阅读说明：本技术 排他地由设计者药物激活的设计者受体在治疗癫痫发作紊乱中的用途 (Use of designer receptors activated exclusively by designer drugs in the treatment of seizure disorders ) 是由 马修·杜林 于 2018-03-15 设计创作，主要内容包括：提供了用于治疗癫痫发作紊乱的方法和组合物,所述方法和组合物包括向患者施用编码修饰的受体的载体,用于将修饰的受体递送至靶位置,该修饰的受体被修饰以被合成配体激活,其中修饰的受体在被激活时抑制神经元放电；以及向患者施用合成配体。还提供了用于治疗需要其的患者中的癫痫发作紊乱的方法和组合物,所述方法和组合物通过向患者施用编码修饰的受体的载体,用于将修饰的受体递送至靶位置,该修饰的受体被修饰以被合成配体激活,其中修饰的受体在被激活时增加神经元放电；以及向患者施用合成配体。所述方法用于改善或减轻癫痫发作紊乱的一种或更多种症状。(Methods and compositions are provided for treating seizure disorders, the methods and compositions comprising administering to a patient a vector encoding a modified receptor, the modified receptor modified to be activated by a synthetic ligand, for delivering the modified receptor to a target location, wherein the modified receptor, when activated, inhibits neuronal discharge; and administering the synthetic ligand to the patient. Also provided are methods and compositions for treating seizure disorders in a patient in need thereof by administering to the patient a vector encoding a modified receptor, the modified receptor modified to be activated by a synthetic ligand, for delivering the modified receptor to a target location, wherein the modified receptor increases neuronal discharge when activated; and administering the synthetic ligand to the patient. The methods are useful for ameliorating or alleviating one or more symptoms of a seizure disorder.)

1. A method of treating a seizure disorder in a patient in need thereof, comprising:

administering to the patient an adeno-associated viral vector encoding hM4Di for delivery of hM4Di to a target location, the vector comprising a human CaMK2A promoter, woodchuck hepatitis virus post-transcriptional regulatory elements, and a bovine growth hormone polyadenylation sequence; and

administering to the patient a synthetic ligand that activates hM4 Di.

2. A method of treating a seizure disorder according to claim 1, wherein said vector comprises two inverted terminal repeats, an SV40 origin of replication, a pUC19 origin of replication, and an ampicillin resistance gene.

3. The method of treating a seizure disorder in accordance with claim 2 wherein said vector is pAM/hCaMK2A-hM4D (Gi) -WPRE-BGHpA.

4. The method of treating a seizure disorder of claim 1, wherein the vector comprises a fluorescent reporter gene.

5. The method of treating a seizure disorder of claim 1, wherein the adeno-associated viral vector is AAV1, AAV2, AAV4, AAV5, AAV7, AAV8, or AAV 9.

6. The method of treating a seizure disorder according to claim 1, wherein said adeno-associated viral vector is AAVRec 3.

7. The method of treating a seizure disorder according to claim 1, wherein said synthetic ligand is clozapine N-oxide.

8. The method of treating a seizure disorder according to claim 1, wherein said synthetic ligand is piperapine.

9. The method of treating a seizure disorder according to claim 1, wherein the vector is delivered to a target location in the brain of the patient.

10. The method of treating a seizure disorder according to claim 1, wherein said carrier is administered via a route selected from the group consisting of oral, buccal, sublingual, rectal, topical, intranasal, vaginal, and parenteral.

11. The method of treating a seizure disorder according to claim 9, wherein the vector is administered directly to the target location through the skull of the patient.

12. The method of treating a seizure disorder according to claim 1, wherein said synthetic ligand is administered via a route selected from the group consisting of oral, buccal, sublingual, rectal, topical, intranasal, vaginal and parenteral.

13. The method of treating a seizure disorder according to claim 1, wherein the seizure disorder is selected from the group consisting of: focal cortical dysplasia, epilepsy with generalized tonic clonic seizures, epilepsy with myoclonic absence, frontal lobe epilepsy, temporal lobe epilepsy, occipital lobe epilepsy, parietal lobe epilepsy, Landau-Kleffner syndrome, Rasmussen syndrome, Dravet syndrome, Doose syndrome, CDKL5 disorder, infantile spasm (West syndrome), Juvenile Myoclonic Epilepsy (JME), vaccine-related encephalopathy, refractory childhood epilepsy (ICE), Lennox-Gastaut syndrome (LGS), Rett syndrome, Ohtahara syndrome, CDKL5 disorder, childhood absence epilepsy, essential tremor, acute recurrent seizures, benign Railando epilepsy, status epilepticus, refractory status epilepticus (SRSE), PCDH19, pediatric tumor-induced seizures, hamartoma induced seizures, drug withdrawal seizures, drug-induced seizures, Alcohol withdrawal induced seizures, increased seizure activity and paroxysmal seizures.

14. The method of treating a seizure disorder in accordance with claim 1, wherein said seizure disorder is characterized by focal seizures.

15. The method of treating a seizure disorder according to claim 1, wherein the method provides an improvement in at least one symptom selected from the group consisting of: ataxia, gait disorders, speech disorders, vocalization, involuntary laughing, impaired cognitive function, abnormal motor activity, clinical seizures, subclinical seizures, hypotonia, hypertonia, watery mouth, orodental behavior, aura, convulsions, repetitive motion, unusual sensations, frequency of seizures, and severity of seizures.

16. A vector comprising a nucleic acid encoding hM4Di under the regulatory control of a human CaMK2A promoter, a woodchuck post-transcriptional regulatory element, and a bovine growth hormone polyadenylation sequence.

17. The vector of claim 16, further comprising two inverted terminal repeats, an SV40 origin of replication, a pUC19 origin of replication, and an ampicillin resistance gene.

18. The vector of claim 16, wherein the vector is pAM/hCaMK2A-hM4D (Gi) -WPRE-BGHpA.

Technical Field

Seizure disorders (seizuredsorders) are treated using Designer Receptors (Designer Receptors explicit Activated by Designer Drugs) and synthetic ligands and ultrasound that are Exclusively Activated by Designer Drugs.

Background

Seizure disorders typically involve abnormal neuronal activity in the brain, resulting in seizures (seizure) which can manifest as abnormal behavior over time, sensations, convulsions, diminished consciousness, and sometimes loss of consciousness. Seizures can be a symptom of many different disorders that can affect the brain. Epilepsy (epilepsy) is a seizure disorder characterized by recurrent seizures. See, e.g., Blume et al, epilepsia.2001; 42:1212-1218. Epileptic seizures (epiepileptic seizures) are usually marked by abnormal discharges in the brain and are usually manifested as sudden transient attacks of altered or diminished consciousness, involuntary movements or convulsions.

Seizures can be classified as focal seizures (also known as partial seizures) and generalized seizures. Focal seizures affect only one side of the brain, while generalized seizures affect both sides of the brain. Specific types of focal seizures include simple focal seizures, complex focal seizures, and secondary generalized seizures. Simple focal seizures may be confined or concentrated to a particular lobe (e.g., temporal, frontal, parietal or occipital). Complex focal seizures usually affect a greater part of one hemisphere than simple focal seizures, but usually originate in the temporal or frontal lobe. When focal seizures spread from one side of the brain (hemisphere) to both sides of the brain, the seizures are referred to as secondary generalized seizures. Specific types of generalized seizures include absence seizures (also known as petit-mal seizures), tonic seizures, dystonic seizures, myoclonic seizures, tonic clonic seizures (also known as grand mal seizures), and clonic seizures.

Examples of seizure disorders include epilepsy, epilepsy with generalized tonic clonic seizures (epilepsy with systemic tonic-tonic seizures), epilepsy with clonic absence (epilepsy with myoclonic absence), frontal lobe epilepsy, temporal lobe epilepsy, Landau-Kleffner syndrome, Rasmussen syndrome, Dravet syndrome, Doose syndrome, CDKL5 disorders, infantile spasms (West syndrome), Juvenile Myoclonic Epilepsy (JME), vaccine-related encephalopathy, intractable childhood epilepsy (intuectable childhood epilepsy) (ICE), Lennox-Gastasy syndrome (LGS), restt syndrome, Ohtahara syndrome, CDKL5 disorders, childhood epilepsy with loss (childhood), essential tremor (acute recurrent epilepsy), epilepsy with acute recurrent seizures (acute status), epilepsy (benign status epilepticus), epilepsy persisting epilepsy (epilepsy), epilepsy persisting epilepsy retention status epilepticus (epilepsy), epilepsy retention status epilepticus (status epilepticus), epilepsy retention status epilepticus, epilepsy (status of epilepsy), epilepsy retention status of epilepsy, epilepsy retention status of epilepsy, epilepsy of childhood, epilepsy retention status of epilepsy, epilepsy of childhood, epilepsy, Hypersensitive status epilepticus (SRSE), PCDH19 pediatric epilepsy, increased seizure activity (impaired seizure activity) or paroxysmal seizures (also known as continuous or cluster seizures). Seizure disorders may be associated with a type 1 sodium channel protein alpha subunit (Scn1 alpha) -related disorder.

All types of brain tumors can be associated with seizure disorders. Some tumors are associated with a greater frequency of seizures. For example, gangliogliomas (gangliogliomas) are slow-growing benign tumors that may occur in the spinal cord and/or temporal lobes. Gangliogliomas include both neoplastic glial cells and ganglion cells, which are disorganized, variably cellular, and non-infiltrating. Gangliogliomas are commonly associated with seizures. Gliomas are brain tumors that develop from glial cells in the brain. Gliomas are classified into four grades (I, II, III and IV), and treatment and prognosis depend on the tumor grade. Low-grade gliomas originate from two different types of brain cells: astrocytes and oligodendrocytes. Low grade gliomas are classified as grade 2 tumors, making them the slowest growing type of glioma. Between 60% and 85% of people with low grade gliomas may experience seizures. High grade gliomas (grade 3 or 4) are rapidly growing gliomas that generally present a poor prognosis. Grade 3 gliomas include anaplastic astrocytomas, anaplastic oligodendrogliomas, anaplastic oligodendroastrocytomas, and anaplastic ependymomas. The glioblastoma is a grade 4 glioma. Seizures occur in more than half of patients with grade III gliomas and about one quarter of patients with grade IV gliomas. Meningiomas are tumors caused by meninges (membranes surrounding the brain and spinal cord). Although not technically located in the brain, meningiomas can press or squeeze adjacent brain, nerves and blood vessels. Meningiomas are the most common type of tumor that forms in the head. Most meningiomas are slow growing. Seizures are associated with meningiomas.

Focal cortical dysplasia is a malformation of cortical development that is a common cause of medically refractory epilepsy in pediatric populations and a common cause of medically refractory seizures in adults. Focal Cortical Dysplasia (FCD) has been classified into three types and another subtype. Type I is often associated with the temporal lobe-malformations exhibiting abnormal cortical lamination, due to abnormal radial migration and maturation of neurons (FCD type Ia) or destruction of the typical 6-layer tangential composition of the cortex with immature neurons (FCD type Ib) or both structural abnormalities, radial and tangential cortical lamination (FCD type Ic). Type II is common in the frontal lobe-malformations caused by disrupted cortical lamination (corticoid) and specific cytological abnormalities, type IIa-heterozygotic neurons (without saccular cells) and type IIb-heterozygotic neurons and saccular cells. Type III-malformations associated with different cortical delaminations (clinical dispiation) and cytological abnormalities, where the main lesions are within the same area/leaf. Type IIIa-in the temporal lobe, cortical stratification (cortical degeneration) with hippocampal atrophy, type IIIb-adjacent gliomas or glial neuron tumors (DNET, gangliogliomas), type IIIc-adjacent vascular malformations (e.g. hemangiomas, arteriovenous malformations, telangiectasias, etc.), type IIId-obtained early (trauma, ischemia or perinatal hemorrhage, infectious or inflammatory disease). See, Kabat and Krol, Pol J radio, 2012,77(2) 35-43. FCDs may relate to any part of the brain, may differ in size and location and may be multifocal. Seizures are the primary symptom of FCD, sometimes associated with mental retardation, particularly in early seizures. Symptoms may occur at any age, primarily in children, but may also occur in adults. The FCD-associated seizures may be drug resistant.

Hamartomas are the primary benign, focal malformations resembling neoplasms (neoplasms) in the tissue from which they originate. They include tissue elements that are typically found at the site, but grow in an unorganized manner. Hamartomas may originate in the brain. Tuberous Sclerosis Complex (TSC) is an inherited seizure disorder characterized by disorganized growth in various organs. Patients with this disorder can exhibit a high rate of epilepsy and cognitive problems caused by a variety of pathologies in the brain. TSC lesions (cortical nodules) typically contain heteromorphic neurons, brightly eosinophilic giant cells, and white matter alterations. The TSC-associated seizure may be refractory. Gray node hamartoma (also known as hypothalamic hamartoma) is a benign tumor in which an unorganized collection of neurons and glia accumulate at the gray node of the hypothalamus. Symptoms include dementia-type seizures, which are disorders characterized by a period of time (shells) of unconscious laughing with intermittent irritability and depressed mood.

