WO2014011768A1

WO2014011768A1 - Anti-anxiety treatment

Info

Publication number: WO2014011768A1
Application number: PCT/US2013/049917
Authority: WO
Inventors: Stephen H. Curry
Original assignee: ADISPELL Inc
Current assignee: ADISPELL Inc
Priority date: 2012-07-10
Filing date: 2013-07-10
Publication date: 2014-01-16
Anticipated expiration: 2015-01-10

Description

ANTI-ANXIETY TREATMENT

[0001] This application claims the priority benefit of U.S. Provisional Patent

Application Serial No. 61/670,030, filed July 10, 2012, which is hereby incorporated by reference in its entirety. FIELD OF THE INVENTION

[0002] The present invention relates to methods for treating and/or preventing anxiety in a subject that involve administering small organic molecules or RNA aptamers.

BACKGROUND OF THE INVENTION

[0003] The standard definition of anxiety is to be found in such compendia as DSM V. Anxiety can be described as a subjective feeling of unease, discomfort, apprehension or fearful concern accompanied by a host of autonomic and somatic manifestations. Anxiety can be a normal response to danger, but if it is prolonged, disproportionate in scale or duration, and interferes with normal daily activities, then a medical diagnosis of Anxiety Disorder is made. Extensive literature draws similarities between anxiety and fear, and also provides clear distinctions between these two different but overlapping phenomena. For example, fear in the absence of threat is anxiety.

Anxiety disorder can be considered to occur when anxiety causes physiological symptoms.

[0004] A patient diagnosed with anxiety experiences one or more of a wide variety of thoughts that can include several of, but not necessarily all of: worry, repetitive thinking, fearfulness, embarrassment, humiliation, phobia, panic, social inadequacy, flash-backs, restlessness, and a feeling of being on edge. These thoughts can vary from being a mild annoyance to a life-modifying pathology.

[0005] The patient also experiences a wide variety of somatic phenomena, such as palpitations, sweating, headache, tachypnea, trembling, chest pain, urinary frequency, motor tension, trouble swallowing, sleep problems, irritability, nausea and dizziness. Emotional and physical sensations together can lead to problems in cognition (learning, memory, concentration, decision-making ability etc.). [0006] Classification of anxiety disorders commonly includes sub-classifications of panic, phobia, post-traumatic symptom disorder, acute stress, substance-abuse linked anxiety, and many others.

[0007] The National Institute of Mental Health provides some information about the occurrence of anxiety related disorders. By their measures (circa 2012), 18.1% of adults in the U.S. suffered some form of anxiety in the prior twelve months. Of these cases, 22.8% are considered severe. Of the population of severe cases, 36.9% are receiving a form of treatment for anxiety.

[0008] Many anxiety disorders affect youth. For adolescents aged 13-18, 25.1 % will have suffered anxiety at some time. For those that do, 5.9% are considered to have a severe case. Historically, anxiety occurs more often with females.

[0009] Most individuals that seek treatment for anxiety-related disorders will work with a licensed psychologist. At times, it is necessary to move to a form of medication; usually, but not always, in conjunction with continued professional consultation. Current medications can relieve some anxiety symptoms. In general, they are known to provide temporary relief, but do not necessarily always treat the underlying cause of anxiety. The medications often come with side effects and safety concerns. Some anxiety medications are habit forming and physically addictive.

[0010] Despite the sedating properties of anti-anxiety drugs, some people who take anti-anxiety medication experience paradoxical excitement. The most common paradoxical reactions are increased anxiety, irritability, and agitation. However, more severe effects can also occur, including mania, hostility, rage, aggressive or impulsive behavior and/or hallucinations.

[0011] Tranquilizers are used to treat anxiety and work by slowing central nervous system activity. Their relaxing and calming effects have made them the most widely prescribed medication for anxiety. Antidepressants were developed to treat depression but are also effective for anxiety. Although these options begin to alter brain chemistry after the very first dose, their full effect requires a series of changes to occur. It can take about 4 to 6 weeks before symptoms start to fade. Therefore, it is necessary to take these medications continuously and long enough to let them work.

[0012] High-potency benzodiazepines combat anxiety and have few side effects, other than drowsiness. Because people can become desensitized to benzodiazepines and may need increasing doses to get the same effect, the drugs are generally prescribed for short periods, especially for people who have abused drugs or alcohol and who become dependent on medication easily. An exception is people with panic disorder, who can take benzodiazepines for up to a year without harm.

[0013] Anti-anxiety drugs like benzodiazepines work by reducing brain activity. While this temporarily relieves anxiety, it can also lead to unwanted side effects. Some people feel sleepy, foggy, and uncoordinated even on low doses of benzodiazepines. Other side-effects could include slurred speech, confusion, lightheadness, memory loss, forgetfulness, nausea and blurred vision.

[0014] Some people experience withdrawal symptoms if they stop taking benzodiazepines abruptly instead of tapering use. Anxiety can return once medication is stopped. These potential problems have led some physicians to avoid using these drugs or have caused them to use benzodiazepines in inadequate doses.

[0015] Other medications for treating anxiety have gained in popularity because of safety concerns with many anti-anxiety drugs. The alternatives to the anti-anxiety tranquilizers include antidepressants, buspirone, and beta blockers.

[0016] Some of the newest antidepressants are called selective serotonin reuptake inhibitors, or SSRIs. These drugs alter the levels of the neurotransmitter serotonin in the brain. Like other neurotransmitters, serotonin is involved in communication between brain cells. SSRIs are used to treat panic disorder when it occurs in combination with obsessive-compulsive disorder (OCD), social phobia, or depression. These medications are started at low doses and gradually increase until a beneficial effect occurs.

[0017] SSRIs have fewer side effects than older antidepressants, but they sometimes produce slight nausea or agitation when people first start to take them.

Sometimes, these symptoms can fade with time. Some people also experience sexual dysfunction with SSRIs.

[0018] Buspirone (Buspar®), an azapirone, is a newer anti-anxiety tranquilizer used to treat General Anxiety Disorder. Possible side effects include drowsiness, dizziness, headaches, constipation, diarrhea, and nausea. Buspirone must be taken consistently for at least 2 weeks to achieve an anti-anxiety effect.

[0019] Tricyclics are older than SSRIs. They are also started at low doses that are gradually increased. They sometimes cause dizziness, drowsiness, dry mouth, and weight gain, which can usually be corrected by changing the dosage or switching to another tricyclic medication. [0020] Monamine oxidase inhibitors (MAOIs) are the oldest class of

antidepressant medications. Those who take MAOIs cannot eat a variety of foods and beverages that contain tyramine (including cheese and red wine), or take certain medications, including some types of birth control drugs, pain relievers, cold and allergy medications, and herbal supplements as these substances can interact with MAOIs to cause dangerous increases in blood pressure. MAOIs can also react with SSRIs to produce a serious condition called "serotonin syndrome," which can cause confusion, hallucinations, increased sweating, muscle stiffness, seizures, changes in blood pressure or heart rhythm, and other potentially life-threatening conditions.

