Guanfacine for fatigue disorders

Abstract

Disclosed herein are methods for treating a fatigue disorder in a subject in need thereof. The method may include administering to the subject by a therapeutically effective amount of a composition comprising an alpha-2 adrenergic agonist, such as guanfacine. Also disclosed herein are methods of diagnosing a fatigue disorder in a subject.

Description

Docket No.093386-9254-WO01 GUANFACINE FOR FATIGUE DISORDERS CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to U.S. Provisional Patent Application No.63 / 470,112, filed May 31, 2023, which is incorporated herein by reference in its entirety. FIELD

[0002] This disclosure relates to methods for treating a fatigue disorder in a subject in need thereof comprising administering to the subject a therapeutically effective amount of an alpha-2 adrenergic agonist such as guanfacine, or a pharmaceutically acceptable salt thereof. The disclosure also relates to methods of diagnosing a fatigue disorder in a subject. INTRODUCTION

[0003] Postural tachycardia syndrome (POTS) mostly affects otherwise healthy young women and is associated with a significantly diminished health-related quality of life, comparable to that of patients with chronic obstructive pulmonary disease or congestive heart failure. An exaggerated increase in standing heart rate (HR) is a hallmark of the disease. This orthostatic tachycardia is due to sympathetic activation, given that it is associated with a significant elevation in plasma norepinephrine and is blunted by even low doses of beta- blockers. However, the precise cause of this sympathetic activation is less clear. Some patients are thought to have partial peripheral denervation (neuropathic POTS), volume depletion, or deconditioning, triggering sympathetic activation as an appropriate compensatory reflex mechanism to maintain upright blood pressure (BP). In contrast, central sympathetic activation has been proposed as the primary pathophysiological mechanism in a subset of patients with “hyperadrenergic POTS” (HyperPOTS). It is proposed that this hyperadrenergic phenotype is reflected in high levels of upright plasma norepinephrine or microneurographic measurements of muscle sympathetic nerve activity (MSNA). Unfortunately, a clinical biomarker that identifies HyperPOTS has not been validated.

[0004] Further, there is a strong association between POTS, characterized by sympathetic overactivity on standing, and chronic fatigue syndrome (CFS). CFS is a debilitating and complex disorder characterized by profound fatigue that is not improved by bed rest and that may be worsened by physical or mental activity. Symptoms affect several body systems and may include weakness, muscle pain, impaired memory and / or mental concentration, andDocket No.093386-9254-WO01 insomnia, which can result in reduced participation in daily activities. It is important to note that fatigue and energy are distinct yet related symptoms. In particular, CFS is a disease of severe and debilitating fatigue that affects between 1-4 million Americans and for which there are no FDA-approved therapies or diagnostic tests. About 75% of CFS patients also have symptoms of autonomic dysfunction, suggesting a role of autonomic nervous system in the pathophysiology of CFS. In particular, increased sympathetic activity has been observed in at least 40% of CFS patients, and this hyperadrenergic phenotype was associated with more severe disease, greater autonomic symptoms, and the lowest quality of life.

[0005] Thus, there is a need for therapies for fatigue disorders, including the foregoing syndromes. SUMMARY

[0006] In an aspect, the disclosure relates to a method for treating a fatigue disorder in a subject, the method may comprise: administering a therapeutically effective amount of a composition comprising an alpha-2 adrenergic agonist, or a pharmaceutically acceptable salt thereof, to the subject. The subject may have at least one symptom selected from the group consisting of chronic fatigue, orthostatic hypertension, increased blood pressure, increased heart rate, and facial flushing. In an embodiment, the subject has a single nucleotide polymorphism in SLC6A2 gene. In another embodiment, the single nucleotide polymorphism is a mutation at Thr99Ile. In another embodiment, the mutation is a missense mutation. In another embodiment, the fatigue disorder is at least one selected from the group consisting of chronic fatigue syndrome (CFS), postural orthostatic tachycardia syndrome (POTS), and fibromyalgia. In another embodiment, the fatigue disorder is chronic fatigue syndrome (CFS) and / or postural orthostatic tachycardia syndrome (POTS). In another embodiment, the subject further having at least one symptom selected from the group consisting of reduced stamina, insomnia, muscle pain, muscle weakness, temperature dysregulation, dizziness, cognitive exhaustion, and physical exhaustion. In another embodiment, the alpha-2 adrenergic agonist comprises at least one selected from the group consisting of clonidine, p-aminoclonidine, tiamenidine, 5-bromo-6-(2-imidazoline-2-yl amino) quinoxaline, dexmedetomidine, detomidine, medetomidine, oxymetazonline, brimonidine, tizanidine, mivazerol, lofexidine, xylazine, guanabenz, guanfacine, guanclofine, guanoxabenz, alpha-methylnorepinephrine, azepexole, indoramin, pharmaceutically acceptable salts thereof, and 6-allyl-2-amino-5,6,7,8-tetrahydro4H- thiazolo [4,5-d]azepine diHCl. In another embodiment, the alpha-2 adrenergic agonistDocket No.093386-9254-WO01 comprises a compound of formula:, or a pharmaceutically acceptable salt thereof, wherein, at each occurrence, R1is independently –Cl or –CH3and R2is –OC1-4alkyl, –. In another embodiment, the alpha-2 adrenergic agonist comprises guanfacine or a pharmaceutically acceptable salt thereof. In another embodiment, the composition comprises about 0.1 mg to about 4 mg of the alpha-2 adrenergic agonist or a pharmaceutically acceptable salt thereof. In another embodiment, the composition comprises about 0.1 mg to about 4 mg guanfacine or a pharmaceutically acceptable salt thereof. In another embodiment, the composition is orally administered to the subject. In another embodiment, the composition is administered to the subject at least once daily. In another embodiment, the composition is in the form of a tablet.

[0007] In a further aspect, the disclosure relates to a method for treating chronic fatigue syndrome (CFS) and / or postural orthostatic tachycardia syndrome (POTS) in a subject, the method may comprise: administering a therapeutically effective amount of a composition comprising guanfacine, or a pharmaceutically acceptable salt thereof, to the subject, the subject having at least one selected from the group consisting of chronic fatigue, orthostatic hypertension, increased blood pressure, increased heart rate, and facial flushing. In an embodiment, the subject has a single nucleotide polymorphism in SLC6A2 gene. In another embodiment, the single nucleotide polymorphism is a mutation at Thr99Ile. In another embodiment, the mutation is a missense mutation. In another embodiment, the composition comprises about 0.1 mg to about 4 mg guanfacine or a pharmaceutically acceptable salt thereof. In another embodiment, the composition is orally administered to the subject. In another embodiment, the composition is administered to the subject at least once daily.

[0008] Another aspect of the disclosure provides a method of diagnosing a fatigue disorder in a subject, the method may comprise: (a) measuring a baseline blood pressure (BP) of the subject, wherein the subject is at rest and BP measurement includes diastolic BP (DBP) and systolic BP (SBP) measurements; (b) instructing the subject to perform a Valsalva maneuver for a period of time; and (c) measuring BP of the subject during late phase 2 of the Valsalva maneuver; wherein when the DBP increases by more than 18 mmHg from the baseline DBP, then the subject has the fatigue disorder; and wherein when the DBP increases by more than 18Docket No.093386-9254-WO01 mmHg from the baseline DBP, then the subject has an increased likelihood of being treated by a composition comprising an alpha-2 adrenergic agonist or a pharmaceutically acceptable salt thereof. In an embodiment, the fatigue disorder is hyperadrenergic postural orthostatic tachycardia syndrome (HyperPOTS) or chronic fatigue syndrome (CFS). In another embodiment, the alpha-2 adrenergic agonist is guanfacine or a pharmaceutically acceptable salt thereof. In another embodiment, the composition comprises about 0.1 mg to about 4 mg guanfacine or a pharmaceutically acceptable salt thereof. In another embodiment, the period of time is from about 10 seconds to about 30 seconds. In another embodiment, the late phase 2 of the Valsalva maneuver is from the first increase in systolic blood pressure (SBP) from baseline SBP after the nadir until phase 3. In another embodiment, the method herein further comprising (d) administering the composition comprising the alpha-2 adrenergic agonist or a pharmaceutically acceptable salt thereof to the subject having a fatigue disorder.

[0009] The disclosure provides for other aspects and embodiments that will be apparent in light of the following detailed description and accompanying figures. BRIEF DESCRIPTION OF THE DRAWINGS

[0010] FIGS.1A-D show correlations between supine resting muscle sympathetic nerve activity (MSNA), upright plasma norepinephrine (NE), and pressor response to phase 2 of the Valsalva maneuver in 28 patients with POTS (Protocol 1). FIG.1A is a graph showing the frequency distribution of supine resting MSNA. Seven patients with POTS (25%) had levels of resting MSNA above the 95% confidence interval limit ^24 bursts / min and were assigned to the “high_MSNA” group. FIG.1B is a graph showing that supine MSNA was positively correlated with upright NE (r2=0.38, p<0.001). FIG.1C is a graph showing that supine MSNA was positively correlated with ¨DBPVM2(r2=0.28, p=0.005). FIG.1D is a graph showing that upright NE had a trend towards a positive correlation with ¨DBPVM2but was not statistically significant (r2=0.14, p=0.057).

[0011] FIGS.2A-F show blood pressure and heart rate responses to head-up tilt tests and the Valsalva maneuver (VM) in the low and high_MSNA groups (Protocol 1). FIG.2A is a graph showing supine and upright heart rates (HR) were similar in both groups. FIG.2B is a graph showing that the high_MSNA group had higher upright systolic blood pressure (SBP) compared to the low_MSNA group. FIG.2C is a graph showing that the high_MSNA group had higher diastolic blood pressure (DBP) compared to the low_MSNA group. FIG.2D is a graph showingDocket No.093386-9254-WO01 HR responses to the VM were similar between groups. FIG.2E is a graph showing that the high_MSNA group had higher SBP during the late phase 2 of the VM (VM2L). FIG.2F is a graph showing that the high_MSNA group had higher DBP during VM2L. Values are mean±SEM. BSL, baseline; VM2e, early phase 2 of the VM; VM4, phase 4 of the VM, *p<0.05.

[0012] FIGS.3A-C show an increase in MSNA spike rate during phase 2 early of the Valsalva maneuver precedes the increase in DBP during phase 2 late in high_MSNA patients. FIG.3A are representative tracings of continuous blood pressure (BP, top) and spike rate of muscle sympathetic nerve activity (MSNA, bottom) during Valsalva maneuver in a high_MSNA patient (higher quartile for MSNA [^24 bursts / min]). FIG.3B is a graph showing the average data for diastolic BP (DBP) during the different phases of the Valsalva maneuver (Protocol 1). FIG.3C is a graph showing the average data for spike rate during the different phases of the Valsalva maneuver (Protocol 1). High_MSNA patients had higher DBP in late phase 2 of the Valsalva maneuver (VM2L) and that was preceded by higher spike rates during early phase 2 (VM2e) compared with low_MSNA patients (lowest quartile of MSNA). Bsl, baseline; VM4, phase 4 of the VM. Data are presented as means±SEM. *p<0.05.

[0013] FIGS.4A-B show that ¨DBPVM2is positively correlated with improvement of the PGIC score. FIG.4A is a graph showing the diastolic blood pressure response from early to late phase 2 of the Valsalva maneuver (¨DBPVM2) versus the patient’s perception of change in overall health with guanfacine (PGIC score 1= “very much improved”, 4= “no change”, and 7= “very much worse”). FIG.4B is a graph showing the proportion of non-HyperPOTS (Hyper –) and HyperPOTS (Hyper +) patients who reported an improvement in their overall health with guanfacine (Protocol 2), assessed by a PGIC score of ^3. The majority of HyperPOTS patients (89%, n=16) reported improved overall health versus 45% (n=9) of non-HyperPOTS patients (p<0.001), and the ¨DBPVM2was negatively correlated with the PGIC score (i.e. a larger ¨DBPVM2was associated with a greater perception of improvement in overall health; r= -0.352, p=0.030 by Spearman’s correlation).

[0014] FIGS.5A-D show patient-reported symptoms before (Pre) and during (Post) guanfacine treatment and their change from baseline in HyperPOTS (Hyper +) and non- HyperPOTS (Hyper –) patients (Protocol 2). FIG.5A is a graph showing that only the HyperPOTS group had a significant decrease in the severity of the spontaneous hyperadrenergic episodes (facial flushing, hypertension and tachycardia, 0= “no symptoms” and 4= “worst imaginable”). FIG.5B is a graph showing only the HyperPOTS group had aDocket No.093386-9254-WO01 significant decrease in the frequency of the spontaneous hyperadrenergic episodes (facial flushing, hypertension and tachycardia, 0= “no symptoms” and 4= “worst imaginable”). FIG.5C is a graph showing orthostatic intolerance, assessed by the OHDAS (0 = “no interference” and 10= “complete interference”). FIG.5D is a graph showing chronic fatigue, assessed by the PROMIS F-SF T-score. Values are mean±SEM. *p<0.05, **p<0.01 and *** p<0.001.\

[0015] FIG.6 is a graph showing autonomic symptom burden (COMPASS 31 total score) before (Pre) and during (Post) guanfacine treatment and their change from baseline in HyperPOTS (Hyper +) and non-HyperPOTS (Hyper –) patients (Protocol 2). Only the HyperPOTS group had a significant decrease in the autonomic symptom burden (the total COMPASS 31 score ranges from 0=normal to 100=the worst condition). Values are expressed as mean±SEM. *p<0.05, and **p<0.01.

[0016] FIG.7 is a graph showing the proportion of non-HyperPOTS (Hyper –) and HyperPOTS (Hyper +) patients who were on guanfacine treatment for 1-4 weeks and >1 month, including patients who stopped or continued taking the medication at the time of the survey (Protocol 2). The duration of treatment was similar between the Hyper- and non-HyperPOTS groups (p=0.947).

[0017] FIG.8 is a graph showing the number of concomitant medications for POTS taken before (Pre) and during (Post) guanfacine therapy and the change Post-Pre (¨) in / non- HyperPOTS (Hyper –) and HyperPOTS (Hyper +) patients (Protocol 2). No differences were observed in the number of concomitant POTS medications taken before (p=0.240) or during (p=0.197) guanfacine treatment or in the number of medications changed at the time guanfacine started (p=0.734).

[0018] FIGS.9A-B show hyperadrenergic patients (MSNA>23.4) had higher upright BP. FIG.9A is a graph showing systolic blood pressure (SBP) for at rest or standing. FIG.9B is a graph showing diastolic blood pressure (DBP) for at rest or standing. MSNA = Muscle sympathetic nerve activity; box = patients with high Q4 MSNA; circle = patients with low Q1 MSNA.

[0019] FIGS.10A-D show that there are similar heart rate (HR) responses to the Valsalva maneuver between groups, but BPs were higher during late Phase 2 in hyperadrenergic patients (MSNA>23.4). FIG.10A is a graph showing HR during the Valsalva maneuver phases. FIG.10B is a graph showing systolic blood pressure (SBP) during the Valsalva maneuverDocket No.093386-9254-WO01 phases. FIG.10C is a graph showing diastolic blood pressure (SBP) during the Valsalva maneuver phases. FIG.10D are representative traces of BP.

[0020] FIG.11 is a graph showing patient global impression of change (not specific for CFS) after empirical treatment with the central sympatholytic guanfacine. N=19 patients with ME / CFS and POTS (26±2 years old, female). Patients were asked to rate how their symptoms have changed while taking guanfacine. Patients reported here met the Fukuda criteria for CFS.

[0021] FIGS.12A-C show survey results from the Fukuda questionnaire. “Before” refers to before individuals began guanfacine treatment, and “during” refers to during guanfacine treatment. FIG.12A is a box and whisker plot showing change in hyperadrenergic symptoms frequency. FIG.12B is a box and whisker plot showing change in hyperadrenergic symptoms intensity. FIG.12C is a box and whisker plot showing change in orthostatic hypotension daily activity scale (OHDAS) improvement.