Status Epilepticus (SE) is characterized by: epileptic seizures greater than five minutes, or more than one seizure over a five minute period without the patient recovering normal in the meantime. If treatment is delayed, SE may be a dangerous condition that may lead to death. SE may be convulsive, with regular arm and leg contraction and extension patterns; or non-convulsive, changes in the level of consciousness of a person of relatively long duration, but without flexion and extension of the large-scale limbs due to seizure activity. Convulsive SE (cse) may also be classified as (a) tonic-clonic SE, (b) tonic SE, (c) clonic SE, and (d) myoclonic SE. Nonconvulsive se (ncse) is characterized by abnormal mental states, unresponsiveness, abnormal eye movement, persistent electrographic seizures, and possible responses to anticonvulsants.

Agents used to treat seizure disorders may be referred to as antiepileptic drugs ("AEDs"). Treatment of recurrent seizures has focused primarily on the use of at least one AED, where in the event of failure of a monotherapy, it is possible to adjunctively use a second or even a third agent. See, Tolman and Faulkner, Ther Clin Risk manag.2011; 7:367-375. However, approximately 30% -40% of epileptic patients have insufficient seizure control with only one AED, and require the use of adjuncts.Same as above. Despite the reasonable dose of many AEDs, a subset of the group will have regular and sustained seizure activity. These seizures are considered refractory to treatment.Same as above. Accordingly, there remains a need for improved and/or additional therapies for treating seizure disorders.

Designer receptors (dreddss) activated exclusively by designer drugs, also known as RASSLs (receptors activated by synthetic ligands only), have recently been proposed for the treatment of epileptic seizures. See, for example, U.S. publication No. US 2016/0375097. Dreddss may involve engineered G protein-coupled signal receptors (GPCRs) activated only by synthetic ligands. These engineered receptors do not respond to endogenous ligands, but can still be activated by specific non-naturally occurring small molecule drugs. Two types of dreafds have been derived from human muscarinic acetylcholine receptors: hM3Dq which activates neuronal firing and hM4Di which inhibits neuronal firing. Typically, natural human muscarinic acetylcholine receptors are activated by the natural ligand acetylcholine. hM3Dq and hM4Di are not activated by natural ligands but by clozapine N-oxide (CNO), a pharmacologically inert ligand.

The Blood Brain Barrier (BBB) prevents many compounds in the blood stream from entering the tissues and fluids of the brain. The BBB is formed by brain-specific endothelial cells and is supported by cells of the neurovascular unit to restrict polar molecules or macromolecules such as proteins and peptides from entering or leaving the brain stroma. However, the BBB also prevents many therapeutic compounds from entering the brain, which can interfere with effective treatment of brain conditions and diseases. The BBB may interfere with the delivery of dredds and/or ligands to the brain, and thus reduce or prevent in vivo therapeutic benefit.

One method of aiding transport of therapeutic agents across the BBB involves the delivery of ultrasonic energy to the BBB, which "opens" the BBB and interferes with the BBB's ability to prevent transport of therapeutic agents into the brain. See, for example, U.S. patent No. 5,752,515, which relates to image-guided ultrasound delivery of compounds through the BBB. In one aspect, the change induced in Central Nervous System (CNS) tissue and/or fluid by ultrasound is by heating or cavitation. Such heating or cavitation can present disadvantages as it can cause damage to tissue and potentially degrade compounds delivered for therapeutic benefit. Sonication also causes degradation of organic compounds. See, for example, Brenner et al, Current Organic Chemistry,15(2):168-Human "). According to Brenner et al, when an aqueous solution is irradiated with ultrasound, H-O bonds in the water are homolytically cleaved to form hydroxyl radicals and hydrogen atoms. This process is the result of cavitation, thereby creating very high temperatures and pressures within the imploding bubble.Same as above. Thus, the use of ultrasound to attempt to open the BBB to cause or increase the delivery of therapeutic compounds such as dredds and ligands discussed above to the brain can degrade them and interfere with or prevent therapeutic treatment.

SUMMARY

Methods of treating a seizure disorder in a patient in need thereof are provided, the methods comprising administering to the patient a vector encoding a modified receptor for delivering the modified receptor to a target location, the modified receptor being modified to be activated by a synthetic ligand, wherein the modified receptor, when activated, inhibits neuronal discharge; and administering the synthetic ligand to the patient. In embodiments, the receptors are derived from human muscarinic acetylcholine receptors. In embodiments, the receptor is a modified G protein-coupled receptor. In embodiments, the modified receptor is hM4 Di. In embodiments, the modified receptor is KORD. Methods of treating a seizure disorder in a patient in need thereof are provided, the methods comprising administering to the patient a vector encoding a modified receptor for delivering the modified receptor to a target location, the modified receptor being modified to be activated by a synthetic ligand, wherein the modified receptor increases neuronal discharge when activated; and administering the synthetic ligand to the patient. In embodiments, the receptors are derived from human muscarinic acetylcholine receptors. In embodiments, the receptor is a modified G protein-coupled receptor. In embodiments, the modified receptor is hM3 Dq.

In embodiments, the vector includes a promoter, such as a CaMKII (also referred to herein as murine CaMKII) promoter. In embodiments, the vector includes a promoter, such as a human CaMKII (also referred to herein as human CaMK2a or hCaMK2a) promoter. In embodiments, the vector includes a post-transcriptional regulatory element, such as a woodchuck hepatitis virus post-transcriptional regulatory element (WPRE). In an embodiment, the vector comprises a polyadenylation sequence, such as the bovine growth hormone polyadenylation sequence (BGHpA).In embodiments, the vector comprises an Inverted Terminal Repeat (ITR). In embodiments, the vector includes an origin of replication, such as SV 40/Ori. In embodiments, the vector includes an origin of replication, such as pUC 19/Ori. In an embodiment, the vector comprises an ampicillin resistance gene (Amp)^R). In embodiments, the vector comprises a fluorescent reporter gene. In embodiments, the vector is pAM/CaMKII-hM4D (Gi) -WPRE-BGHpA. In embodiments, the vector is pAM/hCaMK2A-hM4D (Gi) -WPRE-BGHpA. In embodiments, the vector is an adeno-associated virus. In embodiments, the vector is a lentivirus.

In embodiments, the synthetic ligand is clozapine N-oxide. In embodiments, the synthetic ligand is perlapine (perlapine). In embodiments, the synthetic ligand is administered orally, buccally, sublingually, rectally, topically, intranasally, vaginally, or parenterally.

In embodiments, the vector is delivered to a target location in the brain of the patient. In embodiments, the target site is the frontal, temporal, occipital or parietal lobe. In embodiments, the route of administration of the carrier is oral, buccal, sublingual, rectal, topical, intranasal, vaginal, or parenteral. In embodiments, the vector is administered directly to the target location through the skull of the patient.

In embodiments, ultrasound is applied to a target location in the brain of a patient to enhance permeability of the blood brain barrier of the patient at the target location. A method of treating a seizure disorder in a patient in need thereof is provided, the method comprising applying ultrasound to a target location in the brain of the patient to enhance permeability of the blood-brain barrier of the patient at the target location; administering to the patient a vector encoding a modified receptor, the modified receptor modified to be activated by a synthetic ligand, for delivering the modified receptor to the target location, wherein the modified receptor, when activated, inhibits neuronal firing; and administering the synthetic ligand to the patient. In embodiments, ultrasound is administered prior to administering the carrier to the patient. In embodiments, the carrier is administered to the patient prior to administering ultrasound to the patient. In embodiments, ultrasound is administered to the patient prior to administration of the synthetic ligand. In embodiments, the method includes introducing a contrast agent into the patient, allowing the contrast agent sufficient time to penetrate the blood-brain barrier, and determining whether the contrast agent is present in the target location.

A method of treating a seizure disorder in a patient in need thereof is provided, the method comprising applying ultrasound to a target location in the brain of the patient to enhance permeability of the blood-brain barrier of the patient at the target location; administering to the patient a vector encoding a modified receptor, the modified receptor modified to be activated by a synthetic ligand, wherein the modified receptor increases neuronal firing when activated, for delivery of the modified receptor to the target location; and administering the synthetic ligand to the patient. In embodiments, ultrasound is administered prior to administering the carrier to the patient. In embodiments, the carrier is administered to the patient prior to administering ultrasound to the patient. In embodiments, ultrasound is administered to the patient prior to administration of the synthetic ligand.

In embodiments, administration of the vector and synthetic ligand to a patient having a seizure disorder is associated with reduced symptoms of the seizure disorder. In embodiments, the seizure disorder is characterized by focal seizures. In embodiments, the seizure disorder is focal cortical dysplasia. In embodiments, the seizure disorder is epilepsy, epilepsy with generalized tonic clonic seizures, epilepsy with myoclonic absence, frontal lobe epilepsy, temporal lobe epilepsy, occipital lobe epilepsy, parietal lobe epilepsy, Landau-Kleffner syndrome, Rasmussen syndrome, Dravet syndrome, dosse syndrome, CDKL5 disorder, infantile spasms (West syndrome), Juvenile Myoclonic Epilepsy (JME), vaccine-related encephalopathy, refractory childhood epilepsy (ICE), Lennox-gastatut syndrome (LGS), Rett syndrome, Ohtahara syndrome, CDKL5 disorder, childhood absence epilepsy, essential tremor, acute recurrent seizures, benign rolandolandole epilepsy, status epilepticus, refractory status epilepticus, super-refractory status epilepticus (se), srdh status epilepticus, pc 19 pediatric seizures, brain tumor-induced seizures, hamartures, hamartoma induced seizures, Drug withdrawal-induced seizures, alcohol withdrawal-induced seizures, increased seizure activity, or paroxysmal seizures. In embodiments, the seizure disorder is associated with a type 1 sodium channel protein alpha subunit (Scn1 alpha) -related disorder.

Brief Description of Drawings

FIG. 1 is a plasmid map of pAM/CaMKII-hM4D (Gi) -WPRE-BGHpA.

FIG. 2A, FIG. 2B and FIG. 2C depict the nucleotide sequence of pAM/CaMKII-hM4D (Gi) -WPRE-BGHpA [ SEQ ID NO:1 ].

FIG. 3 depicts the amino acid sequence of AAVRec3 [ SEQ ID NO:2 ].

FIG. 4 is a plasmid map of pAM/hCaMK2A-hM4D (Gi) -WPRE-BGHpA.

FIG. 5A, FIG. 5B and FIG. 5C depict the nucleotide sequence of pAM/hCaMK2A-hM4D (Gi) -WPRE-BGHpA [ SEQ ID NO:5 ].

FIG. 6A, FIG. 6B, FIG. 6C, FIG. 6D, FIG. 6E, FIG. 6F, FIG. 6G, FIG. 6H, FIG. 6I, FIG. 6J and FIG. 6K are schematic diagrams showing the correspondence between the positions and nucleotide sequences of the components of pAM/hCaMK2A-hM4D (Gi) -WPRE-BGHpA.

Detailed Description

Described herein are methods and compositions for treating seizure disorders, the methods and compositions comprising administering to a patient a vector encoding a modified receptor, the modified receptor modified to be activated by a synthetic ligand, for delivering the modified receptor to a target location, wherein the modified receptor, when activated, inhibits neuronal discharge; and administering the synthetic ligand to the patient. Also described herein are methods and compositions for treating seizure disorders in a patient in need thereof by administering to the patient a vector encoding a modified receptor, the modified receptor modified to be activated by a synthetic ligand, for delivering the modified receptor to a target location, wherein the modified receptor increases neuronal discharge when activated; and administering the synthetic ligand to the patient. In embodiments, the modified receptor is hM4 Di. In embodiments, a vector encoding hM4Di is administered to a patient having a seizure disorder. In embodiments, the modified receptor is hM3 Dq. In embodiments, a vector encoding hM3Dq is administered to a patient having a seizure disorder. In embodiments, the modified receptor is KORD. In embodiments, the vector encoding KORD is administered to a patient suffering from a seizure disorder. In embodiments, methods and compositions for treating seizure disorders include applying ultrasound to a target location in the brain of a patient to enhance permeability of the blood-brain barrier of the patient at the target location.

In embodiments, the modified receptor herein is derived from a naturally occurring receptor that is normally activated by an endogenous ligand. Naturally occurring receptors are modified so as not to be activated by endogenous ligands, but by synthetic ligands. These modified receptors are referred to as designer receptors activated exclusively by designer drugs (dreards) or receptors activated only by synthetic ligands (RASSLs). Dreards and RASSLS are referred to herein interchangeably.