[0021] Beta-blockers are often used to treat high blood pressure and heart conditions, but can also prevent the physical symptoms that accompany certain anxiety disorders. When a situation that can produce anxiety can be predicted (such as giving a speech), a doctor may prescribe a beta-blocker to keep physical symptoms of anxiety under control. Beta-blockers carry the potential side effects of light-headedness, sleepiness, nausea, and unusually slow pulse.

SUMMARY OF THE INVENTION

[0022] A first aspect of the present invention is directed to a method that improves, prevents, or treats anxiety and anxiety symptoms in a subject This method involves selecting a subject in need of improvement, treatment, and/or prevention of anxiety and/or anxiety symptons, and administering to the selected subject a ligand that binds to a regulatory site on the extracellular domain of the nicotinic acetylcholine receptor and improves, prevents, or treats anxiety and anxiety symptoms in the subject.

[0023] Current treatments for relieving anxiety can relieve some anxiety symptoms, providing temporary relief, but not necessarily treating the underlying cause of anxiety. Most anti-anxiety medications have a variety of side effects and safety concerns, and some anti-anxiety medications are habit forming or physically addictive. The present invention is directed to ligands, including small organic molecules and RNA aptamers of various types, that bind to specific sites on the nicotinic acetylcholine receptors. These ligands either inhibit (called Class 1 ligands) or alleviate (called Class 2 ligands) the inhibition of receptor channels and so inhibit or restore (despite the continued presence of the inhibitor) the function of the receptor in its role to promote nerve activity and health. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 is a graph showing brain (ng/g) and plasma (ng/ml) concentrations of ecgonine methyl ester ("EME") in rats following intraperitoneal administration of a 10 mg/kg dose at 0, 1, 4, 8, and 24 hours.

[0025] Figures 2A-2C are Morris water maze traces of three individual rats. Figure 2A shows the trace of a young control rat swimming in the Morris water maze bath for approximately 30 seconds searching for the platform, with no drug treatment and before training. Figure 2B shows the trace of an aged anxious rat swimming almost entirely at the perimeter of the water bath with no drug treatment and before training. Figure 2C shows the trace of the aged anxious rat of Figure 2B after 10 days of treatment with EME at lOmg/kg per day. All doses of drug were administered by intraperitoneal injection.

[0026] Figure 3 is a graph measuring thigmotaxis, an index of anxiety in young rats (control), aged anxious rats, and aged anxious rats following treatment with 3mg/kg or 10 mg/kg of EME treatment. The diagram shows percent of time (y-axis) spent at the perimeter of the water bath ("hugging the walls"), with a high percentage indicating severe anxiety. The drug doses reduced the percent of time at the perimeter and therefore the severity of anxiety.

[0027] Figure 4 is a graph showing a decline in the level of thigmotaxis as a function of time in the aged no-treatment control animals, the aged animals treated with 3 mg/kg of EME, and the aged animals treated with 10 mg/kg of EME as described in Figure 3.

DETAILED DESCRIPTION OF THE INVENTION

[0028] A first aspect of the present invention is directed to a method that improves, prevents, or treats anxiety and anxiety symptoms in a subject. This method involves selecting a subject in need of improvement, treatment, and/or prevention of anxiety and/or anxiety symptoms, and administering to the selected subject a ligand that binds to a regulatory site on a nicotinic acetylcholine receptor and improves, prevents, or treats anxiety and anxiety symptoms in the subject.

[0029] As used herein, "improve" and "improvement" indicate a lessening of anxiety in the subject and/or improving the condition of the patient, and not enhancement of the disease state. [0030] In accordance with this and all aspects of the present invention, a subject suitable for treatment using methods of the present invention includes any animal, preferably a mammal, e.g., human, non-human primate, rodent, cow, horse, sheep, pig, goat, deer, elk, bison, etc.

[0031] "Anxiety" as used herein includes any state or condition that causes anxiety behavior or concern by a subject. The intent of this invention is to treat or modify any anxiety behavior and symptoms caused by conditions that include, but are not limited to, the following anxiety-related disorder types in a subject: dissociative, eating, impulse- control, adjustment, anxiety, mood, personality, psychotic, post-traumatic stress, sexual, sleep-related, and somatoform disorders.

[0032] The standard definition of anxiety is found in the Diagnostic and

Statistical Manual of Mental Disorders (DSM V). Anxiety can be described as a subjective feeling of unease, discomfort, apprehension or fearful concern accompanied by a host of autonomic and somatic manifestations. Anxiety can be a normal response to danger, but if it is prolonged, disproportionate in scale or duration, and interferes with normal daily activities, then a medical diagnosis of Anxiety Disorder is made. Extensive literature draws similarities between anxiety and fear, and also provides clear distinctions between these two different but overlapping phenomena. For example, fear in the absence of threat is anxiety. Anxiety disorder can be considered to occur when anxiety causes physiological symptoms.

[0033] A patient diagnosed with anxiety experiences one or more of a wide variety of thoughts that can include several of, but not necessarily all of: worry, repetitive thinking, fearfulness, embarrassment, humiliation, phobia, panic, social inadequacy, flash-backs, restlessness, and a feeling of being on edge. These thoughts can vary from being a mild annoyance to a life-modifying pathology.

[0034] The patient also experiences a wide variety of somatic phenomena, such as palpitations, sweating, headache, tachypnea, trembling, chest pain, urinary frequency, motor tension, trouble swallowing, sleep problems, irritability, nausea and dizziness. Emotional and physical sensations together can lead to problems in cognition (learning, memory, concentration, decision-making ability etc.).

[0035] Classification of anxiety disorders commonly includes sub-classifications of panic, phobia, post-traumatic symptom disorder, acute stress, substance-abuse linked anxiety, and many others. [0036] In accordance with the methods of the present invention anxiety can be improved and anxiety in a subject can be treated or prevented by administering to the subject a ligand that binds to the nicotinic acetylcholine receptor and treats or prevents anxiety or anxiety symptoms in a subject. Laboratory work has demonstrated the existence of a regulatory site within the extracellular domain of the nicotinic

acetylcholine receptor in the brain that is distinct from the binding site of the natural ligand acetylcholine. As used herein, the term "ligand" includes, but is not limited to, small organic molecules, aptamers, and other compounds that similarly bind to this regulatory site on the nicotinic acetylcholine receptor and induce an allosteric change in the receptor, thereby improving or enhancing the flow of inorganic cations through the ion channel of the receptor. The enhanced flow of inorganic cations through the receptor channel improves nerve function and relieves or prevents anxiety. The characteristics and binding function of this regulatory site on the nicotinic acetylcholine receptor have previously been described (see e.g., Hess et al, "Mechanism-Based Discovery of Ligands that Counteract Inhibition of the Nicotinic Acetylcholine Receptor by Cocaine and MK- 801," Proc. Nat. Acad. Set 97(25): 13895-13900 (2000), Chen et al, "Mechanism-Based Discovery of Small Molecules that Prevent Noncompetitive Inhibition by Cocaine and MK-801 Mediated by Two Different Sites on the Nicotinic Acetylcholine Receptor," Biochemistry 43 : 10149-10156 (2004), Hess et al, "Reversing the Action of

