesses treatment response in psychiatric patients. The administration time is 2 minutes. This scale consists of 3 items: severity of illness (item 1); global improvement (item 2); and efficacy index (item 3). Item 1 is evaluated on a 7-point scale (1 = normal, 7 = most severely ill patients), similar to item 2 (1 = much improved, 7 = much worse). Each includes an additional response of "not rated". Item 3 is evaluated on a 4-point scale ("none" to "exceeds treatment effect").

[0140] The Impact of Events Scale (IES) is the most widely used self-report scale for stress reactions to traumatic events. Horowitz et al., Impact of Event Scale: a measure of subjective stress. Psychosom Med. 1979 May; 41(3):209-218. See also Weiss et al., The Impact of Event Scale, revised in Wilson J, Keane T M, eds. Assessing psychological trauma and PTSD. New York: Guilford; 1996:399-411. It measures both intrusion and avoidance. Sundin et al., Impact of Event Scale: psychometric properties. Br J Psychiatry. 2002 March; 180:205-209. Joseph S. Psychometric evaluation of Horowitz’s Impact of Event Scale: a review. J Trauma Stress. 2000 January; 13(1):101-113. The total score can range from 0 to 75.

[0141] The Posttraumatic Stress Disorder Checklist (PCL-5) is a 17-item self-report scale that reflects the DSM-5 symptoms of PTSD. The PCL-5 measures symptoms in response to stressful situations (Weathers, F., et al. (1993). The PTSD checklist (PCL): Reliability, validity, and diagnostic utility. Annual Convention of the International Society for Traumatic Stress Studies, San Antonio, Tex.).

[0142] The Quick Inventory of Depressive Symptomatology, Self Report (QIDS-SR) is a 16-item self-report measure designed to assess the severity of depressive symptoms present within the past 7 days. Rush A J, Trivedi M H, Ibrahim H M et al. The 16-Item quick inventory of depressive symptomatology (QIDS), clinician rating (QIDS-C), and self-report (QIDS-SR): a psychometric evaluation in patients with chronic major depression. Biol. Psychiatry. 2003; 54(5):573-583. These 16 items include 9 symptom domains of major depression and are rated on a scale of 0 to 3. The total score ranges from 0 to 27, including the ranges of 0 to 5 (normal), 6 to 10 (mild), 11 to 15 (moderate), 16 to 20 (moderate to severe), and 21+ (severe).

[0143] The Childhood Trauma Questionnaire (CTQ) is a 28-item self-report measure that assesses childhood trauma in the following domains: physical, sexual, and emotional abuse, as well as physical and emotional neglect. Bernstein D P, Stein J A, Newcomb M D et al. Development and validation of a brief screening version of the Childhood Trauma Questionnaire. Child Abuse Negl.2003 February; 27(2):169-190. Each item is rated on a scale of 1 (not true) to 5 (very much true). Five subscales are then summed, and scores range from 5 to 25 in each trauma category.

[0144] Visual Analogue Scales (VAS) are used to assess subjective changes in condition. Bond A, Lader M. The use of analogue scales in rating subjective feelings. Br J Med Psychol. 1974; 47(3):211 - 218. They are 100 - mm horizontal lines marked in proportion to the perceived intensity of subjective experience (0 = all different, 10 = extreme) for the following conditions: anxiety, depression, sleepiness, high state of excitement, hunger, and nausea.

[0145] The Sheehan Disability Scale (SDS) is a self - reporting disability scale. It has demonstrated sensitivity to disability and change as treatment outcomes in a wide range of mental disorders. The SDS asks only about the approximate current level of disability and provides no indication of whether the person has improved or worsened from the past, thus serving as a reasonable short - term outcome measure not confounded by the impression of the medical history. The independent variable is the total score, which is based on the sum of three 10 - point items (work, social life, and family life), with higher scores reflecting greater disability. Sheehan D. The Anxiety Disease. New York, N.Y.: Scribner; 1983.

[0146] The Wechsler Abbreviated Scale of Intelligence 2 - Subtest (WASI - 2) is a reasonably brief measure of IQ for ages 6 to 89 that includes vocabulary (an estimate of verbal fluency ability) and matrix reasoning (an estimate of non - verbal fluency ability). Wechsler D. Wechsler Abbreviated Scale of Intelligence San Antonio, Tex.: Psychological Corporation; 1999. It is widely used in medical, educational, and research settings. The average reliability coefficient is 0.96, and the retest reliability is 0.88.

[0147] The Hopkins Verbal Learning Test (HVLT) is a repeatable test of verbal memory acquisition and delayed recall. Subjects are presented with the same 12-item list in 3 learning trials and are repeatedly questioned about the items of each list on each trial. Delayed recall and recognition conditions are conducted later. The dependent variables used in this test include total learning over 3 trials (for the acquisition variable) and total delayed recall score (for the recall component). Brandt J, Benedict R. Hopkins Verbal Learning Test, Revised. Odessa, Fla.: Psychological Assessment Resources; 1997.

[0148] The Profile of Mood States - Bipolar (POMS - Bi) scale measures mood and feelings mainly in a clinical rather than a non - clinical setting. It can assist in determining an individual's psychiatric status for treatment purposes or can be used to compare mood profiles associated with various personality disorders. It is also a useful means in identifying the effects of drug treatment.

[0149] The Post - Traumatic Cognitions Inventory (PTCI) is a 33 - item scale that is rated on a Likert - type scale ranging from 1 (strongly disagree) to 7 (strongly agree). The scale score is formed by 3 sub - scales, which show high inter - correlations (rs = 0.57 - 0.75).