In embodiments, the modified receptors herein are derived from naturally occurring muscarinic receptors that are normally activated by the endogenous ligand acetylcholine. Naturally occurring receptors are modified so as not to be activated by endogenous ligands, but by the synthetic ligand clozapine-N-oxide (CNO) or piperapine. These DREADDs allow passage through G_q(hM3Dq) and G_i(hM4Di) non-invasive control of neuronal signaling by the G-protein coupled signaling pathway. hM3Dq originates from the human muscarinic acetylcholine M3 receptor and activates neuronal firing. Without wishing to be bound by any theory, it is believed that hM3Dq activates neuronal firing following CNO or piperapin stimulation, in part, by depolarization and elevation of intracellular calcium levels. Plasmid encoding hM3Dq (pcDNA5/FRT-HA-hM3D (Gq)) is commercially available as plasmid 45547 from Addge, 75 Sidney Street, Cambridge, Massachusetts 02139. hM4Di is derived from the human muscarinic acetylcholine M4 receptor and inhibits neuronal firing. Without wishing to be bound by any theory, it is believed that hM4Di inhibits neuronal firing after CNO or piperapin stimulation via activation of the potassium orthodromic (GIRK) channel within the G protein. When bound by CNO, for example, membrane hyperpolarization is produced by a decrease in cAMP signaling and increased activation of inward rectifying potassium channels. This produces a temporary inhibition of neuronal activity. Plasmid encoding hM4Di (pcDNA5/FRT-HA-hM4D (Gi)) is commercially available as plasmid 45548 from Addge, 75 Sidney Street, Cambridge, Massachusetts 02139.Another modified receptor herein is G derived from the kappa-opioid receptor (KOR)_iConjugated DREADD, the kappa-opioid receptor (KOR) being activated only by the synthetic ligand salvinorin B (SALB). The modified receptor is referred to as KORD. KORD is insensitive to endogenous opioid agonists. Activation of KORD inhibits neuronal firing. See, Vardy et al, Neuron,86(4) (2015)936 and 946.

Many other G-protein coupled receptors are known to activate GIRK and may be used herein, including, for example, M₂-muscarinic receptors, a₁-adenosine receptor, alpha₂-adrenergic receptor, D₂-dopamine receptor, mu-opioid receptor and delta-opioid receptor, 5-HT_1ASerotonin receptor, somatostatin receptor, galanin receptor, m-Glu receptor, GABA_BReceptors and sphingosine-1-phosphate receptors.

Any suitable vector known to those skilled in the art can be used to deliver the modified receptors herein to a target location in the brain. Following such delivery, neurons in the target site are transfected with the modified receptor and can respond to the synthetic ligand. In embodiments, the neurons in the target location are transfected with hM3Dq, hM4Di, or KORD by any suitable vector known to those skilled in the art.

Nucleic acid constructs for expressing hM3Dq, hM4Di, or KORD may be recombinantly produced. Such expression vectors are provided herein. An expression vector is a vector nucleic acid (carrier nucleic acid) into which a nucleic acid sequence can be inserted for introduction into a cell where it can be replicated. Expression vectors include plasmids, cosmids, recombinant viruses such as adeno-associated virus (AAV), adenovirus, retrovirus, poxvirus, and other viruses known in the art (phage, animal, and plant viruses), and artificial chromosomes (e.g., YAC). One of ordinary skill in the art is well equipped to construct expression vectors by standard recombinant techniques. In embodiments, an expression vector comprising a nucleic acid encoding hM3Dq, hM4Di, or KORD is delivered to cells of the patient. The nucleic acid molecules are delivered to the cells of the patient in a form in which they can be taken up and advantageously expressed, such that therapeutically effective levels can be achieved. Following such delivery, neurons in the target site are transfected with hM3Dq, hM4Di, or KORD and can respond to synthetic ligands such that a therapeutically effective level of activation or inhibition can be achieved.

In embodiments, the vector may be a stable integrative vector or a stable non-integrative vector. Examples of suitable vectors are lentiviruses and adeno-associated viruses (AAV). Lentiviruses are a subset of retroviruses. Lentiviruses can integrate into the genome of non-dividing cells such as neurons. Lentiviruses are characterized by high efficiency of infection, long-term stable expression of transgenes, and low immunogenicity. In embodiments, lentiviral vectors may be used to deliver hM3Dq, hM4Di, or KORD receptor genes to the brain.

AAV is a defective parvovirus known to infect a variety of cell types and is not pathogenic to humans. AAV can infect both dividing and non-dividing cells. In embodiments, AAV vectors may be used herein to deliver hM3Dq, hM4Di, or KORD receptor genes to the brain. Any known adeno-associated virus (AAV) can be used herein, for example, AAV1, AAV2, AAV4, AAV5, AAV8, and AAV9 can be used in conjunction with neurons. AAVRec3 can also be used in conjunction with neurons. Additional suitable AAV serotypes have been developed by pseudotyping, i.e. mixing the capsid and genome from different viral serotypes. Thus, for example, AAV2/7 is indicative of a virus containing the genome of serotype 2 packaged in a capsid from serotype 7. Other examples are AAV2/5, AAV2/8, AAV2/9, and the like. Self-complementary adeno-associated viruses (scAAV) can also be used as vectors. Although AAV packages a single strand of DNA and requires a process for second strand synthesis, scAAV packages anneal together to form the two strands of double stranded DNA. scAAV allows rapid expression in cells by skipping second strand synthesis.

Suitable vectors can be constructed by one of ordinary skill in the art using known techniques. Suitable vectors may be selected or constructed to contain, in addition to the gene encoding the modified receptor, appropriate regulatory sequences including promoter sequences, terminator fragments, polyadenylation sequences, marker genes and other appropriate sequences. Those skilled in the art are familiar with appropriate regulatory sequences, including promoter sequences, terminator fragments, polyadenylation sequences, marker genes, and other appropriate sequences. The expression vectors herein include appropriate sequences operably linked to the coding sequence or ORF to facilitate expression thereof in the targeted host cell. "operably linked" sequences include both expression control sequences, such as a promoter adjacent to the coding sequence, and expression control sequences that act in trans or distally to control the expression of the desired product.

Typically, the vector includes a promoter that facilitates expression of the DNA encoding the modified receptor in the target cell. The promoter may be selected from a number of constitutive or inducible promoters that can drive expression of the selected transgene in the brain. Examples of constitutive promoters include the murine or human CaMKII promoter, the CMV immediate early enhancer/chicken β -actin (CBA) promoter-exon 1-intron 1 element, the RSV LTR promoter/enhancer, the SV40 promoter, the CMV promoter, the dihydrofolate reductase (DHFR) promoter, and the phosphoglycerate kinase (PGK) promoter.

Specificity can be achieved exclusively by regional and cell type specific expression of the receptor, for example using tissue or region specific promoters. Viral gene promoter elements may help determine the type of cell expressing a modified receptor such as hM3Dq, hM4Di, or KORD. Some promoters are non-specific (e.g., CAG, a synthetic promoter) while others are neuron specific (e.g., synapsin; hSyn), or are preferential to a particular neuron type, e.g., dynorphin, enkephalin, GFAP (glial fibrillary acidic protein) over astrocytes, or CaMKIIa or hCaM2a over corticoglutaergic cells but may also target gabaergic cells under the cortex. CAG promoters are strong synthetic promoters that can be used to drive high levels of expression. The CAG promoter consists of: 1) cytomegalovirus (CMV) early enhancer element, 2) the promoter, first exon, and first intron of the chicken β -actin gene, and 3) the splice acceptor of the rabbit β -globin gene.

In embodiments, the promoter is a murine or human camkli (α CaM kinase II gene) promoter that can drive expression in the forebrain. In embodiments, the CaMKII promoter is derived from murine α -calponin-dependent kinase ii (CaMKII), a gene that expresses excitatory neurons in the neocortex and hippocampus. As used herein, unless otherwise indicated, "CaMKII" refers to murine CaMKII. In embodiments, the CaMKII promoter is derived from human α -calcium/calmodulin-dependent kinase ii (hcamkii). The hCaMKII promoter is also referred to herein as the "human CaMK2A promoter" or the "hCaMK 2A promoter". Other neuronal cell type specific promoters include the NSE promoter, the tyrosine hydroxylase promoter, the myelin basic protein promoter, the glial fibrillary acidic protein promoter, and the neural filament gene (heavy, medium, light) promoter.

Expression control sequences may also include appropriate transcription initiation, termination and enhancer sequences; highly efficient processing signals, such as splicing and polyadenylation signals; a sequence that stabilizes cytoplasmic nucleic acid; sequences that enhance translation efficiency (e.g., Kozak consensus sequence); sequences that enhance the stability of a nucleic acid or protein; and, when desired, sequences that enhance product processing and/or secretion. Many varied expression control sequences, including natural and non-natural, constitutive, inducible and/or tissue-specific, are known in the art and may be used herein depending on the type of expression desired.

In addition to promoters, expression control sequences for eukaryotic cells typically include enhancers such as those derived from immunoglobulin genes, SV40, CMV, and the like, and polyadenylation sequences that may include splice donor and splice acceptor sites. Polyadenylation sequences will typically be inserted 3' into the coding sequence and 5 ' into the 3' ITR sequence. Illustrative examples of poly a signals that can be used in the vectors herein include a poly a sequence (e.g., AATAAA, ATTAAA, or AGTAAA), a bovine growth hormone poly a sequence (BGHpA), a rabbit β -globin poly a sequence (r β gpA), or another suitable heterologous or endogenous poly a sequence known in the art.

Regulatory sequences useful herein may also comprise introns, such as an intron located between the promoter/enhancer sequence and the coding sequence. One useful intron sequence is derived from SV40 and is referred to as the SV 40T intron sequence. Another includes woodchuck hepatitis virus post-transcriptional element (WPRE). WPRE is a DNA sequence that when transcribed yields a tertiary structure that enhances expression.

The vectors herein may comprise reporter genes, such as those encoding fluorophores. Fluorophores are fluorescent compounds that can re-emit light, usually at a specific frequency, after excitation. They may be used as tags or markers that can be attached to, for example, proteins to allow the proteins to be localized. Many suitable fluorophores are known in the art. The fluorophores may be classified by the color they emit, e.g., blue, cyan, green, yellow, orange, red, and others. For example, mCherry, mRasberry, mTomato and mRuby are red fluorophore proteins; citrine, venus and EYFP are yellow fluorophore proteins. Green Fluorescent Protein (GFP) is a commonly used fluorophore.

The following vectors include hM3Dq or hM4Di modified receptors, which hM3Dq or hM4Di modified receptors are suitable for use herein and are commercially available from addge, 75 Sidney Street, Cambridge, Massachusetts 02139: pAAV-GFAP-hM4D (Gi) -mCherry-Gi-conjugated hM4DDREADD fused to mCherry under the control of the GFAP promoter (50479); pAAV-GFAP-hM3D (Gq) -mCherry-Gq-conjugated hM3D DREADD fused to mCherry under the control of the GFAP promoter (50478); pAAV-CaMKIIa-hM4D (Gi) -mCherry-Gi-coupled hM4D DREADD fused to mCherry under the control of the CaMKIIa promoter (50477); pAAV-CaMKIIa-hM3D (Gq) -mCherry-Gq-conjugated hM3D DREADD fused to mCherry under the control of the CaMKIIa promoter (50476); pAAV-hSyn-hM4D (Gi) -mCherry-Gi-coupled hM4DDREADD fused to mCherry under the control of the human synaptrin promoter (50475); pAAV-hSyn-hM3D (Gq) -mCherry-Gq-coupled hM3D DREADD fused to mCherry under the control of the human synapsin promoter (50474); pAAV-GFAP-HA-hM4D (Gi) -IRES-mCitrine-Gi-conjugated hM4D-IRES-mCitrine (50471) under the control of the GFAP promoter, containing an HA tag; pAAV-GFAP-HA-hM3D (Gq) -IRES-mCitrine-Gq-coupled hM3D-IRES-mCitrine (50470) under control of the GFAP promoter-containing an HA tag; pAAV-CaMKIIa-HA-hM4D (Gi) -IRES-mCitrine-Gi-coupled hM4D-IRES-mCitrine (50467) under the control of the CaMKIIa promoter; pAAV-CaMKIIa-HA-hM3D (Gq) -IRES-mCitrine-Gq-coupled hM3D-IRES-mCitrine (50466) under the control of a CaMKIIa promoter; pAAV-hSyn-HA-hM4D (Gi) -IRES-mCitrine-Gi-conjugated hM4D-IRES-mCitrine (50464) under the control of human synapsin promoter; and pAAV-hSyn-HA-hM3D (Gq) -IRES-mCitrine-Gq-coupled hM3D-IRES-mCitrine (50463) under the control of the human synapsin promoter. Human influenza Hemagglutinin (HA) is a surface glycoprotein required for infectivity of human influenza viruses. The HA tag is derived from an HA-molecule corresponding to amino acids 98-106. It has been widely used as a general epitope tag in expression vectors.

In embodiments, the expression vector is pAM/CaMKII-hM4D (Gi) -WPRE-BGHpA. A plasmid map of pAM/CaMKII-hM4D (Gi) -WPRE-BGHpA is depicted in FIG. 1. The nucleic acid sequence [ SEQ ID NO:1] is shown in FIG. 2A, FIG. 2B and FIG. 2C. Table I notes pAM/CaMKII-hM4D (Gi) -WPRE-BGHpA.

TABLE I

To construct pAM/CaMKII-hM4D (Gi) -WPRE-BGHpA, a fragment containing the full-length hM4D (Gi) coding region (1,437bp) from the pAAV-CaMKIIa-hM4D (Gi) -mCherry plasmid (Cat. No.: 50477, Deposing lab Bryan Roth), commercially available from AddGene,75 Sidney street, Cambridge, Massachusetts 02139, was amplified by PCR using the following primers:

ATC TAG GAA TTC ATG GCC AAC TTC ACA CCT GTC[SEQ ID NO:3]forward direction

ATC TAG AAG CTT CTA CCT GGC AGT GCC GAT GTT CCG[SEQ ID NO:4]Reverse direction

The forward primer was extended with an EcoRI restriction site and with 6 protecting nucleotides. The reverse primer was extended with the original TAG stop codon (reverse complement-CTA) of hM4D, followed by a HindIII restriction site and 6 protected nucleotides. The 1,461bp PCR fragment was cut with EcoRI + HindIII and inserted into the plasmid pAM/CaMKII-pL-WPRE-bGHpA cut with the same restriction enzymes. The integrity of the expression cassette was confirmed by sequencing.