Noncompetitive Inhibitors (MK-801 and Cocaine) on a Protein (Nicotinic Acetylcholine Receptor)-Mediated Reaction," Biochemistry 42:6106-61 14 (2003), Cui et al, "Selection of 2'-Fluoro-modified RNA Aptamers for Alleviation of Cocaine and MK-801 Inhibition of the Nicotinic Acetylcholine Receptor," J. Membrane Biol. 202: 137-149 (2004), Sivaprakasam et al, "Minimal RNA Aptamer Sequences That Can Inhibit or Alleviate Noncompetitive Inhibition of the Muscle-Type Nicotinic Acetylcholine Receptor," J. Membrane Biol. 233 : 1-12 (2010), Ulrich et al, "In Vitro Selection of RNA Molecules that Displace Cocaine from the Membrane-Bound Nicotinic Acetylcholine Receptor," Proc. Nat. Acad. Sci. 95: 14051-14056 (1998), and Grewer et al, "On the Mechanism of Inhibition of the Nicotinic Acetylcholine Receptor by the Anticonvulsant MK-801 Investigated by Laser-Pulse Photolysis in the Microsecond-to-Millisecond Time Region," Biochemistry 38(24):7837-46 (1999), which are all hereby incorporated by reference in their entirety). [0037] Ligands that bind to the regulatory site of the nicotinic acetylcholine receptor comprise two different classes. Both classes modulate the opening and closing of the ion channel of the receptor to control flow of inorganic cations through the ion channel, especially the inflow that is normally stimulated by the natural neurotransmitter acetylcholine. Class 1 ligands are compounds that bind with higher affinity to the regulatory site on the closed-channel form than on the open-channel form of the receptor. Class 1 ligands facilitate closure and/or continued existing closure of the receptor ion channel, which inhibits neurotransmission and impairs cognition. Class 1 ligands include both endogenous and exogenous compounds. Prototypical exogenous Class 1 ligands include, without limitation, cocaine, MK-801, and phencyclidine.

[0038] Broadly, Class 2 ligands are compounds that bind to the regulatory site on the nicotinic acetylcholine receptor and shift the channel-opening equilibrium towards the open channel form of the receptor. For example, in the presence of an activating ligand such as acetylcholine or carbamoylcholine, and in the presence of a deleterious factor such as a Class 1 ligand, a mutation, etc. Class 2 ligands bind with equal or higher affinity to the regulatory site on the open-channel form of the receptor than to the closed- channel form. This binding shifts the channel-opening equilibrium to the open-channel state and alleviates the inhibition and impairment caused by a Class 1 compound, mutation, etc.

[0039] Laboratory work has demonstrated beneficial effects of multiple Class 2 compounds on cell function in vitro. Electrophysiological techniques have shown (i) an increase and then a decay in the current induced by cations moving in and out of a single cell when the cell is stimulated using carbamoylcholine (100 micromolar), a stable analog of the natural ligand acetylcholine; (ii) the decrease in amplitude and duration of prolongation of this current when the effect of the carbamoylcholine is attenuated by certain other compounds (designated as Class 1 compounds with prototypical examples cocaine and MK-801); and (iii) the restoration of the amplitude and duration of the increase in current when an alleviatory Class 2 compound is used to reverse the effect of the Class 1 compound (see Hess et al, "Reversing the Action of Noncompetitive Inhibitors (MK-801 and Cocaine) on a Protein (Nicotinic Acetylcholine Receptor- Mediated Reaction," Biochemistry 42:6106-61 14 (2003), which is hereby incorporated by reference in its entirety). [0040] Exemplary Class 2 ligands suitable for use in accordance with the methods of the present invention include, without limitation, tropane and its derivative, e.g. , ecgonine, ecgonine methyl ester, RTI-4229-70, RCS-III-143, RCS-III- 140A, RCS-III- 218, and RCS-III-202A, piperidine and its derivatives, derivatives of MK801 (but not MK-801), derivatives of phencyclidine (but not phencyclidine), and certain RNA aptamers all of which are described in more detail infra. These Class 2 ligands are the anti-anxiety ligands that are suitable for use in the methods of the present invention.

[0041] According to one embodiment of the present invention, a ligand that binds to nicotinic acetylcholine receptors and improves the condition of anxiety by reducing symptoms related to anxiety comprises an organic compound that is a derivative or analogue of tropane. The general chemical structure of the tropane derivatives are as follows:

where Ri, R2, R3, R4., R5, R«, and R7 are the same or different and are independently selected from the group consisting of hydrogen, hydroxyl, alkyl, cycloalkyl, alkenyl, alkoxy, aryl, alkylaryl, isoxazole, thiophene, indol, naphthalene, heterocyclic ring, halogen, and amine, as well as their esters and ethers, and Xi, X₂, and X3 are

independently selected from the group consisting of N, S, O, and C.

[0042] In addition, other ligands (i.e. Class 2 ligands) that bind to nicotinic acetylcholine receptors and improve anxiety states or treat or prevent conditions of anxiety, including, but not limited to, the following organic compounds: ecgonine;

ecgonine methyl ester; RTI-4229-70; RCS-III-143; RCS-III-140A; RCS-III-218;

RCS-III-202A; and analogues and/or derivatives of these compounds.

[0043] As referred to herein, the organic compound "ecgonine" has the following chemical structure: Ecgonine

[0044] As referred to herein, the organic compound "ecgonine methyl ester" has the following chemical structure:

Ecgonine Methyl Ester

[0045] As referred to herein, the organic compound "RTI-4229-70" has the following chemical structure:

[0046] As referred to herein, the organic compound "RCS-III-143" has the following chemical structure:

RCS-III-143

[0047] As referred to herein, the organic compound "RCS-III-140A" has the following chemical structure: RCS-III-140A

[0048] As referred to herein, the organic compound "RCS-III-218" has the following chemical structure:

RCS-III-218

[0049] As referred to herein, the organic compound "RCS-III-202A" has the following chemical structure:

RCS-III-202A

[0050] In another embodiment, a ligand that binds to nicotinic acetylcholine receptors and improves, treats, or prevents anxiety conditions and symptoms, in a subject, that is useful in carrying out the methods of the present invention includes one of more of the following cocaine analogs and derivatives:

where Ri, R2, R3, R4. R5, R«, R7 Rs. and R9 are the same or different and are

independently selected from the group consisting of hydrogen, hydroxyl, alkyl, cycloalkyl, alkenyl, alkoxy, aryl, alkylaryl, isoxazole, thiophene, indol, naphthalene, heterocyclic ring, halogen, and amine, as well as their esters and ethers, and Xi, X₂, and X₃ are independently selected from the group consisting of N, S, O, and C.

