Combination Therapy for Treating Seizure Disorders - Patent application

Abstract

In certain embodiments, the present disclosure is directed to methods and uses for treating a seizure disorder in a human in need thereof, comprising administering to the human a combination of N-[4-(6-fluoro-3,4-dihydro-1H-isoquinolin-2-yl)-2,6-dimethylphenyl]-3,3-dimethylbutanamide (Compound A) and an anticonvulsant (ASM), in amounts that are therapeutically effective when administered in combination. The present disclosure is further directed to various improved methods of therapy and administration of Compound A.

Description

[Technical field]

[0001] In certain embodiments, the present disclosure is directed to methods and uses for treating a seizure disorder in a human in need thereof, comprising administering to the human a combination of N-[4-(6-fluoro-3,4-dihydro-1H-isoquinolin-2-yl)-2,6-dimethylphenyl]-3,3-dimethylbutanamide (Compound A) and an anticonvulsant (ASM), in amounts that are therapeutically effective when administered in combination. The present disclosure is further directed to various improved methods of therapy and administration of Compound A. [Background technology]

[0002] Epilepsy is a common neurological disorder with an estimated worldwide prevalence of 0.7% of the population (i.e., approximately 50 million people) (see Hirtz, D. et al., Neurology (2007), 68:326-337). Epilepsy is characterized by abnormal electrical activity in the brain that leads to seizures. For epidemiological purposes, the definition requires multiple unprovoked seizures of any type.

[0003] Patients with epilepsy have an elevated risk of death compared to the general population, primarily due to the etiology of their disease. However, in patients with uncontrolled epilepsy, the greatest seizure-related mortality risk is due to sudden unexpected death in epilepsy (SUDEP) (see Hitiris, N. et al., Epilepsy and Behavior (2007), 10:363-376). Patients who participate in investigational anticonvulsant (anti-seizure, anti-epileptic) drug (ASM) clinical trials have generally had epilepsy for more than 10 years and have failed multiple ASM therapies.

[0004] Although the pathophysiology of most forms of epilepsy remains poorly understood, it is known that epileptic seizures result from excessively synchronized and sustained firing of a group of neurons. A persistent increase in neuronal excitability is common to all epilepsy syndromes. Therapeutic strategies in treating epilepsy involve decreasing neuronal excitability through various mechanistic pathways. To broaden the therapeutic spectrum by targeting different mechanisms of action and to improve the risk / benefit profile, several new ASMs have been developed and marketed over the past two decades. Currently available ASMs are believed to act by inhibiting synaptic vesicle glycoproteins, enhancing inhibitory GABAergic neurotransmission, decreasing glutamate-mediated excitatory neurotransmission, or inhibiting voltage-gated sodium or calcium channels. Despite this, up to 30% of patients remain ineffective on conventional treatments and continue to suffer from uncontrolled seizures (see Brown, DA, et al., Nature (1980), 283:673-676, and Elger, CE, et al., Epilepsy Behav. (2008), 12:501-539). The quality of life in refractory patients is poor, patients cannot drive, and patients have difficulty working or living independently. In addition, many patients suffer from behavioral disorders, neurological disorders, and / or intellectual disabilities as sequelae of their seizure disorder. Current drugs have little effect on potassium-dependent channels in neurons, despite the fact that potassium-dependent channels have a major role in controlling neuronal excitability. Therefore, drugs with novel mechanisms of action or that improve on already marketed ASMs are needed to address the significant unmet clinical need for seizure control in patients with treatment-resistant epilepsy.

[0005] The voltage-gated potassium channels Kv7.2 and Kv7.3 (Kv7.2 / Kv7.3) are important in controlling neuronal excitability. Kv7.2 / Kv7.3 underlie the neuronal "M-current", named following early characterization, because the neuronal current is reduced in response to muscarinic / cholinergic agonists (see Brown, DA et al., Nature (1980), 283:673-676). The M-current is a non-inactivating, hyperpolarizing current known to act as a brake on neuronal hyperexcitability. As a result, reduction of Kv7.2-mediated M-current, for example through genetic loss of function, can cause neuronal depolarization and increased membrane and neuronal excitability, which can lead to action potential bursts that manifest as epileptic seizures. In contrast, an increase in Kv7.2-mediated M-current can hyperpolarize the cell membrane, thereby reducing neuronal excitability and preventing the initiation and propagation of action potential bursts and resulting seizures. Enhancing the open state of Kv7.2 / Kv7.3 channels in neurons favors a hyperpolarized resting state, which reduces rapid action potential spikes (i.e., burst firing). Such enhancement can provide a stabilizing effect on excitable, especially hyperexcitable, neurons, and thus may be useful in treating certain seizure disorders. This enhancement has been clinically proven to be effective for treating seizure disorders such as partial onset seizures in adults with epilepsy using retigabine (ezogabine), a known Kv7.2 / Kv7.3 opener. [Prior art documents] [Non-patent literature]

[0006] [Non-Patent Document 1] Hirtz, D. et al., Neurology (2007), 68:326-337 [Non-Patent Document 2] Hitiris, N. et al., Epilepsy and Behavior (2007), 10:363-376 [Non-Patent Document 3] Brown, D. A. et al., Nature (1980), 283:673-676 [Non-Patent Document 4] Elger, CE et al., Epilepsy Behav. (2008), 12:501-539 [Non-Patent Document 5] Kupferberg, H., Epilepsia (1989), 30(Suppl.1):S51-S56 [Non-Patent Document 6] Prescott, JS and Evans, CA, "Pigmentary abnormalities (discoloration) associated with ezogabine / retigabine treatment: nonclinical aspects," presented at the 68th Annual Meeting of the American Epilepsy Society (AES), December 5-9, 2014, Seattle, WA, USA, poster 2.324. Summary of the Invention [Problem to be solved by the invention]

[0007] Although significant progress has been made in this field, particularly with respect to Compound A, defined below, and its use in treating seizure disorders, there remains a substantial need to provide patients with additional options for treating seizure disorders. [Means for solving the problem]

[0008] This disclosure describes certain methods and uses of the small molecule N-[4-(6-fluoro-3,4-dihydro-1H-isoquinolin-2-yl)-2,6-dimethylphenyl]-3,3-dimethylbutanamide (herein referred to as "Compound A").

[0009] In one embodiment, the disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human Compound A and ASM in combination in amounts that are therapeutically effective when administered in combination. Similarly, in some embodiments, the disclosure is directed to the use of Compound A and ASM in therapeutically effective amounts when administered in combination in treating a seizure disorder in a human in need thereof.

[0010] In another embodiment, the disclosure is directed to a method of reducing the amount of ASM required for therapeutic efficacy in a human suffering from a seizure disorder, comprising administering to the human an amount of compound A in combination with ASM that is effective to achieve such reduction when administered with ASM. Similarly, in some embodiments, the disclosure is directed to the use of compound A in reducing the amount of ASM required for therapeutic efficacy in a human suffering from a seizure disorder, for example by administering to the human an amount of compound A in combination with ASM that is effective to achieve such reduction when administered with ASM. In a particular example of these embodiments, the ASM is valproic acid, levetiracetam, phenytoin, lacosamide, cenobamate, or a combination thereof, particularly valproic acid.

[0011] In one embodiment, the disclosure is directed to a method of reducing the amount of Compound A required for therapeutic efficacy in a human suffering from a seizure disorder, comprising administering to the human an amount of ASM in combination with Compound A, effective to achieve such reduction when administered with Compound A. Similarly, in some embodiments, the disclosure is directed to the use of ASM in reducing the amount of Compound A required for therapeutic efficacy in a human suffering from a seizure disorder, for example by administering to the human an amount of ASM in combination with Compound A, effective to achieve such reduction when administered with Compound A. In particular examples of these embodiments, the ASM is valproic acid, levetiracetam, phenytoin, lacosamide, cenobamate, or combinations thereof, particularly phenytoin.

[0012] In some aspects, the methods and uses described herein for treating a seizure disorder in a human in need thereof, reducing the amount of ASM required for therapeutic efficacy, reducing the amount of Compound A required for therapeutic efficacy, or reducing absorption into the plasma or brain of an administered amount of Compound A in a human while effectively treating a seizure disorder, comprise enhancing the opening of Kv7 potassium channels in a human.

[0013] In one embodiment, the disclosure is directed to a method of enhancing the opening of Kv7 potassium channels in a human, comprising administering to the human a combination of Compound A and ASM in therapeutically effective amounts when administered in combination, for example, the human having a seizure disorder. Similarly, in some embodiments, the disclosure is directed to the use of a combination of Compound A and ASM in therapeutically effective amounts when administered in combination in enhancing the opening of Kv7 potassium channels in a human, for example, the human having a seizure disorder.

[0014] In some embodiments, the Kv7 potassium channel is one or more of Kv7.2, Kv7.3, Kv7.4, or Kv7.5. In certain examples, the opening or enhanced opening of one or more of Kv7.2, Kv7.3, Kv7.4, or Kv7.5 potassium channels is selective over Kv7.1. In other examples, the method includes the opening or enhanced opening of Kv7.2 / Kv7.3 (KCNQ2 / 3) potassium channels.

[0015] In some embodiments of the method and use, the ASM is benzodiazepine, carbamazepine, cenobamate, felbamate, gabapentin, lacosamide, lamotrigine, levetiracetam, oxcarbazepine, phenobarbital, phenytoin, pregabalin, rufinamide, tiagabine, topiramate, valproic acid, vigabatrin, zonisamide, or combinations thereof.In certain embodiments, the anticonvulsant is valproic acid, levetiracetam, phenytoin, lacosamide, cenobamate, or combinations thereof.In certain examples, the anticonvulsant does not enhance the opening of Kv7 potassium channel in humans.

[0016] In some embodiments, the ASM reduces neuronal excitation in a human by blocking sodium channels, reduces neuronal excitation in a human by blocking calcium channels, reduces neuronal excitation in a human by binding to synaptic vesicle glycoprotein 2A (SV2A), or increases neuronal inhibition in a human.

[0017] In some embodiments, the ASM is a glutamatergic agent. In other embodiments, the ASM is a GABAergic agent.

[0018] In some instances, the seizure disorder treated by or associated with the method is associated with Kv7 potassium channel dysfunction, hi other instances, the seizure disorder is focal onset epilepsy.

[0019] In some embodiments of the methods and uses, Compound A is orally administered to the human. In certain embodiments of the methods and uses, ASM is orally administered to the human. In further embodiments of the methods and uses, both Compound A and ASM are orally administered to the human.

[0020] In some embodiments of the methods and uses, Compound A is administered, e.g., orally, in combination with ASM at a dose of 1-200 mg of Compound A to a human, at a dose of 2-100 mg of Compound A to a human, at a dose of 5-50 mg of Compound A to a human, at a dose of 5, 10, 15, 20, or 25 mg of Compound A to a human, or at a dose of 20 mg of Compound A to a human. In other embodiments, Compound A is administered, e.g., orally, in combination with ASM at a dose of at least 10 mg of Compound A to a human, at a dose of at least 20 mg of Compound A to a human, or at a dose of at least 50 mg of Compound A to a human. In other embodiments, Compound A is administered, e.g., orally, in combination with ASM at a dose of 5-1000 mg of Compound A to a human per day, at a dose of 5-500 mg of Compound A to a human per day, at a dose of 5-250 mg of Compound A to a human per day, at a dose of 20-150 mg of Compound A to a human per day, or at a dose of 100 mg of Compound A to a human per day. In other examples, Compound A is administered, e.g., orally, in combination with ASM at a dose of 0.01-2.0 mg / kg of Compound A to a human, at a dose of 0.03-1.0 mg / kg of Compound A to a human, or at a dose of 0.05-0.5 mg / kg of Compound A to a human.

[0021] In some embodiments of the methods and uses, Compound A is orally administered to a human between about 30 minutes prior to ingestion of a meal and about 2 hours after ingestion of a meal, for example, Compound A may be orally administered to a human during a meal or within 15 minutes after ingestion of a meal.

[0022] In another embodiment of the method and use, the ASM is valproic acid. In some aspects, the valproic acid is administered in combination with Compound A at a dose of 2 to 16 mg / kg of valproic acid to a human, for example, the valproic acid may be administered at a dose of 4 to 12 mg / kg to a human.

[0023] In another embodiment of the method and use, the ASM is phenytoin. In some embodiments, phenytoin is administered in combination with Compound A at a dose of 0.05 to 5 mg / kg of phenytoin to a human, for example, phenytoin may be administered at a dose of 0.1 to 1 mg / kg to a human.

[0024] In another embodiment of the methods and uses, the ASM is lacosamide. In some embodiments, lacosamide is administered in combination with Compound A at a dose of 0.1 to 5 mg / kg of lacosamide to a human, for example, lacosamide is administered at a dose of 0.5 to 1 mg / kg to a human.

[0025] In another embodiment of the method and use, the ASM is cenobamate. In some aspects, the cenobamate is administered in combination with Compound A at a dose of 0.05 to 5 mg / kg of cenobamate to a human, for example, the cenobamate is administered at a dose of 0.1 to 1 mg / kg to a human.

[0026] In certain embodiments of the methods and uses, the combined administration of Compound A and ASM (e.g., valproic acid, phenytoin, levetiracetam, lacosamide, cenobamate, or a combination thereof) provides improved efficacy (e.g., increases the reduction in the number of seizure (epileptic) episodes or the reduction in the severity of seizure episodes in humans) compared to the individual administration of Compound A or ASM alone. In certain such embodiments, the combined administration provides an additive effect, where additive effect refers to the sum (sum) of the individual effects of administering Compound A and ASM. In some embodiments, the combined administration provides a synergistic effect, where synergistic effect refers to an effect that is greater than the sum of the individual effects of administering Compound A and ASM.

[0027] In other embodiments, the present disclosure provides a pharmaceutical composition comprising Compound A, an anti-seizure medication (ASM), and a pharmaceutically acceptable carrier. In some aspects of the pharmaceutical composition, the ASM is benzodiazepine, carbamazepine, cenobamate, felbamate, gabapentin, lacosamide, lamotrigine, levetiracetam, oxcarbazepine, phenobarbital, phenytoin, pregabalin, rufinamide, tiagabine, topiramate, valproic acid, vigabatrin, zonisamide, or a combination thereof.

[0028] Compound A is a small molecule currently under development for the treatment of seizure disorders, and its use as a potassium channel modulator is disclosed in U.S. Patent No. 8,293,911, U.S. Patent No. 8,993,593, and U.S. Patent Application No. 16 / 409,684 and U.S. Patent Application No. 16 / 410,851, the disclosures of which are hereby incorporated by reference in their entirety.

[0029] These and other aspects of the present disclosure will become apparent with reference to the following detailed description. For this purpose, various references that describe specific background information and procedures in more detail are set forth herein, each of which is hereby incorporated by reference in its entirety.