In embodiments, the expression vector is pAM/hCaMK2A-hM4D (Gi) -WPRE-BGHpA. A plasmid map of pAM/hCaMK2A-hM4D (Gi) -WPRE-BGHpA is depicted in FIG. 4. The nucleic acid sequence [ SEQ ID NO:5] is shown in FIGS. 5A, 5B and 5C. Table II notes pAM/hCaMK2A-hM4D (Gi) -WPRE-BGHpA.

TABLE II

FIGS. 6A-6K are schematic diagrams showing the correspondence between the positions and nucleotide sequences of the components of pAM/hCaMK2A-hM4D (Gi) -WPRE-BGHpA.

In embodiments, the pAM/CaMKII-hM4D (Gi) -WPRE-BGHpA vector or the pAM/hCaMK2A-hM4D (Gi) -WPRE-BGHpA vector described herein is used with a synthetic ligand to treat seizure disorders. Seizure disorders, including those involving complex partial seizures such as Temporal Lobe Epilepsy (TLE), are perhaps one of the most refractory forms of epilepsy. In some cases, one temporal lobe may be defined as the site of seizure origin (epileptogenic region), and the medial temporal lobe, including the anterior hippocampus, may be targeted according to the methods herein. Seizure disorders can be caused by an imbalance of excitation and suppression. The increased antagonism and inhibition of arousal may provide an improvement in at least one symptom of the seizure disorder.

Examples of seizure disorders include epilepsy, epilepsy with generalized tonic clonic seizures, epilepsy with myoclonic loss, frontal lobe epilepsy, temporal lobe epilepsy, Landau-Kleffner syndrome, Rasmussen syndrome, Dravet syndrome, Doose syndrome, CDKL5 disorder, infantile spasms (West syndrome), Juvenile Myoclonic Epilepsy (JME), vaccine-related encephalopathy, refractory childhood epilepsy (ICE), Lennox-Gastaut syndrome (LGS), Rett syndrome, Ohtahara syndrome, CDKL5 disorder, childhood absence epilepsy, essential tremor, acute recurrent seizures, benign Rolando-de-epileptic epilepsy, status epilepticus, refractory status epilepticus, super-refractory status epilepticus (SRSE), PC 19 epilepsy, brain tumor-induced seizures, hamarture-induced seizures, drug withdrawal-induced seizures, alcohol-induced seizures, withdrawal-induced seizures, drug-induced seizures, alcohol-induced seizures, Increased seizure activity or paroxysmal seizures (also known as continuous or clustered seizures). In embodiments, the seizure disorder is associated with a type 1 sodium channel protein alpha subunit (Scn1 alpha) -related disorder. In embodiments, the seizure disorder is characterized by focal seizures. In embodiments, the seizure disorder is focal cortical dysplasia. In embodiments, the FCD is a type I FCD. In embodiments, the FCD is FCD type Ia. In embodiments, FCD is type Ib FCD. In embodiments, the FCD is a type Ic FCD. In embodiments, the FCD is a type II FCD. In embodiments, FCD is FCD type IIa. In an embodiment, the FCD is a type IIb FCD. In embodiments, the FCD is a type III FCD. In embodiments, the FCD is a type IIIa FCD. In embodiments, the FCD is a type IIIb FCD. In embodiments, the FCD is a type IIIc FCD. In embodiments, the seizure disorder is associated with a brain tumor, i.e., a brain tumor-induced seizure, such as a ganglioglioma, glioma-low and high grade (including anaplastic astrocytoma, anaplastic oligodendroglioma, anaplastic oligoastrocytoma, and anaplastic ependymoma), glioblastoma, or meningioma. In embodiments, the seizure disorder is associated with a hamartoma, i.e., a hamartoma-induced seizure, such as Tuberous Sclerosis Complex (TSC) or gray hamartoma.

In embodiments, the seizure disorder is Status Epilepticus (SE). SE is characterized by: epileptic seizures greater than five minutes, or more than one seizure over a five minute period without the patient recovering normal in the meantime. SE may be a dangerous condition that may lead to death if treatment is delayed. SE may be convulsive, with a regular pattern of arm and leg contractions and extensions, or non-convulsive, with a change in the level of consciousness of a person of relatively long duration, but without flexion and extension of the large-scale limbs due to seizure activity. Convulsive SE (cse) may also be classified as (a) tonic-clonic SE, (b) tonic SE, (c) clonic SE, and (d) myoclonic SE. Non-convulsive se (ncse) is characterized by abnormal mental states, unresponsiveness, abnormal eye movement, persistent electrographic seizures, and possible responses to anticonvulsants.

Symptoms of seizure disorders may include, but are not limited to, seizures involving (episode): ataxia, gait disorders, speech disorders, vocalization, involuntary laughing, impaired cognitive function, abnormal motor activity, clinical seizures, subclinical seizures, hypotonia, hypertonia, watery mouth, orodental behavior, aura, repetitive motion, and unusual sensations. In embodiments, the provided methods and compositions can reduce or prevent one or more different types of seizures. Generally, a seizure may include repetitive motion, unusual sensations, and combinations thereof. Seizures can be classified as focal seizures (also known as partial seizures) and generalized seizures. Focal seizures affect only one side of the brain, while generalized seizures affect both sides of the brain. Specific types of focal seizures include simple focal seizures, complex focal seizures, and secondary generalized seizures. Simple focal seizures may be confined or concentrated to a particular lobe (e.g., temporal, frontal, parietal or occipital). Complex focal seizures usually affect a greater part of one hemisphere than simple focal seizures, but usually originate in the temporal or frontal lobe. When focal seizures spread from one side of the brain (hemisphere) to both sides of the brain, the seizures are referred to as secondary generalized seizures. Specific types of generalized seizures include ataxia (also known as petit mal), tonic seizures, dystonia, myoclonic seizures, tonic clonic seizures (also known as grand mal), and clonic seizures. The methods of treatment herein may include providing an improvement in one or more of the foregoing symptoms.

Targeted therapy according to the present disclosure may be implemented after the location or suspected location of the abnormal electrical pulse associated with the seizure disorder in the patient has been determined. Methods of determining the location of abnormal electrical activity in the brain are well known in the art. Although any region in the brain that exhibits abnormal electricity may be targeted for treatment herein, regions of the brain known to be associated with seizure disorders and that may receive targeted therapy include, but are not limited to, the temporal lobe, frontal lobe, occipital lobe, and parietal lobe. For example, the temporal lobe may be a common site of a localized epileptic seizure. In some cases, seizures that begin at the temporal lobe may extend to other parts of the brain. In embodiments, specific regions of the temporal lobe that may be targeted for treatment include structures of the limbic system, such as the hippocampus, auditory-vestibular cortex, medial temporal lobe, and amygdala. In embodiments, specific regions of the occipital lobe, such as the primary visual cortex, may also be targeted. In embodiments, a specific region of the apical lobe, such as the lateral central posterior gyrus, may be targeted. In embodiments, the location of the primary somatosensory cortex may be targeted. In embodiments, specific regions of the frontal lobe, such as the motor cortex, olfactory-gustatory cortex, may be targeted. In embodiments, large regions of the brain that have been identified as exhibiting abnormal electrical activity may be targeted. In some cases, the manifestation of seizure disorders may begin in certain areas of the brain and spread to other areas. For example, the manifestation of seizure disorders may begin in the hippocampus or its surrounding structures. In embodiments, a region determined to be the site of origin of the aberrant electrical activity may be targeted.

Methods for administering materials directly to a target location within the brain are well known. For example, a burr hole, such as a burr hole, may be drilled into the skull and an appropriately sized needle may be used to deliver the vector to the target location. In embodiments, a portion of the skull can be removed to expose dural matter at or near the target location (craniotomy), and the carrier can be administered directly to the target location. In embodiments, the vector is injected intracranially using stereotactic coordinates, micropipettes, and automated pumps for precise delivery of the vector to a desired area with minimal damage to surrounding tissue. In embodiments, the micropump may be used to deliver a pharmaceutical composition comprising a vector encoding hM3Dq, hM4Di, or KORD to a target region in the brain. The composition may be delivered immediately or over an extended period of time, e.g., over 1 minute, 2 minutes, 3 minutes, 4 minutes, 5 minutes, 6 minutes, 7 minutes, 8 minutes, 9 minutes, 10 minutes, or more. After delivery of the vector to the target location in the brain, a sufficient amount of time may be allowed to elapse to allow expression of hM3Dq, hM4Di, or KORD at the target location.

In embodiments, a pharmaceutical composition comprising a carrier herein may be administered systemically. Systemic delivery includes oral, buccal, sublingual, rectal, topical, intranasal, vaginal and parenteral modes of administration. Examples of parenteral administration modes include intravenous administration mode, intraperitoneal administration mode, intramuscular administration mode, and subcutaneous administration mode. In embodiments, the vector will circulate until it contacts a target location in the brain where it delivers a nucleic acid encoding hM3Dq, hM4Di, or KORD and when activated by the synthetic ligand causes hM3Dq, hM4Di, or KORD to be expressed and function, e.g., to modulate a neuronal signaling network.

As previously mentioned, synthetic ligands herein include clozapine N-oxide (CNO), piperapine. CNO can also be referred to as 8-chloro-11- (4-methyl-1-piperazinyl) -5H-dibenzo [ b, e ] (1,4) diazepine N-oxide. In embodiments, the CNO may be administered directly to a target location in the brain by any known means for administering materials to the brain, such as direct injection. In embodiments, the CNO may be administered systemically to the patient. Systemic delivery includes oral, buccal, sublingual, rectal, topical, intranasal, vaginal and parenteral modes of administration. Examples of parenteral administration modes include intravenous administration mode, intraperitoneal administration mode, intramuscular administration mode, and subcutaneous administration mode.

Suitable effective CNO doses may vary based on the expression level of the DREADD receptor, the type of cell infected, the duration of DREADD activation desired, and the species being treated. In embodiments, the CNO administration may be in the range of from 0.1mg/kg to 50 mg/kg. In embodiments, a pharmaceutical composition containing CNO for treating seizure disorders may comprise, for example, CNO in the following amounts: about 0.01mg to 500mg, 0.1mg to 450mg, 0.1mg to 300mg, 0.1mg to 250mg, 0.1mg to 200mg, 0.1mg to 175mg, 0.1mg to 150mg, 0.1mg to 125mg, 0.1mg to 100mg, 0.1mg to 75mg, 0.1mg to 50mg, 0.1mg to 30mg, 0.1mg to 25mg, 0.1mg to 20mg, 0.1mg to 15mg, 0.1mg to 10mg, 0.1mg to 5mg, 0.1mg to 1mg, 0.5mg to 500mg, 0.5mg to 450mg, 0.5mg to 300mg, 0.5mg to 250mg, 0.5mg to 200mg, 0.5mg to 175mg, 0.5mg to 150mg, 0.5mg to 125mg, 0.5mg to 100mg, 0.1mg to 5mg, 0.1mg to 25mg, 0.1mg to 5mg, 0.1mg to 25mg, 0.1, 1mg to 150mg, 1mg to 125mg, 1mg to 100mg, 1mg to 75mg, 1mg to 50mg, 1mg to 30mg, 1mg to 25mg, 1mg to 20mg, 1mg to 15mg, 1mg to 10mg, 1mg to 5mg, 5mg to 500mg, 5mg to 450mg, 5mg to 300mg, 5mg to 250mg, 5mg to 200mg, 5mg to 175mg, 5mg to 150mg, 5mg to 125mg, 5mg to 100mg, 5mg to 75mg, 5mg to 50mg, 5mg to 30mg, 5mg to 25mg, 5mg to 20mg, 5mg to 15mg, 5mg to 10mg, 10mg to 500mg, 10mg to 450mg, 10mg to 300mg, 10mg to 250mg, 10mg to 200mg, 10mg to 175mg, 10mg to 150mg, 10mg to 125mg, 10mg to 100mg, 10mg to 25mg, 10mg to 500mg, 10mg to 15mg, 10mg to 10mg, 10mg to 25mg, 10mg, 5mg to 25mg, 5mg, 15mg to 450mg, 15mg to 300mg, 15mg to 250mg, 15mg to 200mg, 15mg to 175mg, 15mg to 150mg, 15mg to 125mg, 15mg to 100mg, 15mg to 75mg, 15mg to 50mg, 15mg to 30mg, 15mg to 25mg, 15mg to 20mg, 20mg to 500mg, 20mg to 450mg, 20mg to 300mg, 20mg to 250mg, 20mg to 200mg, 20mg to 175mg, 20mg to 150mg, 20mg to 125mg, 20mg to 100mg, 20mg to 75mg, 20mg to 50mg, 20mg to 30mg, 20mg to 25mg, 25mg to 500mg, 25mg to 450mg, 25mg to 300mg, 25mg to 250mg, 25mg to 200mg, 25mg to 175mg, 25mg to 150mg, 25mg to 125mg, 25mg to 100mg, 25mg to 80mg, 25mg to 300mg, 25mg to 30mg, 30mg to 250mg, 25mg to 175mg, 25mg to 25mg, 30mg, 25mg to 25mg, 30mg to 200mg, 30mg to 175mg, 30mg to 150mg, 30mg to 125mg, 30mg to 100mg, 30mg to 75mg, 30mg to 50mg, 40mg to 500mg, 40mg to 450mg, 40mg to 400mg, 40mg to 250mg, 40mg to 200mg, 40mg to 175mg, 40mg to 150mg, 40mg to 125mg, 40mg to 100mg, 40mg to 75mg, 40mg to 50mg, 50mg to 500mg, 50mg to 450mg, 50mg to 300mg, 50mg to 250mg, 50mg to 200mg, 50mg to 175mg, 50mg to 150mg, 50mg to 125mg, 50mg to 100mg, 50mg to 75mg, 75mg to 500mg, 75mg to 450mg, 75mg to 300mg, 75mg to 250mg, 75mg to 200mg, 75mg to 150mg, 75mg to 125mg, 100mg to 100mg, 100mg to 250mg, 100mg, 40mg to 250mg, 40mg to 200mg, 40mg, 100mg to 150mg, 100mg to 125mg, 125mg to 500mg, 125mg to 450mg, 125mg to 300mg, 125mg to 250mg, 125mg to 200mg, 125mg to 175mg, 125mg to 150mg, 150mg to 500mg, 150mg to 450mg, 150mg to 300mg, 150mg to 250mg, 150mg to 200mg, 200mg to 500mg, 200mg to 450mg, 200mg to 300mg, 200mg to 250mg, 250mg to 500mg, 250mg to 450mg, 250mg to 300mg, 300mg to 500mg, 300mg to 450mg, 300mg to 400mg, 300mg to 350mg, 350mg to 500mg, 350mg to 450mg, 350mg to 400mg, 400mg to 500mg, 400mg to 450mg, wherein 0.1mg, 0.25mg, 0.5mg, 0.75mg, 1mg, 2.5mg, 5mg, 7.5mg, 10.5 mg, 12mg, 15.5 mg, 15mg, 5.5 mg, 15.5 mg, 35mg, 45mg, 35.5 mg, 35mg, 70mg, 25mg, 70mg, 25mg, 70mg, 150mg, 80mg, 85mg, 90mg, 95mg, 100mg, 125mg, 150mg 175mg, 200mg, 225mg, 250mg, 275mg, 300mg, 325mg, 350mg, 375mg, 400mg, 425mg, 450mg, 475mg, and 500mg are examples. Suitable doses of CNO may be administered to a patient suffering from seizure disorders once daily, twice daily, three times daily, four times daily, five times daily, or six times daily, every other day, once weekly, or once monthly.