[0051] In another embodiment, a ligand that binds to nicotinic acetylcholine receptors and improves, treats, or prevents anxiety conditions and symptoms, in a subject, that is useful in carrying out the methods of the present invention includes one of more of the following analogs and derivatives of piperidine as follows:

Piperidine Derivatives

where Ri, R2, R3, R4., R5, and R^, are the same or different and are independently selected from the group consisting of hydrogen, hydroxyl, alkyl, cycloalkyl, alkenyl, alkoxy, aryl, alkylaryl, isoxazole, thiophene, indol, naphthalene, heterocyclic ring, halogen, and amine, as well as their esters and ethers, and Xi, X₂, and X3 are independently selected from the group consisting of N, S, O, and C.

[0052] In another embodiment, a ligand that binds to nicotinic acetylcholine receptors and improves, treats, or prevents anxiety conditions and symptoms, in a subject, in accordance with the present invention includes one or more of the following analogues and derivatives of MK-801, but is not dizocilpine. As referred to herein, the general chemical structures of these derivatives are as follows:

MK-801 Derivatives

where R, Ri, and R2, are the same or different and are independently selected from the group consisting of hydrogen, hydroxyl, alkyl, cycloalkyl, alkenyl, alkoxy, aryl, alkylaryl, isoxazole, thiophene, indol, naphthalene, heterocyclic ring, halogen, and amine, as well as their esters and ethers, and Xi, X₂, and X3 are independently selected from the group consisting of N, S, O, and C.

[0053] In another embodiment, a ligand that binds to nicotinic acetylcholine receptors and improves, treats, or prevents anxiety conditions and symptoms, in a subject, in accordance with the present invention includes one of more of the following analogs and derivatives of phencyclidine (PCP), but is not PCP. As referred to herein, the general chemical structures of these derivatives are as follows:

Phencyclidine Derivatives

[0054] In another aspect, the present invention relates to a method that improves, treats, or prevents anxiety conditions and symptoms, in a subject, that involves administering to a subject in need of improvement, treatment, and/or prevention of anxiety an aptamer that binds to nicotinic acetylcholine receptors and improves, treats, and prevents anxiety, in a subject.

[0055] RNA aptamers are preferred types of nucleic acid elements that have affinity for and can bind to a target molecule. Aptamers typically are generated and identified from a combinatorial library (typically in vitro) wherein a target molecule, generally, although not exclusively, a protein or nucleic acid is used to select from a combinatorial pool of molecules, generally although not exclusively oligonucleotides, those that are capable of binding to the target molecule. The selected reagents can be identified as primary aptamers. The term "aptamer" includes not only the primary aptamer in its original form, but also secondary aptamers derived from (i.e., created by minimizing and/or modifying the structure of) the primary aptamer. Aptamers, therefore, behave as ligands, binding to their target molecule.

[0056] Identifying primary aptamers basically involves selecting aptamers that bind a target molecule with sufficiently high affinity (e.g., = 20-50 nM) and specificity from a pool of nucleic acids containing a random region of varying or predetermined length (Shi et al, "A Specific RNA Hairpin Loop Structure Binds the RNA Recognition Motifs of the Drosophila SR Protein B52," Mol. Cell Biol. 17: 1649-1657 (1997), which is hereby incorporated by reference in its entirety).

[0057] Any method known in the art can be used to identify primary aptamers of any particular target molecule. In one embodiment (but not the only method) of the present invention the established in vitro selection and amplification scheme, SELEX, can be used. The SELEX scheme is described in detail in U.S. Patent No. 5,270,163 to Gold et al; Ellington and Szostak, "7w Vitro Selection of RNA Molecules that Bind Specific Ligands," Nature 346:818-822 (1990); and Tuerk and Gold, "Systematic Evolution of Ligands by Exponential Enrichment: RNA Ligands to Bacteriophage T4 DNA

Polymerase," Science 249:505-510 (1990), which are hereby incorporated by reference in their entirety.

[0058] In the case of RNA aptamers, where the structure and sequence of the

RNA has been established, the RNA molecule can either be prepared synthetically or a DNA construct or an engineered gene capable of encoding such an RNA molecule can be prepared.

[0059] Suitable examples of RNA aptamers that can be used in the methods of the present invention, include, but are not limited to, RNA aptamers that have the consensus sequences:

(a) SEQ ID NO: 1 (i.e., ACCG), SEQ ID NO: 2 (i.e., UCCG), SEQ ID NO: 3 (i.e., UUUACCG), SEQ ID NO: 4 (i.e., UUCACCG), and/or SEQ ID NO: 5 (i.e., UUCACCGUAAGG);

(b) SEQ ID NO:6 (i.e., AUCACCGUAAGG (see Aptamer B5)), SEQ ID NO:7 (i.e., UUUACCGUAAGG (see Aptamer B 15)), SEQ ID NO:8 (i.e.,

UUUUCCGUAAGG (see Aptamer B19)), SEQ ID NO:9 (i.e., UUUACCGUAAGG (see Aptamer B27)), SEQ ID NO: 10 (i.e., AUCACCGUAAGG (see Aptamer B28)), SEQ ID NO: 11 (i.e., UCCACCGUAGAU (see Aptamer B36)), SEQ ID NO: 12 (i.e.,

AUCACCGUAAGG (see Aptamer B44)), SEQ ID NO: 13 (i.e., UUUACCGUAAGG (see Aptamer B55)), SEQ ID NO: 14 (i.e., UCCACCGUAAGA (see Aptamer B59)), SEQ ID NO: 15 (i.e., UCCACCGUAAGA (see Aptamer B61)), SEQ ID NO: 16 (i.e.,

UUUACCGUAAGG (see Aptamer B64)), SEQ ID NO: 17 (i.e., UUUACCGUAAGG (see Aptamer B65)), SEQ ID NO: 18 (i.e., UUUACCGUAAGG (see Aptamer B69)), SEQ ID NO: 19 (i.e., UCCACCGUAAGA (see Aptamer B76)), SEQ ID NO:20 (i.e.,

UUUUCCGUAAGG (see Aptamer B78)), SEQ ID NO:21 (i.e., UCCACCGUAAGA (see Aptamer B 108)), SEQ ID NO:22 (i.e., UUUACCGUAAGG (see Aptamer Bl 11)), and/or SEQ ID NO:23 (i.e., AUCACCGUAAGG (see Aptamer B124));

(c) SEQ ID NO: 55 (i.e., GCUGAA);

(d) SEQ ID NO: 66 (i.e., GAAAG); and/or (e) SEQ ID NO: 88 (i.e., GUUAAU).