Brief Description of the Drawings

[0030] [Figure 1] Figure 1 shows the results of PO administration of 1, 2, 4, 8 mg / kg of Compound A or vehicle 2 hours before the assay in CF-1 mice (n = 7 per group). Upper graph: Percentage of mice showing the hindlimb extensor component during the MES test. Each bar represents the mean response ± S.E.M. Lower graph: Percentage of mice protected from tonic seizures. The single asterisk in this figure and all other figures indicates 0.01 < p < 0.05. The “mpk” listed in this figure and all other figures is mg / kg. [Diagram 2]Figure 2 shows the results of PO administration of 1, 2, 4, 8 mg / kg Compound A or vehicle 0.5 hours prior to the assay in CF-1 mice (n=7 per dose group, n=5 for vehicle alone). Top graph: presence or absence of hindlimb tonic extensor component during tonic seizures per animal. Bottom graph: percentage of mice protected. [Diagram 3] Figure 3 shows the pharmacokinetic (PK) and pharmacodynamic (PD) properties of Compound A in the mouse MES assay. Horizontal error bars indicate the SEM of the plasma or brain concentrations. When not visible, this is because the SEM is smaller than the symbols showing the mean. The solid curves through each set of data collected 2 hours (hr) after dosing are the best fits to the concentration-response curves. The IC50s based on brain and plasma concentrations were 275 nM and 154 nM, respectively. The potency measured 0.5 hr after dosing was consistent with the PK-PD relationship determined 2 hr after dosing. [Figure 4] FIG. 4 shows the results of PO administration of 1 mg / kg Compound A 2 hours before the assay and IP administration of 30, 56 or 100 mg / kg valproic acid 0.5 hours before the assay in CF-1 mice (single groups: n=15 1 mg / kg Compound A, n=15 100 mg / kg valproic acid, n=7 30 and 56 mg / kg valproic acid; combined groups: n=15 Compound A + 100 mg / kg valproic acid, n=8 Compound A + 30 and 56 mg / kg valproic acid; n=15 vehicle). Top graph: Presence or absence of hindlimb tonic extensor component during tonic seizures per animal. Bottom graph: Comparison of Compound A and valproic acid administered alone or in combination. Four asterisks indicate p<0.0001. [Diagram 5]Figure 5 shows the PK / PD of valproic acid with and without 1 mg / kg of Compound A in the mouse MES assay. The solid curves show the best fit to the concentration-response curves of valproic acid with and without 1 mg / kg of Compound A. To reflect the efficacy of Compound A alone at this dose, the curves were fit for valproic acid alone at 100% seizures and at the maximum of 73.3% with 1 mg / kg of Compound A. The effect of co-administration with Compound A was to lower the IC50 of valproic acid from 1440 μM to 608 μM. [Figure 6] Figure 6 shows the results of PO administration of 1 or 1.5 mg / kg of Compound A 2 hours before the assay and IP administration of 120 or 150 mg / kg of levetiracetam 2 hours before the assay in mice. Upper graph: presence or absence of the hindlimb extensor component during tonic seizures per animal. Lower graph: comparison of single or combined administration of Compound A and levetiracetam. [Figure 7] Figure 7 shows the results of PO administration of 0.25, 0.75, 1, 1.5 or 2.5 mg / kg of Compound A 2 hours before the assay and IP administration of 2 mg / kg of phenytoin 2 hours before the assay. Single administration: Compound A, n = 8 per dose, phenytoin 2 mg / kg, n = 24; combined administration: n = 8 per group, n = 24 vehicle. Upper graph: presence or absence of the hindlimb extensor component during tonic seizures per animal. Lower graph: comparison of single or combined administration of Compound A and phenytoin. Two asterisks indicate 0.001 < p < 0.01. [Figure 8] Figure 8 shows the PK / PD of Compound A with and without 2 mg / kg of phenytoin in the mouse MES assay. The solid curves show the best fit to the concentration-response curves of Compound A with and without 2 mg / kg of phenytoin. To reflect the efficacy of phenytoin alone at 2 mg / kg, the curves were fit for Compound A alone at 94.1% seizures and at the maximum of 75% with 2 mg / kg of phenytoin. These maxima are indicated by the horizontal dashed lines. The effect of phenytoin co-administered with Compound A is to lower the IC50 of Compound A from 147 nM to 39.7 nM. [Figure 9] Figure 9 shows the percentage of seizures in mice for various doses of Compound A, demonstrating that Compound A provides dose-dependent efficacy in the mouse AC-MES assay (A, B, and C). Compound A was administered PO at 1, 3, 5, 7.5, and 10 mg / kg 0.5 hours prior to assay in CF-1 mice (groups: n=8 1 mg / kg Compound A, n=8 3 mg / kg Compound A, n=16 5 mg / kg Compound A, n=7 7.5 mg / kg Compound A, n=17 10 mg / kg Compound A; n=24 vehicle). Results are expressed as the percentage of seizures in animals in any given dose group. The percentage of seizures in animals in Compound A-treated groups was significantly different from vehicle-treated groups in all three studies (respective p-values ​​are shown in Figure 9A-C). Concentration response curves of Compound A in plasma (D) and brain tissue (E) based on the Hill-Langmuir equation show that efficacy is concentration-dependent. Concentration response curve analysis of Compound A showed an EC50 of 0.30 μM for plasma and 0.47 μM for brain tissue. Final plasma and brain samples were obtained 0.5 hours after administration of Compound A. Each data point in D and E represents the percentage of animals with seizures at the mean concentration level for each dose group. [Figure 10]Figure 10 shows that lacosamide provides dose-dependent efficacy in the mouse AC-MES assay (A, B, C, and D). Lacosamide was administered PO at 6, 8, 10, and 20 mg / kg 2 hours prior to the assay in CF-1 mice (groups: n=16 6 mg / kg lacosamide; n=8 8 mg / kg lacosamide; n=16 10 mg / kg lacosamide; n=8 20 mg / kg lacosamide; n=32 vehicle). Results are expressed as the percentage of seizures in animals in any given dose group (n=8). In Study 2D, the percentage of seizures in animals in the 20 mg / kg lacosamide treatment group was significantly different from the vehicle treatment group (p value shown in Figure 10A). Concentration response curves of lacosamide in plasma (E) and brain tissue (F) based on the Hill-Langmuir equation show that efficacy is concentration-dependent. Concentration response curve analysis of lacosamide showed an EC50 of 21.6 μM for plasma and 22.2 μM for brain tissue. Final plasma and brain samples were obtained 2 hours after administration of lacosamide. Each data point in E and F represents the percentage of animals with seizures at the mean concentration level for each dose group (n=8). [Figure 11]Figure 11 shows the results from the combination of Compound A and lacosamide in mouse AC-MES assay (A). Compound A was administered PO at 3 mg / kg 0.5 hours before the assay in CF-1 mice (single dose groups: n=8 3 mg / kg Compound A, n=8 10 mg / kg Lacosamide; combined dose groups: n=8 3 mg / kg Compound A + 10 mg / kg Lacosamide; n=8 vehicle), and Lacosamide was administered PO at 10 mg / kg 2 hours before the assay. Results are expressed as the percentage of animals in any given dose group (n=8). The difference between the Lacosamide treatment group and the combined group was statistically significant (p value is shown in Figure 11A). The pharmacokinetic-pharmacodynamic relationship (PK / PD) of Compound A with and without lacosamide at 10 mg / kg in the mouse AC-MES assay and the PK / PD of lacosamide with and without Compound A at 3 mg / kg are shown in B and C. Each data point in B and C represents an individual concentration obtained from a single animal and whether tonic seizures were observed in the animal. Final plasma and brain samples were obtained 0.5 hours after PO administration of Compound A and 2 hours after PO administration of lacosamide. [Figure 12] Figure 12 shows the dose response of Compound A in the mouse 6 Hz psychomotor assay 1 hour after a single oral dose. PO administration of Compound A 1 hour prior to the seizure assay showed dose-dependent efficacy (A), reaching a significant effect at a dose of 8 mg / kg (**p=0.0081 vs. vehicle). The dose-response curve based on the Langmuir-Hill equation (B) predicts an ED50 of 6.48 mg / kg and an ED20 of 4.13 mg / kg with a Hill coefficient of n=-3.09. [Figure 13]Figure 13 shows the concentration response of Compound A in the mouse 6 Hz psychomotor assay 1 hour after a single oral dose. Plasma (A) and brain (B) exposures of individual animals 1 hour after a single PO dose of Compound A demonstrate a clear relationship between Compound A tissue concentration and efficacy in the 6 Hz seizure assay. Animals experiencing tremors and being cold to the touch had the highest exposure and are marked with a circle. Concentration response curves for plasma (C) and brain (D) based on the Hill-Langmuir equation predict a plasma EC50 of 0.35 μM with a Hill coefficient of n=-1.95, and a brain EC50 of 0.54 μM with a Hill coefficient of n=-2.17. [Figure 14] Figure 14 shows the efficacy of Compound A and levetiracetam alone and in combination in the 6 Hz psychomotor seizure assay 1 hour after administration. (A) Compound A (administered orally) and levetiracetam (administered intraperitoneally) alone showed different levels of efficacy in each of the two studies (no protection in study 3B, and up to 25% protection in study 3C). In each study, the combination of Compound A and levetiracetam provided significantly increased protection from seizures compared to either compound alone and compared to vehicle (*p=0.034; **p<0.01; ***p=0.0002). (B) Combining both studies, the maximum efficacy of either compound alone was reached with levetiracetam, with 14 / 16 seizures, while the combination of both compounds resulted in seizures in 5 / 15 animals. Thus, the combination of Compound A and levetiracetam protected 66.7% of the animals from seizures, which was significantly different from vehicle (****p<0.0001) and either compound alone (***p<0.001). [Figure 15]FIG. 15 shows the pharmacokinetics of Compound A and levetiracetam alone and in combination 1 hour after administration in CD-1 mice. (A) Comparison of plasma concentrations of Compound A (administered orally) and levetiracetam (administered intraperitoneally) across all experimental groups reveals that 12-fold higher plasma concentrations of Compound A were reached 1 hour after administration in Study 3C compared to Study 3B. Plasma concentrations of levetiracetam were comparable between the two studies. Administration of the combination of Compound A and levetiracetam (Combo) did not significantly alter the exposure of either compound. One animal in the combination group in Study 3C did not have measurable concentrations of levetiracetam in plasma and was excluded (not shown). (B) Comparison of brain concentrations of Compound A (administered orally) and levetiracetam (administered intraperitoneally) across all experimental groups reveals that 11-fold higher brain concentrations of Compound A were reached 1 hour after administration in Study 3C compared to Study 3B. Brain concentrations of levetiracetam were comparable between the two studies. Administration of the combination of Compound A and levetiracetam (Combo) did not significantly alter the exposure of either compound. One animal in the combination group of Study 3C did not have measurable concentrations of levetiracetam in the brain and was excluded (not shown). [Figure 16] Figure 16 shows the pharmacokinetic-pharmacodynamic shift in efficacy of Compound A and Levetiracetam after co-administration. LEV: Levetiracetam. When Compound A (administered orally) is administered in combination with Levetiracetam (administered intraperitoneally; open circles), the rate of seizures in animals is reduced compared to Compound A administered alone (closed circles), but both groups have similar concentrations of Compound A in plasma (A) and brain (B). Similarly, the rate of seizures in animals in the co-administered group (open squares) is reduced compared to Levetiracetam administered alone (closed squares), but both groups have similar concentrations of Levetiracetam in plasma (C) and brain (D). [Figure 17]Figure 17 shows the dose and concentration response of Compound A after a single oral dose in the mouse AC-MES assay. Data for Compound A and vehicle groups are shown from the indicated studies. Compound A shows dose-dependent efficacy in the mouse AC-MES assay (A, B, C, and D). Compound A was administered PO at 1, 2, 3, 5, 7.5, and 10 mg / kg 0.5 hours prior to the assay in CF-1 mice (groups: n=8 1 mg / kg Compound A, n=8 2 mg / kg Compound A, n=8 3 mg / kg Compound A, n=16 5 mg / kg Compound A, n=7 7.5 mg / kg Compound A, n=17 10 mg / kg Compound A; n=24 vehicle). Results are expressed as the percentage of seizures in animals in any given dose group. The percentage of animals with seizures in the Compound A treatment group was significantly different from the vehicle treatment group in three of the four studies (respective p values ​​are shown). Concentration-response curves for Compound A plasma (E) and brain tissue (F) based on the Hill-Langmuir equation show that efficacy was concentration-dependent. Concentration-response curve analysis for Compound A showed an EC50 of 0.296 μM for plasma and 0.471 μM for brain tissue. Final plasma and brain samples were obtained 0.5 hours after administration of Compound A. Each data point in E and F represents the percentage of animals with seizures at the mean concentration level for each dose group. [Figure 18]FIG. 18 shows the dose and concentration response of cenobamate after a single oral dose in the mouse AC-MES assay. Data for cenobamate and vehicle groups are shown from the indicated studies. Cenobamate shows dose-dependent efficacy in the mouse AC-MES assay (A, B, and C). Cenobamate was administered PO at 3, 5, 7.5, 10, and 30 mg / kg 2 hours prior to the assay in CF-1 mice (groups: n=15 3 mg / kg cenobamate; n=15 5 mg / kg cenobamate; n=8 7.5 mg / kg cenobamate; n=8 10 mg / kg cenobamate; n=8 30 mg / kg cenobamate; n=24 vehicle). Results are expressed as the percentage of seizures in animals in any given dose group. The percentage of animals with seizures in the cenobamate-treated groups was significantly different from the vehicle-treated group in two of the three studies (p values ​​shown). Concentration-response curves for cenobamate plasma (D) and brain tissue (E) based on the Hill-Langmuir equation show that efficacy is concentration-dependent. Concentration-response curve analysis for cenobamate showed an EC50 of 70.5 μM for plasma and 25.2 μM for brain tissue. Final plasma and brain samples were obtained 2 hours after administration of cenobamate. Each data point in D and E represents the percentage of animals with seizures at the mean concentration level for each dose group. [Figure 19]Figure 19 shows the anticonvulsant effect of the combination of Compound A and cenobamate in the mouse AC-MES assay. Combination of Compound A and cenobamate (A and B) in the mouse AC-MES assay. Compound A was administered PO at 0.5, 1, and 2 mg / kg 0.5 hours prior to the assay in CF-1 mice (single dose groups: n=8 2 mg / kg Compound A, n=8 5 mg / kg cenobamate; combined dose groups: n=8 2 mg / kg Compound A + 5 mg / kg cenobamate; n=8 1 mg / kg Compound A + 5 mg / kg cenobamate; n=8 0.5 mg / kg Compound A + 5 mg / kg cenobamate; n=8 vehicle), and cenobamate was administered PO at 5 mg / kg 2 hours prior to the assay. Results are expressed as the percentage of seizures in animals in any given dose group. In Study 4F, the difference between the cenobamate-treated group and the combination group was statistically significant, but the difference between the Compound A-treated group and the combination group was not. In Study 4G, the difference between the vehicle group and the combination group was statistically significant. P values ​​are indicated. The pharmacokinetic-pharmacodynamic relationship (PK / PD) of Compound A with and without cenobamate by plasma and brain concentrations is shown in C and D. Each data point in C and D represents the percentage of animals with seizures at the mean concentration level for each dose group. Final plasma and brain samples were obtained 0.5 hours after PO administration of Compound A and 2 hours after PO administration of cenobamate. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0031] The present disclosure relates inter alia to new and improved methods and uses for treating a seizure disorder in a human in need thereof, the methods and uses comprising administering to the human a combination of Compound A and an anticonvulsant (ASM), including by oral administration.

[0032] In the following disclosure, certain specific details are given to provide a thorough understanding of various embodiments. However, one skilled in the art will understand that the methods and uses described herein may be practiced without these details. In other instances, well-known structures have not been shown or described in detail to avoid unnecessarily obscuring the description of the embodiments. Unless otherwise stated, throughout this specification and the appended claims, the word "comprise" and its derivatives, such as "comprises" and "comprising," should be construed in an open-ended, inclusive sense, i.e., "including, but not limited to." Moreover, the subheadings provided herein are for convenience only and do not interpret the scope or meaning of the claimed invention.

[0033] References throughout this specification to "one embodiment" or "an embodiment" mean that a particular feature, structure, or characteristic described in connection with that embodiment is included in at least one embodiment. Thus, the appearances of the phrases "in one embodiment" or "in one embodiment" in various places throughout this specification are not necessarily all referring to the same embodiment. Moreover, particular features, structures, or characteristics may be combined in any suitable manner in one or more embodiments. Also, as used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly contradicts otherwise. It should also be noted that the terms "or" and "or" are generally used in the sense of including "and / or" unless the context clearly contradicts otherwise.

[0034] 1. Definition As used in this specification and the appended claims, unless specifically stated to the contrary, the following terms and abbreviations have the meanings indicated.

[0035] "Compound A" refers to a compound having the formula below and the compound name N-[4-(6-fluoro-3,4-dihydro-1H-isoquinolin-2-yl)-2,6-dimethylphenyl]-3,3-dimethylbutanamide. [ka] The preparation of compound A and its use as a Kv7.2 / Kv7.3 (KCNQ2 / 3) opener are disclosed in US Patent No. 8,293,911 and US Patent No. 8,993,593, and US Patent Application No. 16 / 409,684 and US Patent Application No. 16 / 410,851. Compound A is different from most known anticonvulsants in that compound A enhances or increases the opening of voltage-gated potassium channels Kv7.2 and Kv7.3 (Kv7.2 / Kv7.3), which is important in controlling the excitability of neuronal cells. Compound A is used in the methods and uses described herein. It will be understood that any reference to either Compound A or ASM referred to in this disclosure also includes pharma- ceutically acceptable salts thereof (e.g., valproic acid may be in the sodium valproate or hemisodium valproate form).

[0036] "Administered in combination" as used herein refers to any administration form of two or more different therapeutic compounds, such that the first administered therapeutic compound is still effective in the body while the second administered compound is administered (e.g., the two compounds are effective in the patient at the same time, which may include additive or synergistic effects of the two compounds). For example, Compound A and an anticonvulsant disclosed herein can be administered simultaneously or sequentially, either in the same formulation or in separate formulations. In certain embodiments, Compound A and an anticonvulsant disclosed herein can be administered within 1 hour, 12 hours, 24 hours, 36 hours, 48 ​​hours, 72 hours, or 1 week of each other. Thus, individuals undergoing such treatment can benefit from the combined effects of different therapeutic compounds.

[0037] As used herein, "reduced neuronal excitability" or "reduced neuronal excitation" or "reduced neuronal excitability" or "reduced neuronal excitability" refers to a level of neuronal activity that is reduced to some degree toward the normal physiological state observed in the absence of a seizure disorder in a patient. Certain agents that reduce neuronal excitability include agents that act on channels and receptors expressed on neurons to directly reduce the level of neuronal excitability. Conversely, agents may act indirectly to reduce the level of neuronal excitability by initiating a cascade of cellular events that result in reduced neuronal excitability as a downstream effect. Certain anticonvulsant drugs are described herein that reduce neuronal excitability to restore physiological levels of neuronal activity.

[0038] As used herein, "increased neuronal inhibition" or "increased neuronal inhibition" refers to a level of neuronal inhibition that is elevated some degree toward the normal physiological state observed in the absence of a seizure disorder in a patient. Certain anticonvulsants, such as GABAergic agents, are described herein that increase neuronal inhibition to restore physiological levels of neuronal activity.

[0039] "Seizure disorders" include partial onset (focal) seizures, photosensitive epilepsy (light-sensitive seizures), self-induced syncope, refractory epilepsy, Angelman syndrome, benign rolandic epilepsy, CDKL5 disorder, childhood absence epilepsy and juvenile absence epilepsy, Dravet syndrome, frontal lobe epilepsy, Glut1 deficiency, hypothalamic hamartoma, infantile spasms / West syndrome, juvenile myoclonic epilepsy, Landau-Kleffner syndrome, Lennox-Gastaut syndrome (LGS), myoclonic absence epilepsy, Otawara syndrome, and others. Syndrome, Panayiotopoulos syndrome, PCDH19 epilepsy, progressive myoclonic epilepsy, Rasmussen syndrome, ring chromosome 20 syndrome, reflex epilepsy, temporal lobe epilepsy, Lafora progressive myoclonic epilepsy, neurocutaneous syndromes, tuberous sclerosis, early infantile epileptic encephalopathy, early onset epileptic encephalopathy, generalized epilepsy febrile seizures plus, Rett syndrome, multiple sclerosis, Alzheimer's disease, autism, ataxia, hypotonia, and paroxysmal dyskinesia. In certain embodiments, the term "seizure disorder" refers to focal onset epilepsy, also known as partial onset (focal) epilepsy.

[0040] As used herein, "therapeutically effective amount" refers to an amount of compound A, an amount of ASM, or both an amount of compound A and an amount of ASM sufficient to treat a described disease, disorder, or condition, or to obtain a desired described effect on the disease, disorder, or condition, or one or more mechanisms underlying the disease, disorder, or condition, in a human subject. In certain embodiments, when compound A is administered in combination with ASM for the treatment of a seizure disorder, a therapeutically effective amount refers to an amount of both compound A and an amount of ASM that, upon administration to a human in combination, treats or improves the seizure disorder in the human, or produces a detectable therapeutic or preventive effect in the human with a seizure disorder. This effect can be detected, for example, by a reduction in the number of seizure episodes or a reduction in the severity of the seizure episodes.

[0041] "Treatment" as used herein refers to therapeutic applications involving the combined administration of Compound A and ASM to improve or prevent the indicated disease, disorder or condition or one or more underlying mechanisms of the disease, disorder or condition in a human subject, including slowing or stopping the progression of the indicated disease, disorder or condition or one or more underlying mechanisms of the disease, disorder or condition. In certain embodiments, when Compound A and ASM are administered in combination for the treatment of a seizure disorder, treatment refers to therapeutic applications to slow or stop the progression of the seizure disorder and / or reversing the seizure disorder. Reversal of a seizure disorder differs from therapeutic applications that slow or stop the seizure disorder in that the reversal method not only stops the progression of the seizure disorder, but also moves some cellular behavior toward the normal state that would be observed in the absence of the seizure disorder. In some embodiments, treatment of a seizure disorder comprising the combined administration of Compound A and ASM is accompanied by an alteration in the cellular activity of one or more Kv7 potassium channels (e.g., Kv7.2, Kv7.3, Kv7.4, and / or Kv7.5, particularly Kv7.2 and / or Kv7.3, optionally in preference to Kv7.1) to normal levels that would be observed in the absence of the seizure disorder.

[0042] "Fed conditions" refers to a state in which food has been ingested during the period from about 4 hours before oral administration of an effective amount of Compound A (e.g., within the therapeutically effective dose range described above) to about 4 hours after administration of Compound A. The food may be solid, liquid, or a mixture of solid and liquid foods, provided that the food has sufficient bulk and fat content to prevent rapid dissolution and absorption in the stomach. In some examples, the food is a meal such as breakfast, lunch, or dinner, or baby food (e.g., formula or breast milk). A therapeutically effective amount of Compound A may be orally administered to a subject, for example, between about 30 minutes before and about 2 hours after a meal, and most preferably, a dosage unit of Compound A is orally administered during or within 15 minutes after a meal.

[0043] "Under fasted conditions" refers to the absence of food intake for a period of at least 4 hours prior to oral administration of a therapeutically effective amount of Compound A through about 4 hours after administration of Compound A.

[0044] 2. Embodiment In some embodiments, the present disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human compound A and an anticonvulsant (ASM) in combination (e.g., orally) in amounts that are therapeutically effective when administered in combination. Similarly, in some embodiments, the present disclosure is directed to the use of compound A and ASM in amounts that are therapeutically effective when administered in combination, in treating a seizure disorder in a human in need thereof. It is contemplated that the administration of the combination can be such that compound A is administered simultaneously with, prior to, or after administration of ASM. In certain examples, the seizure disorder for which treatment is administered that includes the administration of a combination of compound A and ASM is focal onset epilepsy.

[0045] In a further embodiment, when a seizure disorder is treated with the present invention, the seizure disorder is selected from the group consisting of focal onset epilepsy, photosensitive epilepsy, self-induced syncope, refractory epilepsy, Angelman syndrome, benign rolandic epilepsy, CDKL5 disorder, childhood absence epilepsy and juvenile absence epilepsy, Dravet syndrome, frontal lobe epilepsy, Glut1 deficiency, hypothalamic hamartoma, infantile spasms / West syndrome, juvenile myoclonic epilepsy, Landau-Kleffner syndrome, Lennox-Gastaut syndrome (LGS), myoclonic epilepsy, and / or myoclonic encephalopathy. The seizure disorder is selected from: clonian absence epilepsy, Otahara syndrome, Panaetopoulos syndrome, PCDH19 epilepsy, progressive myoclonic epilepsy, Rasmussen syndrome, ring chromosome 20 syndrome, reflex epilepsy, temporal lobe epilepsy, Lafora progressive myoclonic epilepsy, neurocutaneous syndrome, tuberous sclerosis, early infantile epileptic encephalopathy, early onset epileptic encephalopathy, generalized epilepsy febrile seizures plus, Rett syndrome, multiple sclerosis, Alzheimer's disease, autism, ataxia, hypotonia, and paroxysmal dyskinesia. In certain embodiments, the seizure disorder is focal onset epilepsy, also known as partial onset (focal) epilepsy.

[0046] In some embodiments, the disclosure is directed to a method of reducing the amount of ASM required for therapeutic efficacy in a human suffering from a seizure disorder, comprising administering to the human an amount of compound A in combination with ASM (e.g., orally) effective to achieve such reduction when administered with ASM. Similarly, in some embodiments, the disclosure is directed to the use of compound A in reducing the amount of ASM required for therapeutic efficacy in a human suffering from a seizure disorder, e.g., by administering to the human an amount of compound A in combination with ASM effective to achieve such reduction when administered with ASM. For example, in some embodiments, the disclosure provides a method of treating a seizure disorder in a subject (e.g., a human) in need thereof, comprising administering compound A and ASM in combination (e.g., orally), wherein the amount of ASM administered is less than the amount of ASM that would be required to achieve the same or similar reduction in the number of seizure episodes or reduction in the severity of seizure episodes in the absence of administration of compound A. In some embodiments, the ASM is valproic acid, levetiracetam, phenytoin, lacosamide, cenobamate, or a combination thereof, particularly valproic acid.

[0047] In some embodiments, the disclosure is directed to a method of reducing the amount of Compound A required for therapeutic efficacy in a human suffering from a seizure disorder, comprising administering to the human an amount of ASM in combination with Compound A (e.g., orally) effective to achieve such reduction when administered with Compound A. Similarly, in some embodiments, the disclosure is directed to the use of ASM in reducing the amount of Compound A required for therapeutic efficacy in a human suffering from a seizure disorder, e.g., by administering to the human an amount of ASM in combination with Compound A effective to achieve such reduction when administered with Compound A. For example, in some embodiments, the disclosure provides a method of treating a seizure disorder in a subject (e.g., a human) in need thereof, comprising administering Compound A and ASM in combination (e.g., orally), wherein the amount of Compound A administered is less than the amount of Compound A that would be required to achieve the same or a similar reduction in the number of seizure episodes or reduction in the severity of seizure episodes in the absence of administration of ASM. In some embodiments, the ASM is valproic acid, levetiracetam, phenytoin, lacosamide, cenobamate, or a combination thereof, particularly phenytoin.