In embodiments, the CNO is administered to a patient suffering from a seizure disorder at a dose of 0.1mg-200 mg/administration twice a day (e.g., morning and evening) or three times a day (e.g., morning, noon, and bedtime). In embodiments, the CNO is administered in one or more doses of 1000 mg/day, 600 mg/day, 550 mg/day, 500 mg/day, 450 mg/day, 400 mg/day, 350 mg/day, 300 mg/day, 250 mg/day, 225 mg/day, 200 mg/day, 190 mg/day, 180 mg/day, 170 mg/day, 160 mg/day, 150 mg/day, 140 mg/day, 130 mg/day, 120 mg/day, 110 mg/day, 100 mg/day, 95 mg/day, 90 mg/day, 85 mg/day, 80 mg/day, 75 mg/day, 70 mg/day, 65 mg/day, 60 mg/day, 55 mg/day, 50 mg/day, 45 mg/day, 40 mg/day, 35 mg/day, 30 mg/day, 25 mg/day, 20 mg/day, 15 mg/day, 10 mg/day, 5 mg/day, 4 mg/day, 3 mg/day, 2 mg/day, 1 mg/day is administered to a patient suffering from a seizure disorder. In embodiments, the adult dose may be about 0.05mg to 500mg per day, and may be increased to 750mg per day. The dosage may be lower for infants and children than for adults. In embodiments, the infant or pediatric dose may be about 0.1mg to 50mg once a day or in 2, 3 or 4 divided doses. In embodiments, a pediatric dose may be 0.75 mg/kg/day to 1.5 mg/kg/day. In embodiments, the patient may start with a low dose and the dose is escalated over time.

Piperapine may also be administered as a synthetic ligand. Piperazine may also be referred to as 6- (4-methyl-1-piperazinyl) -11H-dibenzo [ b, e ] azepine, or 6- (4-methylpiperazin-1-yl) morphidine. In embodiments, the piperapine may be administered directly to a target location in the brain by any known means for administering materials to the brain, such as direct injection. In embodiments, the piperapine may be administered systemically to the patient. Systemic delivery includes oral, buccal, sublingual, rectal, topical, intranasal, vaginal and parenteral modes of administration. Examples of parenteral administration modes include intravenous administration mode, intraperitoneal administration mode, intramuscular administration mode, and subcutaneous administration mode.

Suitable effective dosages of piperapine may vary based on the expression level of the DREADD receptor, the type of cell infected, the desired duration of DREADD activation, and the species being treated. In embodiments, the piperapine administration may be in the range of from 0.01mg/kg to 1 mg/kg.

In embodiments, a pharmaceutical composition containing piperapine for the treatment of seizure disorders may comprise, for example, the following amounts of piperapine: 0.1 to 75mg, 0.1 to 70mg, 0.1 to 65mg, 0.1 to 55mg, 0.1 to 50mg, 0.1 to 45mg, 0.1 to 40mg, 0.1 to 35mg, 0.1 to 30mg, 0.1 to 25mg, 0.1 to 20mg, 0.1 to 15mg, 0.1 to 10mg, 0.5 to 75mg, 0.5 to 70mg, 0.5 to 65mg, 0.5 to 55mg, 0.5 to 50mg, 0.5 to 45mg, 0.5 to 40mg, 0.5 to 35mg, 0.5 to 30mg, 0.5 to 25mg, 0.5 to 20mg, 0.5 to 15mg, 0.5 to 10mg, 1 to 75mg, 1 to 70mg, 1 to 5mg, 1.5 to 5mg, 1 to 5mg, 1.5 to 55mg, 1 to 5mg, 1.5 to 5mg, 1mg to 5mg, 1.5 to 55mg, 1 to 5mg, 1mg to 5mg, 1mg to 55mg, 1 to 5mg, 1mg to 5mg, 1.5 to 45mg, 1.5 to 40mg, 1.5 to 35mg, 1.5 to 30mg, 1.5 to 25mg, 1.5 to 20mg, 1.5 to 15mg, 1.5 to 10mg, 2 to 75mg, 2 to 70mg, 2 to 65mg, 2 to 55mg, 2 to 50mg, 2 to 45mg, 2 to 40mg, 2 to 35mg, 2 to 30mg, 2 to 25mg, 2 to 20mg, 2 to 15mg, 2 to 10mg, 2.5 to 75mg, 2.5 to 70mg, 2.5 to 65mg, 2.5 to 55mg, 2.5 to 50mg, 2.5 to 45mg, 2.5 to 40mg, 2.5 to 35mg, 2.5 to 30mg, 2.5 to 25mg, 3 to 5mg, 3.5 to 35mg, 3 to 5mg, 3.5 to 5mg, 3 to 5mg, 3.5 to 35mg, 3 to 5mg, 3.5 to 5mg, 3mg, 3.5 to 5mg, 3mg to 70mg, 3mg, 5mg, 2 to 5mg, 5mg to 70mg, 5mg, 2 to 5mg, 2 to 70mg, and 70mg, 3mg to 25mg, 3mg to 20mg, 3mg to 15mg, 3mg to 10mg, 3.5mg to 75mg, 3.5mg to 70mg, 3.5mg to 65mg, 3.5mg to 55mg, 3.5mg to 50mg, 3.5mg to 45mg, 3.5mg to 40mg, 3.5mg to 35mg, 3.5mg to 30mg, 3.5mg to 25mg, 3.5mg to 20mg, 3.5mg to 15mg, 3.5mg to 10mg, 4mg to 75mg, 4mg to 70mg, 4mg to 65mg, 4mg to 55mg, 4mg to 50mg, 4mg to 45mg, 4mg to 40mg, 4mg to 35mg, 4mg to 30mg, 4mg to 25mg, 4mg to 20mg, 4mg to 15mg, 4mg to 10mg, 4.5mg to 75mg, 4.5mg to 70mg, 4.5mg to 5mg, 5mg to 5mg, 4.5mg to 5mg, 5mg to 5mg, 4.5mg to, 5mg to 75mg, 5mg to 70mg, 5mg to 65mg, 5mg to 55mg, 5mg to 50mg, 5mg to 45mg, 5mg to 40mg, 5mg to 35mg, 5mg to 30mg, 5mg to 25mg, 5mg to 20mg, 5mg to 15mg, or 5mg to 10mg, or a pharmaceutically acceptable salt thereof.

In embodiments, the pharmaceutical composition comprises 5mg to 20mg, 5mg to 10mg, 4mg to 6mg, 6mg to 8mg, 8mg to 10mg, 10mg to 12mg, 12mg to 14mg, 14mg to 16mg, 16mg to 18mg, or 18mg to 20mg piperapin or a pharmaceutically acceptable salt thereof.

In embodiments, the pharmaceutical composition comprises 0.1mg, 0.25mg, 0.5mg, 1mg, 2.5mg, 3mg, 4mg, 5mg, 7mg, 7.5mg, 10mg, 12.5mg, 15mg, 17.5mg, or 20mg of piperacillin or a pharmaceutically acceptable salt thereof or an amount that is a multiple of such a dose. In an embodiment, the pharmaceutical composition comprises 2.5mg, 5mg, 7.5mg, 10mg, 15mg, 20mg, 25mg, 30mg, 35mg, or 40mg of piperapin or a pharmaceutically acceptable salt thereof.

In embodiments, the composition comprises 0.05mg, 0.1mg, 0.25mg, 0.5mg, 0.75mg, 1mg, 1.25mg, 1.5mg, 1.75mg, 2mg, 2.5mg, 3mg, 3.5mg, 4mg, 4.5mg, 5mg, 7mg, 7.5mg, 10mg, 12.5mg, 15mg, 17.5mg, or 20mg of piperapin or a pharmaceutically acceptable salt thereof or an amount that is a multiple of such a dose.

In embodiments, the pharmaceutical composition comprises from about 0.05mg to about 100mg of piperacillin or a pharmaceutically acceptable salt thereof. In embodiments, the dosage form comprises 0.05mg, 0.1mg, 0.25mg, 0.5mg, 0.75mg, 1mg, 1.25mg, 1.5mg, 1.75mg, 2mg, 2.5mg, 3mg, 3.5mg, 4mg, 4.5mg, 5mg, 6mg, 7mg, 8mg, 9mg, 10mg, 11mg, 12mg, 13mg, 14mg, 15mg, 20mg, 25mg, 30mg, 35mg, 40mg, 45mg, 50mg, 55mg, 60mg, 65mg, 70mg, 75mg, 80mg, 85mg, 90mg, 95mg, or 100mg of piperacillin or a pharmaceutically acceptable salt thereof.

Suitable doses of piperapine may be administered to a patient suffering from a seizure disorder once daily, twice daily, three times daily, four times daily, five times daily, or six times daily, every other day, once weekly, or once monthly. In embodiments, the piperapine is administered to a patient suffering from a seizure disorder at a dose of 0.1mg to 50mg per administration twice a day (e.g., morning and evening) or three times a day (e.g., morning, noon, and bedtime). In embodiments, the piperapine is administered in one or more doses of 250 mg/day, 190 mg/day, 180 mg/day, 170 mg/day, 160 mg/day, 150 mg/day, 140 mg/day, 130 mg/day, 120 mg/day, 110 mg/day, 100 mg/day, 95 mg/day, 90 mg/day, 85 mg/day, 80 mg/day, 75 mg/day, 70 mg/day, 65 mg/day, 60 mg/day, 55 mg/day, 50 mg/day, 45 mg/day, 40 mg/day, 35 mg/day, 30 mg/day, 25 mg/day, 20 mg/day, 15 mg/day, 10 mg/day, 5 mg/day, 4 mg/day, 3 mg/day, 2 mg/day, 1 mg/day is administered to a patient suffering from seizure disorders. In embodiments, the adult dose may be about 0.05mg to 100mg per day, and may be increased to 200mg per day. The dosage may be lower for infants and children than for adults. In embodiments, the infant or pediatric dose may be about 0.1mg to 20mg once a day or in 2, 3 or 4 divided doses. In embodiments, a pediatric dose may be 0.75 mg/kg/day to 1.5 mg/kg/day. In embodiments, the patient may start with a low dose and the dose is escalated over time.

SALB may also be administered as a synthetic ligand that binds to KORD. In embodiments, the SALB may be administered directly to a target location in the brain by any known means for administering materials to the brain, such as direct injection. In embodiments, the SALB may be administered systemically to the patient. Systemic delivery includes oral, buccal, sublingual, rectal, topical, intranasal, vaginal and parenteral modes of administration. Examples of parenteral administration modes include intravenous administration mode, intraperitoneal administration mode, intramuscular administration mode, and subcutaneous administration mode.