[0060] Suitable examples of RNA aptamers that can be used in the methods of the present invention, include, but are not limited to, RNA aptamers having nucleotide sequences:

(a) SEQ ID NO:24 (Aptamer B5), SEQ ID NO:25 (Aptamer B 15), SEQ ID NO:26 (Aptamer B 19), SEQ ID NO:27 (Aptamer B27), SEQ ID NO:28 (Aptamer B28), SEQ ID NO:29 (Aptamer B36), SEQ ID NO:30 (Aptamer B44), SEQ ID NO:31 (Aptamer B55), SEQ ID NO:32 (Aptamer B59), SEQ ID NO:33 (Aptamer B61), SEQ ID NO:34 (Aptamer B64), SEQ ID NO:35 (Aptamer B65), SEQ ID NO:36 (Aptamer B69), SEQ ID NO:37 (Aptamer B76), SEQ ID NO:38 (Aptamer B78), SEQ ID NO:39

(Aptamer B108), SEQ ID O:40 (Aptamer Bi l l), and/or SEQ ID O:41 (Aptamer B124);

(b) SEQ ID NO:42 (Aptamer 01), SEQ ID NO:43 (Aptamer 05), SEQ ID NO:44 (Aptamer 06), SEQ ID NO:45 (Aptamer 07), SEQ ID NO:46 (Aptamer 09), SEQ ID NO:47 (Aptamer 11), SEQ ID NO:48 (Aptamer 13), SEQ ID NO:49 (Aptamer 14), SEQ ID NO:50 (Aptamer 16), SEQ ID NO:51 (Aptamer 18), SEQ ID NO:52 (Aptamer 19), SEQ ID NO:53 (Aptamer 20,21), and/or SEQ ID NO:54 (Aptamer 22);

(c) SEQ ID NO:56 (Aptamer 3), SEQ ID NO:57 (Aptamer 8), SEQ ID NO:58 (Aptamer 23), SEQ ID NO:59 (Aptamer 24), SEQ ID NO:60 (Aptamer 26), SEQ ID NO:61 (Aptamer 30), SEQ ID NO: 62 (Aptamer 31), SEQ ID NO: 63 (Aptamer 38), SEQ ID NO:64 (Aptamer 39), and/or SEQ ID NO:65 (Aptamer 42); (d) SEQ ID NO:67 (Aptamer SI), SEQ ID NO:68 (Aptamer SI 3), SEQ ID

NO: 69 (Aptamer SI 4), SEQ ID NO: 70 (Aptamer S21), SEQ ID NO:71 (Aptamer S24), SEQ ID NO:72 (Aptamer S29), SEQ ID NO:73 (Aptamer S43), SEQ ID NO:74 (Aptamer S44), SEQ ID NO:75 (Aptamer S45), SEQ ID NO:76 (Aptamer S46), SEQ ID NO:77 (Aptamer S47), SEQ ID NO:78 (Aptamer S49), SEQ ID NO:79 (Aptamer S50), SEQ ID NO:80 (Aptamer S53), SEQ ID NO:81 (Aptamer S56), SEQ ID NO:82 (Aptamer S59), SEQ ID NO:83 (Aptamer S62), SEQ ID NO:84 (Aptamer S15), SEQ ID NO:85 (Aptamer S17), SEQ ID NO:86 (Aptamer S28), and/or SEQ ID NO:87 (Aptamer S54); and/or

(e) SEQ ID NO: 89 (Aptamer S5), SEQ ID NO: 90 (Aptamer SI 8), SEQ ID

NO: 91 (Aptamer S20), SEQ ID NO: 92 (Aptamer S25), SEQ ID NO: 93 (Aptamer S48), SEQ ID NO: 94 (Aptamer S51), and/or SEQ ID NO: 95 (Aptamer S57).

[0061] Also included within the invention are modified aptamers, having improved properties such as decreased size, enhanced stability, or enhanced binding affinity. Such modifications of aptamer sequences include adding, deleting or substituting nucleotide residues, and/or chemically modifying one or more residues. Methods for producing such modified aptamers are known in the art and described in, e.g., U.S. Patent Nos. 5,817,785 to Gold et al., and 5,958,691 to Wolfgang et al, which are hereby incorporated by reference in their entirety.

[0062] Chemically modified aptamers include those containing one or more modified bases. For example, modified pyrimidine bases may have substitutions of the general formula 5'-X and/or 2'-Y, and a modified purine bases may have modifications of the general formula 8'-X and/or 2'-Y. The group X includes the halogens I, Br, CI, or an azide or amino group. The group Y includes an amino group, fluorine, or a methoxy group. Other functional substitutions that would serve the same function may also be included. The aptamers of the present invention may have one or more X-modified bases, or one or more Y-modified bases, or a combination of X- and Y-modified bases. The present invention encompasses derivatives of these substituted pyrimidines and purines such as 5'-triphosphates, and 5'-dimethoxytrityl, 3'-beta-cyanoethyl, N,N-diisopropyl phosphoramidites with isobutyryl protected bases in the case of adenosine and guanosine, or acyl protection in the case of cytosine. Further included in the present invention are aptamers bearing nucleotide analogs, e.g., nucleotide analogs modified at the 5 and 2' positions, including 5-(3-aminoallyl)uridine triphosphate (5-AA-UTP), 5-(3-aminoallyl) deoxyuridine triphosphate (5-AA-dUTP), 5 -fluorescein- 12 -uridine triphosphate (5-F-12- UTP), 5 -digoxygenin- 11 -uridine triphosphate (5-Dig-l 1-UTP), 5-bromouridine triphosphate (5-Br-UTP), 2'-amino-uridine triphosphate (2'-NH2-UTP) and 2'-amino- cytidine triphosphate (2'- NH₂-CTP), 2'-fluoro-cytidine triphosphate (2'-F-CTP), and 2'- fluoro -uridine triphosphate (2'-F-UTP).

[0063] The aptamers may also be modified by capping at the 3' and 5' end. For example, the aptamer can be modified by adding to an end a polyethyleneglycol, amino acid, peptide, inverted dT, nucleic acid, nucleosides, myristoyl, lithocolic-oleyl, docosanyl, lauroyl, stearoyl, palmitoyl, oleoyl, linoleoyl, other lipids, steroids, cholesterol, caffeine, vitamins, pigments, fluorescent substances, toxin, enzymes, radioactive substance, biotin and the like. For such alterations, see for example, U.S. Patent Publication No. 2005/0096290 to Adamis et al. and U.S. Patent No. 5,660,985 to Wolfgang et al, which are hereby incorporated by reference in their entirety.