[0150] The New Cognitions scale is a 6-item pilot scale that is rated on a Likert-type scale ranging from 1 (all different) to 4 (many). This scale is based on the Post Traumatic Growth Inventory (PTGI) from which items were directly selected (new items were also added to this scale), and the Brief-COPE (see Carver, C. S. (1997) ”You want to measure coping but your protocol’s too long: Consider the brief COPE.” International Journal of Behavioral Medicine 4; 92-100).

[0151] The Medical Outcomes Study (MOS) Social Support Survey is a 19-item self-report scale designed to assess the level of functional social support. The MOS-SS has two subscales (emotional and instrumental social support) to identify potential social support deficits (Sherbourne, C. D. & Stewart, A. L. (1991). ”The MOS Social Support Survey.” Soc Sci Med 32(6): 705-714).

[0152] The Purpose in Life test-Short Form (PIL-SF) is a brief 4-item form of the 20-item Purpose in Life test. This scale asks respondents to report the degree to which they have achieved their life goals and the degree to which they perceive their life as meaningful or purposeful (Schulenberg et al 2010; Psychotherapy (Chic).2008 December; 45(4):447-63).

[0153] The Posttraumatic Growth Inventory (PTGI)-Short Version is a 10-item short form of the PTGI self-report questionnaire (see reference). It asks respondents to rate the degree to which they have changed as a result of experiencing highly stressful life events. The items cover positive changes in five domains: relating to others, new possibilities, personal strength, spiritual change, and appreciation for life (Cann, A., et al. (2010). A short form of the Posttraumatic Growth Inventory. Anxiety, Stress & Coping, 23, 127-137).

[0154] The Quality of Life Enjoyment and Satisfaction Questionnaire (Q-LES-Q) is a self-report scale that measures the degree of pleasure and satisfaction experienced by subjects in various areas of daily functioning. This global score is a reliable and valid measure of these aspects in groups of depressed subjects (Endicott J, et al. Quality of Life Enjoyment and Satisfaction Questionnaire: A New Measure. Psychopharmacology Bulletin; 1993; 29:321-326).

[0155] In certain embodiments, a self-assessment of the subject to be treated is performed.

[0156] Pharmaceutical composition The agents of the present invention, as well as salts, solvates and physiologically functional derivatives thereof, can be administered as crude chemicals, but it is possible to provide the active ingredient as a pharmaceutical composition. Accordingly, the present invention further provides a pharmaceutical composition comprising a compound of the present invention and / or a salt, solvate and physiologically functional derivative thereof, and one or more pharmaceutically acceptable carriers, diluents or excipients. The carrier(s), diluent(s) or excipient(s) must be acceptable in the sense of being compatible with the other ingredients of the formulation and not harmful to the recipient. According to another aspect of the present invention, there is also provided a process for the preparation of a pharmaceutical composition, which comprises admixing a compound of the present invention, or a salt, solvate and physiologically functional derivative thereof, with one or more pharmaceutically acceptable carriers, diluents or excipients.

[0157] The term "composition" is intended to encompass products containing the active ingredient(s) and the inert ingredients (pharmaceutically acceptable excipients) that make up the carrier, as in a pharmaceutical composition, as well as any combination, complexation or aggregation of two or more starting materials, or any product directly or indirectly resulting from the dissociation of one or more of the starting materials, or from one or more other types of reaction or interaction of the starting materials. Accordingly, the pharmaceutical compositions of the present invention encompass any composition made by admixing compound 20 with a pharmaceutically acceptable excipient.

[0158] Acceptable excipients, diluents, and carriers for therapeutic use are well known in the pharmaceutical art and are described, for example, in Remington: The Science and Practice of Pharmacy. Lippincott Williams & Wilkins (A. R. Gennaro edit. 2005). The choice of pharmaceutical excipients, diluents and carriers can be made with reference to the intended route of administration and standard pharmaceutical practice.

[0159] As used herein, the phrase "pharmaceutically acceptable" refers to molecular substances and compositions that are "generally regarded as safe" which are physiologically tolerable and typically do not produce allergic or similar adverse reactions such as indigestion, dizziness, etc. when administered to humans. Preferably, the term "pharmaceutically acceptable" as used herein means approved by a federal or state government regulatory authority for use in animals, more particularly in humans, or listed in the United States Pharmacopeia or other generally recognized pharmacopeias.

[0160] The pharmaceutical compositions of the present invention may exist in unit dosage forms containing a predetermined amount of the active ingredient per unit dose. Such units may contain, for example, from 5 μg to 1 g, preferably from 1 mg to 700 mg, more preferably from 5 mg to 100 mg of the compound of the present invention, depending on the condition being treated, the route of administration, and the age, weight and condition of the patient. Therefore, such unit doses may be administered more than once a day. Preferred unit dosage compositions contain the daily dose or fractional dose (in the case of administration more than once a day) or an appropriate fraction thereof of the active ingredient listed earlier herein. Further, such pharmaceutical compositions may be prepared by any of the methods well known in the art of pharmacy.

[0161] The pharmaceutical compositions of the present invention may be adapted for administration by any suitable route, such as oral (including buccal or sublingual), inhalation, intranasal, intraocular, or parenteral (including intravenous and intramuscular) routes. The compositions of the present invention may be injected. Such compositions may be prepared by any method known in the art of pharmacy, for example, by bringing about an association of the active ingredient with a carrier(s) or excipient(s).