Suitable effective SALB dosages may vary based on the level of DREADD receptor expression, the type of cell infected, the desired duration of DREADD activation, and the species being treated. In embodiments, the SALB administration may be in the range of from 0.01mg/kg to 20 mg/kg. In embodiments, a pharmaceutical composition containing SALB for treating seizure disorders may comprise, for example, SALB in the following amounts: about 0.01mg to 500mg, 0.1mg to 450mg, 0.1mg to 300mg, 0.1mg to 250mg, 0.1mg to 200mg, 0.1mg to 175mg, 0.1mg to 150mg, 0.1mg to 125mg, 0.1mg to 100mg, 0.1mg to 75mg, 0.1mg to 50mg, 0.1mg to 30mg, 0.1mg to 25mg, 0.1mg to 20mg, 0.1mg to 15mg, 0.1mg to 10mg, 0.1mg to 5mg, 0.1mg to 1mg, 0.5mg to 500mg, 0.5mg to 450mg, 0.5mg to 300mg, 0.5mg to 250mg, 0.5mg to 200mg, 0.5mg to 175mg, 0.5mg to 150mg, 0.5mg to 125mg, 0.5mg to 100mg, 0.1mg to 5mg, 0.1mg to 25mg, 0.1mg to 5mg, 0.1mg to 25mg, 0.1, 1mg to 150mg, 1mg to 125mg, 1mg to 100mg, 1mg to 75mg, 1mg to 50mg, 1mg to 30mg, 1mg to 25mg, 1mg to 20mg, 1mg to 15mg, 1mg to 10mg, 1mg to 5mg, 5mg to 500mg, 5mg to 450mg, 5mg to 300mg, 5mg to 250mg, 5mg to 200mg, 5mg to 175mg, 5mg to 150mg, 5mg to 125mg, 5mg to 100mg, 5mg to 75mg, 5mg to 50mg, 5mg to 30mg, 5mg to 25mg, 5mg to 20mg, 5mg to 15mg, 5mg to 10mg, 10mg to 500mg, 10mg to 450mg, 10mg to 300mg, 10mg to 250mg, 10mg to 200mg, 10mg to 175mg, 10mg to 150mg, 10mg to 125mg, 10mg to 100mg, 10mg to 25mg, 10mg to 500mg, 10mg to 15mg, 10mg to 10mg, 10mg to 25mg, 10mg, 5mg to 25mg, 5mg, 15mg to 450mg, 15mg to 300mg, 15mg to 250mg, 15mg to 200mg, 15mg to 175mg, 15mg to 150mg, 15mg to 125mg, 15mg to 100mg, 15mg to 75mg, 15mg to 50mg, 15mg to 30mg, 15mg to 25mg, 15mg to 20mg, 20mg to 500mg, 20mg to 450mg, 20mg to 300mg, 20mg to 250mg, 20mg to 200mg, 20mg to 175mg, 20mg to 150mg, 20mg to 125mg, 20mg to 100mg, 20mg to 75mg, 20mg to 50mg, 20mg to 30mg, 20mg to 25mg, 25mg to 500mg, 25mg to 450mg, 25mg to 300mg, 25mg to 250mg, 25mg to 200mg, 25mg to 175mg, 25mg to 150mg, 25mg to 125mg, 25mg to 100mg, 25mg to 80mg, 25mg to 300mg, 25mg to 30mg, 30mg to 250mg, 25mg to 175mg, 25mg to 25mg, 30mg, 25mg to 25mg, 30mg to 200mg, 30mg to 175mg, 30mg to 150mg, 30mg to 125mg, 30mg to 100mg, 30mg to 75mg, 30mg to 50mg, 40mg to 500mg, 40mg to 450mg, 40mg to 400mg, 40mg to 250mg, 40mg to 200mg, 40mg to 175mg, 40mg to 150mg, 40mg to 125mg, 40mg to 100mg, 40mg to 75mg, 40mg to 50mg, 50mg to 500mg, 50mg to 450mg, 50mg to 300mg, 50mg to 250mg, 50mg to 200mg, 50mg to 175mg, 50mg to 150mg, 50mg to 125mg, 50mg to 100mg, 50mg to 75mg, 75mg to 500mg, 75mg to 450mg, 75mg to 300mg, 75mg to 250mg, 75mg to 200mg, 75mg to 150mg, 75mg to 125mg, 100mg to 100mg, 100mg to 250mg, 100mg, 40mg to 250mg, 40mg to 200mg, 40mg, 100mg to 150mg, 100mg to 125mg, 125mg to 500mg, 125mg to 450mg, 125mg to 300mg, 125mg to 250mg, 125mg to 200mg, 125mg to 175mg, 125mg to 150mg, 150mg to 500mg, 150mg to 450mg, 150mg to 300mg, 150mg to 250mg, 150mg to 200mg, 200mg to 500mg, 200mg to 450mg, 200mg to 300mg, 200mg to 250mg, 250mg to 500mg, 250mg to 450mg, 250mg to 300mg, 300mg to 500mg, 300mg to 450mg, 300mg to 400mg, 300mg to 350mg, 350mg to 500mg, 350mg to 450mg, 350mg to 400mg, 400mg to 500mg, 400mg to 450mg, wherein 0.1mg, 0.25mg, 0.5mg, 0.75mg, 1mg, 2.5mg, 5mg, 7.5mg, 10.5 mg, 12mg, 15.5 mg, 15mg, 5.5 mg, 15.5 mg, 35mg, 45mg, 35.5 mg, 35mg, 70mg, 25mg, 70mg, 25mg, 70mg, 150mg, 80mg, 85mg, 90mg, 95mg, 100mg, 125mg, 150mg 175mg, 200mg, 225mg, 250mg, 275mg, 300mg, 325mg, 350mg, 375mg, 400mg, 425mg, 450mg, 475mg, and 500mg are examples. Suitable doses of SALB may be administered to a patient suffering from seizure disorders once daily, twice daily, three times daily, four times daily, five times daily, or six times daily, every other day, once weekly, or once monthly.

In embodiments, the SALB is administered to a patient suffering from a seizure disorder at a dose of 0.1mg-200 mg/administration twice a day (e.g., morning and evening) or three times a day (e.g., morning, noon, and bedtime). In embodiments, the SALB is administered in one or more doses of 1000 mg/day, 600 mg/day, 550 mg/day, 500 mg/day, 450 mg/day, 400 mg/day, 350 mg/day, 300 mg/day, 250 mg/day, 225 mg/day, 200 mg/day, 190 mg/day, 180 mg/day, 170 mg/day, 160 mg/day, 150 mg/day, 140 mg/day, 130 mg/day, 120 mg/day, 110 mg/day, 100 mg/day, 95 mg/day, 90 mg/day, 85 mg/day, 80 mg/day, 75 mg/day, 70 mg/day, 65 mg/day, 60 mg/day, 55 mg/day, 50 mg/day, 45 mg/day, 40 mg/day, 35 mg/day, 30 mg/day, 25 mg/day, 20 mg/day, 15 mg/day, 10 mg/day, 5 mg/day, 4 mg/day, 3 mg/day, 2 mg/day, 1 mg/day is administered to a patient suffering from a seizure disorder. In embodiments, the adult dose may be about 0.05mg to 500mg per day, and may be increased to 750mg per day. The dosage may be lower for infants and children than for adults. In embodiments, the infant or pediatric dose may be about 0.1mg to 50mg once a day or in 2, 3 or 4 divided doses. In embodiments, a pediatric dose may be 0.75 mg/kg/day to 1.5 mg/kg/day. In embodiments, the patient may start with a low dose and the dose is escalated over time.

In embodiments, there is provided a method of treating a seizure disorder, the method comprising 1) administering to a patient a vector encoding a modified receptor for delivering the modified receptor to a target location, the modified receptor being modified to be activated by a synthetic ligand, wherein the modified receptor, when activated, inhibits neuronal discharge (e.g., hM4Di or KORD); and administering to the patient a synthetic ligand, or 2) administering to the patient a vector encoding a modified receptor, for delivering the modified receptor to the target location, the modified receptor being modified to be activated by the synthetic ligand, wherein the modified receptor increases neuronal firing when activated (e.g., HM3 dq); and administering the synthetic ligand to the patient, wherein the treatment provides an improvement in one or more symptoms of the disorder that persist for more than 1 hour after administration to the patient. In embodiments, the treatment provides an improvement in one or more symptoms of the disorder that persist for more than 2 hours after administration of the synthetic ligand to the patient. In embodiments, the treatment provides an improvement in one or more symptoms of the disorder that persist for more than 3 hours after administration of the synthetic ligand to the patient. In embodiments, the treatment provides an improvement in one or more symptoms of the disorder that persist for more than 4 hours after administration of the synthetic ligand to the patient. In embodiments, the treatment provides an improvement in one or more symptoms of the disorder that persist for more than 6 hours after administration of the synthetic ligand to the patient. In embodiments, the treatment provides an improvement in one or more symptoms of the disorder that persist for more than 8 hours, 10 hours, 12 hours, 14 hours, 16 hours, 18 hours, 20 hours, 22 hours, or 24 hours after administration of the synthetic ligand to the patient. In embodiments, in accordance with the present disclosure, there is provided an improvement in at least one symptom that persists for 12 hours after administration of a synthetic ligand to a patient. In embodiments, administration of the synthetic ligand provides an improvement in the next day function of the patient. For example, administration of the synthetic ligand can provide an improvement in one or more symptoms of the disorder that persist for more than about, e.g., 2 hours, 4 hours, 6 hours, 8 hours, 10 hours, 12 hours, 14 hours, 16 hours, 18 hours, 20 hours, 22 hours, or 24 hours after administration and waking from overnight sleep.

In embodiments, provided herein are methods of treating seizure disorders comprising administering to a patient in need thereof and having received a modified receptor as described herein, a synthetic ligand to reduce or prevent seizure activity after detection of a warning signal of an impending seizure.

In embodiments, the methods described herein are effective to reduce, delay, or prevent one or more other clinical symptoms of the seizure disorder. For example, the effect of a composition comprising a synthetic ligand or a pharmaceutically acceptable salt thereof on a particular symptom, pharmacological or physiological indicator in a patient having a modified receptor in a target location in the brain, delivery of which is optionally enhanced by ultrasound energy, can be compared to the condition of the untreated patient or the patient prior to treatment. In embodiments, the symptom, pharmacological and/or physiological indicator is measured in the patient prior to treatment and measured one or more times again after treatment begins. In embodiments, the control is a reference level or an average value determined based on measuring a symptom, pharmacological, or physiological indicator in one or more patients (e.g., healthy patients) that do not have the disease or condition to be treated. In embodiments, the effect of the treatment is compared to conventional treatments within the purview of those skilled in the art.

An effective treatment of a seizure disorder (e.g., acute repetitive seizures, status epilepticus) herein may be established by showing a reduction in the frequency or severity of symptoms (e.g., more than 10%, 20%, 30%, 40%, or 50%) after a period of time as compared to baseline. For example, after a baseline period of 1 month, patients with modified receptors may be randomly assigned a synthetic ligand or a pharmaceutically acceptable salt thereof or a placebo as an adjunct therapy to standard therapy during a double-blind period of 2 months. The primary outcome measure may include the percentage of responders to the synthetic ligand, or pharmaceutically acceptable salt thereof, and to placebo, defined as having experienced at least a 10% to 50% reduction in symptoms during the second month of the double-blind period compared to baseline.

In embodiments, the pharmaceutical composition comprising the synthetic ligand may be provided in a conventional release or modified release profile. Pharmaceutical compositions can be prepared using pharmaceutically acceptable "carriers" that contain materials that are considered safe and effective. "carriers" include all components present in a pharmaceutical preparation except for the active ingredient or ingredients. The term "carrier" includes, but is not limited to, diluents, binders, lubricants, disintegrants, fillers and coating compositions. Those skilled in the art are familiar with such pharmaceutical carriers and methods of formulating (compounding) pharmaceutical compositions using such carriers.

In embodiments, the pharmaceutical compositions comprising the vector and/or synthetic ligand are suitable for parenteral administration, including, for example, intramuscular (i.m.), intravenous (i.v.), subcutaneous (s.c.), intraperitoneal (i.p.), or intrathecal (i.t). Parenteral compositions must be sterile for administration by injection, infusion or implantation into the body, and may be packaged in single-dose or multi-dose containers. In embodiments, the liquid pharmaceutical composition for parenteral administration to a patient comprises the active substance, e.g. the carrier and/or the synthetic ligand or a pharmaceutically acceptable salt of the synthetic ligand, in any of the respective amounts described above. In embodiments, the pharmaceutical composition for parenteral administration is formulated to a total volume of about, e.g., 0.1ml, 0.25ml, 0.5ml, 0.75ml, 1ml, 1.25ml, 1.5ml, 1.75ml, 2ml, 2.25ml, 2.5ml, 2.75ml, 3ml, 3.25ml, 3.5ml, 3.75ml, 4ml, 4.25ml, 4.5ml, 4.75ml, 5ml, 10ml, 20ml, 25ml, 50ml, 100ml, 200ml, 250ml, or 500 ml. In an embodiment, the volume of the pharmaceutical composition comprising the carrier is a microliter amount. For example, 0.1 microliter to 10 microliter or more may be injected. For example, 0.1 microliters, 0.2 microliters, 0.3 microliters, 0.4 microliters, 0.5 microliters, 0.6 microliters, 0.7 microliters, 0.8 microliters, 0.9 microliters, 1.0 microliters, 1.25 microliters, 1.5 microliters, 1.75 microliters, 2.0 microliters, 2.25 microliters, 2.5 microliters, 2.75 microliters, 3.0 microliters, 3.25 microliters, 3.5 microliters, 3.75 microliters, 4.0 microliters, 4.25 microliters, 4.5 microliters, 4.75 microliters, 5.0 microliters, 5.25 microliters, 5.5 microliters, 5.75 microliters, 6.0 microliters, 6.25 microliters, 6.5 microliters, 6.75 microliters, 7.0 microliters, 8.25 microliters, 8.5 microliters, 8.75, 9.0 microliters, 9.25 microliters, 9.5, 9.75 microliters, or 10 microliters. In embodiments, the composition is contained in a micropipette, bag, glass vial, plastic vial, or bottle.