[0064] The sequences (consensus and RNA aptamer nucleotide sequences) referenced above by "SEQ ID NO." are identified herein below in Tables 1, 2, 3, 4, 5, and 6. TABLE 1 - Consensus Regions of Selected RNA Aptamers

RELATED CONSENSUS REGION SEQ ID NO: APTAMER

Consensus ACCG 1

Consensus UCCG 2

Consensus UUUACCG 3

Consensus UUCACCG 4

Consensus UUCACCGUAAGG 5

B5 AUCACCGUAAGG 6

B15 UUUACCGUAAGG 7

B19 UUUUCCGUAAGG 8

B27 UUUACCGUAAGG 9

B28 AUCACCGUAAGG 10

B36 UCCACCGUAGAU 11

B44 AUCACCGUAAGG 12

B55 UUUACCGUAAGG 13

B59 UCCACCGUAAGA 14

B61 UCCACCGUAAGA (B61 ) 15

B64 UUUACCGUAAGG (B64) 16

B65 UUUACCGUAAGG (B65) 17

B69 UUUACCGUAAGG (B69) 18

B76 UCCACCGUAAGA (B76) 19

B78 UUUUCCGUAAGG (B78) 20

B108 UCCACCGUAAGA (B108) 21

B1 1 1 UUUACCGUAAGG (B1 1 1 ) 22

B124 AUCACCGUAAGG (B124) 23

TABLE 3 ~ Sequences of Selected RNA Aptamers that Bind to the nACl iR

TABLE 4— Sequences of RNA Aptamers that Alleviate Inhibition of the nAChR.

Table 5— Sequences < )f 2'-Fluoropyrimidine-Modified RNA Aptar ners that Bind to nAChR

Table 6— Sequences of 2'-Fluoropyrimidine-Modified RNA Aptamers that Alleviate

Inhibition of the nAChR Rece tor

[0065] The nicotinic acetylcholine receptor ligands that improve, prevent, or treat anxiety and anxiety symptoms of the present invention can be administered orally, parenterally, for example, subcutaneous ly, intravenously, intramuscularly,

intracerebroventricularly, intraparenchymal (i.e., brain or brain stem), intravascularly, intraperitoneally, by intranasal inhalation, or by application to mucous membranes, such as, that of the nose, throat, and bronchial tubes. The ligands may be administered alone or with suitable pharmaceutical carriers, and can be in solid or liquid form such as, tablets, capsules, powders, solutions, suspensions, or emulsions.

[0066] The nicotinic acetylcholine receptor ligands that improve, prevent, or treat anxiety and anxiety symptoms of the present invention may be orally administered, for example, with an inert diluent, or with an assimilable edible carrier, or they may be enclosed in hard or soft shell capsules, or they may be compressed into tablets, or they may be incorporated directly with the food of the diet. For oral therapeutic

administration, the anxiety enhancing ligands, including compounds and aptamers of the present invention may be incorporated with excipients and used in the form of tablets, capsules, elixirs, suspensions, syrups, and the like. Such compositions and preparations should contain at least 0.1% of active compound. The percentage of anxiety-treatment enhancing ligand of the present invention in these compositions may, of course, be varied and may conveniently be between about 2% to about 60% of the weight of the unit. The concentration of nicotinic acetylcholine receptor ligand in such therapeutically useful composition is such that a suitable dosage will be obtained. Preferred compositions according to the present invention are prepared so that an oral dosage unit contains between about 1 and 250 mg of one or more cognition enhancing ligands of the present invention.

[0067] The tablets, capsules, and the like may also contain a binder such as gum tragacanth, acacia, corn starch, or gelatin; excipients such as dicalcium phosphate; a disintegrating agent such as corn starch, potato starch, alginic acid; a lubricant such as magnesium stearate; and a sweetening agent such as sucrose, lactose, or saccharin. When the dosage unit form is a capsule, it may contain, in addition to materials of the above type, a liquid carrier, such as a fatty oil.

[0068] Various other materials may be present as coatings or to modify the physical form of the dosage unit. For instance, tablets may be coated with shellac, sugar, or both. A syrup may contain, in addition to active ingredient, sucrose as a sweetening agent, methyl and propylparabens as preservatives, a dye, and flavoring such as cherry or orange flavor.

[0069] The nicotinic acetylcholine receptor ligands that improve, prevent, or treat anxiety and anxiety symptoms of the present invention may also be administered parenterally. Solutions or suspensions of these active compounds can be prepared in water suitably mixed with a surfactant, such as hydroxypropylcellulose. Dispersions can also be prepared in glycerol, liquid polyethylene glycols, and mixtures thereof in oils. Illustrative oils are those of petroleum, animal, vegetable, or synthetic origin, for example, peanut oil, soybean oil, or mineral oil. In general, water, saline, aqueous dextrose and related sugar solution, and glycols such as, propylene glycol or polyethylene glycol, are preferred liquid carriers, particularly for injectable solutions. Under ordinary conditions of storage and use, these preparations contain a preservative to prevent the growth of microorganisms.

[0070] The pharmaceutical forms suitable for injectable use include sterile aqueous solutions or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersions. In all cases, the form must be sterile and must be fluid to the extent that easy use in syringes exists. It must be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms, such as bacteria and fungi. The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (e.g., glycerol, propylene glycol, and liquid polyethylene glycol), suitable mixtures thereof, and vegetable oils.

[0071] The nicotinic acetylcholine receptor ligands that improve, prevent, or treat anxiety and/or anxiety symptoms of the present invention may also be administered directly to the airways in the form of an aerosol. For use as aerosols, the compounds of the present invention in solution or suspension may be packaged in a pressurized aerosol container together with suitable propellants, for example, hydrocarbon propellants like propane, butane, or isobutane with conventional adjuvants. The materials of the present invention also may be administered in a non-pressurized form such as in a nebulizer or atomizer. EXAMPLES

[0072] The following examples are provided to illustrate embodiments of the present invention, but they are by no means intended to limit its scope.

Example 1 - Ecgonine Methyl Ester (EME) is Readily Absorbed Following

Intraperitoneal Administration

[0073] Twenty young adult rats were administered intraperitoneal (IP) doses of 10 mg/kg ecgonine methyl ester (EME). The animals were sacrificed at various times after dosing to analyze brain and plasma concentrations of the administered compound. Brain and plasma samples from pre-dose (0-hour), and at 1, 2, 4, 8 and 24 hours after dosing were analyzed by GC-MS and the data are shown in Figure 1. Each data point represents the mean concentration level in four rats. Plasma concentrations had already peaked at one hour after dosing while brain concentrations were maximal at 2 hours. The maximum brain-to-plasma ratio was approximately 10. The data show rapid absorption after an IP dose, a biexponential decay of plasma concentrations, as if the drug confers on the body the characteristics of a two-compartment system, and sufficient persistence in the body to predict a half-life in humans of 6 - 8 hours. Example 2 - Ecgonine Methyl Ester Administration Reduces Anxiety in Treated

Animals

[0074] Thigmotaxis is a commonly used index or measure of anxiety in animals.

In Morris water maze tests with animals, thigmotaxis is noted by the amount of time a rat swims along the wall of a water tank, indicating anxiety and a 'desire' or expectation to find an exit along the edge. The longer the time spent along the edge, the more anxiety is present in the animal.