[0048] In some embodiments, the methods and uses described herein for treating stroke, reducing the amount of ASM required for therapeutic efficacy, or reducing the amount of Compound A required for therapeutic efficacy in a human in need thereof comprise enhancing the opening of Kv7 potassium channels in a human.

[0049] In certain embodiments, the disclosure provides methods or uses comprising opening or enhancing the opening of Kv7 potassium channels, such as Kv7.2, Kv7.3, Kv7.4, and / or Kv7.5 potassium channels, particularly Kv7.2 / Kv7.3 (KCNQ2 / 3) potassium channels, in a human in need thereof by administering an effective amount of Compound A in combination with ASM. In some such embodiments, the human has a seizure disorder, such as those described herein.

[0050] In certain examples, the methods described herein include selectively opening or selectively enhancing the opening of Kv7 potassium channels, such as one or more of Kv7.2, Kv7.3, Kv7.4, or Kv7.5 over Kv7.1. In some embodiments, the methods or uses are selective for Kv7.2 over Kv7.1. In other embodiments, the methods or uses are selective for Kv7.3 over Kv7.1. In still other embodiments, the methods or uses are selective for Kv7.4 over Kv7.1. In still further other embodiments, the methods or uses are selective for Kv7.5 over Kv7.1. In certain embodiments, the methods or uses are selective for Kv7.2 and Kv7.3 over Kv7.1. In certain embodiments, the methods or uses are selective for Kv7.2 and Kv7.3 over other Kv7 potassium channels. In certain embodiments, the method or use is selective for Kv7.2 and Kv7.3 over Kv7.4 and Kv7.5.

[0051] In one embodiment, the methods and uses described herein, e.g., methods of treating or uses in treating a seizure disorder in a human in need thereof, are achieved by administering (e.g., orally) an amount of Compound A, e.g., about 0.01 mg / kg to about 2.0 mg / kg of Compound A, in combination with ASM. More specific representative amounts of Compound A include 0.05 mg / kg, 0.10 mg / kg, 0.20 mg / kg, 0.30 mg / kg, 0.40 mg / kg, 0.5 mg / kg, 0.6 mg / kg, 0.7 mg / kg, 0.80 mg / kg, 0.90 mg / kg, 1.0 mg / kg, 1.1 mg / kg, 1.2 mg / kg, 1.3 mg / kg, 1.4 mg / kg, 1.5 mg / kg, 1.6 mg / kg, 1.7 mg / kg, 1.8 mg / kg, 1.9 mg / kg, and 2.0 mg / kg, or any range of amounts created by using two of the above amounts as endpoints. In some embodiments, the above or methods of use include administering (e.g., orally) 0.03-1.0 mg / kg of Compound A in combination with a disclosed amount of ASM. In some embodiments, the methods include administering (eg, orally) 0.05-0.5 mg / kg of Compound A in combination with a disclosed amount of ASM.

[0052] In some embodiments, the methods and uses described herein, e.g., methods or uses in treating a seizure disorder in a human in need thereof, are accomplished by administering (e.g., orally) an amount of Compound A, e.g., 2-200 mg of Compound A, in a single or divided dose, in combination with ASM. For example, the methods can be administered in a single or divided dose, e.g., about 2 mg, about 3 mg, about 4 mg, about 5 mg, about 6 mg, about 7 mg, about 8 mg, about 9 mg, about 10 mg, about 11 mg, about 12 mg, about 13 mg, about 14 mg, about 15 mg, about 16 mg, about 17 mg, about 18 mg, about 19 mg, about 20 mg, about 21 mg, about 22 mg, about 23 mg, about 24 mg, about 25 mg, about 26 mg, about 27 mg, about 29 mg, about 30 mg, about 31 mg, about 32 mg, about 33 mg, about 34 mg, about 35 mg, about 36 mg, about 37 mg, about 38 mg, about 39 mg, about 40 mg, about 41 mg, about 42 mg, about 43 mg, about 44 mg, about 45 mg, about 46 mg, about 47 mg, about 48 mg, about 49 mg, about 50 mg, about 51 mg, about 52 mg, about 53 mg, about 54 mg, about 55 mg, about 56 mg, about 57 mg, about 58 mg, about 59 mg, about 60 mg, about 61 mg, about 62 mg, about 63 mg, about 64 mg, about 65 mg, about 66 mg, about 67 mg, about 68 mg, about 69 mg, about 70 mg, about 71 mg, about 7 About 36 mg, about 37 mg, about 38 mg, about 39 mg, about 40 mg, about 41 mg, about 42 mg, about 43 mg, about 44 mg, about 45 mg, about 46 mg, about 47 mg, about 48 mg, about 49 mg, about 50 mg, about 51 mg, about 52 mg, about 53 mg, about 54 mg, about 55 mg, about 56 mg, about 57 mg, about 58 mg, about 59 mg, about 60 mg, about 61 mg, about 62 mg, about 63 mg, about 64 mg, about 65 mg, about 66 mg, about 67 mg, about 68 mg, about 69 mg, about 70 mg, about 71 mg g, about 72mg, about 73mg, about 74mg, about 75mg, about 76mg, about 77mg, about 78mg, about 79mg, about 80mg, about 81mg, about 82mg, about 83mg, about 84mg, about 85mg, about 86mg, about 87mg, about 88mg, about 89 mg, about 90mg, about 91mg, about 92mg, about 93mg, about 94mg, about 95mg, about 96mg, about 97mg, about 98mg, about 99mg, about 100mg, about 101mg, about 102mg, about 103mg, about 104mg, about 105mg, about 1 06mg, about 107mg, about 108mg, about 109mg, about 110mg, about 111mg, about 112mg, about 113mg, about 114mg, about 115mg, about 116mg, about 117mg, about 118mg, about 119mg, about 120mg, about 121mg, about 122mg, about 123mg, about 124mg, about 125mg, about 126mg, about 127mg, about 129mg, about 130mg, about 131mg, about 132mg, about 133mg, about 134mg, about 135mg, about 136mg, about 137mg,about 138mg, about 139mg, about 140mg, about 141mg, about 142mg, about 143mg, about 144mg, about 145mg, about 146mg, about 147mg, about 148mg, about 149mg, about 150mg, about 151mg, about 152mg, about 153mg, about 154mg, about 155mg, about 156mg, about 157mg, about 158mg, about 159mg, about 160mg, about 161mg, about 162mg, about 163mg, about 164mg, about 165mg, about 166mg, about 167mg, about 168mg, about 169mg, about 170mg, about 171mg, about 172mg, about 173mg, about 174mg, about 175mg, The method or use may include administering (e.g., orally) about 176 mg, about 177 mg, about 178 mg, about 179 mg, about 180 mg, about 181 mg, about 182 mg, about 183 mg, about 184 mg, about 185 mg, about 186 mg, about 187 mg, about 188 mg, about 189 mg, about 190 mg, about 191 mg, about 192 mg, about 193 mg, about 194 mg, about 195 mg, about 196 mg, about 197 mg, about 198 mg, about 199 mg, or about 200 mg of Compound A, or any range of amounts created by using two of the above amounts as endpoints. In some embodiments, the method or use includes oral administration of 2-100 or 5-50 mg of Compound A in a single or divided dose in combination with a disclosed amount of ASM. In some embodiments, the method or use comprises oral administration of 5, 10, 15, 20, or 25 mg of Compound A in a single or divided dose in combination with a disclosed amount of ASM. In some embodiments, the method or use comprises oral administration of 20 mg of Compound A in a single or divided dose in combination with a disclosed amount of ASM.

[0053] In some aspects, the methods and uses described herein, e.g., methods or uses in treating a seizure disorder in a human in need thereof, are achieved by administering (e.g., orally) at least 10 mg of Compound A, e.g., at least 20, 30, 40, 50, 75, or 100 mg of Compound A, in combination with a disclosed amount of ASM. In some embodiments, the methods and uses described herein, e.g., methods or uses in treating a seizure disorder in a human in need thereof, are achieved by administering (e.g., orally) at least 50 mg of Compound A, e.g., at least 75, 100, 125, 150, 175, or 200 mg of Compound A, in combination with a disclosed amount of ASM.

[0054] In some embodiments, the methods and uses described herein, e.g., methods of treating or uses in treating a seizure disorder in a human in need thereof, are achieved by administering (e.g., orally) an amount of Compound A per day, e.g., 5-1000 mg of Compound A per day, e.g., 5-500 mg or 5-250 mg of Compound A per day, in combination with ASM. For example, the method or use may comprise administering to the patient about 5 mg, about 10 mg, about 15 mg, about 20 mg, about 25 mg, about 30 mg, about 35 mg, about 40 mg, about 45 mg, about 50 mg, about 55 mg, about 60 mg, about 65 mg, about 70 mg, about 75 mg, about 80 mg, about 85 mg, about 90 mg, about 95 mg, about 100 mg, about 105 mg, about 110 mg, about 115 mg, about 120 mg, about 125 mg, about 130 mg, about 135 mg, about 140 mg, about 145 mg, or about 150 mg per day. g, about 150mg, about 155mg, about 160mg, about 165mg, about 170mg, about 175mg, about 180mg, about 185mg, about 190mg, about 195mg, about 200mg, about 205mg, about 210mg, about 215mg, About 220mg, about 225mg, about 230mg, about 235mg, about 240mg, about 245mg, about 250mg, about 255mg, about 260mg, about 265mg, about 270mg, about 275mg, about 280mg, about 285mg, about 29 0mg, about 295mg, about 300mg, about 305mg, about 310mg, about 315mg, about 320mg, about 325mg, about 330mg, about 335mg, about 340mg, about 345mg, about 350mg, about 355mg, about 360m g, about 365 mg, about 370 mg, about 375 mg, about 380 mg, about 385 mg, about 390 mg, about 395 mg, about 400 mg, about 405 mg, about 410 mg, about 415 mg, about 420 mg, about 425 mg, about 430 mg, This can include administering (e.g., orally) about 435 mg, about 440 mg, about 445 mg, about 450 mg, about 455 mg, about 460 mg, about 465 mg, about 470 mg, about 475 mg, about 480 mg, about 485 mg, about 490 mg, about 495 mg, about 500 mg, or about 1000 mg of Compound A per day, or a range created by using two of the above amounts as endpoints.In some embodiments, the method or use comprises orally administering from 5 to 250 mg per day of Compound A, e.g., from 10, 15, 20, 25, 30, 35, or 40 mg per day to 75, 100, 125, 150, 175, or 200 mg per day, e.g., from 20 to 150 mg per day, of Compound A, in combination with a disclosed amount of ASM. In some embodiments, the oral administration comprises from 50, 75, 100, or 125 mg per day, e.g., 100 mg per day of Compound A, in combination with a disclosed amount of ASM.

[0055] In certain examples, the daily dose of Compound A is administered (e.g., orally) as multiple doses per day, such as 2, 3, 4, or 5 doses per day. For example, a 100 mg daily dose may be administered in combination with a disclosed amount of ASM in five 20 mg, four 25 mg, three 33.3 mg, or two 50 mg doses throughout the day.

[0056] In some embodiments, the above daily doses of Compound A are administered (e.g., orally) as a single dose of Compound A or in combination with a single dose of ASM. For example, about 5, 10, 15, 20, 25, or 30 mg to about 50, 65, 75, 100, 125, or 150 mg of Compound A per day can be administered orally as a single dose, including 10-25 mg, 10-30 mg, and 10-40 mg per day as a single dose, e.g., 10-25 mg per day as a single dose. Relatedly, any of the doses of Compound A discussed in the previous paragraphs may be included in a unit dosage form.

[0057] In some embodiments, the dose of Compound A above is administered (e.g., orally) as multiple doses per week, such as 2, 3, 4, 5, 10, 15, or 20 doses per week. For example, a 100 mg weekly dose may be administered in combination with a disclosed amount of ASM in five 20 mg, four 25 mg, or two 50 mg doses throughout the week.

[0058] In some embodiments, the methods and uses described herein achieve a steady state of Compound A within 6 to 9 days, for example within about 1 week, when using the daily dosing regimens disclosed herein.

[0059] In some embodiments, the methods and uses described herein, e.g., methods of treating or uses in treating a seizure disorder in a human in need thereof, are achieved by administering (e.g., orally) an amount of ASM, e.g., about 0.01 mg / kg to about 2.0 mg / kg of ASM, in combination with Compound A. More specific representative amounts of ASM include 0.05 mg / kg, 0.10 mg / kg, 0.20 mg / kg, 0.30 mg / kg, 0.40 mg / kg, 0.5 mg / kg, 0.6 mg / kg, 0.7 mg / kg, 0.80 mg / kg, 0.90 mg / kg, 1.0 mg / kg, 1.1 mg / kg, 1.2 mg / kg, 1.3 mg / kg, 1.4 mg / kg, 1.5 mg / kg, 1.6 mg / kg, 1.7 mg / kg, 1.8 mg / kg, 1.9 mg / kg and 2.0 mg / kg, or any range of amounts created by using two of the above amounts as endpoints. In some embodiments, the method or use comprises administering (e.g., orally) 0.03 to 1.0 mg / kg of ASM in combination with compound A. In some embodiments, the methods involve administering (eg, orally) 0.05-0.5 mg / kg of ASM in combination with Compound A.

[0060] In some embodiments, the methods and uses described herein, e.g., methods of treating or use in treating a seizure disorder in a human in need thereof, are accomplished by administering (e.g., orally) an amount of ASM, e.g., 2-200 mg of ASM, in a single or divided dose, in combination with Compound A. For example, the methods can be administered in a single or divided dose, e.g., about 2 mg, about 3 mg, about 4 mg, about 5 mg, about 6 mg, about 7 mg, about 8 mg, about 9 mg, about 10 mg, about 11 mg, about 12 mg, about 13 mg, about 14 mg, about 15 mg, about 16 mg, about 17 mg, about 18 mg, about 19 mg, about 20 mg, about 21 mg, about 22 mg, about 23 mg, about 24 mg, about 25 mg, about 26 mg, about 27 mg, about 29 mg, about 30 mg, about 31 mg, about 32 mg, about 33 mg, about 34 mg, about 35 mg, about 36 mg, about 37 mg, about 38 mg, about 39 mg, about 40 mg, about 41 mg, about 42 mg, about 43 mg, about 44 mg, about 45 mg, about 46 mg, about 47 mg, about 48 mg, about 49 mg, about 50 mg, about 51 mg, about 52 mg, about 53 mg, about 54 mg, about 55 mg, about 56 mg, about 57 mg, about 58 mg, about 59 mg, about 60 mg, about 61 mg, about 62 mg, about 63 mg, about 64 mg, about 65 mg, about 66 mg, about 67 mg, about 68 mg, about 69 mg, about 70 mg, about 71 mg, about 6mg, about 37mg, about 38mg, about 39mg, about 40mg, about 41mg, about 42mg, about 43mg, about 44mg, about 45mg, about 46mg, about 47mg, about 48mg, about 49mg, about 50mg, about 51mg, about 52mg, about 53mg, about 54mg, about 55mg, about 56mg, about 57mg, about 58mg, about 59mg, about 60mg, about 61mg, about 62mg, about 63mg, about 64mg, about 65mg, about 66mg, about 67mg, about 68mg, about 69mg, about 70mg, about 71mg, about 7 2mg, about 73mg, about 74mg, about 75mg, about 76mg, about 77mg, about 78mg, about 79mg, about 80mg, about 81mg, about 82mg, about 83mg, about 84mg, about 85mg, about 86mg, about 87mg, about 88mg, about 89mg, about 90mg, about 91mg, about 92mg, about 93mg, about 94mg, about 95mg, about 96mg, about 97mg, about 98mg, about 99mg, about 100mg, about 101mg, about 102mg, about 103mg, about 104mg, about 105mg, about 106mg, about 107mg, about 108mg, about 109mg, about 110mg, about 111mg, about 112mg, about 113mg, about 114mg, about 115mg, about 116mg, about 117mg, about 118mg, about 119mg, about 120mg, about 121mg, about 122mg, about 123mg, about 124mg, about 125mg, about 126mg, about 127mg, about 129mg, about 130mg, about 131mg, about 132mg, about 133mg, about 134mg, about 135mg, about 136mg, about 137mg, about 138mg,about 139 mg, about 140 mg, about 141 mg, about 142 mg, about 143 mg, about 144 mg, about 145 mg, about 146 mg, about 147 mg, about 148 mg, about 149 mg, about 150 mg, about 151 mg, about 152 mg, about 153 mg, about 154 mg, about 155 mg, about 156 mg, about 157 mg, about 158 ​​mg, about 159 mg, about 160 mg, about 161 mg, about 162 mg, about 163 mg, about 164 mg, about 165 mg, about 166 mg, about 167 mg, about 168 mg, about 169 mg, about 170 mg, about 171 mg, about 172 mg, about 173 mg, about 174 mg, about 175 mg, about 17 The method or use may include administering (e.g., orally) 6 mg, about 177 mg, about 178 mg, about 179 mg, about 180 mg, about 181 mg, about 182 mg, about 183 mg, about 184 mg, about 185 mg, about 186 mg, about 187 mg, about 188 mg, about 189 mg, about 190 mg, about 191 mg, about 192 mg, about 193 mg, about 194 mg, about 195 mg, about 196 mg, about 197 mg, about 198 mg, about 199 mg, or about 200 mg of ASM, or any range of amounts created by using two of the above amounts as endpoints. In some embodiments, the method or use includes oral administration of 2 to 100 or 5 to 50 mg of ASM in combination with Compound A, in a single or divided dose. In some embodiments, the method or use comprises oral administration of 5, 10, 15, 20, or 25 mg of ASM in a single or divided dose in combination with Compound A. In some embodiments, the method or use comprises oral administration of 20 mg of ASM in a single or divided dose in combination with a disclosed amount of Compound A.

[0061] In some aspects, the methods and uses described herein, e.g., methods or uses in treating a seizure disorder in a human in need thereof, are achieved by administering (e.g., orally) at least 10 mg of ASM, e.g., at least 20, 30, 40, 50, 75, or 100 mg of ASM in combination with Compound A. In some embodiments, the methods and uses described herein, e.g., methods or uses in treating a seizure disorder in a human in need thereof, are achieved by administering (e.g., orally) at least 50 mg of ASM, e.g., at least 75, 100, 125, 150, 175, or 200 mg of ASM in combination with Compound A.

[0062] In some embodiments, the methods and uses described herein, e.g., methods of treating or uses in treating a seizure disorder in a human in need thereof, are achieved by administering (e.g., orally) an amount of ASM per day, e.g., 5-1000 mg of ASM per day, e.g., 5-500 mg or 5-250 mg of ASM per day, in combination with Compound A. For example, the method or use may comprise administering to the patient about 5 mg, about 10 mg, about 15 mg, about 20 mg, about 25 mg, about 30 mg, about 35 mg, about 40 mg, about 45 mg, about 50 mg, about 55 mg, about 60 mg, about 65 mg, about 70 mg, about 75 mg, about 80 mg, about 85 mg, about 90 mg, about 95 mg, about 100 mg, about 105 mg, about 110 mg, about 115 mg, about 120 mg, about 125 mg, about 130 mg, about 135 mg, about 140 mg, about 145 mg, or about 150 mg per day. g, about 150mg, about 155mg, about 160mg, about 165mg, about 170mg, about 175mg, about 180mg, about 185mg, about 190mg, about 195mg, about 200mg, about 205mg, about 210mg, about 215mg, About 220mg, about 225mg, about 230mg, about 235mg, about 240mg, about 245mg, about 250mg, about 255mg, about 260mg, about 265mg, about 270mg, about 275mg, about 280mg, about 285mg, about 2 90mg, about 295mg, about 300mg, about 305mg, about 310mg, about 315mg, about 320mg, about 325mg, about 330mg, about 335mg, about 340mg, about 345mg, about 350mg, about 355mg, about 360 mg, about 365 mg, about 370 mg, about 375 mg, about 380 mg, about 385 mg, about 390 mg, about 395 mg, about 400 mg, about 405 mg, about 410 mg, about 415 mg, about 420 mg, about 425 mg, about 430 mg , about 435 mg, about 440 mg, about 445 mg, about 450 mg, about 455 mg, about 460 mg, about 465 mg, about 470 mg, about 475 mg, about 480 mg, about 485 mg, about 490 mg, about 495 mg, about 500 mg, or about 1000 mg of ASM per day (e.g., orally), or a range created by using two of the above amounts as endpoints.In some embodiments, the method or use comprises orally administering from 5 to 250 mg per day of ASM, such as from 10, 15, 20, 25, 30, 35, or 40 mg per day to 75, 100, 125, 150, 175, or 200 mg per day of ASM, such as from 20 to 150 mg per day, in combination with Compound A. In some embodiments, the oral administration comprises from 50, 75, 100, or 125 mg per day of ASM, such as 100 mg per day, in combination with Compound A.