[0022] FIGS.13A-F show survey results from the Fukuda questionnaire. “Before” refers to before individuals began guanfacine treatment, and “during” refers to during guanfacine treatment. FIG.13A is a box and whisker plot showing change in fatigue Patient-Reported Outcomes Measurement Information System (PROMIS) t-score. FIG.13B is a box and whisker plot showing change in post-exertional malaise (PEM) mean raw score (during-after) frequency. FIG.13C is a box and whisker plot showing change in PEM mean raw score (during-after) severity. FIG.13D is a box and whisker plot showing change in fatigue PROMIS raw score baseline (BSL). FIG.13E are graphs showing the severity and frequency of non-refreshing sleep, memory, and hyperadrenergic symptoms in responders (grey bars) and non-responders (black bars). FIG.13F is a graph showing orthostatic tolerance of standing and walking for short or long periods of time in responders (grey bars) and non-responders (black bars).

[0023] FIGS.14A-D show predictors of response to guanfacine using the Valsalva maneuver. FIG.14A is a graph showing the change in SBP of responders and non-responders during the late phase 2 of the Valsalva maneuver. FIG.14B is a graph showing the change in DBP of responders and non-responders during the late phase 2 of the Valsalva maneuver. FIG. 14C is a plot showing whether DBP improved by 1 minute of head-up tilt (HUT) by PGIC binary. FIG.14D is a plot showing whether DBP improved by 3 minutes of head-up tilt (HUT) by PGIC binary.Docket No.093386-9254-WO01

[0024] FIGS.15A-C show summaries of survey results from the Fukuda and PEM questionnaires. FIG.15A is a graph showing responses from participants that were asked questions regarding their fatigue before and during guanfacine treatment. FIG.15B is a graph showing responses from participants that were asked questions regarding their symptoms before and during guanfacine treatment. FIG.15C is a graph showing responses from participants were asked the frequency and severity of their symptoms before and during guanfacine treatment. Results are graphed based on whether participants had a reduction in frequency of symptoms from before guanfacine to during guanfacine treatment.

[0025] FIG.16 is a graph showing a proposed need for a novel norepinephrine (NE) transporter therapeutic mechanism of action for chronic fatigue.

[0026] FIGS.17A-B show network plots of norepinephrine transporters (NETs). FIG.17A is a network plot showing NET in adult autonomic and cardiac phenotypes. FIG.17B is a network plot showing NET in pediatric cardiac phenotypes.

[0027] FIG.18 is a graph showing GRB0017-day analysis of NET activators in the high throughput screen (HTS) library of FDA-approved compounds. DETAILED DESCRIPTION

[0028] Described herein are methods for treating a fatigue disorder in a subject comprising orally administering a therapeutically effective amount of a composition comprising an alpha-2 adrenergic agonist, or a pharmaceutically acceptable salt thereof, to the subject, the subject having at least one symptom selected from the group consisting of chronic fatigue, orthostatic hypertension, increased blood pressure, increased heart rate, and facial flushing. Also described are methods of diagnosing a fatigue disorder in a subject. The method may comprise: (a) measuring a baseline blood pressure (BP) of the subject, wherein the subject is at rest and BP measurement includes diastolic BP (DBP) and systolic BP (SBP) measurements; (b) instructing the subject to perform a Valsalva maneuver for a period of time; and (c) measuring BP of the subject during late phase 2 of the Valsalva maneuver; wherein when the DBP increases by more than 18 mmHg from the baseline DBP, then the subject has the fatigue disorder; and wherein when the DBP increases by more than 18 mmHg from the baseline DBP, then the subject has an increased likelihood of being treated by a composition comprising an alpha-2 adrenergic agonist.Docket No.093386-9254-WO01 1. Definitions

[0029] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. The meaning and scope of the terms should be clear. In case of conflict, the present document, including definitions, take precedent over any dictionary or extrinsic definition. Further, unless otherwise required by context, singular terms shall include pluralities and plural terms shall include the singular. Preferred methods and materials are described below, although methods and materials similar or equivalent to those described herein can be used in practice or testing of the present invention. All publications, patent applications, patents, and other references mentioned herein are incorporated by reference in their entirety. The materials, methods, and examples disclosed herein are illustrative only and not intended to be limiting.

[0030] The terms “comprise(s),” “include(s),” “having,” “has,” “can,” “contain(s),” and variants thereof, as used herein, are intended to be open-ended transitional phrases, terms, or words that do not preclude the possibility of additional acts or structures. The singular forms “a,” “and,” and “the” include plural references unless the context clearly dictates otherwise. The present disclosure also contemplates other embodiments “comprising,” “consisting of,” and “consisting essentially of,” the embodiments or elements presented herein, whether explicitly set forth or not.

[0031] For the recitation of numeric ranges herein, each intervening number there between with the same degree of precision is explicitly contemplated. For example, for the range of 6-9, the numbers 7 and 8 are contemplated in addition to 6 and 9, and for the range 6.0-7.0, the number 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, and 7.0 are explicitly contemplated.

[0032] The term “about” or “approximately” as used herein as applied to one or more values of interest, refers to a value that is similar to a stated reference value, or within an acceptable error range for the particular value as determined by one of ordinary skill in the art, which will depend in part on how the value is measured or determined, such as the limitations of the measurement system. In certain aspects, the term “about” refers to a range of values that fall within 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less in either direction (greater than or less than) of the stated reference value unless otherwise stated or otherwise evident from the context (except where such number would exceed 100% of a possible value). Alternatively, “about” can mean within 3 or more thanDocket No.093386-9254-WO01 3 standard deviations, per the practice in the art. Alternatively, such as with respect to biological systems or processes, the term “about” can mean within an order of magnitude, preferably within 5-fold, and more preferably within 2-fold, of a value.

[0033] The terms “control,” “reference level,” and “reference” are used herein interchangeably. The reference level may be a predetermined value or range, which is employed as a benchmark against which to assess the measured result. “Control group” as used herein refers to a group of control subjects. The predetermined level may be a cutoff value from a control group. The predetermined level may be an average from a control group. Cutoff values (or predetermined cutoff values) may be determined by Adaptive Index Model (AIM) methodology. Cutoff values (or predetermined cutoff values) may be determined by a receiver operating curve (ROC) analysis from biological samples of the patient group. ROC analysis, as generally known in the biological arts, is a determination of the ability of a test to discriminate one condition from another, e.g., to determine the performance of each marker in identifying a patient having CRC. A description of ROC analysis is provided in P.J. Heagerty et al. (Biometrics 2000, 56, 337-44), the disclosure of which is hereby incorporated by reference in its entirety. Alternatively, cutoff values may be determined by a quartile analysis of biological samples of a patient group. For example, a cutoff value may be determined by selecting a value that corresponds to any value in the 25th-75th percentile range, preferably a value that corresponds to the 25th percentile, the 50th percentile or the 75th percentile, and more preferably the 75th percentile. Such statistical analyses may be performed using any method known in the art and can be implemented through any number of commercially available software packages (e.g., from Analyse-it Software Ltd., Leeds, UK; StataCorp LP, College Station, TX; SAS Institute Inc., Cary, NC.). The healthy or normal levels or ranges for a protein or measurement may be defined in accordance with standard practice. A control may be a subject or cell without an agonist as detailed herein. A control may be a subject, or a sample therefrom, whose disease state is known. The subject, or sample therefrom, may be healthy, diseased, diseased prior to treatment, diseased during treatment, or diseased after treatment, or a combination thereof.

[0034] As used herein “lower alkoxy” refers to a group —OR, wherein R is a lower alkyl as herein defined.

[0035] As used herein “lower alkylthio” refers to a group —SR, wherein R is a lower alkyl as herein defined.Docket No.093386-9254-WO01

[0036] As used herein, “lower alkyl” refers to an unbranched saturated hydrocarbon chain of 1-4 carbons, such as, methyl, ethyl, n-propyl, and n-butyl.

[0037] As used herein, the term “mammal” refers to any member of the class Mammalia including, without limitation, humans and nonhuman primates such as chimpanzees and other apes and monkey species; farm animals such as cattle, sheep, pigs, goats, llamas, camels, and horses; domestic mammals such as dogs and cats; laboratory animals including rodents such as mice, rats, rabbits, guinea pigs, and the like. The term does not denote a particular age or sex. Thus, adult and newborn subjects, as well as fetuses, whether male or female, are intended to be included within the scope of this term.

[0038] As used herein “pharmaceutically acceptable salt” refers to those salts which retain the biological effectiveness and properties of the free bases and which are not biologically or otherwise undesirable, formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid and the like, and organic acids such as acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, malic acid, malonic acid, succinic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, menthanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, and the like.

[0039] “Sample” or “test sample” as used herein can mean any sample in which the presence and / or level of a target is to be detected or determined or any sample comprising a DNA targeting or gene editing system or component thereof as detailed herein. Samples may include liquids, solutions, emulsions, or suspensions. Samples may include a medical sample. Samples may include any biological fluid or tissue, such as blood, whole blood, fractions of blood such as plasma and serum, muscle, interstitial fluid, sweat, saliva, urine, tears, synovial fluid, bone marrow, cerebrospinal fluid, nasal secretions, sputum, amniotic fluid, bronchoalveolar lavage fluid, gastric lavage, emesis, fecal matter, lung tissue, peripheral blood mononuclear cells, total white blood cells, lymph node cells, spleen cells, tonsil cells, cancer cells, tumor cells, bile, digestive fluid, skin, or combinations thereof. In some embodiments, the sample comprises an aliquot. In other embodiments, the sample comprises a biological fluid. Samples can be obtained by any means known in the art. The sample can be used directly as obtained from a patient or can be pre-treated, such as by filtration, distillation, extraction, concentration, centrifugation, inactivation of interfering components, addition of reagents, and the like, to modify the character of the sample in some manner as discussed herein or otherwise as is known in the art.Docket No.093386-9254-WO01

[0040] As used herein, the terms “subject” and “patient” may be used interchangeably to refer to any vertebrate including, but not limited to, a mammal that wants or is in need of the herein described compositions or methods. The subject may be a human or a non-human. The subject may be a vertebrate. The subject may be a mammal. The mammal may be a primate or a non-primate. The mammal can be a non-primate such as, for example, cow, pig, camel, llama, hedgehog, anteater, platypus, elephant, alpaca, horse, goat, rabbit, sheep, hamsters, guinea pig, cat, dog, rat, and mouse. The mammal can be a primate such as a human. The mammal can be a non-human primate such as, for example, monkey, cynomolgous monkey, rhesus monkey, chimpanzee, gorilla, orangutan, and gibbon. The subject may be of any age or stage of development, such as, for example, an adult, an adolescent, or an infant. The subject may be male. The subject may be female. The subject may be undergoing other forms of treatment.

[0041] As used herein, the terms “treat,” “treating” or “treatment” are each used interchangeably to describe reversing, alleviating, or inhibiting the progress of a disease and / or injury, or one or more symptoms of such disease, or completely eliminating a disease, to which such term applies. A treatment may be either performed in an acute or chronic way. The term also refers to reducing the severity of a disease or symptoms associated with such disease prior to affliction with the disease. Depending on the condition of the subject, the term may also refer to preventing a disease, and includes preventing the onset of a disease, or preventing the symptoms associated with a disease. Such prevention or reduction of the severity of a disease prior to affliction refers to administration of a composition or treatment described herein to a subject that is not at the time of administration afflicted with a disease and / or injury. “Preventing” also refers to preventing the recurrence of a disease and / or injury, or of one or more symptoms associated with such disease and / or injury. “Treatment” and “therapeutically,” refer to the act of treating, as “treating” is defined above. Suppressing the disease involves administering a composition of the present invention to a subject after induction of the disease but before its clinical appearance. Repressing or ameliorating the disease involves administering a composition of the present invention to a subject after clinical appearance of the disease. 2. Fatigue Disorders

[0042] Fatigue disorders may be chronic fatigue syndrome (CFS), postural orthostatic tachycardia syndrome (POTS; including hyperadrenergic POTS (HyperPOTS)), or fibromyalgia.Docket No.093386-9254-WO01 A subject with a chronic fatigue disorder may have at least one of the following symptoms: reduced stamina, insomnia, muscle pain, muscle weakness, temperature dysregulation, dizziness, cognitive exhaustion, and physical exhaustion. A subject may have at least one of the foregoing symptoms for at least 6 months.

[0043] A strong association between a particular single nucleotide polymorphism (SNP) in solute carrier family 6 member 2 (SLC6A2) (i.e., rs1805065 (Thr99lle)) and chronic fatigue syndrome was identified. Some patients may experience a ‘chronic fatigue syndrome’-like disease due to an excess of norepinephrine in the synapse and thus may benefit with a norepinephrine reuptake activator. SLC6A2, also known as norepinephrine transporter (NET), is an amine transporter. It terminates the action of noradrenaline by its high affinity sodium- dependent reuptake into presynaptic terminals. This gene encodes a member of the sodium:neurotransmitter symporter family. This member is a multi-pass membrane protein, which is responsible for the reuptake of norepinephrine homeostasis. Mutations in this gene have been associated with orthostatic intolerance, a syndrome characterized by lightheadedness, fatigue, altered mentation, and syncope, and is associated with postural tachycardia and plasma norepinephrine concentrations that are disproportionately high in relation to sympathetic outflow. An opportunity lies in the development of a compound that activates norepinephrine reuptake from the synapse. a. Chronic Fatigue Syndrome

[0044] Chronic fatigue syndrome (CFS), also known as myalgic encephalomyelitis / chronic fatigue syndrome (ME / CFS or CFS / ME or myalgic encephalopathy) or systemic exertion intolerance or systemic exertion intolerance disease (SEID), is a complex, long-term illness that can severely impact a person's ability to live their life. It is characterized by severe fatigue that does not improve with rest, as well as other symptoms that can make physical and mental exertion difficult including, but not limited to: sleep issues (e.g., unrefreshing sleep), memory or concentration problems, cognitive dysfunction, autonomic dysfunction, post-exertional malaise, dizziness such as dizziness that worsens with moving from lying down or sitting to standing, pain (e.g., muscle or joint pain), headaches, sore throats, and tender lymph nodes in the neck or armpits. Subjects with CFS also may become extra sensitive to light, sound, smells, food, and medicines. It is important to note that fatigue and energy are distinct yet related symptoms. The symptoms of CFS may be worsened by physical or mental activity. The illness can also beDocket No.093386-9254-WO01 unpredictable, where symptoms may come and go, or there may be changes in how severe symptoms are over time.

[0045] Autonomic nervous system dysfunction has been implicated in CFS. Studies have found that patients with CFS have higher levels of norepinephrine than that of controls. Patients with orthostatic intolerance or the associated postural tachycardia (both linked to a SLC6A2 mutation) also have similar dysfunction of the autonomic nervous system. Some CFS patients also experience orthostatic intolerance, postural tachycardia, or other associated symptoms (e.g., syncope, lightheadedness, etc.). b. Postural Tachycardia Syndrome

[0046] Postural Tachycardia Syndrome (POTS) is one of a group of conditions characterized by an orthostatic intolerance (OI). OI occurs when a patient has symptoms of low blood volume when they are standing, which are relieved when they lie down. POTS is distinguished from other OI syndromes by a rapid increase in the heartbeat of more than 30 beats per minute, or a heart rate that exceeds 120 beats per minute within 10 minutes of standing. POTS is characterized clinically by symptoms of light-headedness, palpitations, fatigue, dizziness, fainting, and exercise intolerance occurring with standing and relieved by lying down. Symptoms occur in association with an inappropriate rise in heart rate in the absence of a fall in blood pressure with the assumption of standing. Plasma norepinephrine (NE) is often elevated in patients with POTS, resulting in consideration of dysfunction of the norepinephrine transporter (NET) encoded by SLC6A2 gene. There are two main forms of POTS.

[0047] One form is partial dysautonomic where subjects appear to have mild damage to nerves that affect involuntary bodily function (peripheral autonomic neuropathy), such as the heartbeat. This damage affects the peripheral blood vessels and their ability to maintain normal blood pressure when there is more stress from gravity, such as when the patient stands up. This can cause the blood to pool in the lower areas of the body. The onset is abrupt and is autoimmune mediated.