[0075] Thigmotaxis in rats can be studied as a function of age, disease, drug treatment etc., using the Morris water maze test. In each analysis a single rat is place in a circular water bath/swimming pool equipped with a visible or submerged platform, and the time and route taken to reach the platform for the rat is measured and recorded electronically. The internal surface of the wall is decorated with images of symbols of various types such that the rat can accumulate knowledge of the geometry of the pool and when the platform is found, how to subsequently reach the platform as expeditiously as possible. Once on the platform the rat tends to stay there, as this is a state preferable to swimming. Time to reach the platform, distance traveled, speed etc., are recorded for the observer electronically and a permanent image is produced showing the route taken. The whole exercise takes a few seconds in normal circumstances. The rats are trained over a period of time to a steady-state level of competence, and a probe test then measures how well the rats have learned and remembered the position of the platform, which can be visible or submerged in different versions of the test. Also, the rats can be studied during the training program, so that the investigator can gain insight into the speed of, for example, learning. Commonly, learning can be efficient, but then changing the position of the platform can present a whole new challenge, such that the investigator can discriminate between drug effects on the learning process itself and on the ability to perform better in the state created by the learning. Three examples are here presented to illustrate the phenomenon of thigmotaxis as affected by EME treatment, a Class 2 nicotinic acetylcholine receptor ligand.

[0076] Figure 2A shows a Morris water maze trace of a young control rat swimming in the bath for approximately 30 seconds searching for the platform. The trace indicates that the animal covered various quadrants of the pool quite easily, without evidence of anxiety, demonstrating moderately uniform attention to the different areas of the pool, not finding the platform until the last few seconds of the test. Figure 2B shows a Morris water maze trace of an aged and rather anxious rat swimming almost entirely at the perimeter of the water bath, finding the platform briefly at one stage but not staying there as if not certain in his anxious state that staying on the platform would be a good idea. Figure 2C shows a Morris water maze trace of the same rat shown in Figure 2B, following ten days of treatment with EME. Following treatment the rat learned how to find the platform, taking his cues from the wall markings, and lost his state of anxiety previously illustrated by the hugging of the walls phenomenon and the hesitancy to stay on the platform when he had found it.

[0077] In a subsequent study, 15 young and 45 aged rats were first assessed for general health/locomotion/swimming ability with no pharmacological intervention ("Pre- Training" with no drug on board). During this time, the rats were assessed multiple times in the Morris water maze with the platform visible and a flag marking its position. The next stage of the protocol was training of the rats with drug (or vehicle) treatment - this took five days ("Drug-on-Board Training"). This training was conducted with the platform visible and the flag removed. The data showed that the mean ability of each group at the start was the same. The next stage of the protocol was "Acquisition

Training". Drug/vehicle treatment continued without a break, with assessment on ten occasions over 12 days, referred to as Occasions 1 - 10. This was with the platform in a new position, present but submerged, therefore not visible, and with no flag. On days 15 - 19, the protocol for the first 5 days was repeated, with a changed platform position - platform again present but submerged. These stages of the investigation related to the need to incorporate adequate training, with and without drug on board, rewards for learning (finding the submerged platform), and a repeat study with new challenges. [0078] All groups exhibited time-dependent thigmotaxis, or the phenomenon of

"hugging the walls" of the water pool (Figures 3 and 4). This is considered to be associated with anxiety. However, the young rats overcame this anxiety very quickly. The aged controls showed marginal improvement without drug intervention, but EME was associated with a dose-dependent reversal of thigmotaxis. The data in Figure 3 shows mean results in the four groups of rats, including those treated with repeat doses of EME for an extended period of time. The groups were young controls, aged drug- free controls, aged treated with 3 mg/kg of EME, and aged treated with 10 mg/kg of EME. The young rats showed very little thigmotaxis, and recovered from what occurred quite quickly. The aged rats started from the same baseline as the young rats, and recorded the highest overall level of thigmotaxis of the four groups. The three mg/kg group showed a reduction in the thigmotaxis, and thigmotaxis was virtually eliminated in the 10 mg/kg group. The relationship between reversal of thigmotaxis and time is shown in Figure 4. Figure 4 shows the declining level of thigmotaxis in three of the four groups of rats in Figure 4, as a function of time. In this experiment, young rats lost thigmotaxis quickly without drug intervention. The compound EME accelerated the reversal of the problem in the aged rats, illustrating the effect of the drug on the anxiety state as studied by this technique.

[0079] A new challenge (changed platform position) led to reinstatement of impaired performance and a new searching and finding cycle, even with drug on board at the time of the change in platform position.

[0080] Because there were no a priori expectations of effect of the administered compound on thigmotaxis, the data analysis included pooling of all data in a search for trends that would indicate where more detailed testing should be directed. Accordingly, Figure 3 shows the dose-related effect. This data indicates a robust effect on this phenomenon. However, the data were also used to search for, in particular, time- dependent events.

[0081] The young rats showed little or no thigmotaxis, reaching maximum learned performance by the third day. The untreated aged rats learned more slowly, gradually showing improved performance over the 10 days (first trial) and the five days (second trial) reaching a maximum level of performance about one half that of the young rats. The drug-treated aged rats showed faster learning, with the performance of rats receiving 10 mg/kg EME coming close to that of the young controls by the tenth day. A repeat test showed the same results as Figure 4, particularly demonstrating the acute response to the new challenge and a new cycle of searching and learning.

[0082] The pooled data are displayed with standard errors of means (SEMs) calculated. Pooling of such large volumes of data is generally used to only obtain guidance concerning the appropriate testing strategy. Thus, each testing occasion (one rat on one day) involved 4 repeat assessments at 15-minute intervals. Analysis of variance showed no systematic trend in these clusters of data and it was concluded that there was no trend towards changed performance within the 45 minutes of these testing sessions - such a trend could have occurred through learning, tolerance, a pharmacokinetic effect, or something else. Therefore, the mean single day performances of each rat from these clusters of four assessments were assessed. In order to address the question of whether the four groups started from different baselines, non-linear regression analysis on the clusters of data for each treatment after each challenge was done. All four groups showed significant trends in the data over the sequence of days of dosing. There were however no significant differences in the intercepts of the regressions, indicating no differences in the baseline conditions. The rate constants of regression were however different between groups, with faster improvement by young control rats, and aged rats receiving 10 mg/kg doses. The next stage was analysis of variance on the data for the four groups of rats in the blocks of continuous treatment and assessment with the single platform position. There were highly significant differences between the groups, most notably between the control aged rats and the 10 mg/kg treated group. Since the baseline abilities were similar in all groups, and since the final condition would have been the same asymptote, it was not surprising to see that the most significant differences between the groups were at the intermediate times, days 3 - 7 (p<0.05), when the young rats had reached almost perfect scores, and the differences between aged and young controls were greatest.

[0083] The conclusion from this work is that EME caused reversal of thigmotaxis in the aged rats, and that this was associated with reversal of anxiety. These effects were seen as a response to a new challenge, and this was confirmed by repeat studies.