[0063] In certain instances, the daily doses of ASM described above are administered (e.g., orally) as multiple doses per day, such as 2, 3, 4, or 5 doses per day. For example, a daily dose of 100 mg may be administered in combination with Compound A in five doses of 20 mg, four doses of 25 mg, three doses of 33.3 mg, or two doses of 50 mg throughout the day.

[0064] In some embodiments, the above daily doses of ASM are administered (e.g., orally) as a single dose in combination with Compound A. For example, about 5, 10, 15, 20, 25, or 30 mg to about 50, 65, 75, 100, 125, or 150 mg of ASM per day can be administered orally as a single dose, including 10-25 mg, 10-30 mg, and 10-40 mg per day as a single dose, e.g., 10-25 mg per day as a sole dose, in combination with Compound A. Relatedly, any of the doses of ASM discussed in the previous paragraphs may be included in the unit dosage form.

[0065] In some embodiments of the methods and uses described herein, the ASM administered in combination with Compound A is one or more benzodiazepines (e.g., clorazepate, clobazam, clonazepam, diazepam, lorazepam, nitrazepam, etc.), carbamazepine, cenobamate, felbamate, gabapentin, lacosamide, lamotrigine, levetiracetam, oxcarbazepine, phenobarbital, phenytoin, pregabalin, rufinamide, tiagabine, topiramate, valproic acid, vigabatrin, or zonisamide. In some embodiments, the ASM administered in combination with Compound A is valproic acid, levetiracetam, phenytoin, lacosamide, cenobamate, or a combination thereof.

[0066] In some embodiments of the methods and uses described herein, the ASM treats a seizure disorder in a patient by not enhancing the opening of Kv7 potassium channels in humans (e.g., treats a seizure disorder in a patient by a different mechanism than Compound A). In some embodiments, the ASM reduces neuronal excitability by blocking sodium channels in humans. ASMs known to be sodium channel blockers include, for example, carbamazepine, lacosamide, lamotrigine, oxcarbazepine, phenytoin, rufinamide, topiramate, and zonisamide. In some embodiments, sodium channel blockers inhibit neuronal action potential firing and transmission by promoting inactivation and reducing contributions to electrical activity in the axon initial segment (AIS) and the axon itself.

[0067] In some embodiments, the disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human a combination (e.g., orally) of Compound A and carbamazepine in amounts that are therapeutically effective when administered in combination.

[0068] In some embodiments, the present disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human a combination (e.g., orally) of Compound A and lacosamide in amounts that are therapeutically effective when administered in combination.

[0069] In some embodiments, the disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human a combination (e.g., orally) of Compound A and lamotrigine in amounts that are therapeutically effective when administered in combination.

[0070] In some embodiments, the disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human a combination (e.g., orally) of Compound A and oxcarbazepine in amounts that are therapeutically effective when administered in combination.

[0071] In some embodiments, the disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human a combination (e.g., orally) of A and phenytoin in amounts that are therapeutically effective when administered in combination.

[0072] In some embodiments, the disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human a combination (e.g., orally) of Compound A and rufinamide in amounts that are therapeutically effective when administered in combination.

[0073] In some embodiments, the present disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human a combination (e.g., orally) of Compound A and topiramate in amounts that are therapeutically effective when administered in combination.

[0074] In some embodiments, the disclosure is directed to a method of treating a seizure disorder in a human in need thereof comprising administering to the human Compound A and zonisamide in combination (e.g., orally) in amounts that are therapeutically effective when administered in combination.

[0075] In some embodiments of the methods and uses described herein, ASM reduces neuronal excitation by blocking calcium channels in humans.ASMs known to be calcium channel blockers include, for example, gabapentin, phenobarbital, pregabalin, and zonisamide.In some embodiments, calcium channel blockers reduce excitatory transmission by reducing presynaptic neurotransmitter release, which is a calcium-dependent process.

[0076] In some embodiments, the disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human a combination (e.g., orally) of Compound A and gabapentin in amounts that are therapeutically effective when administered in combination.

[0077] In some embodiments, the disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human a combination (e.g., orally) of Compound A and phenobarbital in amounts that are therapeutically effective when administered in combination.

[0078] In some embodiments, the disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human a combination (e.g., orally) of Compound A and pregabalin in amounts that are therapeutically effective when administered in combination.

[0079] In some embodiments, the disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human a combination (e.g., orally) of Compound A and gabapentin in amounts that are therapeutically effective when administered in combination.

[0080] In some embodiments of the methods and uses described herein, the ASM reduces neuronal excitation by binding to synaptic vesicle glycoprotein 2A (SV2A) in humans. ASMs known to bind to SV2A include, for example, levetiracetam. In some embodiments, the SV2A binder reduces excitatory transmission by reducing presynaptic neurotransmitter release.

[0081] In some embodiments, the present disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human a combination (e.g., orally) of Compound A and levetiracetam in amounts that are therapeutically effective when administered in combination.

[0082] In some embodiments of the methods and uses described herein, the ASM increases neuronal inhibition in humans. ASMs known to increase neuronal inhibition include, for example, benzodiazepines (e.g., clorazepate, clobazam, clonazepam, diazepam, lorazepam, nitrazepam, etc.), felbamate, phenobarbital, tiagabine, topiramate, valproic acid, and vigabatrin. In some embodiments, the ASM is a glutamatergic agent. In some aspects, the glutamatergic agent reduces the effect of this neurotransmitter on the AMPA receptor or NMDA receptor on the postsynaptic membrane. In some embodiments, the glutamatergic agent is carbamazepine, felbamate, lamotrigine, pregabalin, phenytoin, pregabalin, tiagabine, or topiramate. In other embodiments, the ASM is a GABAergic agent. In some embodiments, the GABA agonist is a benzodiazepine (e.g., clorazepate, clobazam, clonazepam, diazepam, lorazepam, nitrazepam, etc.), felbamate, phenobarbital, tiagabine, topiramate, valproic acid, or vigabatrin. In some examples, the GABA agonist may affect the GABA receptor by direct positive allosteric modulation of GABA receptor activity (e.g., benzodiazepines). In other examples, the GABA agonist may affect the GABA receptor by indirectly increasing the level of GABA through inhibition of GABA transaminase (e.g., vigabatrin) or GABA transporter-1 (GAT1, e.g., tiagabine).

[0083] In some embodiments, the disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human a combination (e.g., orally) of Compound A and a benzodiazepine (e.g., clorazepate, clobazam, clonazepam, diazepam, lorazepam, nitrazepam, etc.) in amounts that are therapeutically effective when administered in combination.

[0084] In some embodiments, the present disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human a combination (e.g., orally) of Compound A and felbamate in amounts that are therapeutically effective when administered in combination.

[0085] In some embodiments, the disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human a combination (e.g., orally) of Compound A and phenobarbital in amounts that are therapeutically effective when administered in combination.

[0086] In some embodiments, the disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human a combination (e.g., orally) of Compound A and tiagabine in amounts that are therapeutically effective when administered in combination.

[0087] In some embodiments, the present disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human a combination (e.g., orally) of Compound A and topiramate in amounts that are therapeutically effective when administered in combination.

[0088] In some embodiments, the disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human a combination (e.g., orally) of Compound A and valproic acid in amounts that are therapeutically effective when administered in combination.

[0089] In some embodiments, the disclosure is directed to a method of treating a seizure disorder in a human in need thereof, comprising administering to the human a combination (e.g., orally) of Compound A and vigabatrin in amounts that are therapeutically effective when administered in combination.

[0090] In some embodiments of the methods and uses, the ASM administered in combination with Compound A is valproic acid. In some aspects, valproic acid is administered in combination with Compound A at any of the doses of ASM discussed in the paragraph above. In some embodiments, valproic acid is administered to a human in combination with Compound A at a dose of 2 to 16 mg / kg, for example, valproic acid may be administered to a human at a dose of 4 to 12 mg / kg.

[0091] In some embodiments of the methods and uses, the ASM administered in combination with Compound A is phenytoin. In some aspects, phenytoin is administered in combination with Compound A at any of the doses of ASM discussed in the paragraph above. In some embodiments, phenytoin is administered to a human in combination with Compound A at a dose of 0.05-5 mg / kg, for example, phenytoin may be administered to a human (e.g., orally) at a dose of 0.1-1 mg / kg.

[0092] In some embodiments of the methods and uses, the ASM administered in combination with Compound A is lacosamide. In some aspects, lacosamide is administered in combination with Compound A at any of the doses of ASM discussed in the paragraph above. In some embodiments, lacosamide is administered to a human in combination with Compound A at a dose of 0.1-5 mg / kg, for example, lacosamide is administered to a human (e.g., orally) at a dose of 0.5-1 mg / kg.

[0093] In another embodiment of the methods and uses, the ASM administered in combination with Compound A is cenobamate. In some aspects, cenobamate is administered in combination with Compound A at any of the doses of ASM discussed in the paragraph above. In some embodiments, cenobamate is administered to a human in combination with Compound A at a dose of 0.05-5 mg / kg, for example, cenobamate is administered to a human (e.g., orally) at a dose of 0.1-1 mg / kg.

[0094] In certain embodiments of the methods and uses, the combined administration of Compound A and an ASM (e.g., valproic acid, phenytoin, levetiracetam, lacosamide, or cenobamate) provides improved efficacy (e.g., increases the reduction in the number of seizure episodes or the reduction in the severity of seizure episodes in humans) compared to the individual administration of Compound A or an ASM alone. In certain such embodiments, the combined administration provides an additive effect, where additive effect refers to the sum of the individual effects of administering Compound A and administering one or more ASMs. In some embodiments, the combined administration provides a synergistic effect, where synergistic effect refers to an effect that is greater than the sum of the individual effects of administering Compound A and administering one or more ASMs.

[0095] In additional embodiments, the above methods and uses of treating a seizure disorder by administering (e.g., orally) Compound A include orally administering Compound A to a human under fed conditions. In some embodiments, oral administration of Compound A to a human under fed conditions (i.e., with food or in close temporal proximity to the ingestion of food) significantly enhances the bioavailability and exposure of Compound A compared to oral administration of Compound A to a human under fasted conditions (i.e., without food or not in close temporal proximity to the ingestion of food). In some embodiments, oral administration of Compound A to a human under fed conditions enhances one or more pharmacokinetic parameters of Compound A (e.g., C max , AUC inf , T max , t 1 / 2λz etc.) to increase

[0096] In certain embodiments, the methods and uses described herein administer compound A, ASM, or both compound A and ASM in the form of a pharma- ceutically acceptable oral composition comprising compound A, ASM, or both compound A and ASM, and one or more pharma- ceutically acceptable carriers or excipients. The amount of compound A, ASM, or both compound A and ASM contained in these compositions may correspond to one or more of the amounts described herein. In some embodiments, the composition is a unit dose.

[0097] Examples of pharma- ceutically acceptable oral compositions comprising Compound A, ASM, or both Compound A and ASM include solid formulations (such as tablets, capsules, lozenges, dragees, granules, powders, sprinkles, wafers, multiparticulates, and films), liquid formulations (such as aqueous solutions, elixirs, tinctures, tonics, slurries, suspensions, and dispersions), and aerosolized formulations (such as mists and sprays). In one embodiment, the pharma- ceutically acceptable oral composition of Compound A comprises a pediatric suspension or granules. All of the above amounts of Compound A, ASM, or both Compound A and ASM may be included in such formulations, including, for example, a capsule containing 5, 10, 15, 10, 25, 30, or 35 mg of Compound A, a capsule containing 5, 10, 15, 10, 25, 30, 35, 40, 45, 50, 55, 60, 65, 75, 85, 90, 95, or 100 mg of ASM, or a capsule containing 5, 10, 15, 10, 25, 30, or 35 mg of Compound A and 5, 10, 15, 10, 25, 30, 35, 40, 45, 50, 55, 60, 65, 75, 85, 90, 95, or 100 mg of ASM.

[0098] Other suitable routes of administration for Compound A, ASM, or both Compound A and ASM according to the methods and uses described herein include sublingual and buccal (e.g., using a film or other composition that dissolves in the mouth under the tongue or inside the cheek), ocular (e.g., eye drops), aural (e.g., by ear drops), oral or nasal inhalation (e.g., by insufflation or spray), dermal or topical (e.g., by cream or lotion), or transdermal (e.g., by skin patch). In addition to oral administration, other enteral routes of administration can be used for Compound A, ASM, or both Compound A and ASM, including intravaginal and intrarectal (e.g., by ointments, suppositories, enemas).

[0099] Examples of compositions suitable for parenteral administration of Compound A, ASM, or both Compound A and ASM include sterile injectable solutions, suspensions, or dispersions, including aqueous or oily preparations, particularly aqueous preparations. In some embodiments, Compound A, ASM, or both Compound A and ASM are administered according to the methods or uses described herein in sterile injectable aqueous formulations that include parenterally acceptable diluents or solvents, such as water, Ringer's solution, isotonic sodium chloride solution, aqueous buffer solutions, and aqueous solutions containing miscible alcohols, such as 1,3-butanediol. Additional suitable excipients for parenteral formulations of Compound A, ASM, or both Compound A and ASM include mono- or diglycerides; fatty acids, such as oleic acid and its glyceride derivatives; natural pharma- ceutically acceptable oils, such as olive oil or castor oil, including polyoxyethylated versions thereof; long-chain alcohol diluents or dispersants, such as alkylcelluloses, including carboxymethylcellulose; and surfactants, such as Tweens, Spans, and other emulsifiers or bioavailability enhancers.

[0100] In another embodiment, a kit is provided for the combined oral administration of Compound A and ASM for the treatment of depressive disorders by oral administration. Such a kit includes multiple oral dosage unit forms of Compound A, ASM, or both Compound A and ASM, in addition to instructions for the combined oral administration of Compound A and ASM.

[0101] Further embodiments and examples of the present disclosure are described herein. These embodiments and examples are illustrative and should not be construed as limiting the scope of the claimed invention.

[0102] 3. Numbered embodiments Embodiment 1. A method of treating a seizure disorder in a human in need thereof, comprising administering to said human a combination of Compound A and an anticonvulsant (ASM) in amounts that, when administered in combination, are therapeutically effective, where Compound A is N-[4-(6-fluoro-3,4-dihydro-1H-isoquinolin-2-yl)-2,6-dimethylphenyl]-3,3-dimethylbutanamide.

[0103] Embodiment 2. A method of reducing the amount of an anticonvulsant (ASM) required for therapeutic efficacy in a human suffering from a seizure disorder, comprising administering to said human, in combination with said ASM, an amount of Compound A effective to achieve such reduction when administered with said ASM, wherein Compound A is N-[4-(6-fluoro-3,4-dihydro-1H-isoquinolin-2-yl)-2,6-dimethylphenyl]-3,3-dimethylbutanamide.

[0104] Embodiment 3. A method of reducing the amount of Compound A required for therapeutic efficacy in a human suffering from a seizure disorder, comprising administering to said human an anticonvulsant (ASM) in combination with Compound A in an amount effective to achieve such reduction when administered with Compound A, wherein Compound A is N-[4-(6-fluoro-3,4-dihydro-1H-isoquinolin-2-yl)-2,6-dimethylphenyl]-3,3-dimethylbutanamide.

[0105] Embodiment 4. The method of any one of embodiments 1 to 3, wherein said method comprises enhancing opening of Kv7 potassium channels in said human.

[0106] Embodiment 5. A method of enhancing the opening of Kv7 potassium channels in a human, comprising administering to said human a combination of Compound A and an anticonvulsant (ASM) in amounts that, when administered in combination, are therapeutically effective, wherein Compound A is N-[4-(6-fluoro-3,4-dihydro-1H-isoquinolin-2-yl)-2,6-dimethylphenyl]-3,3-dimethylbutanamide, and said human has a seizure disorder.

[0107] Embodiment 6. The method of embodiment 4 or embodiment 5, wherein said Kv7 potassium channel is one or more of Kv7.2, Kv7.3, Kv7.4, or Kv7.5.

[0108] Embodiment 7. The method of embodiment 6, wherein said method is selective for enhancing opening of one or more of Kv7.2, Kv7.3, Kv7.4, or Kv7.5 over Kv7.1.

[0109] Embodiment 8. The method of embodiment 4 or embodiment 5, wherein said method comprises opening of Kv7.2 / Kv7.3 (KCNQ2 / 3) potassium channels.

[0110] Embodiment 9. The method of any one of embodiments 1 to 8, wherein the ASM is a benzodiazepine, carbamazepine, cenobamate, felbamate, gabapentin, lacosamide, lamotrigine, levetiracetam, oxcarbazepine, phenobarbital, phenytoin, pregabalin, rufinamide, tiagabine, topiramate, valproic acid, vigabatrin, zonisamide, or a combination thereof.

[0111] Embodiment 10. The method of any one of embodiments 1 to 9, wherein the ASM is valproic acid, levetiracetam, phenytoin, lacosamide, or cenobamate.

[0112] Embodiment 11. The method of any one of embodiments 1 to 10, wherein said ASM does not enhance the opening of Kv7 potassium channels in said human.

[0113] Embodiment 12. The method of any one of embodiments 1 to 11, wherein the ASM reduces neuronal excitability in the human.

[0114] Embodiment 13. The method of embodiment 12, wherein the ASM reduces neuronal excitability in the human by blocking sodium channels.

[0115] Embodiment 14. The method of embodiment 12, wherein the ASM reduces neuronal excitability in the human by blocking calcium channels.

[0116] Embodiment 15. The method of embodiment 12, wherein said ASM reduces neuronal excitability in said human by binding to synaptic vesicle glycoprotein 2A (SV2A).

[0117] Embodiment 16. The method of any one of embodiments 1 to 11, wherein the ASM increases neuronal inhibition in the human.

[0118] Embodiment 17. The method of any one of embodiments 1 to 11, wherein the ASM is a glutamatergic agent.

[0119] Embodiment 18. The method of any one of embodiments 1 to 11, wherein the ASM is a GABAergic agent.

[0120] Embodiment 19. The method of any one of embodiments 1 to 18, wherein the seizure disorder is associated with Kv7 potassium channel dysfunction.

[0121] Embodiment 20. The method of any one of embodiments 1 to 19, wherein the seizure disorder is focal onset epilepsy.

[0122] Embodiment 21. The method of any one of embodiments 1 to 20, wherein Compound A is administered orally to said human.

[0123] Embodiment 22. The method of any one of embodiments 1 to 21, wherein the ASM is administered orally to the human.

[0124] Embodiment 23. The method of any one of embodiments 1 to 22, wherein Compound A is administered to said human at a dose of 1 to 200 mg.

[0125] Embodiment 24. The method of any one of embodiments 1 to 23, wherein Compound A is administered to said human at a dose of 2 to 100 mg.

[0126] Embodiment 25. The method of any one of embodiments 1 to 24, wherein Compound A is administered to said human at a dose of 5 to 50 mg.

[0127] Embodiment 26. The method of any one of embodiments 1 to 25, wherein Compound A is administered to said human at a dose of 5, 10, 15, 20, or 25 mg.

[0128] Embodiment 27. The method of any one of embodiments 1 to 26, wherein Compound A is administered to said human at a dose of 20 mg.

[0129] Embodiment 28. The method of any one of embodiments 1 to 22, wherein Compound A is administered to said human in a dose of at least 10 mg.

[0130] Embodiment 29. The method of embodiment 28, wherein Compound A is administered to said human in a dose of at least 20 mg.

[0131] Embodiment 30. The method of embodiment 28, wherein Compound A is administered to said human in a dose of at least 50 mg.

[0132] Embodiment 31. The method of any one of embodiments 1 to 22, wherein Compound A is administered to said human at a dose of 5 to 1000 mg per day.

[0133] Embodiment 32. The method of embodiment 31, wherein Compound A is administered to said human being at a dose of 5 to 500 mg per day.

[0134] Embodiment 33. The method of embodiment 31, wherein Compound A is administered to said human being at a dose of 5 to 250 mg per day.

[0135] Embodiment 34. The method of embodiment 31, wherein Compound A is administered to said human being at a dose of 20 to 150 mg per day.

[0136] Embodiment 35. The method of embodiment 31, wherein Compound A is administered to said human at a dose of 100 mg per day.

[0137] Embodiment 36. The method of any one of embodiments 1 to 22, wherein Compound A is administered to said human at a dose of 0.01 to 2.0 mg / kg.

[0138] Embodiment 37. The method of embodiment 36, wherein Compound A is administered to said human at a dose of 0.03 to 1.0 mg / kg.

[0139] Embodiment 38. The method of embodiment 36, wherein Compound A is administered to said human at a dose of 0.05 to 0.5 mg / kg.