[0048] The other form is hyperadrenergic (HyperPOTS), which is a less common type of POTS that appears more gradually and to have a genetic component. The hallmark of HyperPOTS is elevated levels of NE (greater than 600ௗpg / mL on standing), leading to overactivation of the sympathetic nervous system resulting in symptoms of tachycardia, brainDocket No.093386-9254-WO01 fog, and fatigue. Other symptoms include significant tremors, anxiety, cold and / or sweaty extremities, migraines, and increased urination. c. Fibromyalgia

[0049] Fibromyalgia is a condition that causes pain all over the body (i.e., widespread pain). Fibromyalgia also causes sleep problems, fatigue, and emotional and mental distress. People with fibromyalgia may be more sensitive to pain than people without fibromyalgia. This is called abnormal pain perception processing. Fibromyalgia affects about 4 million US adults, about 2% of the adult population. Women are twice as likely to have fibromyalgia as men. Risk factors for developing fibromyalgia include, but are not limited to, stressful or traumatic events, such as adverse childhood experiences, car accidents, or post-traumatic stress disorder (PTSD); repetitive injuries; injury from repetitive stress on a joint, such as frequent knee bending; illness (such as viral infections); genetics; and obesity. 3. Alpha-2 Adrenergic Receptor

[0050] The alpha-2 adrenoreceptor has important roles in signaling within and between both the peripheral and central nervous systems. Stimulation of the alpha-2 adrenoreceptor may suppress calcium entry into the nerve terminal. a. Alpha-2 Adrenergic Receptor Agonists

[0051] Alpha-adrenergic agonists are a class of sympathomimetic agents that selectively stimulates alpha adrenergic receptors. The alpha-adrenergic receptor has two subclassesand Į^. Alpha 2 receptors are associated with sympatholytic properties. Alpha-adrenergic agonists have the opposite function of alpha blockers.

[0052] Alpha-2 adrenergic agonists include, but are not limited to, clonidine, p- aminoclonidine, tiamenidine, 5-bromo-6-(2 imidazolidine-2-ylamino) quinoxaline, dexmedetomidine, detomidine, medetomidine, oxymetazonline, brimonidine, tizanidine, mivazerol (UCB-Pharma, Belgium), lofexidine, xylazine, guanabenz, guanfacine, guanclofine, guanoxabenz, or a derivative or structural analogue thereof, alpha-methylnorepinephrine, azepexole, indoramin, 6-allyl-2-amino-5,6,7,8-tetrahydro4H-thiazolo [4,5-d]azepine diHCl, methyldopa, moxonidine, pharmaceutically acceptable salts thereof and analogs thereof.Docket No.093386-9254-WO01

[0053] Further, alpha-2 adrenergic agonists may include substituted phenylacetyl derivatives of guanidine, O-alkylisoureas, S-alkylisothioureas, and P-nitrobenzylisothiourea as described in U.S. Patent No.3,632,645, which is incorporated herein by reference in its entirety. These compounds and pharmaceutically acceptable salts thereof may be useful in the methods of treatment described herein.

[0054] Also useful for the methods of treatment described herein are acetyl-guanidines, compounds of general Formula I, and pharmaceutically acceptable salts thereof. Formula I follows:in which R1signifies –Cl, –NH2, or –CH3, and their tautomeric compounds of general Formula Ia,in which R1has the above significance, and their acid addition salts.

[0055] In the following description for the sake of brevity, a reference to compounds of general Formula I is intended also to embrace the tautomeric compounds of Formula Ia. The compounds of general Formula I and their pharmaceutically acceptable salts may be obtained by a process characterized either by: (a) reacting a compound of general Formula II,in which R1has the above significance, or a reactive, functional acid derivative thereof, with guanidine, or (b) reacting with ammonia, a compound of general Formula III,Docket No.093386-9254-WO01in which R1has the above significance, and R2signifies a reactive radical capable of being split off together with one of the hydrogen atoms in ammonia, and when the resulting compound of Formula I is obtained in the form of a salt, optionally liberating the free base therefrom, and / or when the resulting compound of Formula I is obtained in the free base form, optionally salifying it.

[0056] In the process described, suitable reactive, functional acid derivatives of the compounds of Formula II include halides preferably chlorides or bromides, and lower alkyl esters. When a halide is used and it is desired to obtain the compound of Formula I in the form of a free base directly, the reaction may be affected in the presence of an acid binding agent, for example, triethyl-amine, or by using at least a 100% excess of guanidine. It will be appreciated that in the absence of such an acid binding agent or excess of guanidine, a salt of the compound of Formula I may be obtained, from which the free base may be liberated in manner known per se, for example, by treatment with dilute aqueous alkali metal hydroxide solutions.

[0057] The reaction described is preferably carried out in the presence of an organic solvent which is inert under the reaction conditions, for example, isopropanol or toluene. It may be carried out at room temperature (from about 20°C to about 25°C) and the reflux temperature of the reaction medium, preferably from about room temperature to about 80°C. The progress of reaction may be followed by, for example, thin layer chromatography.

[0058] In another process, suitable starting materials include, for example, those wherein the reactive radical is a lower alkoxy, lower alkylthio, or a substituted or unsubstituted aralkylthio radical, especially a methoxy, methylthio, or N-nitrobenzylthio radical. The reaction is preferably effected in an organic solvent which is inert under reaction conditions such as isopropanol. The ammonia reactant is suitably in the form of a concentrated aqueous solution and the reaction mixture may be stirred at a temperature from about 20°C to about 80°C for from about 6 hours to about 24 hours in order to complete the reaction.

[0059] The compounds of Formula I produced in accordance with the above processes may be isolated in manner known per se, for example, by extraction, precipitation or salt formation, and may be purified in manner known per se, for example, by recrystallization.Docket No.093386-9254-WO01

[0060] The compounds of general Formula I may be solid, basic compounds at room temperature. They may be converted into their acid addition salts by reacting with suitable organic or inorganic acids. Suitable organic acids for acid addition salts formation include, for example, acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, malic acid, malonic acid, succinic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, menthanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, and the like. Suitable inorganic acids for acid addition salts formation include hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and the like.

[0061] The compounds of general Formula II used as starting materials are known. The functional acid derivatives of compounds of general Formula II used in accordance with the invention are known or may be produced in manner known per se.

[0062] The compounds of general Formula III used as starting materials are known. The compounds of Formula III may be prepared by reacting a halide preferably a chloride or bromide, of a compound of Formula II with a compound of Formula IV, H2N–CR2=NH2(IV) in which R2has the above significance.

[0063] The process may be carried out in an organic solvent which is inert under the reaction conditions such as acetone, preferably at room temperature. Preferred reactants of Formula IV are compounds wherein the carbon atom carries a lower alkoxy, lower alkylthio, or a substituted or unsubstituted aralkylthio radical, especially a methoxy, methylthio, or p- nitrobenzylthio radical as substituent. When said substituent is a substituted or unsubstituted aralkylthio radical, an excess of the reactant is preferably used.

[0064] The resulting compounds of general Formula III may be isolated in manner known per se, for example, by precipitating with a precipitant, and may subsequently be purified in a manner known per se, for example, by recrystallization.

[0065] The compounds of general Formula IV used as starting materials for the above process are known or may be produced in manner known per se.

[0066] The compounds of general Formula I and their pharmaceutically acceptable acid addition salts are useful because they possess pharmacological activity in animals. They areDocket No.093386-9254-WO01 useful as antihypertensives having a beneficial effect particularly with regard to essential and renal hypertonia, as indicated by the "nictitating membrane" test in cats.

[0067] The antihypertensive effect is demonstrated by a subsiding of the contraction of the nictitating membrane in cats in the case of preganglionic and postganglionic stimulus of the neck sympathicus. In order to measure this effect the dose of active compound (ED50) is ascertained with which a 50% reduction of the nictitating membrane contraction occurs. TABLE 1 shows a comparison of the values ascertained for 2,6-disubstituted phenyl-acetylguanidines with those of guanethidine known as the most effective antihypertensive, whereby the superiority of 2,6-disubstituted phenyl-acetylguanidines is demonstrated. TABLE 1. Reduction of the neck sympathicus stimulus

[0068] For the above-mentioned use, the dosage administered will vary depending upon the compound employed, mode of administration, and treatment desired. However, in general, satisfactory results are obtained for the larger mammals at a total daily dosage in the range of from about 1 mg to about 10 mg, and dosage forms suitable for oral administration such as tablets, dragees, or for parenteral administration, such as injectable solutions, comprise from about 1 mg to about 2 mg of the compound administered with a solid or liquid pharmaceutical carrier or diluent.

[0069] Guanfacine is the oldest alpha-2 adrenergic agonist in clinical use in this class. It preferentially binds to Į-2A adrenergic receptor and has 15- to 20-fold greater affinity for Į-2A adrenergic receptor compared to Į-2B and Į-2C adrenergic receptor. Of the alpha-2 agonists, guanfacine has the lowest rate of sedation. Guanfacine is currently approved in tablet form for the treatment of hypertension (from about 1 mg to about 2 mg) and attention deficit hyperactivity disorder (ADHD; from about 1 mg to about 4 mg). Guanfacine has also been used off-label forDocket No.093386-9254-WO01 the treatment of Tourette’s syndrome, autism, and post-trauma stress disorder (PTSD), and has been proposed as a treatment option for withdrawal syndrome of opioids. Guanfacine has the chemical name N-amidino- 2-(2,6-dichlorophenyl) acetamide. Guanfacine is FDA approved in the HCl salt form.

[0070] Guanfacine is readily absorbed with around 80% oral bioavailability and has an elimination half-life of about 17 hours with renal excretion accounting for about 50%. Guanfacine has been shown to have similar in vitro potency as clonidine to inhibit conduction in rat sciatic nerve fibers. Intrathecal guanfacine has a similar antinociceptive effect as that of intrathecal clonidine in a goat model. Guanfacine showed longer antinociceptive effect in comparison to clonidine (8 hours vs.5 hours) due to slower elimination of guanfacine from spinal cord tissue. In a rat model, intrathecal guanfacine has shown anti-nociceptive effects for greater than 18 hours (at concentrations of both 25 mcg and 50 mcg) in comparison to clonidine injection (50 mcg) which declined to baseline within 4 hours. Additionally, a lower rate of elimination of guanfacine from spinal cord tissue compared to clonidine was reported.

[0071] Highly concentrated guanfacine (75 mcg in 15 μl saline) solution administered through an intrathecal route has been shown to be nontoxic in the subarachnoid space, establishing the safe use of guanfacine for perineural blockade. Guanfacine, as studied in animals and used in humans, has a safe risk profile.

[0072] A pharmaceutically acceptable salt of guanfacine useful in the methods described herein may be guanfacine hydrochloride (HCl) with the following structural formula (Formula V):

[0073] Guanfacine hydrochloride may be in powder form that is active pharmaceutical ingredient (API) grade. Guanfacine hydrochloride comes in an immediate release pill form (i.e., Tenex®) and an extended-release tablet (Intuniv®). Tenex® (guanfacine hydrochloride) with the chemical formula N-amidino-2-(2,6-dichlorophenyl) acetamide hydrochloride and molecular formula is C9H9Cl2N3O • HCl, and molecular weight is 282.55 may be used in the methods described herein. Tenex® is a centrally acting antihypertensive with Į2-adrenoceptor agonist properties in tablet form for oral administration and it is typically used to manage high bloodDocket No.093386-9254-WO01 pressure (hypertension). Generic forms of Tenex® may also be used in the methods described herein.

[0074] Intuniv® (guanfacine hydrochloride) with the chemical formula N-amidino-2-(2,6- dichlorophenyl) acetamide monohydrochloride and molecular formula is C9H9Cl2N3O • HCl, and the molecular weight is 282.55 may be used in the methods described herein. Intuniv® is a once-daily, extended-release matrix tablet formulation of Tenex®. Intuniv® is an alpha-2 adrenergic agonist typically used to treat attention deficit hyperactivity disorder (ADHD) in patients older than 6 years of age that is in tablet form for oral administration. Intuniv® is also sometimes used to treat hypertension. Generic forms of Intuniv® may also be used in the methods described herein. 4. Pharmaceutical Compositions

[0075] Further provided herein are compositions comprising the above-described compound(s). The disclosed compositions or compounds may be incorporated into pharmaceutical compositions suitable for administration to a subject (such as a patient, which may be a human or non-human). The pharmaceutical composition may be prepared for administration to a subject. The pharmaceutical compositions can be formulated according to the mode of administration to be used. Such pharmaceutical compositions can be administered in dosages and by techniques well known to those skilled in the medical and pharmaceutical arts taking into consideration such factors as the age, sex, weight, and condition of the particular subject, and the route of administration.

[0076] The pharmaceutical compositions may include pharmaceutically acceptable carriers. As used herein, the term “pharmaceutically acceptable carrier,” refers to a non-toxic, inert solid, semi-solid or liquid filler, diluent, encapsulating material, or formulation auxiliary of any type. Some examples of materials which can serve as pharmaceutically acceptable carriers are sugars such as, but not limited to, lactose, glucose, and sucrose; starches such as, but not limited to, corn starch and potato starch; cellulose and its derivatives such as, but not limited to, sodium carboxymethyl cellulose, ethyl cellulose, and cellulose acetate; powdered tragacanth; malt; gelatin; talc; excipients such as, but not limited to, cocoa butter and suppository waxes; oils such as, but not limited to, peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil, and soybean oil; glycols such as propylene glycol; esters such as, but not limited to, ethyl oleate and ethyl laurate; agar; buffering agents such as, but not limited to, magnesiumDocket No.093386-9254-WO01 hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline; Ringer's solution; ethyl alcohol; and phosphate buffer solutions, as well as other non-toxic compatible lubricants such as, but not limited to, sodium lauryl sulfate and magnesium stearate; as well as coloring agents, releasing agents, coating agents, sweetening agents, flavoring agents, and perfuming agents; preservatives and antioxidants can also be present in the composition, according to the judgment of the formulator. The route by which the composition is administered, and the form of the composition will dictate the type of carrier to be used.

[0077] In cases where pharmaceutical compositions are injectable pharmaceutical compositions, they may be sterile, pyrogen free, and particulate free. An isotonic formulation may be used. Generally, additives for isotonicity may include sodium chloride, dextrose, mannitol, sorbitol, and lactose. In some cases, isotonic solutions such as phosphate buffered saline may be used. Stabilizers include, but are not limited to, gelatin and albumin.

[0078] One aspect of the disclosure described herein encompasses an injectable pharmaceutical composition comprising alpha-2 adrenergic agonist such as guanfacine.

[0079] The pharmaceutical composition may comprise from about 0.1 mg to about 4.0 mg alpha-2 adrenergic agonist. The composition may comprise from about 0.5 mg to about 4.0 mg, about 1.0 mg to about 4.0 mg, about 1.1 mg to about 4.0 mg, about 1.3 mg to about 4.0 mg, about 1.5 mg to about 4.0 mg, about 1.7 mg to about 4.0 mg, about 1.9 mg to about 4.0 mg, about 2.1 mg to about 4.0 mg, about 2.3 mg to about 4.0 mg, about 2.5 mg to about 4.0 mg, about 2.7 mg to about 4.0 mg, about 2.9 mg to about 4.0 mg, about 3.1 mg to about 4.0 mg, about 3.3 mg to about 4.0 mg, about 3.5 mg to about 4.0 mg, about 3.7 mg to about 4.0 mg, about 3.9 mg to about 4.0 mg, about 0.1 mg to about 3.9 mg, about 0.1 mg to about 3.7 mg, about 0.1 mg to about 3.5 mg, about 0.1 mg to about 3.3 mg, about 0.1 mg to about 3.1 mg, about 0.1 mg to about 2.9 mg, about 0.1 mg to about 2.7 mg, about 0.1 mg to about 2.5 mg, about 0.1 mg to about 2.3 mg, about 0.1 mg to about 2.1 mg, about 0.1 mg to about 1.9 mg, about 0.1 mg to about 1.7 mg, about 0.1 mg to about 1.5 mg, about 0.1 mg to about 1.3 mg, about 0.1 mg to about 1.1 mg, about 0.1 mg to about 1.0 mg, or about 0.1 mg to about 0.5 mg alpha-2 adrenergic agonist. The composition may comprise at least about 0.1 mg, at least about 0.5 mg, at least about 1.0 mg, at least about 1.1 mg, at least about 1.3 mg, at least about 1.5 mg, at least about 1.7 mg, at least about 1.9 mg, at least about 2.1 mg, at least about 2.3 mg, at least about 2.5 mg, at least about 2.7 mg, at least about 2.9 mg, at least about 3.1 mg, at least about 3.3 mg, at least about 3.5 mg, at least about 3.7 mg, or at least about 3.9 mg alpha-Docket No.093386-9254-WO01 2 adrenergic agonist. The composition may comprise less than about 4.0 mg, less than about 3.9 mg, less than about 3.7 mg, less than about 3.5 mg, less than about 3.3 mg, less than about 3.1 mg, less than about 2.9 mg, less than about 2.7 mg, less than about 2.5 mg, less than about 2.3 mg, less than about 2.1 mg, less than about 1.9 mg, less than about 1.7 mg, less than about 1.5 mg, less than about 1.3 mg, less than about 1.1 mg, less than about 1.0 mg, or less than about 0.5 mg alpha-2 adrenergic agonist.