[0162] In a further embodiment, the present invention provides a pharmaceutical composition adapted for oral administration and for the treatment of stress-induced mood disorders.

[0163] The pharmaceutical composition of the present invention adapted for oral administration may be provided as discrete units such as capsules or tablets; powders or granules; solutions or suspensions in aqueous or non-aqueous liquids; edible forms or whips; or water-in-oil liquid emulsions or oil-in-water liquid emulsions.

[0164] For example, in the case of oral administration in the form of tablets or capsules, the active drug ingredient can be combined with an orally non-toxic pharmaceutically acceptable inert carrier such as ethanol, glycerol, water and the like. Powders are prepared by grinding the compound to an appropriate fine size and mixing with a similar ground pharmaceutical carrier such as an edible carbohydrate such as starch or mannitol. Flavoring agents, preservatives, dispersing agents and coloring agents may also be present.

[0165] Capsules are made by preparing the powder mixture described above and filling the formed gelatin coating. Lubricants and lubricating agents such as colloidal silica, talc, magnesium stearate, calcium stearate, or solid polyethylene glycol can be added to the powder mixture prior to the filling operation. Disintegrants or solubilizing agents such as agar, calcium carbonate, sodium carbonate can also be added to improve the availability of the medicine when the capsule is ingested.

[0166] In addition, if desired or necessary, suitable binders, lubricants, disintegrants and colorants may also be incorporated into the mixture. Suitable binders include starches, gelatin, glucose or beta-lactose, natural sugars such as corn sweeteners, natural and synthetic gums such as gum arabic, tragacanth or sodium alginate, carboxymethyl cellulose, polyethylene glycol, waxes and the like. Lubricants used in these dosage forms include sodium oleate, sodium stearate, magnesium stearate, sodium benzoate, sodium acetate, sodium chloride and the like. Disintegrants include, but are not limited to, starches, methyl cellulose, agar, bentonite, xanthan gum and the like. Tablets are formulated, for example, by preparing a powder mixture, granulating or slugging, adding lubricants and disintegrants, and tableting. The powder mixture is prepared by mixing the previously described diluents or bases, and optionally binders such as carboxymethyl cellulose, alginates, gelatin or polyvinylpyrrolidone, solution retardants such as paraffin, absorption promoters such as quaternary salts, and / or absorbents such as bentonite, kaolin or dicalcium phosphate, with the appropriately milled compound. The powder mixture can be granulated by wetting it with a binder such as syrup, starch paste, gum arabic solution, or a solution of cellulose or polymeric material, and forcing it through a screen. Instead of granulating, the powder mixture may be passed through a tableting machine to yield incompletely formed slugs which are broken down into granules. The granules can be lubricated by the addition of stearic acid, stearates, talc, or mineral oil to prevent adhesion to the tablet forming die. The lubricated mixture is then tabletted. The compounds of the present invention may also be combined with a free-flowing inert carrier and directly tabletted without undergoing a granulating or slugging step. Transparent or opaque protective coatings consisting of shellac seal coatings, sugar or polymeric material coatings, and wax gloss coatings may be provided.Pigments can be added to these coatings to distinguish different unit dosage forms.

[0167] Oral fluids such as solutions, syrups, and elixirs can be prepared as dosage unit forms such that a given quantity contains a predetermined amount of the compound. Syrups can be prepared by dissolving the compound in an aqueous solution with an appropriate flavor, while elixirs are prepared through the use of a non-toxic alcoholic vehicle. Suspensions can be formulated by dispersing the compound in a non-toxic vehicle. Solubilizing and emulsifying agents such as ethoxylated isostearyl alcohol and polyoxyethylene sorbitol ethers, preservatives, flavoring additives such as peppermint oil, or natural sweeteners or saccharin or other artificial sweeteners can also be added.

[0168] In addition to the components described in detail above, the composition may contain other agents conventionally known in the art having a relationship to the type of formulation in question, for example, as suitable for oral administration, flavoring agents can be mentioned.

[0169] The therapeutically effective amount of the compounds of the present invention will depend on a plurality of factors including, for example, the age and weight of the subject, the exact condition and its severity that requires treatment, the nature of the formulation, and the route of administration, and will ultimately be at the discretion of the attending physician or veterinarian.

[0170] Kit Also provided is a kit for use in the method of the present invention for prophylactically treating stress-induced mood disorders.

[0171] The kit may include the agent or composition provided herein and instructions for providing information to a healthcare provider in relation to the specifications according to the methods of the present invention. The kit may optionally contain a second agent or composition. The instructions may be provided in printed form, or in the form of an electronic medium such as a floppy disk, CD or DVD, or in the form of a website address from which such instructions can be obtained. The unit dosage of the compound or composition provided herein, or of the second agent or composition, may include an amount such that a therapeutically or prophylactically effective plasma level of the compound or composition is maintained when administered to a subject. In some embodiments, the compound or composition may include a sterile aqueous pharmaceutical composition or a composition in the form of a dry powder (e.g., lyophilized). In some embodiments, appropriate packaging is provided. As used herein, "packaging" includes a solid matrix or material that is customarily used in the art and that can hold within defined limits the compounds provided herein and / or a second agent suitable for administration to a subject. Such materials include bottles, vials, paper, plastic, and plastic-foil laminated envelopes of glass and plastic (e.g., polyethylene, polypropylene, and polycarbonate).