[0140] Embodiment 39. The method of any one of embodiments 1 to 38, wherein Compound A is orally administered to said human between about 30 minutes prior to ingestion of a meal and about 2 hours after ingestion of a meal.

[0141] Embodiment 40. The method of embodiment 39, wherein Compound A is orally administered to said human during a meal or within 15 minutes of ingesting a meal.

[0142] Embodiment 41. The method of any one of embodiments 1 to 40, wherein the ASM is valproic acid.

[0143] Embodiment 42. The method of embodiment 41, wherein the valproic acid is administered to the human being at a dose of 2 to 16 mg / kg.

[0144] Embodiment 43. The method of embodiment 41, wherein the valproic acid is administered to the human being at a dose of 4 to 12 mg / kg.

[0145] Embodiment 44. The method of any one of embodiments 1 to 40, wherein the ASM is phenytoin.

[0146] Embodiment 45. The method of embodiment 44, wherein the phenytoin is administered to the human being at a dose of 0.05 to 5 mg / kg.

[0147] Embodiment 46. The method of embodiment 44, wherein the phenytoin is administered to the human being at a dose of 0.1 to 1 mg / kg.

[0148] Embodiment 47. The method of any one of embodiments 1 to 40, wherein the ASM is lacosamide.

[0149] Embodiment 48. The method of embodiment 47, wherein the lacosamide is administered to the human being at a dose of 0.1 to 5 mg / kg.

[0150] Embodiment 49. The method of embodiment 47, wherein the lacosamide is administered to the human at a dose of 0.5 to 1 mg / kg.

[0151] Embodiment 50. The method of any one of embodiments 1 to 40, wherein the ASM is cenobamate.

[0152] Embodiment 51. The method of embodiment 50, wherein the cenobamate is administered to the human being at a dose of 0.05 to 5 mg / kg.

[0153] Embodiment 52. The method of embodiment 50, wherein the cenobamate is administered to the human being at a dose of 0.1 to 1 mg / kg.

[0154] Embodiment 53. The method of any one of embodiments 1 to 52, wherein the combined administration of compound A and said ASM provides improved efficacy compared to the administration of compound A alone or said ASM individually. EXAMPLES

[0155] A study was conducted to determine the effects of Compound A versus other ASMs. 1. Example 1. Anticonvulsant effects of Compound A alone and in combination with a common ASM The interactions of Compound A with other ASMs were evaluated following oral administration in the mouse maximal electric shock (MES) assay to determine whether some combinations were favorable or unfavorable. Efficacy was quantified by calculating the percentage of animals with tonic hind limb seizures following corneal stimulation.

[0156] 1.1 Compound A When administered alone, Compound A (1, 2, 4, or 8 mg / kg) reduced the incidence of tonic seizures in the MES assay at both 0.5 and 2 hours after oral dosing. Concentration-response relationships were fitted to binding isotherms, yielding an IC at a plasma concentration of 154 nM. 50 and IC at brain concentration of 275 nM. 50 (n=7 per dose, per time point) were obtained.

[0157] Compound preparation: Compound A was first solubilized in DMSO. This solution was then added to a 0.5% methylcellulose solution to create a more homogenous and less aggregated compound suspension. Serial dilutions were prepared from the tube containing the highest concentration of Compound A, and the compound suspension was further vortexed before administration to the animals. The final concentration of DMSO was 5%, an amount that had no apparent toxicity or neuroprotection in the MES assay. Phenytoin and valproic acid were solubilized in 100% saline (0.9%). Levetiracetam was solubilized in 0.5% methylcellulose and 0.2% Tween 80 in deionized water.

[0158] Animals: Adult male CF-1 albino mice (25-35 g) were purchased from Harlan-Envigo. Mice were housed 4 per cage and had free access to filtered water and food throughout the experiment.

[0159] MES Assay: Compounds were administered orally by gavage prior to testing unless otherwise stated. During MES testing, a 60 Hz alternating current (50 mA) was delivered to mice through corneal electrodes for 0.2 seconds. A drop of 0.5% Alcaine solution was placed on the eye prior to current delivery. The electrodes were then gently placed over the animal's eye(s) and the electric shock was initiated by pulling the trigger via a foot-operated activator. Animals were manually restrained and gently released once the shock was delivered and seizures had begun. Animals were monitored for tonic extension of the hind limbs as the endpoint of this test.

[0160] Statistical analysis: All statistics were calculated using Prism version 7 software (Graphpad Software). The methods used for each experiment are presented in the Results section. Concentration-response curves were calculated as best fits to the following equation: Y=Min+(Max-Min) / (1+10^((LogIC50-X)*Hill Slope)) where X is the logarithm of the plasma concentration. Unless otherwise stated, the minimum value was constrained to be 0. The maximum value was constrained to be the value experimentally determined by vehicle control measurements.

[0161] Dose response of Compound A: The dose response after oral administration of 1, 2, 4, or 8 mg / kg of Compound A was evaluated in the mouse MES assay at two different time points: 0.5 and 2 hours after oral administration (Figures 1 and 2). Animals were randomly assigned to vehicle (n=7 at 2 hours, n=5 at 0.5 hours) or different dose groups (1 dose, n=7 per time point) and the MES assay was performed by an experimenter blinded to the treatment conditions.

[0162] At 2 hours, Compound A showed a dose-dependent reduction in the number of tonic seizures as assessed by hindlimb extension. The 8 mg / kg dose provided significant protection compared to vehicle-treated mice (number of mice with tonic seizures / total tested: 1 mg / kg: 5 / 7 (p=0.939), 2 mg / kg: 4 / 7 (p=0.598), 4 mg / kg: 2 / 7 (p=0.085), 8 mg / kg: 1 / 7 (p=0.023), vehicle: 6 / 7 (p values ​​calculated by one-way ANOVA with Dunnett's multiple comparison test). Using the same data, hindlimb tonic seizures were evaluated. The percentage of mice protected from sexual extension was calculated (1 mg / kg: 28.6% (p>0.999), 2 mg / kg: 42.9% (p>0.999), 4 mg / kg: 71.4% (p=0.103), 8 mg / kg: 85.7% (p=0.029), vehicle: 14.3%; p values ​​were calculated by Fisher's exact test). Brain and plasma samples were taken from mice immediately after efficacy testing and analyzed using UHPLC-ESI-MS / MS (Table 1).

[0163] [Table 1]

[0164] Total concentrations in brain and plasma were 1 mg / kg: 0.19 μM (70.2 ng / g) and 0.1 μM (35.8 ng / mL), 2 mg / kg: 0.21 μM (75.8 ng / g) and 0.13 μM (46.7 ng / mL), 4 mg / kg: 0.36 μM (131.3 ng / g) and 0.23 μM (83 ng / mL), and 8 mg / kg: 0.56 μM (207.7 ng / g) and 0.43 μM (157.8 ng / mL), respectively.

[0165] At 0.5 hours, doses of 1, 2 and 4 mg / kg showed a dose-dependent reduction in the number of tonic seizures with hind limb extension, with 4 mg / kg providing significant protection compared to vehicle-treated mice (1 mg / kg: 4 / 7 (p=0.449), 2 mg / kg: 2 / 7 (p=0.088), 4 mg / kg: 1 / 7 (p=0.032), 8 mg / kg: 3 / 7 (p=0.215), vehicle: 5 / 5; one-way ANOVA calculated using Dunnett's multiple comparison test). At 8 mg / kg, 1 / 7 animals had 2 tonic seizures with hind limb extension after stimulation, an occurrence not seen in the other dose groups or in vehicle-treated mice. A similar dose-dependent efficacy was observed for mice protected from the hindlimb tonic extensor component of tonic seizures (1 mg / kg: 42.9% (p>0.205), 2 mg / kg: 71.4% (p=0.028), 4 mg / kg: 85.7% (p=0.015), 8 mg / kg: 71.4% (p=0.028), vehicle: 0%; calculated using Fisher's exact test). Brain and plasma samples were taken from mice immediately after efficacy testing and analyzed using UHPLC-ESI-MS / MS (Table 2).

[0166] [Table 2]

[0167] The mean brain and plasma total concentrations for each dose group were as follows: 1 mg / kg: 0.27 μM (97.9 ng / g) and 0.13 μM (48.9 ng / mL), 2 mg / kg: 0.36 μM (132 ng / g) and 0.22 μM (80.7 ng / mL), 4 mg / kg: 0.64 μM (234.7 ng / g) and 0.42 μM (154.4 ng / mL), and 8 mg / kg: 1.09 μM (402 ng / g) and 0.71 μM (261.3 ng / mL), respectively.

[0168] Figure 3 shows concentration-response relationships summarizing all of the PK-PD data for Compound A in the MES assay. The top panel shows inhibition of tonic seizures as a function of plasma concentration, and the bottom panel shows inhibition as a function of brain concentration. Compared to the 2-hour pretreatment time, higher brain and plasma exposures were observed at 0.5 hours, but the efficacy observed at each exposure level reflected brain or plasma concentrations regardless of time after dosing (Figure 3). Due to the clear dose-response with 2-hour pretreatment of Compound A, this time point was selected for further studies. Data collected 2 hours after dosing were fitted to a concentration-response curve to determine the IC 50 Only the and Hill coefficients were varied for best fitting. The dashed horizontal line indicates the incidence of tonic seizures in vehicle: 94.1 ± 0.03% (mean ± SEM, n = 85). This value was taken as the maximum of the curve and the minimum was constrained to 0 (complete inhibition). IC based on brain and plasma concentrations 50 The estimated values ​​were 275 nM and 154 nM, respectively.

[0169] 1.2 Combination of Compound A and Valproic Acid The doses of 1 mg / kg PO with 2 hours pretreatment for Compound A and 100 mg / kg IP with 0.5 hours pretreatment for Valproic acid were selected for the first combination study.To quantify the degree of additional efficacy provided by Compound A (1 mg / kg PO), a second experiment was performed to repeat the findings of the first study and test additional doses of Valproic acid (30 and 56 mg / kg IP).The combined results of these two experiments are discussed below.

[0170] Animals were randomly assigned to receive oral doses of Compound A or vehicle 2 hours prior to efficacy testing, followed by IP doses of valproic acid or vehicle 0.5 hours prior to efficacy testing. Based on this experimental design, animals in the single-dose control group or vehicle control group also received two doses each: (Compound A + vehicle, vehicle + valproic acid, Compound A + valproic acid, or vehicle + vehicle).

[0171] Co-administration of Compound A and 100 mg / kg valproic acid resulted in a significant increase in the number of mice without tonic seizures with hind limb extension compared to mice treated with Compound A alone (Compound A + 100 mg / kg valproic acid: 10 / 15 vs. 1 mg / kg Compound A + vehicle: 4 / 15 (p=0.021) and vehicle + 100 mg / kg valproic acid: 5 / 15 (p=0.059); p values ​​calculated by two-way ANOVA followed by Dunnett's multiple comparison test). There was a trend towards increased protection against tonic seizures when Compound A was administered in combination with 56 mg / kg valproic acid, but no difference was observed between Compound A and 30 mg / kg valproic acid: Compound A + 56 mg / kg valproic acid: 4 / 8 vs. 1 mg / kg Compound A + vehicle: 4 / 15 (p=0.396) and vs. vehicle + 56 mg / kg valproic acid: 1 / 7 (p=0.226); Compound A + 30 mg / kg valproic acid: 2 / 8 vs. 1 mg / kg Compound A + vehicle: 4 / 15 (p=0.995) and vs. vehicle + 56 mg / kg valproic acid: 0 / 8 (p=0.463) (Figure 4).

[0172] Both combination treatment groups of Compound A (1 mg / kg) + 56 and 100 mg / kg valproic acid provided significant protection compared to vehicle-treated animals: Compound A + 100 mg / kg valproic acid: 10 / 15 (p=0.0001) and Compound A + 56 mg / kg valproic acid: 4 / 8 (p=0.0381) vs. vehicle + vehicle: 0 / 15; p values ​​were calculated by one-way ANOVA followed by Dunnett's multiple comparison test.

[0173] Plasma and brain samples were taken from the mice immediately following the efficacy study and analyzed using UHPLC-ESI-MS / MS (Table 3).

[0174] [Table 3]

[0175] There were no significant differences in plasma or brain concentrations of Compound A when administered alone or co-administered with valproic acid. Mean total plasma and brain concentrations of Compound A at 1 mg / kg when administered alone: ​​0.066 μM (24 ng / mL) and 0.16 μM (59 ng / g), when administered with valproic acid (30 mg / kg): 0.038 μM (14 ng / mL) and 0.075 μM (28 ng / g), when administered with valproic acid (56 mg / kg): 0.050 μM (18 ng / mL) and 0.117 μM (43 ng / g), and when administered with valproic acid (100 mg / kg): 0.042 μM (15 ng / mL) and 0.097 μM (36 ng / g). Similarly, there were no significant changes in plasma or brain concentrations of valproic acid at the three doses tested when combined with Compound A.

[0176] Coadministration of Compound A with VA also demonstrated IC values ​​for inhibition of tonic seizures by VA. 50 To quantify the effect of Compound A on VA, the effect of Compound A on the concentration-response relationship of VA was analyzed (Figure 5).

[0177] 1.3 Combination of Compound A with Levetiracetam Levetiracetam is inactive in the MES assay (ED 50 >500mg / kg) was associated with ED 50 =19.4 mg / kg (Barton et al., Epilepsy Res. 2001, 47:217-227). A high dose of levetiracetam was chosen because it is known to have biological activity.

[0178] Levetiracetam was evaluated in the MES assay by IP dosing at 120 and 150 mg / kg, which resulted in plasma concentrations of 1001 μM (170.3 μg / mL) and 1154 μM (196.4 μg / mL) and brain concentrations of 583 μM (99.2 μg / g) and 540 μM (91.9 μg / g), respectively (Table 4).

[0179] [Table 4]

[0180] None of the doses produced a significant increase in the percentage of mice free of tonic seizures compared to vehicle-treated mice: 120 mg / kg: 0 / 6 (p=0.98), 150 mg / kg 2 / 8 (p=0.93), vehicle: 2 / 18; p values ​​were calculated by one-way ANOVA with Dunnett's multiple comparison test (Figure 6).

[0181] Co-administration with 1 or 1.5 mg / kg Compound A did not affect plasma or brain levels of levetiracetam: 957 μM (163 μg / mL) plasma and 554 μM (943 μg / g) brain (LEV 120 mg / kg + 1 mg / kg Compound A); 1235 μM (210 μg / mL) plasma and 531 μM (90.4 μg / g) brain (LEV 150 mg / kg + 1.5 mg / kg Compound A). Exposure values ​​for Compound A at 1 mg / kg alone or co-administered with levetiracetam at 120 mg / kg were 0.03 μM (11.1 ng / mL) and 0.03 μM (11.1 ng / mL) in plasma and 0.06 μM (20.9 ng / g) and 0.05 μM (20.0 ng / g) in brain, respectively. Exposure values ​​for Compound A at 1.5 mg / kg alone or co-administered with levetiracetam at 150 mg / kg were 0.04 μM (15 ng / mL) and 0.07 μM (25 ng / mL) in plasma and 0.08 μM (31 ng / g) and 0.14 μM (52 ng / g) in brain, respectively.

[0182] At none of the dose combinations did the increase in the percentage of mice free of tonic seizures significantly exceed the effect of Compound A alone: ​​Compound A at 1 mg / kg: 2 / 11 vs. Compound A (1 mg / kg) + LEV (120 mg / kg): 2 / 5, p=0.578; and Compound A at 1.5 mg / kg: 3 / 8 vs. Compound A (1.5 mg / kg) + LEV (150 mg / kg): 4 / 8, p=0.805; p values ​​were calculated by two-way ANOVA with Dunnett's multiple comparison test (Figure 6).

[0183] 1.4 Combination of Compound A with phenytoin K V 7 channels are located at the axon initial region and are V 1.6 and Na V Compound A colocalizes with the K1.2 channel. V Because phenylalanine activates naphthalene-7 channels and phenytoin inhibits voltage-gated sodium channels, a favorable interaction was predicted, but the magnitude was unknown.

[0184] The dose response of Compound A (0.25, 0.75, 1, 1.5, and 2.5 mg / kg) administered PO with a 2 hour pretreatment period was tested in the MES assay in the presence or absence of a fixed dose of phenytoin (2 mg / kg, IP, 2 hour pretreatment).

[0185] Compared to vehicle-treated animals, administration of Compound A alone resulted in a dose-dependent increase in the proportion of animals free of tonic seizures: 0.25 mg / kg Compound A: 0 / 8 (p=0.999), 0.75 mg / kg: 2 / 8 (p=0.948), 1 mg / kg: 2 / 8 (p=0.948), 1.5 mg / kg: 2 / 8 (p=0.948), 2.5 mg / kg: 4 / 8 (p=0.12); 2 mg / kg phenytoin: 6 / 24 (p=0.72); vehicle 2 / 24. When co-administered with phenytoin, Compound A at doses of 0.75, 1, 1.5 and 2.5 mg / kg resulted in a significant increase in the percentage of animals free of tonic seizures compared to vehicle-treated animals: Compound A 0.25 mg / kg + phenytoin 2 mg / kg: 3 / 8 (p=0.48), Compound A 0.75 mg / kg + phenytoin 2 mg / kg: 5 / 8 (p=0.018), Compound A 1 mg / kg + phenytoin 2 mg / kg: 6 / 8 (p=0.002), Compound A 1.5 mg / kg + phenytoin 2 mg / kg: 6 / 8 (p=0.002), Compound A 2.5 mg / kg + phenytoin 2 mg / kg: 7 / 8 (p=0.0001); p values ​​were calculated by one-way ANOVA and Dunnett's multiple comparison test (Figure 7).

[0186] Comparison of animals administered Compound A alone with groups co-administered with both Compound A and phenytoin showed a significant increase in the percentage of animals free of tonic seizures in the co-administered groups at doses of 1 and 1.5 mg / kg. A similar trend was observed when comparing phenytoin alone with the co-administration groups, with significant seizure freedom observed with Compound A at 1, 1.5 and 2.5 mg / kg: Compound A 0.25 + phenytoin 2 mg / kg combination: 3 / 8 vs. Compound A alone (0.25 mg / kg): 0 / 8 (p=0.165) and 6 / 24 phenytoin (2 mg / kg; p=0.715), Compound A 0.75 + phenytoin 2 mg / kg combination: 5 / 8 vs. Compound A alone (0.75 mg / kg): 2 / 8 (p=0.165) and 6 / 24 phenytoin (2 mg / kg; p=0.074), Compound A1 + phenytoin 2 mg / kg combination: 6 / 8 vs. Compound A alone (1 mg / kg): 2 / 8 (p=0.048) and 6 / 24 Phenytoin (2 mg / kg; p=0.012), Compound A1.5 + phenytoin 2 mg / kg combination: 6 / 8 vs. Compound A administered alone (1.5 mg / kg): 2 / 8 (p=0.048) and 6 / 24 (phenytoin at 2 mg / kg; p=0.012), Compound A2.5 + phenytoin 2 mg / kg combination: 7 / 8 vs. Compound A administered alone (2.5 mg / kg): 4 / 8 (p=0.165) and 6 / 24 phenytoin (2 mg / kg; p=0.001); p values ​​were calculated by two-way ANOVA followed by Dunnett's multiple comparison test (Figure 7).

[0187] No significant changes in plasma or brain concentrations were observed for phenytoin when administered with or without Compound A (Table 4).

[0188] [Table 5]

[0189] Plasma and brain exposures were 4.1 μM (1019 ng / mL) and 4.2 μM (1052 ng / g) for phenytoin 2 mg / kg alone; 4.0 μM (1014 ng / mL) and 4.2 μM (1055 ng / g) when combined with 0.25 mg / kg Compound A; and 4.1 μM (1025 ng / mL) and 2.9 μM (107 0 ng / g): 3.9 μM (989 ng / mL) and 5.0 μM (1263 ng / g) when combined with 1 mg / kg Compound A: 4.8 μM (1210 ng / mL) and 4.8 μM (1196 ng / g) when combined with 1.5 mg / kg Compound A, and 3.9 μM (971 ng / mL) and 2.9 μM (1051 ng / g) when combined with 2.5 mg / kg Compound A.

[0190] Similarly, there are no significant changes in plasma and brain concentrations of Compound A when administered alone or in combination with phenytoin (Table 5).

[0191] [Table 6]

[0192] Plasma and brain showed the following concentrations of Compound A at 0.25 mg / kg alone vs. in combination with phenytoin (2 mg / kg): 0.01 μM (4.3 ng / mL) and 0.04 μM (14.8 ng / g) vs. 0.02 μM (5.6 ng / mL) and 0.05 μM (18.4 ng / g); at 0.75 mg / kg alone vs. in combination: 0.03 μM (11.7 ng / mL) and 0.12 μM (43.8 ng / g) vs. 0.04 μM (16.0 ng / mL) and 0.15 μM (56.7 ng / g); at 1 mg / kg alone vs. in combination: 0.05 μM (17.0 ng / mL) and 0.15 μM (18.4 ng / g); 1.5 mg / kg alone vs. combination: 0.05 μM (19.5 ng / mL) and 0.08 μM (30.8 ng / g) vs. 0.06 μM (23.4 ng / mL) and 0.11 μM (42 ng / g); 2.5 mg / kg alone vs. combination: 0.12 μM (45.1 ng / mL) and 0.39 μM (144.1 ng / g) vs. 0.16 μM (58.4 ng / mL) and 0.51 μM (186.5 ng / g).