[0080] In some embodiments, the composition may comprise about 0.1 mg to about 4.0 mg guanfacine. The composition may comprise from about 0.4 mg to about 4.0 mg, about 0.8 mg to about 4.0 mg, about 1.2 mg to about 4.0 mg, about 1.4 mg to about 4.0 mg, about 1.6 mg to about 4.0 mg, about 1.8 mg to about 4.0 mg, about 2.0 mg to about 4.0 mg, about 2.2 mg to about 4.0 mg, about 2.4 mg to about 4.0 mg, about 2.6 mg to about 4.0 mg, about 2.8 mg to about 4.0 mg, about 3.0 mg to about 4.0 mg, about 3.2 mg to about 4.0 mg, about 3.4 mg to about 4.0 mg, about 3.6 mg to about 4.0 mg, about 3.8 mg to about 4.0 mg, about 0.1 mg to about 3.8 mg, about 0.1 mg to about 3.6 mg, about 0.1 mg to about 3.4 mg, about 0.1 mg to about 3.2 mg, about 0.1 mg to about 3.0 mg, about 0.1 mg to about 2.8 mg, about 0.1 mg to about 2.6 mg, about 0.1 mg to about 2.4 mg, about 0.1 mg to about 2.2 mg, about 0.1 mg to about 2.0 mg, about 0.1 mg to about 1.8 mg, about 0.1 mg to about 1.6 mg, about 0.1 mg to about 1.4 mg, about 0.1 mg to about 1.2 mg, about 0.1 mg to about 0.8 mg, or about 0.1about 0.4 mg guanfacine. The composition may comprise at least about 0.1 mg, at least about 0.4 mg, at least about 0.8 mg, at least about 1.0 mg, at least about 1.2 mg, at least about 1.4 mg, at least about 1.6 mg, at least about 1.8 mg, at least about 2.0 mg, at least about 2.2 mg, at least about 2.4 mg, at least about 2.6 mg, at least about 2.8 mg, at least about 3.0 mg, at least about 3.2 mg, at least about 3.4 mg, at least about 3.6 mg, or at least about 3.8 mg guanfacine. The composition may comprise less than about 4.0 mg, less than about 3.8 mg, less than about 3.6 mg, less than about 3.4 mg, less than about 3.2 mg, less than about 3.0 mg, less than about 2.8 mg, less than about 2.6 mg, less than about 2.4 mg, less than about 2.2 mg, less than about 2.0 mg, less than about 1.8 mg, less than about 1.6 mg, less than about 1.4 mg, less than about 1.2 mg, less than about 0.8 mg, or less than about 0.4 mg guanfacine. In some embodiments, the pharmaceutical compositions may comprise about 2.5 mg guanfacine.

[0081] The pharmaceutical composition may further comprise a solvent such as water. 5. AdministrationDocket No.093386-9254-WO01

[0082] The compositions or compounds disclosed herein can be administered therapeutically. In therapeutic applications, the composition or compound is administered to a subject in need thereof in an amount sufficient to elicit a therapeutic effect. An amount adequate to accomplish this is defined as “therapeutically effective dose.” Amounts effective for this use will depend on, for example, the particular composition of the conjugate regimen administered, the manner of administration, the stage and severity of the disease, the general state of health of the patient, and the judgment of the prescribing physician. For purposes of all of the methods described herein, the dose should be sufficient to affect a therapeutic response in the subject over a reasonable time frame.

[0083] The compositions or compounds disclosed herein can be administered by methods well known in the art as described in Donnelly et al. (Ann. Rev. Immunol.1997, 15, 617-648); Felgner et al. (U.S. Patent No.5,580,859, issued Dec.3, 1996); Felgner (U.S. Patent No. 5,703,055, issued Dec.30, 1997); and Carson et al. (U.S. Patent No.5,679,647, issued Oct.21, 1997). One skilled in the art would know that the choice of a pharmaceutically acceptable carrier, including a physiologically acceptable compound, depends, for example, on the route of administration.

[0084] The compositions or compounds disclosed herein may conveniently be presented in a single dose or as divided doses administered at appropriate intervals, for example, as two, three, four, or more sub-doses per day. The sub-dose itself may be further divided, for example, into a number of discrete loosely spaced administrations.

[0085] The determination of effective dosage levels, that is the dosage levels necessary to achieve the desired result, can be accomplished by one skilled in the art using routine methods, for example, human clinical trials, in vivo studies, and in vitro studies.

[0086] An attending physician would know how to and when to terminate, interrupt, or adjust administration due to toxicity or organ dysfunctions. Conversely, the attending physician would also know to adjust treatment to higher levels if the clinical response were not adequate (precluding toxicity). The magnitude of an administrated dose in the management of the disorder of interest may vary with the severity of the symptoms to be treated and the route of administration. Further, the dose, and perhaps dose frequency, may also vary according to the age, body weight, and response of the individual patient. A program comparable to that discussed above may be used in veterinary medicine.Docket No.093386-9254-WO01

[0087] A therapeutically effective amount of the compositions or compounds may be administered alone or in combination with a therapeutically effective amount of at least one additional therapeutic agent. In some embodiments, effective combination therapy is achieved with a single composition or pharmacological formulation that includes both agents, or with two distinct compositions or formulations, administered at the same time, wherein one composition includes a compound of described herein, and the other includes second agent(s). Alternatively, in other embodiments, the therapy may precede or follow the other agent treatment by intervals ranging from minutes to months.

[0088] The dose of the compositions or compounds may also be determined by the existence, nature, and extent of any adverse side effects that might accompany the administration of a particular agent. Typically, the attending physician will decide the dosage with which to treat each individual patient, taking into consideration a variety of factors, such as age, body weight, general health, diet, sex, therapeutic agent to be administered, route of administration, and the severity of the condition being treated.

[0089] For veterinary use, the compositions or compounds may be administered as a suitably acceptable formulation in accordance with normal veterinary practice. The veterinarian may readily determine the dosing regimen and route of administration that is most appropriate for a particular animal.

[0090] The compositions or compounds disclosed can be administered using standard administration techniques, including oral, intravenous, intraperitoneal, subcutaneous, pulmonary, transdermal, intramuscular, intranasal, buccal, sublingual, rectal, vaginal, or suppository administration. The compositions or compounds disclosed may be administered by injection. In some embodiments, the administration is oral administration. 6. Methods a. Methods of Treating a Fatigue Disorder

[0091] Provided herein are methods of treating a fatigue disorder in a subject in need thereof. The methods may include administering a therapeutically effective amount of a composition comprising an alpha-2 adrenergic agonist, or a pharmaceutically acceptable salt thereof, to the subject.Docket No.093386-9254-WO01

[0092] The method may comprise administering to the subject by oral administration a therapeutically effective amount of a composition comprising an alpha-2 adrenergic agonist described herein. The alpha-2 adrenergic agonist may comprise at least one of the group consisting of clonidine, p-aminoclonidine, tiamenidine, 5-bromo-6-(2-imidazoline-2-yl amino) quinoxaline, dexmedetomidine, detomidine, medetomidine, oxymetazonline, brimonidine, tizanidine, mivazerol (UCB-Pharma, Belgium), lofexidine, xylazine, guanabenz, guanfacine, guanclofine, guanoxabenz, alpha-methylnorepinephrine, azepexole, indoramin, 6-allyl-2-amino- 5,6,7,8-tetrahydro4H-thiazolo [4,5-d]azepine diHCl, methyldopa, moxonidine, pharmaceutically acceptable salts thereof, analogs thereof, and combinations thereof. In one embodiment, the alpha-2 adrenergic agonist may include guanfacine or a pharmaceutically acceptable salt thereof.

[0093] The subject may have at least one symptom selected from the group consisting of orthostatic hypertension, increased blood pressure, increased heart rate, and facial flushing. The subject may further have at least one symptom selected from the group consisting of reduced stamina, insomnia, muscle pain, muscle weakness, temperature dysregulation, dizziness, cognitive exhaustion, and physical exhaustion. The methods may improve or decrease the severity of any of the foregoing symptoms. The symptoms may be acute or chronic. Some acute symptoms can develop into chronic symptoms. In some embodiments, the methods may prevent the transition of acute symptoms from developing into chronic symptoms.

[0094] The subject may have a single nucleotide polymorphism in SLC6A2 gene. The single nucleotide polymorphism may be a mutation at Thr99Ile. The mutation may be a missense mutation.

[0095] The fatigue disorder may be at least one selected from the group consisting of chronic fatigue syndrome (CFS), postural orthostatic tachycardia syndrome (POTS), and fibromyalgia. In some embodiments, the fatigue disorder is CFS. In some embodiments, the fatigue disorder is POTS. In some embodiments, the fatigue disorder is hyperadrenergic POTS (HyperPOTS).

[0096] The composition comprising alpha-2 adrenergic agonist may be in the form of a tablet. The tablet may be an extended-release tablet. The composition comprising alpha-2 adrenergic agonist may be administered to the subject as described herein. In particular, theDocket No.093386-9254-WO01 composition comprising alpha-2 adrenergic agonist may be administered to the subject at least once daily. b. Methods of Diagnosing a Fatigue Disorder

[0097] Provided herein are methods of diagnosing a fatigue disorder in a subject. The methods may include: (a) measuring a baseline blood pressure (BP) of the subject, wherein the subject is at rest and BP measurement includes diastolic BP (DBP) and systolic BP (SBP) measurements; (b) instructing the subject to perform a Valsalva maneuver for a period of time; and (c) measuring BP of the subject during late phase 2 of the Valsalva maneuver. When the DBP increases by more than 18 mmHg from the baseline DBP, then the subject may have the fatigue disorder. In addition, when the DBP increases by more than 18 mmHg from the baseline DBP, then the subject may have an increased likelihood of being treated by a composition comprising an alpha-2 adrenergic agonist as described herein.

[0098] The fatigue disorder may be at least one selected from the group consisting of chronic fatigue syndrome (CFS), postural orthostatic tachycardia syndrome (POTS), and fibromyalgia. In some embodiments, the fatigue disorder is CFS. In some embodiments, the fatigue disorder may be POTS. In some embodiments, the fatigue disorder may be hyperadrenergic postural orthostatic tachycardia syndrome (HyperPOTS).