In embodiments, the pharmaceutical composition for parenteral administration comprises the respective amounts described above for the synthetic ligand or a pharmaceutically acceptable salt thereof. In embodiments, a pharmaceutical composition for parenteral administration comprises from about 0.0001mg to about 500mg of the active substance, e.g., a carrier or synthetic ligand or a pharmaceutically acceptable salt of a synthetic ligand. In embodiments, the pharmaceutical composition for parenteral administration to a patient comprises the active substance, e.g., a carrier or synthetic ligand or a pharmaceutically acceptable salt of a synthetic ligand, at a corresponding concentration of about 0.001mg/ml to about 500 mg/ml. In embodiments, the pharmaceutical composition for parenteral administration comprises the active agent, e.g., a carrier or synthetic ligand or a pharmaceutically acceptable salt of a synthetic ligand, at a corresponding concentration of, e.g., about 0.005mg/ml to about 50mg/ml, about 0.01mg/ml to about 50mg/ml, about 0.1mg/ml to about 10mg/ml, about 0.05mg/ml to about 25mg/ml, about 0.05mg/ml to about 10mg/ml, about 0.05mg/ml to about 5mg/ml, or about 0.05mg/ml to about 1 mg/ml. In embodiments, the pharmaceutical composition for parenteral administration comprises the active substance, e.g., a carrier or a synthetic ligand or a pharmaceutically acceptable salt of a synthetic ligand, at a corresponding concentration of, e.g., about 0.05mg/ml to about 15mg/ml, about 0.5mg/ml to about 10mg/ml, about 0.25mg/ml to about 5mg/ml, about 0.5mg/ml to about 7mg/ml, about 1mg/ml to about 10mg/ml, about 5mg/ml to about 10mg/ml, or about 5mg/ml to about 15 mg/ml.

In an embodiment, a pharmaceutical composition for parenteral administration is provided, wherein the pharmaceutical composition is stable for at least six months. In embodiments, a pharmaceutical composition for parenteral administration exhibits a reduction of the active agent (e.g., the carrier or synthetic ligand or pharmaceutically acceptable salt of the synthetic ligand) of no more than about 5%, for example, for at least 3 months or 6 months. In embodiments, the amount of carrier or synthetic ligand or pharmaceutically acceptable salt of synthetic ligand does not degrade more than about, e.g., 2.5%, 1%, 0.5%, or 0.1%. In embodiments, the degradation is less than about, e.g., 5%, 2.5%, 1%, 0.5%, 0.25%, 0.1% for at least six months.

In an embodiment, a pharmaceutical composition for parenteral administration is provided, wherein the pharmaceutical composition remains soluble. In embodiments, pharmaceutical compositions for parenteral administration are provided that are stable, soluble, locally-compatible, and/or ready-to-use. In embodiments, the pharmaceutical compositions herein are ready-to-use for direct administration to a patient in need thereof.

Pharmaceutical compositions provided herein for parenteral administration may comprise one or more excipients, such as solvents, solubility enhancers, suspending agents, buffers, isotonic agents, stabilizers, or antimicrobial preservatives. When used, the excipients of the parenteral composition will not adversely affect the stability, bioavailability, safety and/or efficacy of the carrier, synthetic ligand or pharmaceutically acceptable salt of the synthetic ligand used in the composition. Accordingly, parenteral compositions are provided in which there is no incompatibility between any of the components of the dosage form.

In embodiments, the parenteral composition comprising a carrier or synthetic ligand or pharmaceutically acceptable salt of a synthetic ligand comprises a stabilizing amount of at least one excipient. For example, the excipient may be selected from the group consisting of: buffers, solubilizers, tonicity agents, antioxidants, chelating agents, antimicrobial agents and preservatives. One skilled in the art will appreciate that excipients may have more than one function and be classified into one or more defined groups.

In embodiments, the parenteral composition comprises a carrier or synthetic ligand or a pharmaceutically acceptable salt of a synthetic ligand and an excipient, wherein the excipient is present at a weight percentage (w/v) of less than about, e.g., 10%, 5%, 2.5%, 1%, or 0.5%. In embodiments, the excipient is present at a weight percentage of between about, e.g., 1.0% to 10%, 10% to 25%, 15% to 35%, 0.5% to 5%, 0.001% to 1%, 0.01% to 1%, 0.1% to 1%, or 0.5% to 1%. In embodiments, the excipient is present at a weight percentage of between about, e.g., 0.001% to 1%, 0.01% to 1%, 1.0% to 5%, 10% to 15%, or 1% to 15%.

In embodiments, the parenteral composition can be administered as needed, e.g., once daily, twice daily, three times daily, four times daily, five times daily, six or more times daily, or continuously depending on the needs of the patient.

In an embodiment, a parenteral composition of an active substance, such as a carrier or synthetic ligand or a pharmaceutically acceptable salt of a synthetic ligand, is provided, wherein the pH of the composition is between about 4.0 to about 8.0. In embodiments, the pH of the composition is between, for example, about 5.0 to about 8.0, about 6.0 to about 8.0, about 6.5 to about 8.0. In embodiments, the pH of the composition is between, for example, about 6.5 to about 7.5, about 7.0 to about 7.8, about 7.2 to about 7.8, or about 7.3 to about 7.6. In embodiments, the pH of the aqueous solution is, for example, about 6.8, about 7.0, about 7.2, about 7.4, about 7.6, about 7.7, about 7.8, about 8.0, about 8.2, about 8.4, or about 8.6.

It is to be understood that the dosages of the synthetic ligand or the pharmaceutically acceptable salt of the synthetic ligand provided herein are applicable to all dosage forms described herein, including conventional dosage forms, modified dosage forms, any first and second pharmaceutical compositions having different release profiles, and the parenteral formulations described herein. The appropriate amount of carrier and/or synthetic ligand will be determined by one skilled in the art based on criteria such as dosage form, route of administration, patient tolerance, efficacy, therapeutic objectives and therapeutic benefits, and other pharmaceutically acceptable criteria.

Combination therapies utilizing more than one modified receptor, e.g., hM3Dq, hM4Di, or KORD, and their corresponding synthetic ligands are contemplated herein. For example, hM4Di may be co-administered with KORD to a target location in the brain. Either or both receptors may be activated by their corresponding synthetic ligands as desired. Likewise, hM3Dq may be co-administered with KORD to a target location in the brain. Either or both receptors may be activated by their corresponding synthetic ligands as desired. Combination therapies utilizing hM3Dq, hM4Di, or KORD, their corresponding synthetic ligands in combination with one or more AEDs are contemplated herein. Furthermore, different pharmaceutical compositions having different release profiles may include administration of the active agents together in the same mixture or in separate mixtures.

In embodiments, provided herein are methods for treating a seizure disorder, the method comprising applying ultrasound to a target location in the brain of a patient to enhance permeability of the blood-brain barrier of the patient at the target location; administering to the patient a vector encoding a modified receptor, the modified receptor modified to be activated by a synthetic ligand, for delivering the modified receptor to the target location, wherein the modified receptor, when activated, inhibits neuronal firing; and administering the synthetic ligand to the patient, wherein the patient exhibits an improvement in at least one symptom of the seizure disorder.

In embodiments, provided herein are methods for treating a seizure disorder, the method comprising applying ultrasound to a target location in the brain of a patient to enhance permeability of the blood-brain barrier of the patient at the target location; administering to the patient a vector encoding a modified receptor, the modified receptor modified to be activated by a synthetic ligand, for delivering the modified receptor to the target location, wherein the modified receptor increases neuronal firing when activated; and administering the synthetic ligand to the patient, wherein the patient exhibits an improvement in at least one symptom of the seizure disorder.

In embodiments, sonication is used to enhance delivery of hM3Dq, hM4Di, or KORD to a target location in the brain by disrupting the blood brain barrier. The use of focused ultrasound energy herein disrupts the BBB without adversely affecting the vector, hM3Dq, hM4Di or KORD, their respective synthetic ligands, and/or the brain tissue itself. This may be considered surprising in view of the potential damage of the ultrasonic energy to organic compounds and tissues. The use of ultrasound energy herein can increase the rate of delivery of the vector and/or synthetic ligand to a target location in the brain, reduce side effects that may be associated with delivery of the vector and/or synthetic ligand to a target location in the brain, reduce the amount of the dose while concentrating the vector and/or synthetic ligand at the target location, and can allow for controlled release of the amount of the vector and/or synthetic ligand at the target location.

In accordance with the present disclosure, in embodiments, the ultrasound energy assists and/or advances penetration of the vehicle carrying the modified receptor and/or synthetic ligand to a target location in the brain. In embodiments, the ultrasound energy is used to render the blood brain barrier permeable to the carrier herein. Thus, in embodiments, the ultrasonic energy may be applied to the target site prior to administration of the carrier. In embodiments, the vectors herein may be administered to a target region in the brain concurrently with the administration of ultrasound energy. In embodiments, the vectors herein may be administered to a target region in the brain following administration of ultrasound energy.

As mentioned previously, the vectors herein may be administered systemically. Systemic delivery includes oral, buccal, sublingual, rectal, topical, intranasal, vaginal and parenteral modes of administration. Examples of parenteral administration modes include intravenous administration mode, intraperitoneal administration mode, intramuscular administration mode, and subcutaneous administration mode. In this way, the vector circulating in the bloodstream is delivered to a target location in the brain through a portion of the BBB disrupted by ultrasound energy. In embodiments, the vectors herein may be administered systemically following ultrasound energy treatment to the target location, and the vector penetrates the disrupted BBB to become located at the target location. In embodiments, the vectors herein can be administered directly to a target location in the brain. In embodiments, the vectors herein may be administered directly to a target location in the brain after treatment with ultrasound energy at the target location to become located at the target location. In embodiments, the vectors herein can be administered directly to a target location in the brain without ultrasound therapy.

To activate the modified receptor, a synthetic ligand that activates the modified receptor is administered to the patient. In embodiments, ultrasound energy is applied to a target region in the brain to disrupt the BBB to allow, assist and/or advance penetration of the synthetic ligand to a target location where the synthetic ligand can interact with the modified receptor. In embodiments, the synthetic ligand may be administered directly to a target location in the brain by any known means for administering materials to the brain, such as by direct injection through, for example, a drilled hole. In embodiments, the synthetic ligand may be administered systemically to the patient. Systemic delivery includes oral, buccal, sublingual, rectal, topical, intranasal, vaginal and parenteral modes of administration. Examples of parenteral administration modes include intravenous administration mode, intraperitoneal administration mode, intramuscular administration mode, and subcutaneous administration mode. In this way, synthetic ligands circulating in the bloodstream are delivered to target locations in the brain through a portion of the BBB destroyed by ultrasound energy. In embodiments, the ultrasound energy may be applied to a target region in the brain prior to administration of the synthetic ligand. In embodiments, the ultrasound energy may be applied to a target region in the brain concurrently with the administration of the synthetic ligand. In embodiments, the ultrasound energy may be applied to a target region in the brain following administration of the synthetic ligand.

In embodiments, the ultrasound energy may be administered directly to the target area by removing a portion of the skull (craniotomy) to expose dural matter at or near the target location, and delivering the ultrasound energy at or below the exposed dural matter. In embodiments, the ultrasound energy may be administered to the target location through the skull, which eliminates the need for surgery associated with delivering the ultrasound energy to the target location. Methods for delivering ultrasound energy through the skull are known in the art. See, for example, U.S. patent No. 5,752,515 and U.S. publication No. 2009/0005711, both of which are hereby incorporated by reference in their respective entireties. See also, Hynynen et al, NeuroImage 24(2005) 12-120.