[0172] The kits described herein contain one or more containers containing the described compounds, signaling molecules, biomolecules and / or particles. The kit also contains instructions for mixing, diluting and / or administering the compounds. The kit also includes other containers having one or more solvents, surfactants, preservatives and / or diluents (e.g., saline (0.9% NaCl), or 5% dextrose), and containers for mixing, diluting the components, or for administering to a sample or to a patient in need of such treatment.

[0173] The composition of the kit may be provided in any suitable form, for example, as a liquid solution or as a dry powder. If the composition provided is a dry powder, the powder may be reconstituted by the addition of a suitable solvent, which may also be provided. In embodiments where a liquid form of the composition is used, the liquid form may be concentrated or ready for immediate use. The solvent will depend on the compound and the mode of use or administration. Suitable solvents for pharmaceutical compositions are well known and available in the literature. The solvent will depend on the compound and the mode of use or administration.

[0174] The kit includes a compartmentalized carrier configured to receive one or more containers, such as vials, test tubes, and the like, each of the containers containing one of the other elements used in the method of the present invention. For example, one of the containers may contain a positive control in the assay. Additionally, the kit may include other components, such as containers for buffers useful in the assay.

[0175] Unless otherwise indicated, all numbers expressing quantities of ingredients, properties, and so forth used in the specification and claims of this disclosure are to be understood as being modified in all instances by the term “about.” Accordingly, unless indicated to the contrary, the numerical parameters set forth in the specification and claims are approximations that may vary depending upon the desired properties sought to be obtained by the embodiments of the present invention. At the very least, and not as an attempt to limit the application of the doctrine of equivalents to the scope of the claims, each numerical parameter should be construed in light of the reported number of significant digits and in reference to the normal rounding techniques. It should be noted that when “about” is placed at the beginning of a numerical listing, it modifies each number in that numerical listing. Further, in some numerical listings of ranges, some of the lower limits listed may be greater than some of the upper limits listed. Those skilled in the art will recognize that the selected subset will require the selection of upper limits that exceed the selected lower limits. The term “about” refers to ±10% of the referenced value. In other words, the numerical value can be in the range of 90% to 110% of the stated value.

[0176] The present invention will be better understood from the following examples. However, it will be immediately apparent to those skilled in the art that the specific methods and results discussed are merely illustrative of the invention, which is more fully described in the claims that follow.

[0177] Examples Example 1: Preventive Efficacy of 5-HT4R Agonists against Stress Enhancing stress resilience can protect against stress-induced mental disorders in at-risk populations. The inventors have previously reported that (R,S)-ketamine acts as a prophylactic against stress when administered one week before stress. The inventors have shown that the selective 5-hydroxytryptamine (5-HT) (serotonin) reuptake inhibitor (SSRI) fluoxetine (Flx) is ineffective as a prophylactic, but that the serotonin 4 receptor (5-HT 4It was hypothesized that other serotonergic compounds such as R agonists could act as prophylactic agents. The inventors utilized chronic corticosterone (CORT) administration or contextual fear conditioning (CFC) to test whether three 5-HT 4 R agonists with various affinities could defend against stress in two mouse strains. Mice were administered saline, (R,S)-ketamine, Flx, RS-67,333, prucalopride, or PF-04995274 at various doses, and one week later, the mice were subjected to chronic CORT or CFC. In C57BL / 6N mice, chronic Flx administration attenuated CORT-induced body weight fluctuations and increased open arm entries in the elevated plus maze (EPM). Chronic RS-67,333 administration attenuated CORT-mediated body weight fluctuations and defended against depressive- and anxiety-like behaviors. In 129S6 / SvEv mice, RS-67,333 attenuated learned fear in males but not in females. RS-67,333 was ineffective against stress-induced depressive-like behavior in the forced swim test (FST) but prevented anxiety-like behavior in both genders. Prucalopride and PF-04995274 attenuated learned fear and decreased stress-induced depressive-like behavior. Electrophysiological recordings following (R,S)-ketamine or prucalopride administration revealed that both drugs modified AMPA receptor-mediated synaptic transmission in CA3. These data indicate that, in addition to (R,S)-ketamine, 5-HT 4 R agonists are also effective prophylactic agents against stress, suggesting that 5-HT 4 R may be a novel target for prophylactic drug development.

[0178] Here, the inventors hypothesized that 5-HT 4 R, which is highly associated with depression and anxiety disorders, may have a role in stress resilience. The inventors utilized three 5-HT 4Trials were targeted at 5-HT4 agonists. First, RS-67,333 (1-(4-amino-5-chloro-2-methoxyphenyl)-3-[1-(n-butyl)-4-piperidinyl]-1-propanone HCl) is a high-affinity 5-HT4 partial agonist

[22] . This drug is effective in improving behavioral deficits, reducing the number of amyloid plaques and the levels of amyloid-beta (Aβ) species, as well as reducing hippocampal astrogliosis and microgliosis in a 5×FAD mouse model of Alzheimer's disease (AD)

[23] . Second, prucalopride (4-amino-5-chloro-2,3-dihydro-N-[1-(3-methoxypropyl)-4-piperidinyl]-7-benzofuran carboxamide monohydrochloride) is a selective high-affinity 5-HT4 agonist

[24] . It received FDA approval for chronic constipation in 2018 and is currently being tested for chronic pseudo-obstruction. Prucalopride was tested in two separate clinical trials to investigate its effect on emotional processing in healthy volunteers after acute (e.g., single-dose) or chronic (e.g., 1-week) administration [25, 26]. Third, PF-04995274 (4-[4-[4-(tetrahydrofuran-3-yloxy)-benz[d]isoxazol-3-yloxymethyl]-piperidin-1-ylmethyl]-tetrahydropyran-4-ol) is a potent partial 5-HT4 agonist