[0099] The alpha-2 adrenergic agonist may be administered to the subject after the subject has been diagnosed with a fatigue disorder. The alpha-2 adrenergic agonist may comprise at least one of the group consisting of clonidine, p-aminoclonidine, tiamenidine, 5-bromo-6-(2- imidazoline-2-yl amino) quinoxaline, dexmedetomidine, detomidine, medetomidine, oxymetazonline, brimonidine, tizanidine, mivazerol (UCB-Pharma, Belgium), lofexidine, xylazine, guanabenz, guanfacine, guanclofine, guanoxabenz, alpha-methylnorepinephrine, azepexole, indoramin, 6-allyl-2-amino-5,6,7,8-tetrahydro4H-thiazolo [4,5-d]azepine diHCl, methyldopa, and moxonidine, analogs thereof, pharmaceutically acceptable salts thereof, and combinations thereof. In one embodiment, the alpha-2 adrenergic agonist may include guanfacine. [000100] Blood pressure may be measured using standard clinical practices. For example, a blood pressure reading may be taken while a subject is seated with their feet flat on the floor and their arm resting at heart level; a blood pressure cuff that is attached to a pump may be applied around the top part of the subject’s arm where the bottom portion of the cuff is justDocket No.093386-9254-WO01 above the subject’s elbow; the blood pressure reading may be measured using a machine (e.g., automatic or digital) or manually (e.g., using a stethoscope and a gauge); and the blood pressure is recorded with the top number (systolic or SBP) being the pressure of the blood flow when the heart muscle squeezes (contracts) / pumping blood and the bottom number (diastolic or DBP) being the pressure measured between heartbeats. [000101] The period of time for performing the Valsalva maneuver may be from about 10 seconds to about 30 seconds. [000102] The late phase 2 of the Valsalva maneuver may be from the first increase in SBP from baseline SBP after the nadir until phase 3. 7. Examples [000103] The foregoing may be better understood by reference to the following examples, which are presented for purposes of illustration and are not intended to limit the scope of the invention. The present disclosure has multiple aspects and embodiments, illustrated by the appended non-limiting examples. Example 1 Materials and Methods [000104] Protocol 1: Autonomic Characterization of the Hyperadrenergic Phenotype in POTS. Twenty-eight patients with POTS (“Phenotyping Cohort”) were recruited from referrals to the Vanderbilt Autonomic Dysfunction Center (ADC). Patients had ^6 months of orthostatic symptoms accompanied by an increase in HR ^30 bpm after a 10-minute orthostatic challenge, either during head-up tilt or standing still (Raj et al., Can J Cardiol.2020; 36: 357–372; Sheldon et al., Heart Rhythm.2015; 12: e41–e63; Garland et al., Journal of the American College of Cardiology.2021; 77: 2174–2184; Freeman et al., Autonomic Neuroscience.2011; 161: 46–48). Patients were excluded if other medical conditions explained their illness, had orthostatic hypotension, were chair- or bedridden, or were unable to stop their POTS medications. The Vanderbilt University Institutional Review Board approved the study protocol, and written informed consent was obtained from each subject. [000105] Patients were evaluated at the Vanderbilt Clinical Research Center. Medications affecting the autonomic nervous system, BP, and HR were discontinued for ^5 half-lives.Docket No.093386-9254-WO01 Subjects had a history, physical examination, ECG, laboratory assessments, and autonomic function tests. The autonomic characterization was conducted on a separate day, >2 hours after meals, and included supine measurements of muscle sympathetic nerve activity (MSNA) at rest, hemodynamic responses to the Valsalva maneuver (VM) and 10 minutes of upright posture, and supine and upright plasma norepinephrine levels (as detailed below). [000106] The phase 2 of the VM was divided into 2 periods: an early phase 2 (VM2e), from the first decrease in systolic BP (SBP) after phase 1 until the SBP nadir, and a late phase 2 (VM2L), from the first increase in SBP after the nadir until phase 3. The DBP increase during phase 2 of the VM (¨DBPVM2), calculated as the difference in DBP between the end of VM2L and VM2e, was used as a marker of sympathetic-mediated vasoconstriction. [000107] Tests for the autonomic characterization were conducted on a separate day, 2 hours or more after a light meal, and included measurements of muscle sympathetic nerve activity (MSNA) at supine rest, hemodynamic responses to the Valsalva maneuver (VM) and 10 minutes of upright posture and supine and upright plasma norepinephrine levels. [000108] During the MSNA and VM measurements, ECG, HR, respiration, and brachial oscillometric BP were recorded using a Vital-Guard 450C monitor (Ivy Biomedical Systems, Inc., Branford, CT). Finger BP was estimated by the Penaz volume clamp method (BMEYE Nexfin Amsterdam, Netherlands). MSNA was recorded from the peroneal nerve of the left leg using the 662C-4 Nerve Traffic Analysis System (Bioengineering, University of IOWA). Recordings were performed while supine and during controlled VM with exhaled strength up to 40 mmHg for 15 seconds, as detailed elsewhere (Costa and Biaggioni. J Clin Invest.1994; 93: 1654–1660). Data collected during the VM were digitized and recorded using a WINDAQ data acquisition system (14 Bit, 5 kHz; DI720; DATAQ Inc, Acron OH) and processed offline. MSNA data were analyzed with custom software packages "Physiowave” written in PV-Wave language (PVWave; Visual Numerics, Boulder, CO) and “Physiowavelab” written in Matlab (MathWorks Inc.Natick, Massachusetts, NE). Bursts were detected with an automated detection algorithm with artifact elimination, dynamic noise level detection, and signal-to-noise estimation. Satisfactory recordings of MSNA were defined by (1) heart pulse synchronicity; (2) activation during VM straining and suppression during the hypertensive overshoot phase after VM release; (3) increases in response to breath-holding; and (4) no change during tactile or auditory stimulation (Delius et al., Acta Physiol Scand.1972; 84: 65–81; Diedrich et al., IEEE Trans Biomed Eng. 2003; 50: 41–50).Docket No.093386-9254-WO01 [000109] For the orthostatic tests, an indwelling catheter was placed in an antecubital vein at least 30 min before testing. BP and HR measurements were then obtained after at least 30 minutes of supine rest and at 1, 3, 5, and 10 minutes in the upright position. Blood samples for plasma norepinephrine were obtained at the end of the supine period and after 10 minutes of standing. Blood samples were collected in plastic syringes, immediately transferred to chilled vacuum tubes with sodium heparin (BD, Franklin Lakes, NJ), and placed on ice. Plasma was separated by centrifugation at –4°C and stored at –80°C in collection tubes with 6% GSH (Sigma) until the assay was performed. Concentrations of norepinephrine were measured by batch alumina extraction followed by high-performance liquid chromatography for separation with electrochemical detection and quantification (Goldstein et al., J Clin Invest.1988; 81: 213– 220). [000110] Protocol 2: Association Between the Hyperadrenergic POTS Phenotype and Clinical Response to Guanfacine. Adult patients were identified that were treated at the Vanderbilt ADC outpatient clinic with a diagnosis of POTS, who had had autonomic testing off medications and before receiving guanfacine and were prescribed guanfacine by an autonomic specialist for presumed hyperadrenergic POTS based on a history of spontaneous “hyperadrenergic episodes” (facial flushing, hypertension, and tachycardia), increased BP on standing, or an “exaggerated” pressor response to the phase 2 or 4 of the VM. Patients were excluded if they were prescribed guanfacine for medical conditions other than POTS or took guanfacine for <1 week. The Vanderbilt University Institutional Review Board approved the study, and written informed consent was obtained from each subject. [000111] Thirty-eight of these patients (“Treatment Cohort”) completed an online survey using REDCap electronic data capture that included demographic information, POTS-related medical history, symptoms before and after initiation of guanfacine, and the use and side effects of guanfacine. The survey included instruments (outlined in TABLE 2 and detailed in the below) to assess the impact of guanfacine treatment on patient-reported change in overall health (Patient Global Impression of Change, PGIC; Rampakakis et al., RMD Open.2015; 1: e000146); frequency and severity of spontaneous hyperadrenergic episodes (facial flushing, hypertension, and tachycardia); the impact of POTS symptoms on orthostatic tolerance (Orthostatic Hypotension Daily Activities Scale, OHDAS; Kaufmann et al., Clin Auton Res.2012; 22: 79–90); autonomic symptoms burden (COMPASS 31; Sletten et al., Mayo Clinic Proceedings.2012; 87: 1196–1201); and impact of chronic fatigue on daily activities (PROMIS Fatigue Short Form 7a (PROMIS F-SF; Yang et al., Qual Life Res.2019; 28: 3375–3384).Docket No.093386-9254-WO01 TABLE 2. Survey Instrument Used to Assess Clinical Impact of Guanfacine Treatment Test Range Score Patient Global Impression of Change 1-7 1, very much improved; 4, no change; (PGIC)a7, very much worse Frequency and Severity of 0-4 0, no symptoms; 4, worse imaginable hyperadrenergic symptomsbOrthostatic Hypotension Daily Activities 0-10 0, no interference; 10, complete Scale (OHDAS)cinterference COMPASS 31 autonomic symptom 0-100 0, normal; 100, worst burdendPROMIS F-SF (Fatigue Short Form)e0-100 T-score >50, greater fatigue than the US norm. a Patient-reported change in overall health b Patient rating of frequency and severity of spontaneous hyperadrenergic symptoms of facial flushing, hypertension and tachycardia. c Self-report of how much orthostatic symptoms interfered with patients’ daily upright activities (standing and walking for a short and long time) over the previous week. The score is calculated by averaging the scores of each item. Activities marked as zero or ‘cannot be done for other reasons’ at baseline were not scored. d Self-reported symptoms in 6 domains of autonomic function: orthostatic intolerance, vasomotor, secretomotor, gastrointestinal, bladder, and pupillomotor. The total score is calculated by adding the weighted scores of each domain. e Total scores are estimated on a T-score metric, where a T-score of 50 represents the mean score of the US general population matching the marginal distributions of gender, age, race, and education. [000112] In addition, the participant’s medical records were reviewed to collect data on past medical history relevant to POTS and the results of autonomic testing, and a 10-minute head-up tilt test at 75º, done off medications affecting BP or HR and before guanfacine treatment. [000113] Patient-reported change in overall health with guanfacine treatment was assessed using the Patient Global Impression of Change (PGIC; 7-point Likert scale where 1 is “very much improved”, 4 is “no change”, and 7 is “very much worse”; Rampakakis et al., RMD Open. 2015; 1: e000146). Patients rated the frequency and severity of their spontaneous events characterized by facial flushing, hypertension, and tachycardia before and during guanfacine treatment using a 5-point Likert scale where 0 is “no symptoms” and 4 is “worst imaginable”. [000114] Patients assessed the impact of POTS symptoms on orthostatic tolerance using the Daily Activities Scale (OHDAS), Part II of the Orthostatic Hypotension Questionnaire (OHQ; Kaufmann et al., Clin Auton Res.2012; 22: 79–90). This is a 4-item scale that allows patients to self-report how much their orthostatic symptoms interfered with daily upright activities (standing and walking for a short and long time) over the previous week. Each question is scored on anDocket No.093386-9254-WO01 11-point scale from 0 (“no interference”) to 10 (“complete interference”), and the option of selecting “cannot be done for other reasons”. The OHDAS score was calculated by averaging the scores of each item, and did not include activities marked as zero or ‘cannot be done for other reasons’ at baseline (Kaufmann et al., Clin Auton Res.2012; 22: 79–90). Thus, OHDAS scores of 28 patients (out of 38) were included in the analysis. Minimally important changes in the OHDAS score have been estimated as 0.71 to 0.89 points (Kaufmann et al., Clin Auton Res. 2012; 22: 79–90). [000115] Autonomic symptom burden before and during guanfacine treatment was assessed with the COMPASS-31 (Sletten et al., Mayo Clinic Proceedings.2012; 87: 1196–1201), which evaluates self-reported symptoms in 6 domains of autonomic function: orthostatic intolerance, vasomotor, secretomotor, gastrointestinal, bladder, and pupillomotor. The total score, calculated by adding the weighted scores of each domain, ranges from 0 (normal) to 100 (the worst condition; Sletten et al., Mayo Clinic Proceedings.2012; 87: 1196–1201). The minimally clinically important difference (MCID) for the COMPASS-31 total score has not been determined yet. However, a previous randomized clinical trial in POTS defined clinical improvement as a decrease from baseline in the COMPASS-31 total score of 15 points (Vernino et al., Clin Auton Res.2024; 34: 153–163). Fifteen patients had complete responses and were included in the analysis. [000116] Chronic fatigue and the interference of fatigue on daily activities over the previous week were assessed before and during guanfacine using the PROMIS Fatigue Short Form 7a (PROMIS F-SF; Yang et al., Qual Life Res.2019; 28: 3375–3384). This is a 7-item questionnaire scored on a 5-point Likert scale (1=”never” to 5=”always”). Total scores are estimated on a T-score metric (mean=50, SD= 10), where a T-score of 50 represents the mean score of the US general population matching the marginal distributions of gender, age, race, and education in the 2000 US Census (Goldstein et al., J Clin Invest.1988; 81: 213–220). The MCID for the PROMIS F-SF has been reported as a 3 to 7-point change in T-scores (Bartlett et al., Arthritis Care Res (Hoboken).2022; 74: 588–597). [000117] Statistical Methods. Frequency tables were generated for categorical variables. Continuous variables are expressed as mean±SEM. Normal distribution of data was assessed using the Kolmogorov–Smirnov test.Docket No.093386-9254-WO01 [000118] In Protocol 1, the relationships between MSNA, plasma norepinephrine, and ¨DBPVM2were evaluated using univariate regression analyses. MSNA recordings were then categorized into quartiles. The 1st and 4th quartile of the frequency distribution of MSNA in the Phenotyping Cohort were defined as the low and high_MSNA groups, respectively. The hemodynamic responses to the VM and orthostatic tests and supine and upright plasma norepinephrine levels were compared between these groups using Student’s t-tests if they had normal distribution. Otherwise, a Mann–Whitney U-test was used. Intraindividual comparisons were tested using a paired t-test or the Wilcoxon test, and 2-way repeated -measures ANOVA was used for multiple comparisons, followed by paired t tests with Bonferroni correction as post hoc test when overall significant differences were found. The ^2or Fisher’s exact tests were used to compare categorical data. Receiver operating characteristic (ROC) curves were calculated to differentiate the two MSNA groups based on the sensitivity and specificity of the ¨DBPVM2. The areas under the curve (AUC), SE, 95% CI, and cutoff values were calculated. The optimum cutoff values from the ROC curves were obtained by applying the Youden index. ROC curves, AUC and cutoff values of ¨DBPVM2were also estimated in an independent and complementary analysis of patients in the Treatment Cohort (Protocol 2) to validate ¨DBPVM2as a biomarker that differentiates HyperPOTS vs. non-HyperPOTS (defined as responders [PGIC score ^3] and non-responders [PGIC score ^4] to guanfacine, respectively). [000119] In Protocol 2, subjects in the Treatment Cohort were categorized into Hyper- and nonHyper POTS based on the ¨DBPVM2. Demographic and clinical data and survey responses were compared between the 2 groups using the same statistical approach described in Protocol 1. All tests were two-tailed, and a p<0.05 was considered significant. Analyses and graphing were performed with SPSS 28.0.1.1 (IBM Corp) and Prism 9 (GraphPad Software, LLC). Example 2 Correlations Between Supine Resting MSNA, Upright Plasma Norepinephrine, and Pressor Response to Phase 2 of the VM (Protocol 1, Phenotyping Cohort) [000120] Hyperadrenergic POTS (HyperPOTS) patients tend to have exaggerated pressor responses to the Valsalva maneuver (VM). The increase in intrathoracic pressure induced by the VM leads to a reduction in venous return, stroke volume, and an initial decrease in blood pressure (early Phase 2), triggering sympathetic activation in ways analogous to upright posture. Blood pressure recovers because of sympathetic-mediated vasoconstriction (lateDocket No.093386-9254-WO01 Phase 2). The inventors hypothesized that this pressor response, particularly in diastolic BP (DBP), would be exaggerated in HyperPOTS patients, would correlate with measurements of sympathetic activation such as upright plasma norepinephrine and resting MSNA, and would identify patients that improved to treatment with the central sympatholytic guanfacine. [000121] Patients with POTS in the Phenotyping Cohort (n=28, age 32±2 years, BMI 23.6±0.7 Kg / m2; 25 females; TABLE 3) had a wide range of sympathetic activity, measured directly by MSNA recordings while resting supine (18±3 bursts / min, range 1 to 54, 95% CI 13 and 23, FIG. 1A), and biochemically by supine plasma norepinephrine (244±36 pg / ml, range 89 to 716, 95% CI 170 to 318) and upright plasma norepinephrine (upright NE, 765±70 pg / ml, range 292 to 1650, 95% CI 619 to 909). Supine MSNA showed a positive correlation with upright NE (r2=0.38, p<0.001, FIG.1B) and a borderline significant correlation with supine norepinephrine levels (r2=0.14, p=0.054). When assessing the relationship between the hemodynamic responses to the VM and these markers of sympathetic activity, it was found that ¨DBPVM2(13±2 mm Hg, range 0 to 37, 95% CI 9 to 17) had a stronger and positive correlation with supine MSNA (r2=0.28, p=0.005, FIG.1C) than with upright NE (r2=0.14, p=0.057, FIG.1D). TABLE 3. Clinical Characteristics MeasurementProtocol 1Protocol 2 Age, yBody mass index, Kg / m223.6 ± 0.7 27.9 ± 1.4SupineHeart rate, bpm 73 ± 3 82±2 Systolic BP, mm Hg 109 ± 3 126±2 Diastolic BP, mm Hg 67 ± 2 78±1 UprightHeart rate, bpm 107 ± 4 119±3 Systolic BP, mm Hg 115 ± 5 129±3 Diastolic BP, mm Hg 73 ± 2 84±2 Orthostatic Change (standing-supine)Heart rate, bpm 34 ± 4 37±2 Systolic BP, mm Hg 7 ± 3 3±2 Diastolic BP, mm Hg 6 ± 2 7±1Data are presented as mean±SEM. Bpm, beats per min.Docket No.093386-9254-WO01 Example 3 Hemodynamic Responses to Orthostatic Challenge and Valsalva Maneuver in POTS Patients with Low and High Supine Resting MSNA (Protocol 1, Phenotyping Cohort) [000122] BP and HR responses to orthostatic challenge and the VM were compared between subjects in the 1st and 4th quartiles of MSNA (“low_MSNA” [^7 bursts / min] and “high_MSNA” [^24 bursts / min], respectively; n=7 in each group. TABLE 4 and FIGS.2A-F). Supine and upright HR were similar in both MSNA groups (FIG.2A and TABLE 4), with no difference in orthostatic tachycardia (low_MSNA: 34±8 vs. high_MSNA: 34±5 bpm; p=0.988). Supine BP was also similar in both groups (FIG.2B and FIG.2C and TABLE 4). However, the high_MSNA group had significantly higher upright BP (122±8 / 76±5 vs.99±5 / 62±3 mmHg in the low_MSNA group; p<0.05 for comparisons of SBP and DBP (FIG.2B and FIG.2C). Thus, BP increased on standing (i.e., orthostatic change) in the high_MSNA group and decreased in the low_MSNA group (5±5 / 6±3 vs. -5±3 / -1±2 mmHg in the low_MSNA group), but the differences in the orthostatic changes between groups did not reach statistical significance (p=0.117 and p=0.070 for comparisons of SBP and DBP). TABLE 4. Protocol 1: Clinical Characteristics MSNA Measurement Low MSNA High p (n=7) (n=7) value Age, y 26 ± 3 38 ± 3 0.022 Body mass index, Kg / m220.8 ± 0 24.8 ± 2 0.034 SupineHeart rate, bpm75±8 68±50.504Systolic blood pressure, mm Hg104±4 116±70.163Diastolic blood pressure, mm Hg63±2 70±50.200UprightHeart rate, bpm109±9 103±60.559Systolic blood pressure, mm Hg99±5 122±80.034Diastolic blood pressure, mm Hg62±3 76±50.027Orthostatic Change (standing-supine)Heart rate, bpm34±8 34±50.988Systolic blood pressure, mm Hg-5±3 5±50.117Diastolic blood pressure, mm Hg-1±2 6±30.070Valsalva ManeuverHeart rate, bpmDocket No.093386-9254-WO01 Baseline 66±3 75±4 0.118Early phase 2 97±5 94±6 0.629Late phase 2 104±3 104±4 0.979Phase 4 52±2 59±3 0.107Systolic blood pressure, mmHgBaseline 107±5 125±6 0.037Early phase 2 76±7 91±5 0.116Late phase 2 81±8 126±11 0.007Phase 4 139±5 160±8 0.054¨SBP late-early 5±2 34±7 0.002Diastolic blood pressure, mm HgBaseline 61±5 69±3 0.179Early phase 2 57±6 63±2 0.423Late phase 2 62±6 84±6 0.026Phase 4 78±5 87±30.189¨DBP late-early 5±2 21±4 0.002Data are presented as mean±SEM. Bpm, beats per min. [000123] HR responses to the VM were similar in both groups (FIG.2D and TABLE 4). The high_MSNA group, however, had greater SBP and DBP during late phase 2 compared to the low_MSNA group (126±11 / 84±6 vs.81±8 / 62±6 mmHg, respectively; p<0.05 for comparisons of SBP and DBP; FIG.2E and FIG.2F, TABLE 4). The increase in BP during late phase 2 in the high_MSNA group was preceded by higher MSNA spike rates during early phase 2 (81±9 vs. 44±6 spikes / second in the low_MSNA groups; p<0.01. FIGS.3A-C). Furthermore, the high_MSNA group had a significantly greater ¨DBPVM2(21±4 vs.5±2 in the low_MSNA group; p=0.002). Example 4 Determination of the Pressor Response to Phase 2 of the VM as a Biomarker of HyperPOTS (Phenotyping and Treatment Cohorts) [000124] In the Phenotyping Cohort (Protocol 1), ROC analysis was performed to determine if the ¨DBPVM2could discriminate between patients with low or high MSNA. It was found that a ¨DBPVM2 >18 mmHg was a conservative cutoff value that predicted high_MSNA patients with the highest specificity (100%) and 70% sensitivity (AUC 0.939 [0.807-1.000]; p=0.006.). ForDocket No.093386-9254-WO01 simplicity, only the analyses between the lowest and highest quartiles of MSNA are presented, but analyses including the middle quartiles did not change the conclusions. [000125] As an independent and complementary approach, a similar analysis was performed in the patients with POTS in the Treatment Cohort studied in Protocol 2 (n=38) to determine if ¨DBPVM2could predict responsiveness to guanfacine therapy (assessed by the PGIC score). It was found that ¨DBPVM2was positively correlated with improvement of the PGIC score (i.e., a larger ¨DBPVM2was associated with a greater perception of improvement in overall health; r= - 0.352, p=0.030. FIG.4A). The ROC analysis using PGIC also showed that a conservative cutoff of ¨DBPVM2>18 mmHg predicted a response to guanfacine with 77% specificity and 68% sensitivity (AUC 0.774 [0.627-0.920]; p=0.006). [000126] Thus, both supine MSNA and self-reported improvement to treatment with the central sympatholytic guanfacine (PGIC score) independently validated ¨DBPVM2as a clinical biomarker of the HyperPOTS phenotype. Then, a ¨DBPVM2>18 mmHg was used to explore the clinical characteristics of the HyperPOTS phenotype in the patients studied in Protocol 2. Example 5 Response to Guanfacine in POTS Patients with and without Hyperadrenergic Phenotype (Protocol 2, Treatment Cohort) [000127] Thirty-eight female patients with POTS in the Treatment Cohort (26±1 years, BMI 27.9±1.4 Kg / m2; TABLE 3) were included. Patients were divided into HyperPOTS or non- HyperPOTS based on the increase in ¨DBPVM2>18 mmHg. Both groups had similar age, BMI, and duration of POTS (estimated from diagnosis; TABLE 5 and TABLE 6); most patients (n=23, 61%) had a diagnosis of POTS for 1-5 years, and 10 (26%) for ^6 years. Supine and upright BP, HR, and orthostatic changes were similar in both groups (TABLE 6). The VM showed similar HR responses and VM ratios between patients with and without HyperPOTS. As expected, the HyperPOTS group had a greater BP response to late phase 2 of the VM compared to the non-HyperPOTS group (48±7 / 33±3 vs.19±3 / 10±1 mmHg; p<0.001 for both comparisons of SBP and DBP, TABLE 6). TABLE 5. Protocol 2: Clinical Characteristics Measurementnon-HyperPOTSHyperPOTS valueDocket No.093386-9254-WO01 Age, y 25 ± 2 27 ± 2 0.446 27. 