[0193] IC for inhibition of tonic seizures by Compound A 50 Co-administration experiments with Compound A and phenytoin were also analyzed to quantify the effect of phenytoin on the concentration-response relationship of Compound A (FIG. 8).

[0194] 1.5 Conclusion Compound A showed dose-dependent efficacy in the mouse MES assay that correlated well with plasma and brain exposure values. The combination of Compound A and 100 mg / kg valproic acid or 2 mg / kg phenytoin resulted in higher efficacy observed in the MES assay than observed with either compound alone. Co-administration of Compound A with levetiracetam did not provide a significant improvement in efficacy in the MES assay when compared to Compound A alone.

[0195] Valproic acid (30, 56 or 100 mg / kg IP, administered 0.5 hours prior to MES): Compound A (1 mg / kg PO, 2 hours prior to MES) combined with 100 mg / kg valproic acid (VA) produced greater inhibition than Compound A administered alone (1 mg / kg Compound A + 100 mg / kg valproic acid: 10 / 15 vs. 1 mg / kg Compound A + vehicle: 4 / 15 (p=0.021)) (single groups: n=15 1 mg / kg Compound A, n=15 100 mg / kg valproic acid, n=7 30 and 56 mg / kg valproic acid; combination groups: n=15 Compound A 1 mg / kg + 100 mg / kg valproic acid, n=8 Compound A + 30 and 56 mg / kg valproic acid; n=15 vehicle). Compound A administered at 1 mg / kg reduced the plasma levels of valproic acid required for protection against tonic seizures in the MES assay. IC 50 The IC value of valproic acid in combination with 1 mg / kg of Compound A was 1440 μM. 50 The mean total plasma concentration of Compound A at 1 mg / kg when administered alone was 0.07 μM (24.2 ng / mL) and when administered with valproic acid (30 mg / kg): 0.04 μM (14.0 ng / mL), when administered with valproic acid (56 mg / kg): 0.03 μM (12.0 ng / mL), and when administered with valproic acid (100 mg / kg): 0.04 μM (15.0 ng / mL).

[0196] Levetiracetam (120 and 150 mg / kg IP, administered 2 hours prior to MES): Compound A at a dose of 1 mg / kg produced plasma concentrations of 66 ± 14 nM (mean ± SEM, n = 15) and reduced the proportion with tonic seizures by 26.7%. Co-administration of Compound A with levetiracetam (LEV) did not provide improved protection against tonic seizures when compared to Compound A alone: ​​Compound A at 1 mg / kg: 2 / 11 vs. Compound A (1 mg / kg) + LEV (120 mg / kg): 2 / 5, p = 0.578; Compound A at 1.5 mg / kg: 3 / 8 vs. Compound A (1.5 mg / kg) + LEV (150 mg / kg): 4 / 8, p = 0. 805. Plasma concentrations of Compound A administered at 1 mg / kg alone or co-administered with 120 mg / kg LEV were 0.03 μM (11.1 ng / mL) and 0.03 μM (11.1 ng / mL), and those administered at 1.5 mg / kg alone or co-administered with 150 mg / kg LEV were 0.041 μM (15 ng / mL) and 0.068 μM (25 ng / mL), respectively.

[0197] Phenytoin (2 mg / kg IP, given 2 hours prior to MES): Combining Compound A with phenytoin given at 2 mg / kg produced greater inhibition than Compound A given alone. When co-administered with phenytoin 2 hours prior to MES, oral doses of 0.75, 1, 1.5 and 2.5 mg / kg Compound A resulted in a significant increase in the percentage of mice free of tonic seizures compared to vehicle-treated animals: Compound A 0.25 mg / kg + phenytoin 2 mg / kg: 3 / 8 (p=0.48), Compound A 0.75 mg / kg + phenytoin 2 mg / kg: 5 / 8 (p=0.013), Compound A 1 mg / kg + phenytoin 2 mg / kg: 6 / 8 (p=0.002), Compound A 1.5 mg / kg + phenytoin 2 mg / kg: 6 / 8 (p=0.002), Compound A 2.5 mg / kg + phenytoin 2 mg / kg: 7 / 8 (p=0.0001). IC of Compound A alone 50 The IC of Compound A in combination with 2 mg / kg phenytoin was 147 nM. 50The plasma concentration of Compound A was 39.7 nM. There were no significant changes in plasma concentrations of Compound A when administered alone or in combination with phenytoin: Compound A at 0.25 mg / kg alone vs. in combination with phenytoin (2 mg / kg): 0.01 μM (4.3 ng / mL) vs. 0.02 μM (5.6 ng / mL), and at 0.75 mg / kg alone vs. in combination: 0.03 μM (11.7 ng / mL) vs. 0. 0.04 μM (16 ng / mL), 1 mg / kg alone vs. combination: 0.05 μM (17 ng / mL) vs. 0.05 μM (19 ng / mL), 1.5 mg / kg alone vs. combination: 0.05 μM (19.5 ng / mL) vs. 0.06 μM (23.4 ng / mL), 2.5 mg / kg alone vs. combination: 0.12 μM (45.1 ng / mL) vs. 0.16 μM (58.4 ng / mL).

[0198] 2. Example 2. Anticonvulsant Effects of Compound A Alone and in Combination with Lacosamide The efficacy of Compound A and its pharmacological interaction with lacosamide were evaluated after oral administration in the mouse maximal electroconvulsive seizure (AC-MES) assay.

[0199] The aim of these studies was to characterize the dose-dependent anticonvulsant activity of Compound A in the AC-MES assay in mice after a single oral dose of Compound A and its pharmacological interaction with lacosamide. The AC-MES assay is typically used as a translational animal model of partial epileptic seizures (partial seizure onset) in response to nonselective sodium channel blockers and potassium channel openers. The MES assay has been widely used to screen and characterize novel antiseizure compounds (Loescher et al., Epilepsy Res. 1991, 8(2):79-94; Piredda et al., J Pharmacol Exp Ther. 1985, 232(3):741-745; and White et al., Ital J Neurol Sci. 1995, 16(1-2):73-77). After electroshock stimulation with a sufficiently high current, mice and rats show tonic extension followed by clonus of the hind limbs. A test compound was considered protective if it was able to prevent tonic extension.

[0200] The anticonvulsant efficacy of Compound A and lacosamide, as well as the effect of combining Compound A with lacosamide, were examined in the AC-MES assay in male CF-1 mice after a single PO dose. Plasma and brain samples were obtained to understand the relationship between drug concentration and efficacy.

[0201] 2.1 Materials and Methods Test compound - Compound A [Table 7]

[0202] Test compound - Lacosamide [Table 8]

[0203] Vehicle F1: 0.5% methylcellulose and 0.2% Tween-80 in deionized (DI) water. 0.8 L of DI water was heated to 70°C-80°C. 5 grams of methylcellulose was weighed and slowly added in small portions to the heated DI water. The mixture was stirred until it formed a homogenous milky suspension. The suspension was transferred to a cold room and stirred overnight to obtain a clear solution. 2 mL of Tween-80 was added to the clear solution and diluted to 1 L with DI water. This vehicle solution was stored at 2°C-8°C.

[0204] Vehicle F2: 5% dimethylsulfoxide (DMSO) and F1. 5% DMSO was added to F1 vehicle.

[0205] Dosage formulation: Compound A and lacosamide were weighed into separate vials. Compound A was formulated in F2 vehicle, and lacosamide was formulated in F1 vehicle. The appropriate amount of vehicle was added to Compound A and lacosamide powder, then mixed with an IKA T-18 Ultra-Turrax homogenizer to create a homogenous suspension of the desired concentration. The vial was then wrapped in aluminum foil to protect it from light, and placed on a stir plate until administration.

[0206] Test system [Table 9]

[0207] Experimental Design: Animals were assigned to treatment groups as shown in Tables 6, 7, 8, and 9.

[0208] Three studies were conducted with Compound A and four with lacosamide. The dates of each study are shown in Tables 6 to 9.

[0209] [Table 10] PO: per os, orally.

[0210] [Table 11] PO: per os, orally.

[0211] [Table 12] PO: per os, orally.

[0212] [Table 13] PO: per os, orally.

[0213] Blinding and Randomization: The experimenter administering the compound randomly assigned each animal to a treatment group. A different experimenter blinded to the treatment group assignment performed the testing. In addition, compound administration and MES testing were performed in different rooms to ensure that the experimenter performing the testing was completely blinded to the treatments. All animals tested in a given experiment had an equal chance of being assigned to any treatment group.

[0214] Clinical Observations: All animals were observed for abnormal behavior for 10 minutes after administration of Compound A and lacosamide by the investigator who administered the animals, and again for 10 minutes at the time of stimulation by the investigator who tested the animals. Any qualitative changes from normal behavior were recorded.

[0215] AC-MES Assay: The MES test is widely used to discover anticonvulsants (Piredda et al., Loescher et al., and White et al.). The MES test is considered a model of generalized tonic-clonic (GTC) seizures and provides an index of the ability of compounds to prevent seizure spread. In the AC-MES model, an alternating current (60 Hz, 40 mA) electric shock was delivered for 0.2 seconds by a corneal electrode (HSE-HA Rodent Shocker, Harvard Apparatus, model number: 73-0105). CF-1 mice were administered vehicle or compound A PO (as per Standard Operating Procedure (SOP) TECH-006) 0.5 hours before the electric shock assay. Just before the electric shock stimulation, the eyes of the animals were anesthetized with a topical application of 0.5% Alcaine solution (Proparacaine hydrochloride, 1 drop / eye). The mice were then restrained, corneal electrodes were attached, and shocks were administered. In naive animals, seizures are characterized by an initial generalized tonic seizure with a hindlimb tonic extensor component. Animals are considered protected from MES-induced seizures upon disappearance of the hindlimb tonic extensor component of the seizure, in which case they are scored as "0". If the mouse shows tonic hindlimb extension, it is scored as "1". Mice were euthanized immediately after the initial seizure score assessment after the electric shock for collection of plasma and brain.

[0216] Sample collection and preparation: Mice were anesthetized by isoflurane inhalation until they reached a surgical plane of anesthesia (per SOP TECH-018). A syringe (1 mL syringe with a 22-gauge needle) was then inserted substernally into the heart (per SOP TECH-031). Approximately 0.5 mL of blood was collected, placed in a K2EDTA tube, and stored on ice. The animals were then euthanized by cervical dislocation. The brains were removed, placed in pre-weighed vials, and snap frozen on dry ice. At the end of sample collection, blood was centrifuged at 4000 rpm for 10 minutes at 4°C, and plasma was transferred by pipette into labeled tubes. All samples were stored in a -80°C freezer until the time of bioanalysis.

[0217] Plasma samples: Extraction of plasma samples was performed by protein precipitation with acetonitrile. Diluted plasma samples (50 μL) were mixed with 50 μL of internal standard (IS) solution in 1:1 acetonitrile:water (v:v), followed by the addition of 200 μL of acetonitrile. Samples were vortex mixed for 30 s, centrifuged at 13,000 rpm for 20 min, decanted into a 96-well plate, and centrifuged for an additional 20 min at 4000 rpm. Samples were analyzed by ultra-high performance liquid chromatography electrospray ionization tandem mass spectrometry (UHPLC-ESI-MS / MS) as described in the bioanalytical procedures below.

[0218] Brain samples: Prior to extraction, pre-weighed whole brains were homogenized in 1:1 acetonitrile:water (v:v) (2 mL per mouse brain) using an IKA T18 Ultra-Turrax homogenizer for approximately 1 min at setting 4. The homogenate was centrifuged at 13,000 rpm for 20 min and 50 μL of the supernatant was treated exactly as described above for the plasma samples.

[0219] Bioanalytical Procedure: All samples, including plasma and brain homogenate extracts (including calibration standards and quality control (QC) samples prepared in K2EDTA mouse plasma), were extracted by protein precipitation. To each 50 μL aliquot of sample, 50 μL of IS solution (2500 ng / mL lacosamide in water:acetonitrile (1:1)) and 50 μL of 6% (v:v) aqueous phosphoric acid were added, followed by 200 μL of acetonitrile. Samples in 1.7 mL tubes were vortex mixed and then centrifuged at 13,000 rpm for 20 min. Fifteen microliters of the supernatant were mixed with 150 μL of water:acetonitrile (1:1) in a 96-well plate and centrifuged at 4000 rpm for 20 min after mixing. Samples were then subjected to UHPLC-MS / MS analysis.

[0220] Samples were analyzed for Compound A by a research grade UHPLC-MS / MS method using the conditions below.

[0221] [Table 14]

[0222] Samples were analyzed for lacosamide by a research grade UHPLC-MS / MS method using the conditions below.

[0223] [Table 15]

[0224] Sample concentrations were determined using a best-fit model, either a linear or quadratic calibration function (weighted by 1 / x), generated by regression of analyte-to-IS peak area ratios in standard samples to their respective concentrations. Acceptance criteria for analytical runs required that back-calculated values ​​of standards and QC samples fall within ±20% of their nominal values, except for the lowest standard or lower limit of quantification (LLOQ), for which the acceptance criteria was ±25%. At least six of the twelve standard points had to show back-calculated values ​​within ±20% of the nominal concentration for an acceptable calibration. At least three QC samples, one at each concentration, had to show back-calculated values ​​within ±20% of the nominal concentration for the entire sample batch to be valid.

[0225] Data processing and analysis: All statistics were calculated using GraphPad Prism software, version 8. Concentration-response curves were constructed using the Hill-Langmuir equation. formula 1 Y = B + (TB) × x n / (I C 50 n +x n ) In the above formula 1, B = the lowest value, set to 0; T = maximum value, set as 1, n = Hill coefficient, constrained to be less than zero, ·I C 50 = concentration of compound required for 50% inhibition in vitro It is.

[0226] All group data are expressed as the mean. Differences between groups were analyzed using the Kruskal-Wallis followed by Dunn's multiple comparison test. Statistical significance was achieved at a value of p<0.05.

[0227] Bioanalysis: All system suitability tests (SSTs), QC, matrix, and solvent blanks met the acceptance criteria described in SOP MTD-066. The LLOQs for Compound A and lacosamide were 2.34 ng / mL in Study 2C, 4.69 ng / mL for both compounds in Studies 2D and 2A, and 2.34 ng / mL and 4.69 ng / mL for lacosamide and Compound A, respectively, in Study 2B. The upper limit of quantification (ULOQ) was 4800 ng / mL for both compounds in all studies.

[0228] Efficacy in AC-MES assay: The results of the AC-MES induced seizure study in mice are shown in Tables 10, 11, 12, and 13 and in Figures 9, 10, and 11.

[0229] Compound A dose and concentration response: Compound A dose and concentration response is shown in Figure 9. In all three efficacy studies (2A, 2B, and 2C), 100% of vehicle-treated CF-1 mice exhibited tonic seizures with hindlimb extension. In study 2A, the percentage of animals treated with Compound A that exhibited tonic seizure responses to AC-MES stimulation decreased from 8 / 8 at 1 mg / kg to 7 / 8 at 5 mg / kg to 3 / 8 at 10 mg / kg, suggesting a dose-dependent increase in efficacy. In study 2B, the 5 mg / kg and 10 mg / kg doses showed greater efficacy than in study 2A, with 0 / 8 and 1 / 9 animals experiencing seizures, respectively, which may have resulted from the higher plasma concentrations achieved in study 2B (mean plasma concentrations: 0.470 μM at 5 mg / kg and 0.553 μM at 10 mg / kg) than in study 2A (mean plasma concentrations: 0.276 μM at 5 mg / kg and 0.366 μM at 10 mg / kg). In addition, studies 2A and 2B were conducted one week apart from each other, which may also explain the difference in efficacy. In study 2C, the 3 mg / kg dose of compound A showed 2 / 8 animals experiencing seizures, with a mean plasma concentration of 0.284 μM. In all three studies (2A, 2B, and 2C), the difference between the compound A-treated group and the vehicle-treated group reached statistical significance (p values ​​are shown in FIG. 9).

[0230] One mouse from the 10 mg / kg group in study 2A and three mice from study 2B (two from the 10 mg / kg group and one from the 7.5 mg / kg group) showed behavioral signs (tremors, reduced activity and hind limb spreading) (plasma concentrations: 2A: 0.391 μM; 2B: 10 mg / kg: 0.608 and 0.516 μM, 7.5 mg / kg: 0.746 μM). The concentrations of Compound A reached in brain tissue and plasma were linear with respect to dose (Figure 9).

[0231] The combined concentration-response curves for Compound A from three studies showed an EC 50 and an EC of 0.471 μM for brain tissue. 50 was shown (Figure 9).

[0232] 2.2 Results Prior to testing Compound A in combination with lacosamide in the AC-MES model, lacosamide was tested at different doses in a separate study (2D) and in separate groups of mice within the study summarized above for Compound A to establish a full dose response. The combined dose and concentration response of lacosamide from these four studies is shown in Figure 10. Lacosamide showed a dose- and concentration-dependent effect on AC-MES-induced tonic seizures, with a maximal effect at 20 mg / kg, where only 3 / 8 animals had a seizure (mean plasma concentration 23.9 μM). The proportion of animals with seizures in the 20 mg / kg lacosamide treatment group was significantly different from the vehicle treatment group (p value = 0.0052). Concentration response curve analysis of lacosamide showed an EC of 21.6 μM for plasma. 50 and an EC of 22.2 μM for brain tissue. 50 For Study 2C, the Compound A + lacosamide combination study, doses of 10 mg / kg lacosamide and 3 mg / kg Compound A were selected, both of which showed minimal efficacy when administered alone in the AC-MES model.

[0233] Compound A (3 mg / kg administered PO 0.5 hours prior to AC-MES) in combination with lacosamide (10 mg / kg administered PO 2 hours prior to AC-MES) abolished tonic seizures (Table 12). The difference between the lacosamide treatment group (10 mg / kg lacosamide alone, 6 / 8 (6 out of 8) animals with seizures) and the combination group (Compound A + lacosamide, 0 / 8 animals with seizures) was statistically significant (p value = 0.0189), but the difference between the Compound A treatment group (3 mg / kg Compound A alone, 2 / 8 animals with seizures) and the combination group was not statistically significant. The plasma concentrations of Compound A (administered at 3 mg / kg) and lacosamide (administered at 10 mg / kg) were similar when administered alone or in combination, suggesting that the greater efficacy observed in the combination group was not the result of higher exposure of either compound. The mean plasma concentrations achieved were: 0.284 μM for Compound A when administered alone at 3 mg / kg; 0.279 μM for Compound A when administered in combination with lacosamide at 3 mg / kg; 17.1 μM for lacosamide when administered alone at 10 mg / kg; and 15.5 μM for lacosamide when administered in combination with Compound A at 10 mg / kg.

[0234] [Table 16] B / P: brain-to-plasma ratio; n / a: not applicable.

[0235] [Table 17] B / P: brain-to-plasma ratio; n / a: not applicable.

[0236] [Table 18] B / P: brain-to-plasma ratio; n / a: not applicable.

[0237] [Table 19] B / P: brain-to-plasma ratio; n / a: not applicable.

[0238] 2.1 Conclusion Compound A and lacosamide demonstrated concentration-dependent efficacy in the CF-1 mouse AC-MES assay. EC values ​​of Compound A in plasma and brain predicted from concentration-response curve analysis. 50 The values ​​were 0.30 μM and 0.47 μM, respectively. The combination of 3 mg / kg Compound A and 10 mg / kg lacosamide resulted in complete suppression of tonic seizures in the mouse AC-MES assay, compared to the partial suppression observed with Compound A or lacosamide administered alone.

[0239] Five oral (PO) doses of Compound A were tested in the AC-MES model, respectively. Compound A was administered by oral gavage at 1 mg / kg (n=8), 3 mg / kg (n=8), 5 mg / kg (n=16), 7.5 mg / kg (n=7), and 10 mg / kg (n=17) to male CF-1 mice (body weight 28.2-43.8 g) 30 min after which these mice were subjected to 60 Hz corneal electrical stimulation (0.2 s duration, 40 mA). This stimulation induced tonic hindlimb extension in all vehicle-treated animals. Any animals that did not show hindlimb extension in response to the electrical stimulation were considered protected. Observational neurological evaluation (qualitative test) was also performed at the time of testing as a tolerability screen. Terminal plasma and brain samples were collected from all animals to obtain the levels of Compound A concentration and understand the relationship between efficacy and drug concentration.