[27] . Clinical trials were conducted to evaluate PF-04995274 alone or in combination with donepezil for scopolamine-induced deficits in psychomotor and cognitive function in healthy adults, but this trial was terminated, not due to safety concerns

[28] . Currently, a clinical trial is underway to test whether adjunctive administration of PF-04995274 has a positive effect on emotional processing and neuroactivity in treatment-resistant depression (TRD) patients mediated compared to placebo

[29] . 4 4 4 4 4 4

[0179] 5-HT4 4To determine whether R agonists can be potential prophylactic agents against stress, the inventors utilized two different stress models (acute and chronic) in two different strains of mice (C57BL / 6NTac and 129S6 / SvEv). The inventors found that RS-67,333, prucalopride, and PF-04995274 attenuate learned fear. RS-67,333 prevented depressive-like behavior when administered chronically and prevented stress-induced anxiety-like behavior in both genders when administered acutely. Prucalopride and PF-04995274 decreased stress-induced depressive-like behavior in the FST. Prophylactic (R,S)-ketamine and 5-HT 4 To investigate the common or different mechanisms of action of R agonists, the inventors utilized slice electrophysiological examinations to investigate spontaneous glutamatergic transmission in CA3. The inventors found that (R,S)-ketamine and prucalopride attenuate bursts of large-amplitude AMPA receptor-mediated synaptic currents. These data, in addition to (R,S)-ketamine, 5-HT 4 suggest that R agonists may also be effective prophylactic agents against stress and can modify AMPA-related glutamatergic transmission to enhance stress resilience.

[0180] Materials and Methods Mice: All mice were housed in a 12-hour (6:00 - 18:00) light-dark colony room at 22°C. Food and water were provided ad libitum. Behavioral tests were performed during the light period. C57BL / 6NTac mice: Male C57BL / 6NTac mice were purchased at 8 weeks of age from Taconic Farms (Lille Skensved, Denmark) and housed at 5 mice / cage before the start of CORT treatment. All tests were performed according to the guidelines for the care of laboratory animals and a protocol approved by the Institutional Animal Care and Use Committee (IACUC) (European Directive 2010 / 63 / EU for the protection of animals used for scientific purposes, authorization number 92 - 256B, authorization ethical committee CEEA n°262012_098). 129S6 / SvEv mice: Male and female 129S6 / SvEvTac mice were purchased at 7 - 8 weeks of age from Taconic (Hudson, NY). The procedures described herein were performed in accordance with the National Institutes of Health (NIH) guidelines and approved by the IACUC of the New York State Psychiatric Institute (NYSPI).

[0181] Stress model: Corticosterone (CORT) model: In this model, glucocorticoid levels are externally increased in C57BL / 6NTac mice. This chronic CORT elevation dysregulates the hypothalamic - pituitary - adrenal (HPA) axis in a manner similar to that observed in clinical depression. The dose and duration of CORT treatment were selected based on previous studies [20, 30]. CORT (35 μg / ml, equivalent to approximately 5 mg / kg / day) dissolved in 0.45% hydroxypropyl - β - cyclodextrin (β - CD) or vehicle (VEH) (0.45% β - CD) was freely available in drinking water contained in opaque bottles to protect from light. VEH and CORT - treated water were changed every 3 days to prevent possible degradation. Contextual fear conditioning (CFC): The 3-shock CFC procedure was performed as previously described [31, 32]. Briefly, mice were placed in context A, and 32 s of shock (0.75 mA) was delivered 180 s, 240 s, or 300 s after placement in context A. Mice were removed from that context 15 s after the end of the shock (at the 317 s mark). For context recovery, mice were returned to context A for 300 s. Electrophysiological examination: Electrophysiological examinations were performed as previously described

[33] . Statistical analysis: Results from data analysis were presented as mean ± SEM. Alpha was set at 0.05 for all analyses. Data were analyzed using GraphPad Prism v7.0 or v8.0. In all experiments, unless otherwise stated, one-way or two-way repeated measures analysis of variance was applied to the data as appropriate. Significant main effects and / or interactions were followed up by Fisher's PLSD post hoc analysis or unpaired t-tests. All main effects, interactions, and p-values are listed in Table 2. Drugs: All drugs were prepared in physiological saline, and all injections were administered intraperitoneally (i.p.) at a volume of 0.1 cc / 10 mg body weight, unless otherwise stated. Fluoxetine hydrochloride (Flx): Flx (BioTrend Chemicals AG, BG197, Zurich, Germany) was administered in the drinking water (18 mg / kg / day) 3 weeks before the start of CORT. RS-67,333 (RS): RS-67,333 (Tocris Bioscience, 0989, Bristol, UK) was administered either chronically or as a single injection. In the chronic experiment, RS-67,333 (1.5 mg / kg / day) was administered via an ALZET osmotic minipump (ALZET, Model 2004, Cupertino, CA)