2. Body mass index, Kg / m228.7 ± 2.0 1 ± 0 0.578 Diagnosis of POTS, n(%) a 0.887< 1 year 3 (15%) 2 (11%)1-5 years 11 (55%) 12 (67%)6-10 years 5 (25%) 3 (17%)>10 years 1 (5%) 1 (6%)Currently on Guanfacine, %(n) 0.914Yes 13 (65%) 12 (67%)No 7 (35%) 6 (33%)Reasons for Stopping, %(n)Not effective / Not needed 3 (15%) 3 (17%) 0.723 Side effects 3 (15%) 1 (6%) 0.344 Stopped by physician 3 (15%) 3 (17%) 0.888 Other 2 (10%) 1 (6%) 0.612 Data are presented as mean±SEM. aEstimated from the diagnosis of POTS. TABLE 6. Protocol 2: Autonomic Testing Before Guanfacine Treatment non- Measurement HyperPOTS HyperPOTSp valueSupine Heart rate, bpm81±3 82±20.882Systolic BP, mm Hg124±3 128±30.309Diastolic BP, mm Hg77±2 79±20.531Upright Heart rate, bpm118±4 120±50.723Systolic BP, mm Hg126±4 132±40.275Diastolic BP, mm Hg82±2 87±20.161Orthostatic Change (standing-supine) Heart rate, bpm36±3 38±40.769Systolic BP, mm Hg2±3 5±30.535Diastolic BP, mm Hg5±2 9±20.166Valsalva Maneuver Heart rate, bpm Baseline 84±3 88±2 0.321Early phase 2 121±4 125±4 0.537Docket No.093386-9254-WO01 Late phase 2 122±4 125±4 0.586Phase 4 61±3 62±2 0.794Systolic BP, mmHg Baseline 140±5 140±4 0.929Early phase 2 106±7 98±7 0.378Late phase 2 126±6 146±6 0.034Phase 4 175±7 182±7 0.512¨SBP late-earlya19±3 48±7 <0.001Diastolic BP, mmHg Baseline 77±5 79±4 0.794Early phase 2 73±6 67±5 0.496Late phase 2 83±6 100±5 0.044Phase 4 91±5 96±5 0.423¨DBP late-earlya10±1 33±3 <0.001Valsalva ratio 2.11±0.10 2.05±0.10 0.718Data are presented as mean±SEM. Bpm, beats per min. aBP recovery from early to late phase II of the Valsalva maneuver. [000128] Before guanfacine treatment, all patients reported spontaneous hyperadrenergic episodes that felt like “adrenaline rushes” associated with facial flushing, an increase in BP and HR, and feeling extreme and prolonged tiredness after an episode. These episodes resemble those seen in patients with mast cell activation disorders (MCAD). Indeed, the association of MCAD and POTS in patients with a hyperadrenergic clinical phenotype responsive to central sympatholytics was previously reported. In the present study, however, there was no difference in the proportion of Hyper and non-HyperPOTS patients with a possible diagnosis of MCAD (6 [33%] vs.7 [35%] patients, respectively; p=0.914), but this diagnosis was not systematically diagnosed or excluded. [000129] Hyper- and non-HyperPOTS patients had a similar pre-treatment severity (3.5±0.2 vs.3.2±0.2 a.u.; p=0.282. FIG.5A) and frequency (3.4±0.1 vs.3.2±0.2 a.u.; p=0.467. FIG.5B) of hyperadrenergic episodes. Similarly, Hyper- and non-HyperPOTS patients did not differ in the impact of their POTS symptoms on daily upright activities (OHDAS mean score 6.9±0.6 vs. 6.1±0.5; p= 0.296. FIG.5C), chronic fatigue (PROMIS F-SF [T-score] 71.4±1.8 vs.69.3±1.8; p= 0.476. FIG.5D) or the burden of autonomic symptoms (COMPASS-31 total score 56.7±4.7 vs. 45.6±3.7; p=0.142. FIG.6 and TABLE 7) before guanfacine treatment.Docket No.093386-9254-WO01 TABLE 7. Protocol 2: COMPASS-31 Scores Before and During Guanfacine Treatment COMPASS-31 Domainsnon-HyperPOTS HyperPOTS P-Orthostatic intoleranceBaseline 30.7 ± 3.0 34.0 ± 2.1 0.414 Post-treatment 21.3 ± 4.2 13.5 ± 2.9 0.228 ¨ (post-baseline) -9.3 ± 6.3 -20.5 ± 2.9 0.142 Vasomotor functionBaseline 3.5 ± 0.3 3.0 ± 0.6 0.491 Post-treatment 3.2 ± 0.1 2.1 ± 0.4 0.013 ¨ (post-baseline) -0.3 ± 0.3 -0.9 ± 0.3 0.228 Secretomotor functionBaseline 1.1 ± 0.5 5.1 ± 1.6 0.081 Post-treatment 3.6 ± 1.7 5.1 ± 1.5 0.573 ¨ (post-baseline) 2.5 ± 1.6 0.0 ± 1.3 0.181 Gastrointestinal functionBaseline 6.8 ± 1.7 10.8 ± 1.7 0.108 Post-treatment 9.2 ± 1.6 7.6 ± 1.7 0.662 ¨ (post-baseline) 2.4 ± 2.5 -3.2 ± 1.5 0.059 Bladder function Baseline 1.9 ± 1.6 1.4 ± 0.5 0.491 Post-treatment 2.0 ± 1.6 0.7 ± 0.3 0.95 ¨ (post-baseline) 0.2 ± 0.2 -0.7 ± 0.6 0.282 Pupillomotor functionBaseline 1.7 ± 0.4 2.4 ± 0.4 0.491 Post-treatment 1.8 ± 0.5 1.7 ± 0.3 1 ¨ (post-baseline) 0.1 ± 0.2 -0.7 ± 0.3 0.081 Total COMPASS 31Baseline 45.6 ± 3.7 56.7 ± 4.7 0.142 Post-treatment 41.2 ± 6.8 30.7 ± 3.3 0.228 ¨ (post-baseline) -4.4 ± 7.1 -26.1 ± 4.5 0.029 Data are presented as mean±SEM. [000130] At the time of the survey, a similar proportion of Hyper- and non-HyperPOTS patients were still taking guanfacine (12 [67%] vs.13 [65%]; p=0.914). Of all participants (n=38), 28 (74%) took guanfacine for >1 month and 10 (26%) between 1-4 weeks. There were no differences in the dose of guanfacine between Hyper- and non-HyperPOTS patients (median 1 mg / d, range 0.5-2 mg in both groups), the duration of guanfacine treatment (FIG.7) or in the reasons for stopping treatment (TABLE 5). The majority of patients were taking otherDocket No.093386-9254-WO01 medications for POTS during guanfacine treatment. However, there were no differences in the number of concomitant POTS medications between Hyper- and non-HyperPOTS patients (median 1, range 0-3 in both groups; FIG.8) or the number of concomitant POTS medications added or stopped during guanfacine treatment (median 0, range -2 to 1 in both groups). Beta- blockers were the most common concomitant medication for POTS and were taken by a similar proportion of patients in both groups (9 [50%] HyperPOTS and 8 [40%] non-HyperPOTS patients; p=0.536). [000131] During guanfacine treatment, a greater percentage of HyperPOTS patients reported an improvement in their overall health (PGIC score ^ 3) (16 [89%]) than non-HyperPOTS (9 [45%]; p<0.001, FIG.4B). Only the HyperPOTS group had significant improvements in hyperadrenergic symptoms severity (-1.4± 0.2 vs. -0.7±0.2 in non-HyperPOTS; p= 0.019. FIG. 5A) and frequency (-1.5± 0.2 vs. -0.7±0.2 in non-HyperPOTS; p= 0.014. FIG.5B), in the impact of POTS symptoms on daily upright activities (¨OHDAS mean score -2.0± 0.9 vs.0.1±0.5 in non-HyperPOTS; p=0.021. FIG.5C), chronic fatigue (PROMIS F-SF [T-score] -13.2±2.9 vs. - 3.1±2.1; p= 0.006. FIG.5D), and burden of autonomic symptoms (COMPASS-31 total score - 26.1±4.5 vs. -4.4±7.1; p=0.029. FIG.6 and TABLE 7). This improvement in symptoms in HyperPOTS patients was not only statistically significant but also clinically meaningful given that these changes were larger than the minimal important differences previously reported. Example 6 Discussion [000132] Taken together, the results herein are consistent with the concept that POTS is caused by a primary central sympathetic activation in a subset of patients, which can be identified clinically by an exaggerated DBP increase during phase 2 of the Valsalva maneuver and improved by central sympatholytic therapy. These results support the need for clinical trials to determine the safety and efficacy of guanfacine in patients with POTS enriched for the presence of this clinical biomarker. [000133] POTS is recognized as a syndrome caused by diverse underlying pathophysiologies that have in common orthostatic intolerance associated with tachycardia. The inventors have previously reported congenital causes of POTS related to NET deficiency, but these are rare. More commonly proposed causes include partial peripheral neuropathy (“neuropathic POTS”), a deficit in blood volume, abnormalities in the renin-angiotensin-aldosterone system (RAAS),Docket No.093386-9254-WO01 physical deconditioning, autoantibodies targeting peripheral adrenergic receptors, and a hyperadrenergic state resulting from central activation of the sympathetic nervous system (“HyperPOTS”). It has been estimated that this “hyperadrenergic” phenotype is present in 30- 60% of patients with POTS. [000134] The heterogeneous nature of POTS represents a challenge not only in understanding the disease but also in developing an effective treatment. In particular, therapies aimed at targeting sympathetic activation, e.g., central sympatholytics, would be expected to worsen patients in whom the orthostatic tachycardia is an adequate compensatory mechanism to maintain upright BP, e.g., due to volume depletion. Finding a clinical biomarker that identifies HyperPOTS would be essential for precision medicine. There is, however, no agreement about the criteria used to identify them. [000135] An upright plasma NE ^600 pg / mL is the criterion most often used to define hyperadrenergic POTS, and this threshold has been adapted in recent expert consensus statements. It is a useful biomarker in that it identifies patients who respond better to beta blockers. The inventors were among the first to introduce this threshold in the literature, but cautioned that a high plasma norepinephrine could be due not only to a primary hyperadrenergic state but could also reflect a compensatory sympathetic activation secondary to a partial neuropathy, a concern shared by others. Indeed, plasma NE >600 pg / ml has been documented in healthy subjects with volume depletion, in patients with diabetic neuropathy, and even in patients with neuropathic orthostatic hypotension. Others have used a paradoxical increase in BP with upright posture as a clinical indicator of HyperPOTS, alone or in combination with elevated norepinephrine. POTS patients have been characterized according to calf blood flow response to standing, with low-flow patients probably representing those with a hyperadrenergic phenotype. Thus, current biomarkers of sympathetic activity used to identify “hyperadrenergic” POTS either provide only indirect evidence of sympathetic activation or cannot differentiate between primary or appropriate compensatory sympathetic activation. [000136] Several publications have reported direct recordings of central sympathetic outflow to skeletal muscle (MSNA) in POTS, and the variability in these results likely reflects the heterogeneity of this condition. Most studies have shown either normal or even decreased supine MSNA and a normal or exaggerated MSNA response to stimulation by upright posture and other stressors. It is likely that an exaggerated increase in MSNA triggered by standing will have the same limitations as upright norepinephrine in differentiating between a primaryDocket No.093386-9254-WO01 hyperadrenergic state or a compensatory response to e.g., volume depletion. Therefore, supine resting MSNA was focused on. Similarly, to other studies, a wide range of resting supine MSNA in POTS was observed, but the results herein suggest that high resting MSNA identifies POTS patients with a hyperadrenergic phenotype. These patients have significantly higher upright BPs compared to those with the lowest MSNA despite having similar severity of orthostatic tachycardia. Furthermore, these patients have an exaggerated sympathetic activation in response to the VM, and this is translated as a greater increase in DBP, reflecting sympathetic- mediated vasoconstriction. The inventors found that an increase in DBP>18 mmHg during phase 2 of the VM (late-early) to be a useful clinical biomarker of HyperPOTS. It is reassuring that the identical DBP threshold (>18 mmHg) resulted from ROC analysis using MSNA characteristics and, independently, using patients’ reported responsiveness to guanfacine (PGIC). It correlated with other clinical characteristics suggestive of a hyperadrenergic phenotype, like a paradoxical increase in upright BP and spontaneous episodes that resemble “adrenaline rushes,” with facial flushing, hypertension, and tachycardia. It is more readily available than direct measurement of MSNA, and it’s the clinical reflection of sympathetic vasoconstriction. More importantly, this clinical biomarker predicted a beneficial response to treatment with the central sympatholytic guanfacine. Thus, this is prima facie evidence that these patients have a primary central sympathetic activation driving their POTS because patients in whom sympathetic activation is a compensatory mechanism needed to maintain upright BP would be expected to get worse if treated with a central sympatholytic. [000137] Notably, there was a lack of association between the severity of postural tachycardia and the HyperPOTS phenotype. Patients in the highest and lower quartiles of MSNA had indistinguishable orthostatic tachycardia and identical HR changes to the VM (FIGS.2A-F). Even though orthostatic tachycardia reflects sympathetic activation in POTS, it does not inform us whether this activation is due to a primary central sympathetic overactivity (as in HyperPOTS) or an adequate compensatory reflex response (e.g., to volume depletion). [000138] The pathophysiological mechanisms causing this hyperadrenergic phenotype have not been explored. The inventors have previously reported that spontaneous hyperadrenergic episodes with flushing, hypertension, and tachycardia are associated with MCAD in patients with POTS. MCAD was suspected in a minority of patients in the cohort herein, but making a definitive diagnosis is often difficult and may require measuring mast cell mediators during or shortly after an episode. Clinically indistinguishable episodes are also seen in patients with pseudopheochromocytoma. The magnitude of these episodes can be dramatic, with acuteDocket No.093386-9254-WO01 increases in SBP in the 180 mmHg range. A similar clinical picture can be seen in panic attacks, and a non-specific anxiolytic effect of guanfacine cannot be excluded, which has been observed in children and adolescents and adults with anxiety complicating cardiac surgery. However, patients with POTS typically deny a sensation of doom or panic and accept the transient nature of these episodes. Furthermore, formal assessment for psychiatric disorders has shown that patients with POTS do not have an increased prevalence of anxiety disorders, including panic disorder, compared with the general population. Importantly, the observation that patients with panic disorders have normal resting MSNA levels argues against a panic disorder underlying the hyperadrenergic state in the HyperPOTS patients herein. Further studies are required to explore the underlying cause of hyperadrenergic POTS. Regardless, the studies herein suggest the involvement of brain alpha-2 receptors in its pathophysiology. [000139] Clonidine was used to treat patients with mitral valve prolapse syndrome and hyperdynamic circulatory state with orthostatic tachycardia, likely a condition similar to POTS. The inventors used guanfacine instead of clonidine because the latter has a short half-life, and the initial sympathetic inhibition is followed by rebound sympathetic activation. For this reason, short-acting clonidine was avoided in other hyperadrenergic conditions, such as afferent baroreflex failure. Slow-release clonidine and transdermal patches may be alternative therapies. Also, guanfacine has fewer CNS adverse effects than clonidine, such as sedation, headache, orthostatic hypotension and impotence, and a lower incidence of withdrawal symptoms such as high BP, headache, tremor, restlessness, and nausea (3% vs.50-80% with clonidine). Methyldopa has been used in the treatment of POTS associated with mast cell activation disorders that have a similar hyperadrenergic phenotype. Methyldopa, however, may be associated with a higher incidence of depression, and there is a chronic shortage of the drug in the American market. [000140] The results herein are encouraging in that only the group of patients that met the clinical biomarker of HyperPOTS had a significant improvement on guanfacine even though all were treated with this medication because of an apparent hyperadrenergic clinical picture. Furthermore, patients were referred to the inventors after having tried multiple unsuccessful therapies. Establishing the efficacy and safety of guanfacine in the treatment of HyperPOTS will require a placebo-controlled, randomized clinical trial. Such a study can also be used to validate ¨DBPVM2as a clinical biomarker of HyperPOTS.Docket No.093386-9254-WO01 [000141] The clinical biomarker developed herein to identify HyperPOTS patients, ¨DBPVM2, uses a conservative threshold of >18 mmHg to ensure high specificity (100%) over sensitivity (70%). This may explain why some overlap was observed herein in the individual responses to guanfacine between groups, with patients who did not meet the strict diagnostic criteria still reporting clinical benefit in the global impression of change score after guanfacine treatment. It is possible that the clinical biomarker criteria could be refined by studying a larger group of patients. Therefore, patients who do not meet the strict DBP criteria may still improve on guanfacine. [000142] The results herein are consistent with the concept that a subset of patients with POTS has a primary central sympathetic activation as their underlying pathophysiology. This can be documented by direct measurements of resting sympathetic activity, and this HyperPOTS phenotype can be identified clinically by an exaggerated increase in DBP during phase 2 of the VM. Importantly, this parameter predicted a greater improvement in symptoms of orthostatic intolerance and fatigue in response to the central sympatholytic guanfacine. Example 7 Identifying Predictors of Response to Guanfacine in Myalgic Encephalomyelitis / Chronic Fatigue Syndrome (ME / CFS) Patients [000143] Clinical characteristics of CFS patients with hyperadrenergic phenotype were used as the study population. Patients who meet criteria for ME / CFS and have a combination of the following putative hyperadrenergic features: orthostatic hypertension; spontaneous episodes described as “adrenaline rushes” by the patients: facial flushing + increased BP + increased HR; and hyperadrenergic responses to the Valsalva maneuver. This phenotype is associated with increased sympathetic activity, measured directly with intraneuronal recordings. MSNA is a reflection of central sympathetic outflow. Patients were characterized by an increase in MSNA at rest and standing. Responses to orthostatic stress are shown in FIGS.9A-B. Patients with high MSNA had higher resting blood pressure and a significantly larger increase in blood pressure during standing. FIGS.10A-D show that patients with high MSNA had a higher BP response to the late phase 2 of the Valsalva maneuver. [000144] Two surveys were conducted. The first survey contained general assessment of disease diagnosis, associated symptoms, and effects of guanfacine administration. The goal was to determine initial perception of drug (PGIC) and serve as a screening tool for the secondDocket No.093386-9254-WO01 survey. Enrollment details: 170 subjects were identified; 59 subjects were enrolled; and 48 subjects completed the survey. The second survey contained a detailed assessment of autonomic and CFS-related symptoms, quality of life, and response to guanfacine administration. The goal was to gain in-depth knowledge on CFS-related symptoms and perception of drug. Enrollment details: 27 subjects were identified; 20 subjects were enrolled; and 19 subjects completed the survey. [000145] Patient-reported impressions of the drug guanfacine using the PGIC are shown in FIG.11. The PGIC assesses the change in the patient's overall status after an intervention. It integrates symptomatic, functional, mood and quality of life changes, and any treatment effects. It is tied to the conceptual framework of overall improvement or non-improvement. [000146] FIGS.12A-C show survey results from the Fukuda questionnaire where participants were asked questions regarding their fatigue before and during guanfacine treatment. Compared to non-responders, overall improvement was associated with significant improvement in the frequency and severity of hyperadrenergic symptoms and significant improvement in orthostatic tolerance. [000147] FIGS.13A-C show survey results from the Fukuda questionnaire where participants were asked questions regarding their fatigue before and during guanfacine treatment. Compared to non-responders, overall improvement was associated with significant improvement in chronic fatigue and post-exertional malaise (PEM). Responders vs non- responders had similar: age (responders: 27±2 years vs non-responders: 23±1 years); disease duration, the majority had ^1-5 years of diagnosis; and PEM (severity and frequency). Fatigue (PROMIS 7a raw score) tended