[0084] Although preferred embodiments have been depicted and described in detail herein, it will be apparent to those skilled in the relevant art that various modifications, additions, substitutions, and the like can be made without departing from the spirit of the invention and these are therefore considered to be within the scope of the invention as defined in the claims which follow.

Claims

WHAT IS CLAIMED

1. A method that improves, prevents, or treats anxiety and anxiety symptoms in a subject, said method comprising:

selecting a subject in need of improvement, treatment, and/or prevention of anxiety and/or anxiety symptoms, and

administering to the subject a ligand that binds to a regulatory site on a nicotinic acetylcholine receptor and improves, prevents, or treats anxiety and anxiety symptoms in the subject.

2. The method according to claim 1, wherein said ligand comprises tropane or a derivative thereof having one of the following structures:

wherein Ri, R2, R3, R4, R5, R«, and R7 are the same or different and are independently selected from the group consisting of hydrogen, hydroxyl, alkyl, cycloalkyl, alkenyl, alkoxy, aryl, alkylaryl, isoxazole, thiophene, indol, naphthalene, heterocyclic ring, halogen, and amine, as well as their esters and ethers, and Xi, X₂, and X3 are

independently selected from the group consisting of N, S, O, and C.

3. The method of claim 2, wherein said ligand is selected from the group consisting of ecgonine, ecgonine methyl ester, RTI-4229-70, RCS-III-143, RCS-III-140A, RCS-III-218, and RCS-III-202A.

4. The method according to claim 1 , wherein said ligand comprises a cocaine analogue selected from the group consisting of

wherein

Ri, R2, R₃, R4, R5, Re, R7, Rs, and R9 are the same or different and are independently selected from the group consisting of hydrogen, hydroxyl, alkyl, cycloalkyl, alkenyl, alkoxy, aryl, alkylaryl, isoxazole, thiophene, indol, naphthalene, heterocyclic ring, halogen, and amine, as well as their esters and ethers, and

X is independently selected from a group consisting of N, S, O, and C.

5. The method according to claim 1, wherein said ligand comprises piperidine or a derivative thereof havin the structure

wherein

Ri, R2, R₃, R4, R5, and 5 are the same or different and are independently selected from a group consisting of hydrogen, hydroxyl, alkyl, cycloalkyl, alkenyl, alkoxy, aryl, alkylaryl, isoxazole, thiophene, indol, naphthalene, heterocyclic ring, halogen, and amine, as well as their esters and ethers, and Xi and X2 are independently selected from a group consisting of N, S, O, and C.

6. The method according to claim 1, wherein said ligand comprises a structure selected from the group consisting of:

wherein R, Ri, and R2 are the same or different and are independently selected from the group consisting of hydrogen, hydroxyl, alkyl, cycloalkyl, alkenyl, alkoxy, aryl, alkylaryl, halogen, and amine, as well as their esters and ethers, and X is N or C.

7. The method according to claim 1, wherein said ligand comprises:

R3

(W erein

8. The method of claim 1, wherein said ligand which opens the ion channel

by binding to the nicotinic acetylcholine receptor comprises an RNA aptamer.

9. The method of claim 8, wherein the RNA aptamer comprises a consensus sequence selected from the group of nucleotide sequences consisting of SEQ ID NO: l, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, and SEQ ID NO:5.

10. The method of claim 8, wherein the RNA aptamer comprises a consensus sequence selected from the group of nucleotide sequences consisting of SEQ

ID NO:6, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO:18, SEQ ID NO: 19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, and SEQ ID NO:23.

11. The method according to claim 8, wherein said RNA aptamer comprises a nucleotide sequence selected from the group consisting of SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:27, SEQ ID NO:28, SEQ ID NO:29, SEQ ID NO:30, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34, SEQ ID NO:35, SEQ ID NO:36, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, and SEQ ID NO:41.

12. The method according to claim 8, wherein said RNA aptamer comprises a nucleotide sequence selected from the group consisting of SEQ ID NO:42, SEQ ID NO:43, SEQ ID NO:44, SEQ ID NO:45, SEQ ID NO:46, SEQ ID NO:47, SEQ ID NO:48, SEQ ID NO:49, SEQ ID NO:50, SEQ ID NO:51, SEQ ID NO:52, SEQ ID NO:53, and SEQ ID NO:54.

13. The method of claim 8, wherein said RNA aptamer comprises a consensus sequence comprising a nucleotide sequence of SEQ ID NO: 55.

14. The method according to claim 13, wherein said RNA aptamer comprises a nucleotide sequence selected from the group consisting of SEQ ID NO:56, SEQ ID NO:57, SEQ ID NO:58, SEQ ID NO:59, SEQ ID NO:60, SEQ ID N0:61, SEQ ID NO: 62, SEQ ID NO: 63, SEQ ID NO: 64, and SEQ ID NO: 65.

15. The method of claim 8, wherein said RNA aptamer comprises a consensus sequence comprising a nucleotide sequence of SEQ ID NO: 66.

16. The method of claim 15, wherein said RNA aptamer comprises a nucleotide sequence selected from the group consisting of SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:72, SEQ ID NO:73, SEQ ID NO:74, SEQ ID NO:75, SEQ ID NO:76, SEQ ID NO:77, SEQ ID NO:78, SEQ ID NO:79, SEQ ID NO:80, SEQ ID NO:81, SEQ ID NO:82, SEQ ID NO:83, SEQ ID NO:84, SEQ ID NO:85, SEQ ID NO:86, and SEQ ID NO:87.

17. The method of claim 8, wherein said RNA aptamer comprises a consensus sequence comprising a nucleotide sequence of SEQ ID NO: 88.

18. The method according to claim 17, wherein said RNA aptamer comprises a nucleotide sequence selected from the group consisting of SEQ ID NO: 89, SEQ ID NO: 90, SEQ ID NO: 91, SEQ ID NO: 92, SEQ ID NO: 93, SEQ ID NO: 94 and SEQ ID NO: 95.

19. The method of claim 8, wherein said RNA aptamer is chemically modified.

20. The method according to claim 19, wherein said chemically modified RNA aptamer comprises one or more modified nucleotides.

21. The method according to claim 1 , wherein said administering is carried out orally, parenterally, nasally, subcutaneously, intravenously, intramuscularly, intracerebroventricularly, intraparenchymal, intraperitoneally, by intranasal inhalation, by implantation, by intracavitary or intravesical instillation, intraocularly, intraarterially, intralesionally, trans dermally, or by application to mucous membranes.

22. The method according to claim 1, wherein the subject is any animal, preferably a mammal.

23. The method of claim 1 , wherein anxiety and any anxiety symptoms are improved in the selected subject.

24. The method of claim 1, wherein anxiety and any anxiety symptoms are prevented in the selected subject.

25. The method of claim 1, wherein anxiety and any anxiety symptoms are treated in the selected subject.