[30] . In the acute experiment, RS-67,333 was administered as a single dose of 1.5, 10, or 30 mg / kg body weight one week before the start of CFC. RS-67,333 was dissolved in saline using an ultrasonic homogenizer (BioLogics, Model 3000, Manassas, VA). (R,S)-Ketamine (K): (R,S)-Ketamine (Ketaset III ketamine HCl injection, Fort Dodge Animal Health, Fort Dodge, IA) was administered as a single dose of 30 mg / kg body weight one week before the start of CFC. The dose of 30 mg / kg body weight was selected in the 129S6 / SvEv experiment because past studies had shown that it was an effective dose for preventive efficacy [S1]. Prucalopride: Prucalopride (Sigma, 179474-81-8, St. Louis, MO) was administered as a single dose of 3 or 10 mg / kg body weight one week before the start of CFC. Prucalopride was dissolved in saline using an ultrasonic homogenizer (BioLogics, Model 3000, Manassas, VA). PF-04995274: PF-04995274 (Sigma, Catalog No. 1331782-27-4, St. Louis, MO) was administered as a single dose of 3 or 10 mg / kg body weight one week before the start of CFC. PF-04995274 was dissolved in saline using an ultrasonic homogenizer (BioLogics, Model 3000, Manassas, VA). Osmotic minipump implantation: An ALZET osmotic minipump (model 2004, 0.25 μl / hour, 28 days) was implanted subcutaneously under isoflurane anesthesia as previously described [S2]. The osmotic minipump was rotated subcutaneously 2 - 3 times / week. Behavioral assays: All experiments were approved by the Institutional Animal Care and Use Committee (IACUC) of the New York Psychiatric Institute (NYSPI). Elevated plus maze (EPM): Tests were conducted as previously described [S3]. Briefly, the maze is a cruciform apparatus consisting of four arms, two open arms and two enclosed arms, connected by a central platform 50 cm above the floor. Mice were individually placed in the center of the maze facing an open arm and allowed to explore the maze for 5 min. The time spent and the number of entries into the open arms were used as an index of anxiety. Videos were scored using ANY-maze behavioral tracking software (Stoelting, Wood Dale, IL). Food intake suppression by novel environment: NSF is a conflict test that induces competing motivations, namely the urge to eat and the fear of venturing into the center of a brightly lit arena. Since classical anxiolytics reduce this measure, the latency to initiate food intake is used as an index of anxiety-like behavior. NSF tests were conducted for an 8-min period as previously described [S3]. Briefly, the test apparatus consisted of a plastic box (50 × 50 × 20 cm), the floor of which was covered with approximately 2 cm of bedding. In 129S6 / SvEv experiments, mice were food restricted for 12 h. In C57BL / 6N experiments, mice were food restricted for 24 h. At the time of the test, one pellet of chow (regular diet) was placed on a paper platform located in the center of the box. Each animal was placed in a corner of the box and a stopwatch was immediately started. The time to initiate food intake (defined as the mouse taking a bite of the pellet) was recorded. Immediately thereafter, the animal was transferred to its home cage and the amount of food consumed by the mouse over the following 5 min was measured as a control for possible confounding factors related to changes in appetite. Sucrose splash test: This test consists of spraying 200 μl of a 10% sucrose solution onto the nose of the mouse. The grooming period was quantified using Stopwatch+ (Center for Behavioral Neuroscience, Georgia State University). Forced swim test (FST): The FST is typically used in rodents to screen for potential human antidepressants [S4, S5]. Indeed, many papers that test ketamine only in mouse models have observed an effect in the FST [S6 - S8]. In the FST, as opposed to swimming, the time spent in a motionless state is used as a measure of depressive behavior. The FST was performed as described previously [1]. Briefly, mice were placed in a transparent plastic bucket 20 cm in diameter and 23 cm deep filled to 2 / 3 with water at 22°C. Mice were filmed from the side for 6 minutes and exposed to the swim test for 2 consecutive days. The immobility time was scored by an experimenter blinded to the experimental group. Open field (OF): The OF assay was performed as described previously [3]. Briefly, locomotor activity was quantified in four Plexiglas open field boxes 43×43 cm 2 (MED Associates, Georgia, Vermont) by two sets of 16 pulsed modulated infrared beams to opposite walls 2.5 cm apart, recording the x - y walking behavior. The activity chamber was connected with a computer interface for data acquisition at 100 - millisecond resolution. The computer defined grid lines dividing the central and peripheral regions, and the central square consisted of four lines 11 cm from the walls. Electrophysiological examination: One week after injection of saline, (R,S)-ketamine (30 mg / kg), or prucalopride (3 mg / kg), mice were anesthetized by isoflurane inhalation, decapitated, and the brains were rapidly removed. CA3 slices (350 μm) were placed in ice - cold partial sucrose artificial cerebrospinal fluid (ACSF) solution equilibrated with 95% O 2 / 5% CO 2 : (in mM) 80 NaCl, 3.5 KCl, 4.5 MgSO4 , 0.5 CaCl 2 , 1.25 H 2 PO 4 , 25 NaHCO 3 , 10 glucose and 90 sucrose, were cut out by a vibratome (Leica VT1000S), stored in the same solution at 37 °C for 30 minutes, and then stored at room temperature until use. Recordings were made in ACSF (in mM: 124 NaCl, 8.5 KCl, 1 NaH 2 PO 4 , 25 NaHCO 3 , 20 glucose, 1 MgCl 2 , 2 CaCl 2 ) at 30 - 32 °C (TC324 - B; Warner Instrument Corp). Whole - cell voltage - clamp recordings (-70 mV) were obtained using a patch pipette (4 - 6 MΩ) containing (in mM): 135 K - gluconate, 5 KCl, 0.1 EGTA - Na, 10 HEPES, 2 NaCl, 5 ATP, 0.4 GTP, 10 phosphocreatine (pH 7.2; 280 - 290 mOsm). Bicuculline (5 μM) was also included in the bath solution to inhibit GABA A R. NBQX (20 mM) was added in the second half of the recording to inhibit AMPAR synaptic currents. The patch pipettes were made from borosilicate glass (A - M Systems, Sequim, WA) using a micropipette puller (Model P - 1000; Sutter Instruments). Recordings were made without correction of the junction potential. Pyramidal cells were visualized and targeted via an infrared differential interference contrast microscope (IR - DIC; 40x objective) lens on an Axioskop - 2 FS (Zeiss).