to be higher in responders (FIG.13D). Responders vs non- responders also had similar: impairment in sleep and memory; frequency and severity of sleep and memory impairment and hyperadrenergic symptoms (FIG.13E-F). The overall impact of orthostatic intolerance on activities of daily living tended to be higher in responders, and they had more difficulties performing activities that require being standing for short and long time. [000148] The following were tests used to determine predictors of a response to guanfacine. Compared to non-responders, responders had a greater BP increase during the late phase 2 of the Valsalva maneuver (FIGS.14A-B). Compared to non-responders, responders had greater DBP increase at 1 and 3 min of head-up tilt (HUT; FIGS.14C-D). Predictors of having aDocket No.093386-9254-WO01 response to guanfacine were greater orthostatic hypertension; exaggerated pressor response to the VM at baseline; and more severe baseline fatigue (PROMIS score 31 vs 28 (p=0.07)). [000149] The majority of participants believed their POTS symptoms improved while taking guanfacine. Participants believed there was minimal improvement or no change to their CFS symptoms while taking guanfacine. Numerous symptoms and their severity were reduced while taking guanfacine. A few participants believed guanfacine decreased their adrenaline rushes, while others believed it caused low blood pressure. Summaries of the responses to the two surveys are shown in FIGS.15A-C. Example 8 Improving Symptoms of Myalgic Encephalomyelitis / Chronic Fatigue Syndrome (ME / CFS) [000150] In the studies discussed in Example 7, a subset of ME / CFS patients with a hyperadrenergic phenotype (HyperME / CFS) characterized by an exaggerated pressor response to the vasoconstrictive phase of the Valsalva maneuver was identified. More importantly, in feasibility studies herein, these HyperME / CFS patients improved their fatigue, post-exertional malaise (PEM) and other ME / CFS-related symptoms during treatment with the central sympatholytic guanfacine, whereas those with non-hyperadrenergic phenotype did not. This suggests that increased sympathetic activation is indeed a primary disease-driving phenomenon in this subset of HyperME / CFS patients. Thus, the overarching hypothesis is that there is a subset of ME / CFS patients in whom sympathetic activation plays a pathophysiological role (“HyperME / CFS”) and that their ME / CFS-related symptoms will improve with the central sympatholytic guanfacine. [000151] This hypothesis will be tested in a double-blind, placebo-controlled, randomized, parallel-group exploratory study using the central sympatholytic guanfacine in a cohort of ME / CFS patients enriched for this hyperadrenergic phenotype by their exaggerated pressor response to the vasoconstrictive phase of the Valsalva maneuver. If the hypothesis is true, the administration of guanfacine to patients with HyperME / CFS will improve their fatigue and reduce PEM, the two hallmark symptoms of ME / CFS, and these effects will be associated with decreased markers of sympathetic activity. The efficacy and safety of guanfacine will be compared using subjective and objective measures of fatigue, PEM, exercise capacity, and orthostatic tolerance.Docket No.093386-9254-WO01 [000152] To test the hypothesis that guanfacine improves fatigue in HyperME / CFS compared to placebo, the effects of guanfacine on validated instruments that measure fatigue will be determined. The effects of guanfacine on subjective and objective measures of physical function, orthostatic tolerance, and quality of life, and whether these effects are associated with a decrease in biomarkers of sympathetic activity will also be determined. [000153] To test the hypothesis that guanfacine improves post-exertional malaise in HyperME / CFS compared to placebo, a standardized submaximal exercise challenge will be used as a standardized stressor to induce PEM before and after treatment. [000154] An Exploratory Clinical Trial with two phases will be used. At the end of the initial phase, an interim analysis with a go / no-go decision based on safety, tolerability, and feasibility of meeting enrollment and retention milestones will be performed. The second phase will provide an assessment of short-term efficacy, and tolerability and inform about the design of subsequent efficacy studies. [000155] If successful, these studies have the potential to contribute to the understanding of the pathophysiology of ME / CFS and provide a novel therapeutic target for this disease by overcoming two important hurdles in the treatment of these patients: the lack of specific therapies and the lack of clinical biomarkers to select patients most likely to benefit. Furthermore, because an FDA-approved medication will be repurposed, the results can have an immediate impact in the quality of life of patients. Example 9 Phenome Wide Association Study (PheWAS) Analysis [000156] The focus of the Phenome Wide Association Study (PheWAS) analysis is to find potential links between mutations in the Solute carrier family 6 member 2 (SLC6A2) gene and phenotypes that are currently not well treated or understood. This method identifies phenotypes associated with SNPs in various genes, including those producing proteins targeted by different pharmacologic agents. Gene mutation recapitulates the effect of the disease / drug of interest. The goal is to lend support for the development of a norepinephrine reuptake activator or potentially repurpose existing drugs that may aid in treating some of these diseases or symptoms.Docket No.093386-9254-WO01 [000157] A PheWAS was performed in a disease-agnostic cohort of 40,000 BioVU patients in order to identify potential novel genotype-phenotype associations related to the SNP(s) of interest. PheWAS leverages Vanderbilt’s informatics and data mining expertise and robust de- identified health records with associated genomic data in BioVU (>205,000 DNA samples). [000158] The Exomechip contains ~250,000 coding variants across the protein coding region of the genome. The coding variants were discovered through exome and whole genome sequencing in more than 12,000 individuals. It was designed to serve as an intermediate step between current genotyping arrays, which are designed to study common variants, and exome sequencing which can discover rare variants. Nearly all non-synonymous, splice and stop altering variants detected in an average genome through exome sequencing were included on the Exomechip. The Exomechip also includes single nucleotide polymorphisms (SNPs) with known disease associations as well as unpublished associations from consortia working on diabetes, blood lipids, blood pressure, lung function, myocardial infraction, anthropometric traits, psychiatric traits, Crohn's disease and age-related macular degeneration. Approximately 35,000 BioVU subjects have genotyping data on this platform based on inclusion criteria defined in StarBRITE. It is important to note that the European ancestry data was focused on because other ethnicities were underpowered. [000159] The SNP of interest in SLC6A2 is rs1805065 (Thr99lle). Information about this SNP’s predicted consequences and minor allele frequencies is presented in TABLE 8. TABLE 8. Relevant SNP for NET* Minor allele frequency reflects the MAF in the Exomechip European ancestry population. [000160] Sorting Intolerant from Tolerant (SIFT) scores at or below 0.05 are considered to be deleterious; those above 0.05 are considered to be tolerated. Polyphen2 (PP2) scores below 0.447 are considered benign; those higher than 0.908 are considered probably damaging; andDocket No.093386-9254-WO01 those in between possibly damaging. As shown in TABLE 8, the SNP is predicted to be tolerated by SIFT and PolyPhen2. An expression study found no effects of this SNP on kinetic constants of norepinephrine uptake or on potency of desipramine for inhibition of norepinephrine uptake. [000161] Given the concordance of phenotypes revealed in the PheWAS, the rs1805065 minor allele essentially has (or is indirectly associated with) the opposite effect of the drug, (i.e. it inhibits, reduces, or prevents norepinephrine reuptake from the synapse). Thus, phenotypes for which this SNP is statistically associated may yield new opportunities for therapeutic use of a novel norepinephrine reuptake activator. The phenotypes expected to be associated with the NET gene are confirmed with T99Ile: a constellation of phenotypes affecting the cardiovascular system (as are observed with known alterations in NET function, e.g. A457P SNP), particularly in the pediatric subpopulation (TABLES 9-10). TABLE 9. Adult PheWAS Results Validating Current UseTABLE 10. Pediatric PheWAS Results Validating Current UseDocket No.093386-9254-WO01[000162] NLP-PheWAS based on mapped concepts from clinical text also revealed hits that match the validating associations reported above (e.g., drooling, cardiomyopathies), as well as associations with the novel phenotype that will be explored further in the next section (chronic fatigue). TABLE 11 reports some results of the NLP-PheWAS. TABLE 11. NLP-PheWAS Results[000163] Chronic fatigue syndrome is associated with the same SNP (T99I) in adults (p=0.0000157; TABLE 12). TABLE 12. Potentially Novel AssociationDocket No.093386-9254-WO01[000164] Upon in depth chart review of minor allele carriers in the dataset herein, the presence of chronic fatigue in cases was verified, and it was also noted that there was apparent preponderance of cardiovascular comorbidities. Natural language processing based PheWAS analysis of the medical records associated with this SNP yielded further evidence of a connection between this SNP and presence of fatigue (p=0.0002296), as well an intriguing concept, “physically well but worried” (p=0.003707). [000165] Hypotheses herein are that: 1) some patients may experience a ‘chronic fatigue syndrome’-like disease due to an excess of norepinephrine in the synapse, and thus 2) may potentially benefit with a norepinephrine reuptake activator that 3) once designed would have efficacy in humans reducing these levels (FIG.5). [000166] Further, NET-deficient transgenic mice are noted to be affected by elevated blood pressure, heart rate, and catecholamine concentrations. One study of healthy volunteers found that NET functional polymorphisms (rs28386840 (upstream) and rs2242446 (intronic)) were associated with greater blood pressure response to exercise challenge. [000167] Interestingly, the package insert for Strattera®(atomoxetine), a medication sometimes used off label for chronic fatigue and a norepinephrine reuptake inhibitor, specifically comments on this condition: “Pheochromocytoma: Serious reactions, including elevated blood pressure and tachyarrhythmia, have been reported in patients with pheochromocytoma or a history of pheochromocytoma who received STRATTERA®. Therefore, STRATTERA®should not be taken by patients with pheochromocytoma or a history of pheochromocytoma.” [000168] Related to the chronic fatigue phenotype of interest, the Strattera®insert also notes adverse events that actually parallel some of the chronic fatigue symptoms of interest herein: “In placebo-controlled registration studies involving adult patients, tachycardia was identified as an adverse event for 1.5% (8 / 540) of STRATTERA®patients compared with 0.5% (2 / 402) of placebo patients…The most commonly observed adverse reactions in patients treated with STRATTERA®(incidence of 5% or greater and at least twice the incidence in placebo patients, for either BID or QD dosing) were: nausea, vomiting, fatigue…Additional data from ADHDDocket No.093386-9254-WO01 clinical trials (controlled and uncontrolled) has shown that approximately 5 to 10% of adult patients experienced potentially clinically important changes in heart rate (^20 beats per min) or blood pressure (^15 to 20 mm Hg).” TABLE 13 shows the phenotypes associated with CFS. TABLE 13. Phenotypes associated with chronic fatigue syndrome.[000169] The SNP carriers identified herein might have: 1) NET impairment not detectable by (S)-3,5-Dihydroxyphenylglycine (DHPG), 2) some leaking effect that masks what might otherwise be decreased DHPG, or 3) be susceptible to epigenetic changes in SLC6A2 in response to stress. [000170] A paper exploring the differences between how eye symptoms are reported in the electronic medical record (EMR) vs. patient report on an Eye Symptom Questionnaire (ESQ) found that there were large inconsistencies in symptom reporting between the ESQ and EMR. The authors conclude that symptom reporting varies between methods, with patients reportingDocket No.093386-9254-WO01 more symptoms on self-reported questionnaires. This is of interest, where symptoms may not be adequately captured in the EMR, therefore shoring up the need to directly survey patients about their experiences. In preparation for the biomarker study, a survey was administered through MyResearch at Vanderibilt (MRAV), to learn more about the discourse course and symptoms. These data were used to inform the biomarker study design and identify the patient population for recruitment. Some findings from the MRAV survey are presented in TABLE 14.MRAV Volunteer Reported IdeasDocket No.093386-9254-WO01[000171] Almost 3,000 compounds were screened in fluorescent substrate NET assay using a brief compound incubation protocol (2-hour compound incubation). Benzbromarone was a hit that was not promising in the orthogonal follow-up radioactive NET assay. 1,312 compounds (FDA collection) + 5 research compounds (CFR2 agonists) identified from a literature search were subsequently screened using a longer compound incubation protocol (1–7-day compound incubation) in fluorescent assay. Dose-response curves showed a reliable, dose-dependent increase in NET activity with guanfacine after treatment for 7 days. [000172] Given the sex differences in stress response and CFS prevalence, a straight to humans (no preclinical efficacy work in animals) is necessary since sex differences are highly species-specific, which would make any animals models very poor predictors of human response to treatment. Therefore, another high throughput screen (HTS) was conducted, and that screen identified guanfacine as the most promising candidate (FIG.18). Therefore, guanfacine was chosen to be further analyzed as shown in the above examples. *** [000173] The foregoing description of the specific aspects will so fully reveal the general nature of the invention that others can, by applying knowledge within the skill of the art, readily modify and / or adapt for various applications such specific aspects, without undue experimentation, without departing from the general concept of the present disclosure. Therefore, such adaptations and modifications are intended to be within the meaning and range of equivalents of the disclosed aspects, based on the teaching and guidance presented herein.Docket No.093386-9254-WO01 It is to be understood that the phraseology or terminology herein is for the purpose of description and not of limitation, such that the terminology or phraseology of the present specification is to be interpreted by the skilled artisan in light of the teachings and guidance. [000174] The breadth and scope of the present disclosure should not be limited by any of the above-described exemplary aspects, but should be defined only in accordance with the following claims and their equivalents. [000175] All publications, patents, patent applications, and / or other documents cited in this application are incorporated by reference in their entirety for all purposes to the same extent as if each individual publication, patent, patent application, and / or other document were individually indicated to be incorporated by reference for all purposes. [000176] For reasons of completeness, various aspects of the invention are set out in the following numbered clauses: [000177] Clause 1. A method for treating a fatigue disorder in a subject, the method comprising: administering a therapeutically effective amount of a composition comprising an alpha-2 adrenergic agonist, or a pharmaceutically acceptable salt thereof, to the subject, the subject having at least one symptom selected from the group consisting of chronic fatigue, orthostatic hypertension, increased blood pressure, increased heart rate, and facial flushing. [000178] Clause 2. The method of clause 1, wherein the subject has a single nucleotide polymorphism in SLC6A2 gene. [000179] Clause 3. The method of clause 2, wherein the single nucleotide polymorphism is a mutation at Thr99Ile. [000180] Clause 4. The method of clause 3, wherein the mutation is a missense mutation. [000181] Clause 5. The method of any of clauses 1-4, wherein the fatigue disorder is at least one selected from the group consisting of chronic fatigue syndrome (CFS), postural orthostatic tachycardia syndrome (POTS), and fibromyalgia. [000182] Clause 6. The method of any of clauses 1-5, wherein the fatigue disorder is chronic fatigue syndrome (CFS) and / or postural orthostatic tachycardia syndrome (POTS).Docket No.093386-9254-WO01 [000183] Clause 7. The method of any of clauses 1-6, the subject further having at least one symptom selected from the group consisting of reduced stamina, insomnia, muscle pain, muscle weakness, temperature dysregulation, dizziness, cognitive exhaustion, and physical exhaustion. [000184] Clause 8. The method of any of clauses 1-7, wherein the alpha-2 adrenergic agonist comprises at least one selected from the group consisting of clonidine, p-aminoclonidine, tiamenidine, 5-bromo-6-(2-imidazoline-2-yl amino) quinoxaline, dexmedetomidine, detomidine, medetomidine, oxymetazonline, brimonidine, tizanidine, mivazerol, lofexidine, xylazine, guanabenz, guanfacine, guanclofine, guanoxabenz, alpha-methylnorepinephrine, azepexole, indoramin, pharmaceutically acceptable salts thereof, and 6-allyl-2-amino-5,6,7,8-tetrahydro4H- thiazolo [4,5-d]azepine diHCl. [000185] Clause 9. The method of any of clauses 1-7, wherein the alpha-2 adrenergic agonist comprises a compound of formula:, or a pharmaceutically acceptable salt thereof, wherein, at each occurrence, R1is independently –Cl or –CH3and R2is –OC1-4alkyl, –[000186] Clause 10. The method of any of clauses 1-9, wherein the alpha-2 adrenergic agonist comprises guanfacine or a pharmaceutically acceptable salt thereof. [000187] Clause 11. The method of any of clauses 1-10, wherein the composition comprises about 0.1 mg to about 4 mg of the alpha-2 adrenergic agonist or a pharmaceutically acceptable salt thereof. [000188] Clause 12. The method of any of clauses 1-11, wherein the composition comprises about 0.1 mg to about 4 mg guanfacine or a pharmaceutically acceptable salt thereof. [000189] Clause 13. The method of any of clauses 1-12, wherein the composition is orally administered to the subject. [000190] Clause 14. The method of any of clauses 1-13, wherein the composition is administered to the subject at least once daily.Docket No.093386-9254-WO01 [000191] Clause 15. The method of any of clauses 1-14, wherein the composition is in the form of a tablet. [000192] Clause 16. A method for treating chronic fatigue syndrome (CFS) and / or postural orthostatic tachycardia syndrome (POTS) in a subject, the method comprising: administering a therapeutically effective amount of a composition comprising guanfacine, or a pharmaceutically acceptable salt thereof, to the subject, the subject having at least one selected from the group consisting of chronic fatigue, orthostatic hypertension, increased blood pressure, increased heart rate, and facial flushing. [000193] Clause 17. The method of clause 16, wherein the subject has a single nucleotide polymorphism in SLC6A2 gene. [000194] Clause 18. The method of clause 17, wherein the single nucleotide polymorphism is a mutation at Thr99Ile. [000195] Clause 19. The method of clause 18, wherein the mutation is a missense mutation. [000196] Clause 20. The method of any of clauses 16-19, wherein the composition comprises about 0.1 mg to about 4 mg guanfacine or a pharmaceutically acceptable salt thereof. [000197] Clause 21. The method of any of clauses 16-20, wherein the composition is orally administered to the subject. [000198] Clause 22. The method of any of clauses 16-21, wherein the composition is administered to the subject at least once daily. [000199] Clause 23. A method of diagnosing a fatigue disorder in a subject, the method comprising: (a) measuring a baseline blood pressure (BP) of the subject, wherein the subject is at rest and BP measurement includes diastolic BP (DBP) and systolic BP (SBP) measurements; (b) instructing the subject to perform a Valsalva maneuver for a period of time; and (c) measuring BP of the subject during late phase 2 of the Valsalva maneuver; wherein when the DBP increases by more than 18 mmHg from the baseline DBP, then the subject has the fatigue disorder; and wherein when the DBP increases by more than 18 mmHg from the baseline DBP, then the subject has an increased likelihood of being treated by a composition comprising an alpha-2 adrenergic agonist or a pharmaceutically acceptable salt thereof.Docket No.093386-9254-WO01 [000200] Clause 24. The method of clause 23, wherein the fatigue disorder is hyperadrenergic postural orthostatic tachycardia syndrome (HyperPOTS) or chronic fatigue syndrome (CFS). [000201] Clause 25. The method of clause 23 or clause 24, wherein the alpha-2 adrenergic agonist is guanfacine or a pharmaceutically acceptable salt thereof. [000202] Clause 26. The method of clause 25, wherein the composition comprises about 0.1 mg to about 4 mg guanfacine or a pharmaceutically acceptable salt thereof. [000203] Clause 27. The method of any one of clauses 23-26, wherein the period of time is from about 10 seconds to about 30 seconds. [000204] Clause 28. The method of any one of clauses 23-27, wherein the late phase 2 of the Valsalva maneuver is from the first increase in systolic blood pressure (SBP) from baseline SBP after the nadir until phase 3. [000205] Clause 29. The method of any one of clauses 23-28, further comprising (d) administering the composition comprising the alpha-2 adrenergic agonist or a pharmaceutically acceptable salt thereof to the subject having a fatigue disorder.