In embodiments, the ultrasound energy may be applied to a target location in the brain at a frequency in the range from about 20kHz to about 5MHz, and with a sonication duration in the range from 100 nanoseconds to 1 minute. In embodiments, the ultrasound energy may be operated in a continuous wave or burst mode to be applied to a target location in the brain at a frequency in the range from about 20kHz to about 10MHz, for a sonication duration in the range from about 100 nanoseconds to about 30 minutes, wherein the burst mode repeats varying from about 0.01Hz to about 1 MHz. In embodiments, the ultrasonic energy may be at a frequency in the range from about 200kHz to about 10MHz, toAnd a sonication duration in the range from about 100 milliseconds to about 30 minutes is applied to the target location in the brain. In embodiments, the ultrasonic energy may be applied to a target location in the brain at a frequency in the range from about 250kHz to about 10MHz, and with a sonication duration in the range from about 0.10 microseconds to about 30 minutes. In an embodiment, the ultrasound energy may be applied to a target location in the brain at a frequency of about 1.525 MHz. In an embodiment, the ultrasound energy may be applied to a target location in the brain at a frequency of about 0.69 MHz. In embodiments, the pressure amplitude generated by the ultrasonic energy may be from about 0.5MPa to about 2.7 MPa. In embodiments, the pressure amplitude generated by the ultrasonic energy may be from about 0.8MPa to about 1 MPa. In embodiments, ultrasound energy is applied to a target location in the brain at a focal region having a size consistent with the volume of tissue and/or fluid to which the carrier or synthetic ligand is to be delivered, e.g., from about 0.1mm³To about 5mm³。

In embodiments, the target location and proximity thereto is confirmed by: introducing a contrast agent into the patient before, during, or after applying ultrasound energy to the target location, allowing sufficient time for the contrast agent to penetrate the BBB, and determining whether the contrast agent is present in the target location. Contrast agents are well known and include, for example, iodine-based compounds, barium-based compounds, and lanthanide-based compounds. Iodine-based agents include, for example, iohexol (iohexol), iopromide (iopromide), iodixanol (iodixanol), iomenol (iosimenol), iodixanic acid (ioxaglate), iophthalate (iothalamate), and iopamidol (iopamidol). The barium-based compound includes barium sulfate. Lanthanide-based compounds include, for example, gadolinium-based chelates, such as gadoformide (gadoforsetamide), gadopentetate dimeglumine (gadopentetate dimeglumine), gadobutrol (gadobutrol), gadobenate dimeglumine (gadobenate meglumine), gadotetate meglumine (gadobenate meglumine), and gadobenate disodium (gadobetate disodium). Detection modalities include 2-dimensional radiography, X-ray computed tomography, and magnetic resonance imaging, which are well known techniques that can be used to confirm the presence or absence of a contrast agent in a target location.

In an embodiment, a method of treating a seizure disorder is provided, the method comprising applying ultrasound to a target location in the brain of a patient to enhance permeability of the blood-brain barrier of the patient at the target location; administering to the patient a vector encoding a modified receptor, the modified receptor modified to be activated by a synthetic ligand, for delivering the modified receptor to the target location, wherein the modified receptor, when activated, inhibits neuronal firing; and administering the synthetic ligand to the patient, wherein the treatment provides an improvement in one or more symptoms of the disorder that persist for more than 1 hour after administration to the patient. In embodiments, the treatment provides an improvement in one or more symptoms of the disorder that persist for more than 2 hours after administration of the synthetic ligand to the patient. In embodiments, the treatment provides an improvement in one or more symptoms of the disorder that persist for more than 3 hours after administration of the synthetic ligand to the patient. In embodiments, the treatment provides an improvement in one or more symptoms of the disorder that persist for more than 4 hours after administration of the synthetic ligand to the patient. In embodiments, the treatment provides an improvement in one or more symptoms of the disorder that persist for more than 6 hours after administration of the synthetic ligand to the patient. In embodiments, the treatment provides an improvement in one or more symptoms of the disorder that persist for more than 8 hours, 10 hours, 12 hours, 14 hours, 16 hours, 18 hours, 20 hours, 22 hours, or 24 hours after administration of the synthetic ligand to the patient. In embodiments, in accordance with the present disclosure, there is provided an improvement in at least one symptom that persists for 12 hours after administration of a synthetic ligand to a patient. In embodiments, the synthetic ligand provides an improvement in the next day function of the patient. For example, the synthetic ligand can provide an improvement in one or more symptoms of the disorder that persist for more than about, e.g., 2 hours, 4 hours, 6 hours, 8 hours, 10 hours, 12 hours, 14 hours, 16 hours, 18 hours, 20 hours, 22 hours, or 24 hours after administration and waking from overnight sleep.

In an embodiment, a method of treating a seizure disorder is provided, the method comprising applying ultrasound to a target location in the brain of a patient to enhance permeability of the blood-brain barrier of the patient at the target location; administering to the patient a vector encoding a modified receptor, the modified receptor modified to be activated by a synthetic ligand, for delivering the modified receptor to the target location, wherein the modified receptor increases neuronal firing when activated; and administering the synthetic ligand to the patient, wherein the treatment provides an improvement in one or more symptoms of the disorder that persist for more than 1 hour after administration to the patient. In embodiments, the treatment provides an improvement in one or more symptoms of the disorder that persist for more than 2 hours after administration of the synthetic ligand to the patient. In embodiments, the treatment provides an improvement in one or more symptoms of the disorder that persist for more than 3 hours after administration of the synthetic ligand to the patient. In embodiments, the treatment provides an improvement in one or more symptoms of the disorder that persist for more than 4 hours after administration of the synthetic ligand to the patient. In embodiments, the treatment provides an improvement in one or more symptoms of the disorder that persist for more than 6 hours after administration of the synthetic ligand to the patient. In embodiments, the treatment provides an improvement in one or more symptoms of the disorder that persist for more than 8 hours, 10 hours, 12 hours, 14 hours, 16 hours, 18 hours, 20 hours, 22 hours, or 24 hours after administration of the synthetic ligand to the patient. In embodiments, in accordance with the present disclosure, there is provided an improvement in at least one symptom that persists for 12 hours after administration of a synthetic ligand to a patient. In embodiments, the synthetic ligand provides an improvement in the next day function of the patient. For example, the synthetic ligand can provide an improvement in one or more symptoms of the disorder that persist for more than about, e.g., 2 hours, 4 hours, 6 hours, 8 hours, 10 hours, 12 hours, 14 hours, 16 hours, 18 hours, 20 hours, 22 hours, or 24 hours after administration and waking from overnight sleep.

In embodiments, provided herein are methods of treating seizure disorders comprising administering ultrasound energy and a synthetic ligand to a patient in need thereof and who has received a modified receptor as described herein after detection of a warning signal of an impending seizure to reduce or prevent seizure activity.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs.

The terms "about" or "approximately" as used herein mean 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, i.e., the limitations of the measurement system. For example, "about" can mean within 3 or more than 3 standard deviations, as practiced in the art. Alternatively, "about" may mean a range of up to 20%, up to 10%, up to 5%, and/or up to 1% of a given value. Alternatively, in particular with respect to biological systems or processes, the term may mean within an order of magnitude of the value, preferably within 5 times the value, and more preferably within 2 times the value.

"ameliorating" refers to the treatment of a seizure disorder, such as focal epilepsy, refractory focal epilepsy, focal cortical dysplasia, epilepsy with generalized tonic clonic seizures, epilepsy with myoclonic absence, frontal lobe epilepsy, temporal lobe epilepsy, Landau-Kleffner syndrome, Rasmussen syndrome, Dravet syndrome, Doose syndrome, CDKL5 disorders, infantile spasms (West syndrome), Juvenile Myoclonic Epilepsy (JME), vaccine-related encephalopathy, refractory childhood epilepsy (ICE), Lennox-Gastaut syndrome (LGS), Rett syndrome, Ohtahara syndrome, CDKL5 disorders, childhood psychogenic epilepsy, essential tremor, acute recurrent seizures, benign Ralanduo epilepsy, status epilepticus, refractory status epilepticus, and essential tremor, as measured relative to at least one symptom of the foregoing disorders, Hypersensitive status epilepticus (SRSE), PCDH19 pediatric epilepsy, brain tumor-induced seizures, hamartoma-induced seizures, drug withdrawal-induced seizures, alcohol withdrawal-induced seizures, increased seizure activity, or paroxysmal seizures (also known as continuous or cluster seizures).

"improvement in next day function" or "wherein there is improvement in next day function" refers to improvement upon waking from an overnight sleep period, wherein the beneficial effects of administering one or more synthetic ligands to a patient are tailored to at least one symptom of a syndrome or disorder herein, and are objectively discernible by the patient or observer within a certain period of time, e.g., 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 12 hours, 24 hours, etc., after waking.

"treating", "treatment" or "treatment" may refer to the following: reducing or delaying the appearance of clinical symptoms of a disease or condition in a patient who may be suffering from or susceptible to the disease or condition but has not yet experienced or exhibited clinical or subclinical symptoms of the disease or condition. In certain embodiments, "treating" or "treatment" may refer to preventing the appearance of clinical symptoms of a disease or condition in a patient who may be suffering from or susceptible to the disease or condition but does not yet experience or display clinical symptoms or subclinical symptoms of the disease or condition. "treating", "treatment" or "treatment" also refers to inhibiting a disease or condition, e.g., preventing or reducing its development or at least one clinical or subclinical symptom thereof. "treating", "treating" or "treatment" also refers to alleviating a disease or condition, e.g., causing regression of the disease or condition or at least one of its clinical or subclinical symptoms. The benefit to the patient to be treated may be statistically significant, mathematically significant, or at least perceptible to the patient and/or physician. Nevertheless, prophylactic (preventative) and therapeutic (curative) treatments are two separate embodiments of the disclosure herein.

By "pharmaceutically acceptable" is meant "molecular entities and compositions that are generally considered safe", e.g., physiologically tolerable and do not typically produce allergic or similar untoward reactions (such as gastric upset, etc.) when administered to a human. In embodiments, the term refers to molecular entities and compositions approved by a regulatory agency of the Federal or a state government as a GRAS list (approved by the FDA under sections 204(s) and 409 of the Federal Food, Drug and Cosmetic Act) or similar list (the U.S. pharmacopeia or another generally recognized pharmacopeia for use in animals, and more particularly in humans).

An "effective amount" or "therapeutically effective amount" can mean a dose sufficient to alleviate one or more symptoms of a syndrome, disorder, disease or condition being treated or to otherwise provide a desired pharmacological and/or physiological effect. An "effective amount" or a "therapeutically effective amount" may be used interchangeably herein.

A combination of (a combination of) or (an administered along with) may be used interchangeably and means that two or more agents are administered during the course of therapy. The agents may be administered together at the same time or separately at spaced intervals. The agents may be administered in a single dosage form or in separate dosage forms.

A "patient in need thereof" may include an individual who has been diagnosed as having a seizure disorder, e.g., a mammal such as a human, or a canine, feline, porcine, rodent, etc., such as epilepsy, epilepsy with generalized tonic clonic seizures, epilepsy with clonic loss of consciousness, focal epilepsy, refractory focal epilepsy, focal cortical dysplasia, frontal epilepsy, temporal epilepsy, Landau-Kleffner syndrome, Rasmussen's syndrome, Dravet syndrome, Doose syndrome, CDCDKL 5 disorder, infantile spasms (West syndrome), Juvenile Myoclonic Epilepsy (JME), vaccine-related encephalopathy, refractory childhood epilepsy (ICE), Lennox-stam syndrome (LGS), Rehart syndrome, Ohtaa syndrome, KL5 disorder, childhood absence epilepsy, essential tremor, acute recurrent seizures, Benign loratadine epilepsy, status epilepticus, refractory status epilepticus, super-refractory status epilepticus (SRSE), PCDH19 pediatric epilepsy, brain tumor-induced seizures, hamartoma-induced seizures, drug withdrawal-induced seizures, alcohol withdrawal-induced seizures, increased seizure activity, or paroxysmal seizures (also known as continuous or clustered seizures). Seizure disorders may be associated with a type 1 sodium channel protein alpha subunit (Scn1 alpha) -related disorder. The methods can be provided to any individual, including for example where the patient is a neonate, an infant, a pediatric patient (6 months to 12 years old), an adolescent patient (12-18 years old), or an adult (over 18 years old).

"PK" refers to the pharmacokinetic profile. C_maxIs defined as the highest plasma drug concentration (ng/ml) estimated during the experiment. T is_maxIs defined as when C_maxEstimated time (min). AUC_0-∞Is the total area under the plasma drug concentration-time curve (ng-hr/ml or μ g-hr/ml) from drug administration until drug is expelled. The area under the curve is determined by the clearance. Clearance is defined as the volume of blood or plasma (ml/min) per unit time that is completely cleared of its drug content.

"prodrug" refers to a pharmacological substance (drug) that is administered to a subject in an inactive (or significantly less active) form. Upon administration, the prodrug is metabolized in the body (in vivo) to a compound having the desired pharmacological activity.

"analog" and "derivative" are used interchangeably and refer to a compound that has the same core as the parent compound but can differ from the parent compound by bond order, the absence or presence of one or more atoms and/or groups of atoms, and combinations thereof. Enantiomers are examples of derivatives. Derivatives may differ from the parent compound, for example, in one or more substituents present on the nucleus, which may include one or more atoms, functional groups or substructures. In general, it is contemplated that derivatives may be formed from the parent compound, at least in theory, via chemical and/or physical processes.

The term "pharmaceutically acceptable salt" as used herein refers to derivatives of a compound as defined herein, wherein the parent compound is modified by making acid or base salts thereof. Examples of pharmaceutically acceptable salts include, but are not limited to, non-toxic base addition salts with inorganic bases. Suitable inorganic bases such as alkali metal bases and alkaline earth metal bases include metal cations such as sodium, potassium, magnesium, calcium, and the like. Pharmaceutically acceptable salts can be synthesized from the parent compound by conventional chemical methods.

Examples

The examples provided herein are included solely to enhance the disclosure herein and should not be considered limiting in any respect.