[0182] Results Chronic administration of RS - 67,333 is prophylactic against stress in male mice We previously reported that chronic Flx administration (for 3 weeks) was not preventive in 129S6 / SvEv mice [3]. However, whether other serotonergic agents could act as preventive agents had not yet been determined. Here, we administered Flx (18 mg / kg / day) in the drinking water or RS-67,333 (1.5 mg / kg / day) by osmotic minipump 3 weeks prior to CORT administration in C57Bl / 6NTac male mice, and subsequently conducted a series of behavioral tests including the EPM, novel environment-induced feeding suppression (NSF), and sucrose splash test (ST) (Figures 1A-1B). CORT increased body weight over the 6-week behavioral protocol as previously observed

[34] , (Figures 1C-1F), which was attenuated by Flx and RS-67,333 administration.

[0183] In the EPM, CORT+Veh, CORT+Flx, and CORT+RS-67,333 administrations did not modify the time spent in the open arms when compared to VEH+Veh administration (Figure 1G). However, CORT+Veh mice showed a significant decrease in the number of entries into the open arms of the EPM when compared to VEH+Veh mice (Figure 1H). CORT+Flx and CORT+RS-67,333 mice had significantly more entries into the open arms of the EPM when compared to CORT+Veh mice. There was no difference in the total distance traveled in the EPM among any of the groups (Figure 1I).

[0184] Next, the NSF task was performed to assay anxiety-like behavior (Figures 1J-1K). CORT+Veh mice showed an increase in the latency to approach the pellet diet when compared to VEH+Veh mice. CORT+RS-67,333 mice showed a significant decrease in the latency to approach the pellet when compared to CORT+Veh mice, while CORT+Flx mice did not.

[0185] Finally, in the ST, CORT+Veh mice showed a decrease in grooming duration compared to VEH+Veh mice (Figure 1L). CORT+RS-67,333 showed an increase in grooming duration compared to CORT+Veh mice, but CORT+Flx mice did not. These data suggest that chronic RS-67,333 is prophylactic against a wide range of CORT-induced behavioral abnormalities, while chronic Flx administration is not.

[0186] Single injection of RS-67,333 attenuates learned fear and protects against stress-induced feeding suppression in male mice Previously, we showed that a single injection of (R,S)-ketamine is prophylactic against stress-induced depressive-like behavior and attenuates learned fear in 129S6 / SvEv mice [3]. Here, we sought to determine whether a single injection of RS-67,333 could also prevent various maladaptive behaviors following a single acute stressor. Male 129S6 / SvEv mice were injected with saline or RS-67,333 (1.5, 10, or 30 mg / kg) (Figure 2A). One week later, the mice were subjected to 3-shock CFC. Mice administered 30 mg / kg of RS-67,333 showed significantly less freezing behavior during CFC training compared to mice administered saline, but not at 1.5 or 10 mg / kg (Figure 2B). Five days later, the mice were re-exposed to the training context. Mice administered 1.5 or 10 mg / kg of RS-67,333 showed significantly less freezing behavior compared to mice administered saline, but not at 30 mg / kg (Figures 2C-2D).

[0187] Following CFC, the mice were subjected to the FST. On day 1, mice administered 10 mg / kg were significantly less immobile compared to saline mice, but not at 1.5 or 30 mg / kg (Figure 2E). However, on day 2, the immobility time was equivalent among all groups (Figures 2F-2G).

[0188] Next, mice administered with saline or RS-67,333 (10 mg / kg) were tested in the OF. Mice in both groups moved the same distance (Figure 2H) and spent the same amount of time in the center of the arena (Figure 2I). Next, the mice were tested in the EPM, and neither the time spent in the open arms of the maze nor the entries into the open arms differed significantly between the saline or RS-67,333 mice (Figures 2J - 2K).

[0189] Finally, the mice were subjected to NSF. Mice administered prophylactic RS-67,333 (10 mg / kg) exhibited significantly reduced pellet approach latency (Figures 2L - 2M). However, neither the food eaten in the home cage nor the weight loss after food deprivation differed between the groups (Figures 2N - 2O). Overall, these data indicate that a single injection of RS-67,333 is effective as a prophylactic agent in attenuating learned fear and preventing stress-induced feeding suppression as measured by the FST in male 129S6 / SvEv mice, but is not effective in depressive-like behavior.

[0190] A single prophylactic injection of RS-67,333 protects against stress-induced anxiety-like behavior in female mice The inventors then sought to determine whether a single injection of RS-67,333 could also be prophylactic in female mice. Female 129S6 / SvEv mice were injected with saline or RS-67,333 (1.5 or 10 mg / kg) (Figure 3A). One week later, the mice were subjected to 3-shock CFC. The entire group of mice exhibited the same level of freezing behavior during CFC training (Figure 3B). Five days later, the mice were re-exposed to the training context. Again, the entire group of mice exhibited the same level of freezing behavior (Figures 3C - 3D). Following CFC, the mice were subjected to the FST. On day 1 (Figure 3E) and day 2 (Figures 3F - 3G) of the FST, the entire group of mice had the same level of immobility.