Claims

Docket No.093386-9254-WO01 CLAIMS What is claimed is:

1. A method for treating a fatigue disorder in a subject, the method comprising: administering a therapeutically effective amount of a composition comprising an alpha-2 adrenergic agonist, or a pharmaceutically acceptable salt thereof, to the subject, the subject having at least one symptom selected from the group consisting of chronic fatigue, orthostatic hypertension, increased blood pressure, increased heart rate, and facial flushing.

2. The method of claim 1, wherein the subject has a single nucleotide polymorphism in SLC6A2 gene.

3. The method of claim 2, wherein the single nucleotide polymorphism is a mutation at Thr99Ile.

4. The method of claim 3, wherein the mutation is a missense mutation.

5. The method of any of claims 1-4, wherein the fatigue disorder is at least one selected from the group consisting of chronic fatigue syndrome (CFS), postural orthostatic tachycardia syndrome (POTS), and fibromyalgia.

6. The method of any of claims 1-5, wherein the fatigue disorder is chronic fatigue syndrome (CFS) and / or postural orthostatic tachycardia syndrome (POTS).

7. The method of any of claims 1-6, the subject further having at least one symptom selected from the group consisting of reduced stamina, insomnia, muscle pain, muscle weakness, temperature dysregulation, dizziness, cognitive exhaustion, and physical exhaustion.

8. The method of any of claims 1-7, wherein the alpha-2 adrenergic agonist comprises at least one selected from the group consisting of clonidine, p-aminoclonidine, tiamenidine, 5-bromo- 6-(2-imidazoline-2-yl amino) quinoxaline, dexmedetomidine, detomidine, medetomidine, oxymetazonline, brimonidine, tizanidine, mivazerol, lofexidine, xylazine, guanabenz, guanfacine,Docket No.093386-9254-WO01 guanclofine, guanoxabenz, alpha-methylnorepinephrine, azepexole, indoramin, pharmaceutically acceptable salts thereof, and 6-allyl-2-amino-5,6,7,8-tetrahydro4H-thiazolo [4,5-d]azepine diHCl.

9. The method of any of claims 1-7, wherein the alpha-2 adrenergic agonist comprises a compound of formula:, or a pharmaceutically acceptable salt thereof, wherein, at each occurrence, R1is independently10. The method of any of claims 1-9, wherein the alpha-2 adrenergic agonist comprises guanfacine or a pharmaceutically acceptable salt thereof.

11. The method of any of claims 1-10, wherein the composition comprises about 0.1 mg to about 4 mg of the alpha-2 adrenergic agonist or a pharmaceutically acceptable salt thereof.

12. The method of any of claims 1-11, wherein the composition comprises about 0.1 mg to about 4 mg guanfacine or a pharmaceutically acceptable salt thereof.

13. The method of any of claims 1-12, wherein the composition is orally administered to the subject.

14. The method of any of claims 1-13, wherein the composition is administered to the subject at least once daily.

15. The method of any of claims 1-14, wherein the composition is in the form of a tablet.

16. A method for treating chronic fatigue syndrome (CFS) and / or postural orthostatic tachycardia syndrome (POTS) in a subject, the method comprising: administering a therapeutically effective amount of a composition comprising guanfacine, or a pharmaceutically acceptable salt thereof, to the subject, the subject having at least oneDocket No.093386-9254-WO01 selected from the group consisting of chronic fatigue, orthostatic hypertension, increased blood pressure, increased heart rate, and facial flushing.

17. The method of claim 16, wherein the subject has a single nucleotide polymorphism in SLC6A2 gene.

18. The method of claim 17, wherein the single nucleotide polymorphism is a mutation at Thr99Ile.

19. The method of claim 18, wherein the mutation is a missense mutation.

20. The method of any of claims 16-19, wherein the composition comprises about 0.1 mg to about 4 mg guanfacine or a pharmaceutically acceptable salt thereof.

21. The method of any of claims 16-20, wherein the composition is orally administered to the subject.

22. The method of any of claims 16-21, wherein the composition is administered to the subject at least once daily.

23. A method of diagnosing a fatigue disorder in a subject, the method comprising: (a) measuring a baseline blood pressure (BP) of the subject, wherein the subject is at rest and BP measurement includes diastolic BP (DBP) and systolic BP (SBP) measurements; (b) instructing the subject to perform a Valsalva maneuver for a period of time; and (c) measuring BP of the subject during late phase 2 of the Valsalva maneuver; wherein when the DBP increases by more than 18 mmHg from the baseline DBP, then the subject has the fatigue disorder; and wherein when the DBP increases by more than 18 mmHg from the baseline DBP, then the subject has an increased likelihood of being treated by a composition comprising an alpha-2 adrenergic agonist or a pharmaceutically acceptable salt thereof.Docket No.093386-9254-WO01 24. The method of claim 23, wherein the fatigue disorder is hyperadrenergic postural orthostatic tachycardia syndrome (HyperPOTS) or chronic fatigue syndrome (CFS).

25. The method of claim 23 or claim 24, wherein the alpha-2 adrenergic agonist is guanfacine or a pharmaceutically acceptable salt thereof.

26. The method of claim 25, wherein the composition comprises about 0.1 mg to about 4 mg guanfacine or a pharmaceutically acceptable salt thereof.

27. The method of any one of claims 23-26, wherein the period of time is from about 10 seconds to about 30 seconds.

28. The method of any one of claims 23-27, wherein the late phase 2 of the Valsalva maneuver is from the first increase in systolic blood pressure (SBP) from baseline SBP after the nadir until phase 3.

29. The method of any one of claims 23-28, further comprising (d) administering the composition comprising the alpha-2 adrenergic agonist or a pharmaceutically acceptable salt thereof to the subject having a fatigue disorder.

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