[0240] After a single oral dose, Compound A showed a concentration-dependent effect on AC-MES-induced tonic seizures, with the maximal effect of seizures in 0 / 8 animals being obtained at a plasma concentration of 0.47 μM (from Study 2B at 5 mg / kg). In Study 2A (1 mg / kg, n=8; 5 mg / kg, n=8; and 10 mg / kg, n=8; study date March 28, 2019), the percentage of animals that showed a tonic seizure response to AC-MES stimulation after 30 min pretreatment with Compound A was reduced to 8 / 8 (mean plasma concentration of 0.0815 μM; 1 mg / kg), 7 / 8 (mean plasma concentration of 0.28 μM; 5 mg / kg), and 3 / 8 (mean plasma concentration of 0.37 μM; 10 mg / kg). In Study 2B (5 mg / kg, n=8; 7.5 mg / kg, n=7 and 10 mg / kg, n=9; study date June 5, 2019), the 5 mg / kg (mean plasma concentration of 0.47 μM) and 10 mg / kg (mean plasma concentration of 0.55 μM) doses of Compound A showed strong efficacy (0 / 8 and 1 / 9 animals seizures, respectively). In Study 2C (3 mg / kg, n=8; study date June 11, 2019), the 3 mg / kg dose of Compound A showed 2 / 8 animals seizures with a plasma concentration of 0.28 μM. In all three efficacy studies (2A, 2B, and 2C), the differences between Compound A-treated and vehicle-treated groups reached statistical significance. Data from all three studies were combined for concentration-response curve analysis of Compound A when administered alone. The concentration-response curve for Compound A was calculated with a half-maximum effective concentration (EC 50 ) and an EC of 0.47 μM for brain tissue. 50 showed.

[0241] One mouse from the 10 mg / kg group in Study 2A and three mice in Study 2B (two mice from the 10 mg / kg group and one mouse from the 7.5 mg / kg group) showed behavioral signs (tremors, decreased activity and hindlimb spreading) (plasma concentrations: 2A: 0.39 μM; 2B: 10 mg / kg: 0.61 and 0.52 μM, 7.5 mg / kg: 0.75 μM).

[0242] Prior to testing Compound A in combination with lacosamide in the AC-MES model (Study 2C), lacosamide was tested alone at different doses to establish a full dose response. Each of the four PO doses of lacosamide was tested separately in the AC-MES model (Study 2D: 6 mg / kg and 20 mg / kg, n=8 per group, Study Date March 14, 2019; Study 2A: 6 mg / kg and 8 mg / kg, n=8 per group, Study Date May 28, 2019; Study 2B: 10 mg / kg, n=8, Study Date June 5, 2019; and Study 2C: 10 mg / kg, n=8, Study Date June 11, 2019) and as part of the study. Lacosamide was administered by oral gavage to male CF-1 mice (body weight 28.2-43.8 g) at 6 mg / kg (n=16), 8 mg / kg (n=8), 10 mg / kg (n=16), and 20 mg / kg (n=8) 2 hours prior to AC-MES stimulation. Lacosamide demonstrated dose- and concentration-dependent effects on AC-MES-induced tonic seizures, with a maximal effect at 20 mg / kg with 3 / 8 animals seizing (mean plasma concentration of 23.9 μM) and minimal effects at the lower doses of 6, 8, and 10 mg / kg. Data from all four studies (2A, 2B, 2C, and 2D) were combined for concentration-response curve analysis of lacosamide when administered alone. The concentration-response curve for lacosamide showed an EC of 21.6 μM for plasma. 50 and an EC of 22.2 μM for brain tissue. 50 Based on dose-response studies, doses of 10 mg / kg lacosamide and 3 mg / kg Compound A, which showed minimal efficacy when administered alone, were selected for combination testing in the AC-MES model.

[0243] In male CF-1 mice, the combination of Compound A (3 mg / kg PO, 0.5 hours prior to AC-MES) with Lacosamide (10 mg / kg PO, 2 hours prior to AC-MES) completely suppressed tonic seizures compared to the partial suppression when Compound A or Lacosamide were administered alone (Compound A alone at 3 mg / kg: 2 / 8 animals seizures; Lacosamide alone at 10 mg / kg: 6 / 8 animals seizures; Compound A + Lacosamide: 0 / 8 animals seizures; Vehicle: 8 / 8 animals seizures). Plasma concentrations of Compound A (3 mg / kg) and Lacosamide (10 mg / kg) were similar when administered alone or in combination, suggesting that the maximum efficacy observed in the combination group was not the result of higher exposure of either compound. The mean total plasma concentrations achieved were as follows: Compound A when administered alone at 3 mg / kg: 0.283 μM; Compound A when administered in combination at 3 mg / kg: 0.279 μM; Lacosamide when administered alone at 10 mg / kg: 17.1 μM; Lacosamide when administered in combination at 10 mg / kg: 15.5 μM.

[0244] 3. Example 3. Compound A alone and in combination with levetiracetam in the 6 Hz psychomotor seizure assay The purpose of this study was to evaluate the interaction between Compound A and levetiracetam in a mouse model of seizures sensitive to each compound, the 6Hz psychomotor seizure assay (Barton et al., Epilepsy Res. 2001;47(3):217-227). Compound A and levetiracetam were administered alone or in combination at doses that yielded submaximal efficacy on their own to evaluate whether the combination of both compounds favored or unfavored efficacy. Compound A and levetiracetam were administered alone at doses that yielded submaximal efficacy in the 6Hz assay, allowing detection of changes in efficacy in either direction when these compounds were combined. Plasma and brain samples were analyzed to understand how the combination of Compound A and levetiracetam may affect pharmacokinetics and pharmacodynamics.

[0245] 3.1 Materials and Methods Test compound - Compound A [Table 20]

[0246] Reference compound – Levetiracetam [Table 21]

[0247] Vehicle: Compound A Formulation F2: 5% dimethylsulfoxide (DMSO), 0.5% methylcellulose in deionized (DI) water. Levetiracetam Formulation F1: 0.5% w:w methylcellulose, 0.2% v:v Tween 80 in DI water.

[0248] Dosing formulation: An appropriate amount of Compound A (uncorrected for purity) was weighed and dissolved in DMSO at 20x the intended final concentration. This 20x DMSO stock solution of Compound A was diluted 20x with 0.5% methylcellulose in DI water to achieve the final desired concentration. The resulting Compound A suspension was stirred or vortexed to obtain a homogenous suspension. The formulation was kept at room temperature and continuously stirred or vortexed before each dose administration.

[0249] For levetiracetam, DI water (0.8 L) was heated to 70°C-80°C. Five grams of methylcellulose was weighed and slowly added in small portions to the heated DI water. The mixture was stirred until it formed a homogenous milky suspension. The suspension was transferred to a cold room and stirred overnight to obtain a clear solution. 2 mL of Tween 80 was added to the clear solution and diluted to 1 L with DI water. This vehicle solution was stored at 2°C-8°C.

[0250] Levetiracetam powder was weighed into a vial. The appropriate amount of vehicle was added to the powder and then mixed with an IKA-T18 ULTRA TURRAX homogenizer to create a homogenous suspension of the desired concentration. The vial was then wrapped in aluminum foil to protect it from light and placed on a stir plate until administration.

[0251] Test system [Table 22]

[0252] Experimental Design: Animals were assigned to treatment groups as shown in Table 14. All mice received either vehicle or Compound A via PO gavage (Standard Operating Procedure (SOP) TECH-006) and either vehicle or levetiracetam by IP injection (TECH-004) 1 hour prior to seizure induction. ED of Compound A 20 (4 mg / kg) was selected for the combination experiment (based on dose-response experiment, Study 3A). A levetiracetam dose of 300 mg / kg was selected based on previous experiments which yielded an average efficacy of 35% at this dose.

[0253] [Table 23] Admin: administration; Conc.: concentration; Cpmd.: compound; IP: intraperitoneal; Lev: levetiracetam; n / a: not applicable; PO: per os, orally.

[0254] Randomization and blinding: The experimenter administering the compound randomly assigned each animal to a treatment group. A different experimenter blinded to the treatment group assignment performed the tests. All animals tested in a given experiment had an equal chance of being assigned to any treatment group.

[0255] Observation of clinical signs: All animals administered compounds were observed for abnormal behavior for 10 minutes after administration by the experimenter who administered the animal, and again for 10 minutes at the time of stimulation by the experimenter who tested the animal. Any qualitative changes from normal behavior were recorded.

[0256] 6Hz Psychomotor Seizure Assay: All animals were placed in the experimental room at least 1 hour before electrical stimulation. Just before the assay, a drop of Alcaine (Proparacaine Hydrochloride, 0.5%) was applied to each of the mouse's eyes. The animals were then securely restrained and a pair of electrodes was attached to the eyes (Electro Convulsive Therapy Unit 57800, Ugo Basile). A 3-second stimulation was induced with a foot pedal at 34 milliamps, 6 Hz, and 0.2 ms pulse width. Immediately after completion of stimulation, the animals were placed in a Plexiglas cylinder and seizure behaviors (chin clonus, forelimb clonus, and Straub's tail) were recorded (by TECH 036). Animals were considered to be protected from seizures if they did not show any of the three typical psychomotor seizure behaviors (chin clonus, forelimb clonus, Straub's tail) induced by this stimulation protocol in control animals.

[0257] Sample collection and preparation: Mice were anesthetized by isoflurane inhalation until they reached a surgical plane of anesthesia (TECH-018). A syringe (1 mL syringe with a 22-gauge needle) was then inserted substernally into the heart (TECH-031). Approximately 0.5 mL of blood was collected, placed in a K2EDTA tube, and stored on ice. Animals were then euthanized by cervical dislocation (TECH-018). Brains were removed, placed in pre-weighed vials, and snap frozen on dry ice. At the end of sample collection, blood was centrifuged at 4000 rpm for 10 min at 4°C, and plasma was transferred by pipette into labeled tubes. All samples were stored in a -80°C freezer until bioanalysis (8 days after sample collection for Study 3A and 3B; 20 days after sample collection for Study 3C).

[0258] Analysis of plasma and tissue samples: Bead mill polypropylene tubes containing weighed brain tissue were thawed at room temperature and 3 mL of homogenization solvent (water:acetonitrile (1:1, v:v)) was added. The tubes were placed in a bead mill homogenizer (Bead Ruptor Elite Model, Omni International) and shaken for one cycle lasting 30 seconds at a speed of 3.70 m / s. The homogenized tubes were centrifuged at 4000 rpm for 20 minutes and the supernatants were transferred to 1.5 mL Eppendorf tubes and stored frozen at -80 °C until analysis. All samples, including plasma and brain homogenate extracts (including calibration and quality control (QC) samples prepared in K2EDTA rat plasma), were extracted by protein precipitation. To each 50 μL aliquot of sample, 50 μL of internal standard solution ((S)-5-chloro-4-((1-(5-chloro-2-fluorophenyl)ethyl)amino)-2-fluoro-N-(pyrazin-2-yl)benzenesulfonamide and 4-((2-((7-azabicyclo[2.2.1]heptan-7-yl)methyl)-6-fluorobenzyl)amino)-2,6-difluoro-N-(isothiazol-3-yl)benzenesulfonamide at 2500 ng / mL in water / acetonitrile (1:1) for Compound A and levetiracetam, respectively), 50 μL of 6% (v / v) aqueous phosphoric acid solution, followed by 200 μL of acetonitrile. Samples in 1.5 mL tubes were vortex mixed and then centrifuged at 13,000 rpm for 20 min. Fifteen microliters of the supernatant was mixed with 150 μL of water / acetonitrile (1:1) in a 96-well plate and centrifuged at 4000 rpm for 20 min after mixing, and then was ready for analysis by liquid chromatography tandem mass spectrometry (LC-MS / MS).

[0259] Samples were analyzed by a research grade LC-MS / MS method as follows. [Table 24(1)] [Table 24(2)]

[0260] Data processing and analysis: Statistical data analysis was performed using GraphPad Prism (version 8.2.1). Dose-response data were analyzed using the Kruskal-Wallis test followed by Dunn's multiple comparisons. A p-value <0.05 was considered significant. Dose-response and concentration-response curves were generated using the Hill-Langmuir equation. formula 1 Y = B + (TB) × x n / (I C 50 n +x n ) In the above formula 1, B = the lowest value, set to 0; T = maximum value, set as 1, n = Hill coefficient, constrained to be less than zero, ·I C 50 = concentration of compound required for 50% inhibition in vitro It is.

[0261] Unless otherwise stated, all data are reported to three significant figures and all group data are reported as the mean ± SD.

[0262] 3.2 Results Bioanalysis: All system suitability tests, QC samples, matrices, and solvent blanks met the acceptance criteria. Analytical parameters are summarized in Table 15. QC samples at each concentration were analyzed in triplicate.

[0263] [Table 25] Lev: levetiracetam; LLOQ: lower limit of quantification; n / a: not applicable; ULOQ: upper limit of quantification.

[0264] Clinical Observations: One animal dosed with 8 mg / kg Compound A (plasma: 1.16 μM; brain: 1.65 μM) exhibited tremors and was cold to the touch when tested.

[0265] Dose response of Compound A in the 6 Hz psychomotor seizure assay (Study 3A): Compound A administered PO 1 hour prior to corneal stimulation showed dose-dependent efficacy with 8 / 8 animals experiencing seizures at 1 mg / kg and 3 mg / kg, 5 / 8 animals experiencing seizures at 5 mg / kg, and 3 / 8 animals experiencing seizures at 8 mg / kg. Seizure protection was significantly different from vehicle at 8 mg / kg Compound A (p=0.0081; FIG. 12A). The dose-response curve showed an ED of 6.48 mg / kg with a Hill coefficient of n=-3.09. 50 and ED of 4.13 mg / kg. 20 (Figure 12B).

[0266] The plasma and brain pharmacokinetic-pharmacodynamic relationships of individual animals are shown in Figures 12A and 12B, respectively. Results and exposure by group are summarized in Table 16. The efficacy of Compound A was concentration-dependent, with plasma EC 50 is 0.35 μM, and the brain EC 50 The α-to-α concentration was 0.54 μM, with Hill coefficients of n=-1.95 and n=-2.17, respectively (FIGS. 12C and 12D).

[0267] [Table 26] n / a: Not applicable.

[0268] Combination of Compound A and Levetiracetam in the 6 Hz Psychomotor Seizure Assay (Studies 3B and 3C): Submaximal doses were selected for each compound to assess whether the combination of Compound A with Levetiracetam was favorable or unfavorable for efficacy in the 6 Hz Psychomotor Seizure Assay. ED of Compound A 20 was determined to be 4 mg / kg in the dose-response experiment described above. In previous studies, an average efficacy of 35% was achieved with 300 mg / kg levetiracetam, with considerable variability between studies. In two similarly designed experiments (Studies 3B and 3C, see above and Table 14), the combination of 4 mg / kg Compound A and 300 mg / kg levetiracetam was significantly more effective than either compound alone in the 6 Hz psychomotor seizure assay (Table 16).

[0269] Study 3B: In Study 3B, Compound A or levetiracetam administered alone did not provide protection from seizures (plasma and brain concentrations of Compound A were much lower than expected), but combining both compounds protected 3 / 8 animals from seizures (Figure 14A, Table 12). The benefit of combination administration was significant compared to vehicle and compared to either compound administered alone (p=0.034). Compound A and levetiracetam concentrations achieved in plasma (Compound A: 0.014±0.009 μM; Levetiracetam: 1500±320 μM) and brain (Compound A: 0.03±0.02 μM; Levetiracetam: 861±120 μM) were comparable between the single and combined groups (Figure 14, Table 17). Therefore, the beneficial effect of the combination of Compound A and levetiracetam cannot be explained by increased exposure of either compound (Figure 15).

[0270] Study 3C: In Study 3C, the design of Study 3B was repeated. This time, the same dose of 4 mg / kg Compound A resulted in more than 10-fold higher concentrations of Compound A in plasma (0.17±0.09 μM vs. 0.01±0.01 μM) and brain (0.41±0.23 μM vs. 0.03±0.02 μM) than in Study 3B (Figure 15, Table 17). The concentration of levetiracetam was also slightly higher than in Study 3B (brain: 1130±130 μM vs. 861±120 μM, plasma: 1770±287 μM vs. 1500±320 μM; Figure 15, Table 17). The increased exposure led to increased efficacy of either compound administered alone compared to Study 3B: Compound A at 4 mg / kg protected 1 / 7 animals, and levetiracetam at 300 mg / kg protected 2 / 8 animals from seizures (Figure 14A, Table 17). The combination of Compound A and levetiracetam protected all animals tested in this study (Figure 14A, Table 17; n=7; one animal was excluded as it did not have measurable levels of levetiracetam in plasma or brain). Full efficacy was reached after the combination of Compound A and levetiracetam, so the effect on efficacy was significantly different from vehicle (p=0.0002) as well as from either compound administered alone (p<0.01). As in Study 3B, the concentrations of Compound A and levetiracetam reached in plasma and brain were comparable between the single and combination groups (Figure 15, Table 17). Therefore, the beneficial effect of the combination of Compound A and levetiracetam cannot be explained by increased exposure of either compound (Figure 16).

[0271] Combination of Study 3B and 3C: Combining the dose response of both experiments, the maximum efficacy of Compound A or Levetiracetam administered alone was achieved with Levetiracetam in 14 / 16 seizures, while the combination of both compounds resulted in seizures in 5 / 15 animals. Thus, the combination of Compound A and Levetiracetam protected 66.7% of animals from seizures, which was significantly different from vehicle (p<0.0001) and either compound alone (p<0.001, FIG. 14B, Table 17).

[0272] [Table 27]

[0273] 3.3 Conclusion Compound A demonstrated dose-dependent efficacy in the mouse 6 Hz psychomotor assay, with ED 50 The efficacy was a plasma EC50 of 0.35 μM. 50 and 0.54 μM brain EC 50 The plasma and brain concentrations of Compound A correlated well with those of the control.

[0274] The combination of 4 mg / kg Compound A and 300 mg / kg Levetiracetam resulted in significantly greater efficacy in this assay than that observed with either compound alone in two separate identically designed experiments (Study 3B: p=0.034; Study 3C: p<0.01). Coadministration did not significantly affect plasma or brain exposure of either compound. At comparable brain concentrations of Levetiracetam (1130±130 μM vs. 1100±265 μM; Study 3C), the addition of Compound A to Levetiracetam increased efficacy from 25% (Levetiracetam alone) to 100% (Compound A + Levetiracetam).

[0275] First, a dose-response study (3A) was performed with Compound A in the 6 Hz psychomotor assay to determine the dose at which 20% of animals were protected from seizures (ED 20 ) was determined. Compound A was administered by oral gavage to male CF-1 mice (body weight: 32.7-46.0 g) at 1, 3, 5, or 8 mg / kg, eight animals per group, and one hour later psychomotor seizures were induced via 3-second corneal stimulation at 34 milliamps (6 Hz, 0.2 ms pulse width). Efficacy in this assay was quantified by calculating the percentage of animals in each group of eight that were protected from seizure behavior (chin clonus, forelimb clonus, or Straub's tail lift) after 6 Hz stimulation. Plasma and brain samples were analyzed to assess concentration-response relationships.

[0276] Compound A at 1 mg / kg and 3 mg / kg did not provide protection from seizures, but the percentage of animals with seizures was reduced to 5 / 8 in animals receiving 5 mg / kg and 3 / 8 in the 8 mg / kg group. At 8 mg / kg (plasma: 0.47 ± 0.29 μM; brain: 0.70 ± 0.41 μM; mean ± SD), Compound A provided significant protection compared to the vehicle group (8 / 8 seizures) (p = 0.0081). Dose and concentration response curves for Compound A showed a dose at which 50% of animals were protected from seizures (ED 50 ) was 6.48 mg / kg, and the plasma concentration at which 50% of the animals were protected from seizures (EC 50 ) at 0.35 μM, and brain EC 50 The ED of Compound A in this assay is predicted to be 0.54 μM. 20 was calculated to be 4.13 mg / kg.

[0277] One animal dosed with 8 mg / kg Compound A exhibited tremors and was cold to the touch. With a plasma concentration of 1.16 μM and a brain concentration of 1.65 μM, this animal had the highest exposure to Compound A of all animals in this study.

[0278] The combination of Compound A and levetiracetam at doses resulting in submaximal efficacy was evaluated in two separate, similarly designed experiments (Studies 3B and 3C). Previous experiments had established that an intraperitoneal (IP) injection of 300 mg / kg levetiracetam 1 hour prior to stimulation resulted in 35% efficacy at 34 milliamps of 6 Hz stimulation. A dose-response experiment with Compound A (Study 3A) demonstrated an ED of 4 mg / kg when Compound A was administered orally (PO) 1 hour prior to stimulation. 20The following experimental groups (n=8) were then evaluated in each of two identically designed experiments: 1. Vehicle control (PO and IP); 2. Compound A at 4mg / kg PO and vehicle IP; 3. Levetiracetam at 300mg / kg IP and vehicle PO; 4. Compound A at 4mg / kg PO and levetiracetam at 300mg / kg IP. A 3-second corneal stimulation at 34 milliamps (6Hz, 0.2ms pulse width) was administered 1 hour after compound administration. Efficacy was evaluated based on the percentage of animals protected from seizure behavior (chin clonus, forelimb clonus, or Straub tail lift).