[0191] Next, the mice were tested in the OF and EPM. All groups of mice moved equal distances and spent equal amounts of time in the center of the arena in the OF (Figs. 3H - 3I). Similarly, in the EPM, the mice spent equal amounts of time in the open arms of the maze (Fig. 3J) and had equal numbers of entries into the open arms (Fig. 3K).

[0192] Finally, the mice were assayed in the NSF paradigm. Prophylactic RS - 67,333 (10 mg / kg) significantly reduced the latency to initiate feeding, while RS - 67,333 (1.5 mg / kg) did not (Figs. 3L - 3M). Neither the amount of food eaten in the home cage nor the weight loss after feeding disruption differed between the groups (Figs. 3N - 3O). Overall, these data indicate that RS - 67,333 does not attenuate learned fear or protect against stress - induced depressive - like behavior in female 129S6 / SvEv mice, but can prevent stress - induced feeding suppression in NFS.

[0193] Single prophylactic injection of prulopride or PF - 04995274 is prophylactic against stress in male mice The inventors next sought to determine whether other 5 - HT 4 R agonists could also be prophylactic in male 129S6 / SvEv mice. Male 129S6 / SvEv mice were injected with saline, (R,S) - ketamine (30 mg / kg), prulopride (3 or 10 mg / kg), or PF - 04995274 (3 or 10 mg / kg) (Fig. 4A). One week later, the mice were given 3 - shock CFC. All groups of mice exhibited equal levels of freezing behavior during CFC training (Fig. 4B). Five days later, the mice were re - exposed to the training context. As the inventors have previously published, (R,S) - ketamine attenuated learned fear (Figs. 4C - 4D). Interestingly, 3 mg / kg of prulopride and 10 mg / kg of PF - 04995274 attenuated learned fear compared to saline administration, while 10 mg / kg of prulopride and 3 mg / kg of PF - 04995274 did not.

[0194] Subsequent to CFC, mice were subjected to FST. On day 1, the entire group of mice had an equivalent level of immobility (Figure 4E). On day 2, administration of (R,S)-ketamine decreased the immobility time when compared to saline administration (Figures 4F - 4G). Moreover, 3 mg / kg of prucalopride and 10 mg / kg of PF-04995274 decreased the immobility time when compared to saline administration, while 10 mg / kg of prucalopride and 3 mg / kg of PF-04995274 did not.

[0195] Next, stress-induced anxiety-like behavior was quantified. In the OF, the entire group of mice moved an equivalent distance (Figure 4H). In the EPM, the entire group of mice spent an equivalent amount of time in the open arms (Figure 4I) and entered the open arms an equivalent number of times (Figure 4J). In the NSF paradigm, the entire group of mice approached the pellet for an equivalent amount of time (Figures 4K - 4L). Finally, all mice lost an equivalent amount of body weight during the NSF paradigm (Figure 4M). In summary, these data indicate that single injections of prucalopride or PF-04995274 provide prophylactic efficacy in attenuating learned fear and reducing stress-induced depressive-like behavior. However, these drugs are not prophylactic against stress-induced anxiety-like behavior.

[0196] (R,S)-Ketamine and prucalopride exhibit a common mechanism by reducing bursts of large AMPA receptor-driven synaptic currents in CA3 Next, the inventors studied 5-HT such as (R,S)-ketamine and prucalopride 4An attempt was made to elucidate the potential common mechanism among R agonists. Specifically, since the present inventors have previously reported that prophylactic (R,S)-ketamine modifies the activity in the ventral CA3 (vCA3) of the abdomen but not in the DG [6], it was hypothesized that there might be a similarity between the effects of (R,S)-ketamine and prucalopride...

Claims

1. A pharmaceutical composition comprising prucalopride, PF-04995274, or a combination thereof, for use in preventing or delaying the onset of stress-induced fear and depressive-like behavior in a subject prior to a stressor, the pharmaceutical composition being administered to the subject in a prophylactically effective amount 48 hours to 3 weeks prior to the stressor.

2. The pharmaceutical composition according to claim 1, wherein the pharmaceutical composition is administered to the subject about 72 hours to about 2 weeks prior to the stressor.

3. The pharmaceutical composition according to claim 1 or 2, wherein the pharmaceutical composition is administered to the subject about 1 week prior to the stressor.

4. The pharmaceutical composition according to claim 1 or 2, wherein the pharmaceutical composition is administered to the subject once prior to the stressor.

5. The pharmaceutical composition according to claim 1 or 2, wherein the pharmaceutical composition is administered to the subject orally, intravenously, intranasally, or via injection.

6. The pharmaceutical composition according to any one of claims 1 to 5, wherein the use further comprises administering an effective amount of an antidepressant, an antianxiety drug, or a combination thereof.

7. The pharmaceutical composition according to any one of claims 1 to 6, wherein the use further comprises administering an effective amount of a selective serotonin reuptake inhibitor (SSRI), or a pharmaceutically acceptable salt or derivative thereof.

8. The pharmaceutical composition according to any one of claims 1 to 7, wherein the use further comprises administering an effective amount of fluoxetine, paroxetine, sertraline, lithium, riluzole, prazosin, lamotrigine, ifenprodil, or a combination thereof.

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