[0279] In study 3B, none of the compounds administered alone to male CF-1 mice (body weight: 29.0-37.3 g) provided protection from seizures, whereas the combination of both compounds protected 3 / 8 animals from seizures (p=0.034). Compound A and levetiracetam concentrations reached in plasma (compound A: 0.014±0.009 μM; levetiracetam: 1500±320 μM) and brain (compound A: 0.03±0.02 μM; levetiracetam: 861±120 μM) were comparable between the single and combined groups, but overall were much lower than would be expected for a 4 mg / kg dose.

[0280] In study 3C, performed in male CF-1 mice (body weight: 31.0-40.0 g), 4 mg / kg compound A resulted in more than 10-fold higher concentrations of compound A in plasma (0.167 ± 0.0897 μM vs. 0.014 ± 0.009 μM) and brain (0.41 ± 0.23 μM vs. 0.03 ± 0.02 μM) than in study 3B. Concentrations of levetiracetam were also slightly higher than in 3B (brain: 1130 ± 130 μM vs. 861 ± 120 μM; plasma: 1770 ± 287 μM vs. 1500 ± 320 μM). The increased exposure led to increased efficacy of either compound administered alone: ​​4 mg / kg compound A protected 1 / 7 animals and 300 mg / kg levetiracetam protected 2 / 8 animals from seizures. The combination of Compound A and levetiracetam protected all animals tested in this group compared to either compound alone (7 / 7 protection; p<0.01). At comparable brain concentrations of levetiracetam (1130±130 μM vs. 1100±265 μM, Study 3C), the addition of Compound A to levetiracetam increased efficacy from 25% (levetiracetam alone) to 100% (Compound A + levetiracetam).

[0281] 4. Example 4. Anticonvulsant Effects of Compound A Alone and in Combination with Cenobamate in the AC-MES Assay in CF-1 Mice The efficacy of Compound A and its pharmacological interaction with cenobamate were evaluated after oral administration in the mouse alternating current maximal electroshock convulsion (AC-MES) assay (see Example 2).

[0282] The objective of this study was to characterize the dose-dependent anticonvulsant activity of Compound A in the AC-MES assay in mice after a single oral dose of the compound and its pharmacological interaction with cenobamate. The anticonvulsant potency of Compound A and cenobamate, as well as the effect of combining Compound A with cenobamate, were tested in the AC-MES assay in male CF-1 mice after a single PO dose. Plasma and brain samples were obtained to understand the relationship between drug concentration and efficacy.

[0283] 4.1 Materials and Methods The Compound A, vehicle, dosing formulation, and test system used in Example 4 were the same as those used in Example 2, with cenobamate replacing lacosamide.

[0284] Test Compound - Cenobamate [Table 28]

[0285] Experimental Design: Animals were assigned to treatment groups as shown in Tables 18-19. Four studies were conducted, each with Compound A and cenobamate. The dates of each study are shown in Tables 18-19.

[0286] [Table 29] PO: per os, orally.

[0287] [Table 30] PO: per os, orally.

[0288] [Table 31] PO: per os, orally.

[0289] [Table 32] PO: per os, orally.

[0290] [Table 33] PO: per os, orally.

[0291] [Table 34] PO: per os, orally.

[0292] [Table 35] PO: per os, orally.

[0293] Blinding and randomization, clinical observations, AC-MES assays, sample collection and preparation, analysis of plasma and brain samples, and bioanalytical procedures in Example 4 were the same as those used in Example 2.

[0294] Samples were analyzed for Compound A by a research grade UHPLC-MS / MS method using the conditions below. [Table 36]

[0295] Samples were analyzed for cenobamate by a research grade UHPLC-MS / MS method using the conditions below. [Table 37]

[0296] The determination of sample concentrations and data processing and analysis in Example 4 were the same as in Example 2.

[0297] 4.2 Results Bioanalysis: All system suitability tests (SSTs), QC, matrix, and solvent blanks met the acceptance criteria described in SOP MTD-066. The lower limit of quantitation (LLOQ) and upper limit of quantitation (ULOQ) for Compound A and cenobamate are shown in Table 25.

[0298] [Table 38] LLOQ: Lower limit of quantification. ULOQ: Upper limit of quantification

[0299] Efficacy in AC-MES assay: The results of the AC-MES induced seizure study in mice are shown in Tables 26-32 and Figures 17, 18 and 19.

[0300] Compound A alone: ​​Data from the dose and concentration response of Compound A are summarized in Figure 17, Table 26, Table 27 and Table 28. In all three efficacy studies (4A, 4B, 4C and 4F), 100% of vehicle-treated CF-1 mice exhibited tonic seizures with hind limb extension. In Study 4A (1 mg / kg, n=8; 5 mg / kg, n=8; and 10 mg / kg, n=8), the percentage of animals exhibiting tonic seizure responses to AC-MES stimulation after 30 minutes pretreatment with Compound A decreased from 8 / 8 (mean plasma concentration of 0.08 μM; 1 mg / kg), to 7 / 8 (mean plasma concentration of 0.28 μM; 5 mg / kg), and 3 / 8 (mean plasma concentration of 0.37 μM; 10 mg / kg). In Study 4B (5 mg / kg, n=8; 7.5 mg / kg, n=7 and 10 mg / kg, n=9), the 5 mg / kg (mean plasma concentration of 0.47 μM) and 10 mg / kg (mean plasma concentration of 0.55 μM) doses of Compound A showed strong efficacy (0 / 8 and 1 / 9 animals had seizures, respectively). In Study 4C (3 mg / kg, n=8), the 3 mg / kg dose of Compound A showed 2 / 8 animals had seizures, with a plasma concentration of 0.28 μM. In Study 4F (2 mg / kg, n=8), the 2 mg / kg dose showed 5 / 8 animals had seizures, with a plasma concentration of 0.11 μM. In all efficacy studies except 4F, the difference between the Compound A-treated group and the vehicle-treated group reached statistical significance (p values ​​are shown in FIG. 17).

[0301] One mouse from the 10 mg / kg group in Study 4A and three mice in Study 4B (two mice from the 10 mg / kg group and one mouse from the 7.5 mg / kg group) showed behavioral signs (tremors, decreased activity, and hindlimb spreading) (plasma concentrations: 4A: 0.39 μM; 4B: 10 mg / kg: 0.61 and 0.52 μM, 7.5 mg / kg: 0.75 μM).

[0302] The combined concentration-response curves for Compound A from three studies showed an EC50 and an EC of 0.47 μM for brain tissue. 50 The concentrations of Compound A achieved in brain tissue and plasma were linear with respect to dose (Figure 17).

[0303] Cenobamate alone: ​​Before testing Compound A in combination with cenobamate in the AC-MES model (Studies 4F and 4G), we tested cenobamate alone at different doses to establish a complete dose response. Data from the dose and concentration response of cenobamate are summarized in Figure 18, Table 29, and Table 30. Seven PO doses of cenobamate were each tested in the AC-MES model separately (Study 4D: 3 mg / kg, 10 mg / kg, and 30 mg / kg, n=8 per group; Study 4E: 3 mg / kg, 5 mg / kg, and 7.5 mg / kg, n=8; and Study 4F: 5 mg / kg, n=8) and as part of this study. Cenobamate was administered by oral gavage to male CF-1 mice at 3 mg / kg (n=15), 5 mg / kg (n=7), 7.5 mg / kg (n=7), 10 mg / kg (n=8), and 30 mg / kg (n=8) 2 hours prior to AC-MES stimulation. Cenobamate demonstrated dose- and concentration-dependent effects on AC-MES-induced tonic seizures, with 3 / 7 animals at 7.5 mg / kg having seizures (mean plasma concentration of 78.1 μM), 1 / 8 animals at 10 mg / kg having seizures (mean plasma concentration of 87 μM), and 0 / 8 animals at 30 mg / kg having seizures (mean plasma concentration of 24 μM), with minimal effects at the lower doses of 3 and 5 mg / kg. Data from all three studies (4D, 4E, and 4F) were combined for concentration-response curve analysis of cenobamate when administered alone (FIG. 18). The concentration-response curve for cenobamate showed an EC 50 and an EC of 25.2 μM for brain tissue. 50 showed.

[0304] Combination of Compound A and Cenobamate: Based on dose-response studies, a dose of 5 mg / kg cenobamate and doses of 0.5, 1, and 2 mg / kg Compound A, which showed minimal efficacy when administered alone, were selected for combination studies in the AC-MES model.

[0305] Data from the combination studies are summarized in Figure 19, Table 31, and Table 32. We performed two combination studies: 4F (2 mg / kg Compound A combined with 5 mg / kg cenobamate) and 4G (0.5 and 1 mg / kg Compound A combined with 5 mg / kg cenobamate). Combining Compound A (0.5, 1, and 2 mg / kg PO, 0.5 hr prior to AC-MES) with cenobamate (5 mg / kg PO, 2 hr prior to AC-MES) in male CF-1 mice showed 2 / 8 animals with seizures at 0.5 mg / kg Compound A, 1 / 8 animals with seizures at 1 mg / kg Compound A, and 0 / 8 animals with 2 mg / kg Compound A, compared to partial or no effect with Compound A (5 / 8 animals with Compound A alone at 2 mg / kg) or cenobamate (8 / 8 animals with cenobamate alone at 5 mg / kg). In Study 4F, the plasma concentrations of Compound A (mean plasma concentration at 2 mg / kg: 0.11 μM when administered alone, 0.29 μM when administered in combination) and cenobamate (mean plasma concentration at 5 mg / kg: 31.3 μM when administered alone, 38.9 μM when administered in combination) were slightly higher when administered in combination compared to when administered alone. In Study 4G, the mean total plasma concentrations achieved were as follows: Compound A administered in combination at 0.5 mg / kg: 0.03 μM; Compound A administered in combination at 1 mg / kg: 0.11 μM; cenobamate administered in combination (+0.5 mg / kg Compound A) at 5 mg / kg: 41.1 μM; cenobamate administered in combination (+1 mg / kg Compound A) at 5 mg / kg: 39.7 μM. Concentration-response curves for Compound A and cenobamate administered in combination showed an extrapolated EC for Compound A of 0.01 μM for plasma and 0.03 μM for brain tissue.50 showed.

[0306] [Table 39] B / P: brain-to-plasma ratio; n / a: not applicable.

[0307] [Table 40] B / P: brain-to-plasma ratio; n / a: not applicable.

[0308] [Table 41] B / P: brain-to-plasma ratio; n / a: not applicable.

[0309] [Table 42] B / P: brain-to-plasma ratio; n / a: not applicable.

[0310] [Table 43] B / P: brain-to-plasma ratio; n / a: not applicable.

[0311] [Table 44] B / P: brain-to-plasma ratio; n / a: not applicable.

[0312] [Table 45]

[0313] 4.3 Conclusion Compound A and cenobamate demonstrated concentration-dependent efficacy in the CF-1 mouse AC-MES assay. EC values ​​of Compound A in plasma and brain predicted from concentration-response curve analysis. 50The combined concentration-response curves for Compound A and cenobamate administered in combination showed an EC value of 0.01 μM in plasma for Compound A. 50 and an EC of 0.03 μM for brain tissue. 50 These results predict an increase in potency of Compound A of 33.3-fold and 14.2-fold for plasma and brain tissue, respectively, when administered in combination with cenobamate.

[0314] Six oral (PO) doses of Compound A were tested in the AC-MES assay, respectively. Compound A was administered by oral gavage to male CF-1 mice at 1 mg / kg (n=8), 2 mg / kg (n=8), 3 mg / kg (n=8), 5 mg / kg (n=16), 7.5 mg / kg (n=7), and 10 mg / kg (n=17) 30 min after administration to the mice, they were subjected to a 60 Hz corneal electrical stimulation (0.2 sec duration, 40 mA). This stimulation induced tonic hind limb extension in all vehicle-treated animals. Any animals that did not show hind limb extension in response to the electrical stimulation were considered protected. Observational neurological evaluation (qualitative test) was also performed at the time of testing as a tolerability screen. Terminal plasma and brain samples were collected from all animals to obtain the levels of Compound A and cenobamate concentrations and understand the relationship between efficacy and drug concentration.

[0315] After a single oral dose, Compound A showed a concentration-dependent effect on AC-MES-induced tonic seizures, with the maximal effect of seizures in 0 / 8 animals being obtained at a plasma concentration of 0.470 μM (from Study 4B at 5 mg / kg). In Study 4A (1 mg / kg, n=8; 5 mg / kg, n=8; and 10 mg / kg, n=8), the percentage of animals that showed a tonic seizure response to AC-MES stimulation after 30 min pretreatment with Compound A decreased from 8 / 8 (mean plasma concentration of 0.08 μM; 1 mg / kg), to 7 / 8 (mean plasma concentration of 0.28 μM; 5 mg / kg), and 3 / 8 (mean plasma concentration of 0.37 μM; 10 mg / kg). In Study 4B (5 mg / kg, n=8; 7.5 mg / kg, n=7 and 10 mg / kg, n=9), Compound A at 5 mg / kg (mean plasma concentration of 0.47 μM) and 10 mg / kg (mean plasma concentration of 0.55 μM) doses showed strong efficacy (0 / 8 and 1 / 9 animals seizures, respectively). In Study 4C (3 mg / kg, n=8), Compound A at 3 mg / kg dose showed 2 / 8 animals seizures with a plasma concentration of 0.28 μM. In Study 4F (2 mg / kg, n=8), the 2 mg / kg dose showed 5 / 8 animals seizures with a plasma concentration of 0.11 μM. In all efficacy studies except 4F, the differences between Compound A-treated and vehicle-treated groups reached statistical significance (p values ​​are shown in FIG. 17). Data from all four studies were combined for concentration-response curve analysis of Compound A when administered alone. The concentration-response curve for Compound A was determined to be 0.30 μM in plasma at half-maximal effective concentration (EC 50 ) and an EC of 0.47 μM for brain tissue. 50 showed.

[0316] One mouse from the 10 mg / kg group in Study 4A and three mice in Study 4B (two mice from the 10 mg / kg group and one mouse from the 7.5 mg / kg group) showed behavioral signs (tremors, decreased activity and hindlimb spreading) (plasma concentrations: 4A: 0.39 μM; 4B: 10 mg / kg: 0.61 and 0.52 μM, 7.5 mg / kg: 0.75 μM).

[0317] Before testing Compound A in combination with cenobamate in the AC-MES model, cenobamate was tested alone at different doses to establish a complete dose response. Seven PO doses of cenobamate were tested separately and as part of the AC-MES model (Study 4D: 3 mg / kg, 10 mg / kg, and 30 mg / kg, n=8 per group; Study 4E: 3 mg / kg, 5 mg / kg, and 7.5 mg / kg, n=8; and Study 4F: 5 mg / kg, n=8). Male CF-1 mice were administered 3 mg / kg (n=15), 5 mg / kg (n=7), 7.5 mg / kg (n=7), 10 mg / kg (n=8), and 30 mg / kg (n=8) of cenobamate by oral gavage 2 hours before receiving the AC-MES stimulation. Cenobamate demonstrated dose- and concentration-dependent efficacy against AC-MES-induced tonic seizures, with 3 / 7 animals experiencing seizures at 7.5 mg / kg (mean plasma concentration of 78.1 μM), 1 / 8 animals experiencing seizures at 10 mg / kg (mean plasma concentration of 87 μM), and 0 / 8 animals experiencing seizures at 30 mg / kg (mean plasma concentration of 237 μM). Lower doses of 3 and 5 mg / kg had minimal effects, with 7 / 8 animals experiencing seizures. The percentage of animals experiencing seizures in the cenobamate-treated group was significantly different from the vehicle-treated group in two of the three studies (p values ​​are shown in FIG. 18). Data from all three studies (4D, 4E, and 4F) were combined for concentration-response curve analysis of cenobamate when administered alone. The concentration-response curve for cenobamate showed an EC of 70.5 μM for plasma. 50 and an EC of 25.2 μM for brain tissue. 50 showed.

[0318] Based on dose-response studies, doses of 5 mg / kg cenobamate and 0.5, 1, and 2 mg / kg Compound A, which showed minimal efficacy when administered alone, were selected for combination testing in the AC-MES model.

[0319] Two combination studies were performed: 4F (2 mg / kg Compound A combined with 5 mg / kg cenobamate) and 4G (0.5 and 1 mg / kg Compound A combined with 5 mg / kg cenobamate). Combining Compound A (0.5, 1, and 2 mg / kg PO, 0.5 hr prior to AC-MES) with cenobamate (5 mg / kg PO, 2 hr prior to AC-MES) in male CF-1 mice showed 2 / 8 animals with seizures at 0.5 mg / kg Compound A, 1 / 8 animals with seizures at 1 mg / kg Compound A, and 0 / 8 animals with 2 mg / kg Compound A, compared to 5 / 8 animals with Compound A administered alone at 2 mg / kg and 8 / 8 animals with cenobamate administered alone at 5 mg / kg. In Study 4F, the difference between the cenobamate-treated group and the combination group was statistically significant, but the difference between the Compound A-treated group and the combination group was not. In Study 4G, the difference between the vehicle group and the combination group was statistically significant (Note: vehicle control and Compound A and cenobamate alone control groups were not used in this study; p values ​​are shown in Figure 19). In Study 4F, the plasma concentrations of Compound A (mean plasma concentration at 2 mg / kg: 0.11 μM when administered alone and 0.29 μM when administered in combination) and cenobamate (mean plasma concentration at 5 mg / kg: 31.3 μM when administered alone and 38.9 μM when administered in combination) were slightly higher when administered in combination compared to when administered alone. In Study 4G, the mean total plasma concentrations achieved were as follows: Compound A when administered in combination at 0.5 mg / kg: 0.03 μM; Compound A when administered in combination at 1 mg / kg: 0.11 μM; Cenobamate when administered in combination at 5 mg / kg (0.5 mg / kg Compound A): 41.1 μM; Cenobamate when administered in combination at 5 mg / kg (1 mg / kg Compound A): 39.7 μM.

[0320] The integrated concentration-response curves for Compound A and cenobamate administered in combination showed an extrapolated EC for Compound A of 0.01 μM for plasma and 0.03 μM for brain tissue. 50These results show an increase in potency of Compound A of 33.3-fold and 14.2-fold for plasma and brain tissue, respectively, when administered in combination with cenobamate.

[0321] All U.S. patents, U.S. patent application publications, U.S. patent applications, foreign patents, foreign patent applications, and non-patent publications mentioned herein, including U.S. Provisional Application No. 63 / 147,736, filed February 9, 2021, are hereby incorporated by reference in their entireties.

[0322] Although the above compositions, methods, and uses have been described in some detail for ease of understanding, it will be apparent that certain changes and modifications may be practiced within the scope of the appended claims. Accordingly, the described embodiments are to be considered as illustrative rather than restrictive, and the claimed invention is not to be limited to the details given herein, but may be modified within the scope and equivalents of the appended claims.

Claims

1. An agent for use in treating a seizure disorder in a human in need thereof, said treatment comprising administering to said human a combination of said agent and an anticonvulsant (ASM), in amounts that are therapeutically effective when administered in combination; The agent for use, wherein the agent is N-[4-(6-fluoro-3,4-dihydro-1H-isoquinolin-2-yl)-2,6-dimethylphenyl]-3,3-dimethylbutanamide.

2. 1. An agent for use in reducing the amount of an anticonvulsant (ASM) required for therapeutic efficacy in a human suffering from a seizure disorder, said reduction comprising administering to said human an amount of said agent in combination with said ASM effective to achieve such reduction when administered with said ASM; The agent for use, wherein the agent is N-[4-(6-fluoro-3,4-dihydro-1H-isoquinolin-2-yl)-2,6-dimethylphenyl]-3,3-dimethylbutanamide.

3. 1. An agent for use in reducing the amount of an agent required for therapeutic efficacy in a human suffering from a seizure disorder, said reduction comprising administering to said human an anticonvulsant (ASM) in combination with said agent in an amount effective to achieve such reduction when administered with said agent; The agent for use, wherein the agent is N-[4-(6-fluoro-3,4-dihydro-1H-isoquinolin-2-yl)-2,6-dimethylphenyl]-3,3-dimethylbutanamide.

4. 4. The method of claim 1, further comprising enhancing the opening of Kv7 potassium channels in said human.

5. 1. An agent for use in enhancing the opening of Kv7 potassium channels in a human, said enhancing comprising administering to said human in combination said agent and an anticonvulsant (ASM) in amounts that are therapeutically effective when administered in combination; The agent is N-[4-(6-fluoro-3,4-dihydro-1H-isoquinolin-2-yl)-2,6-dimethylphenyl]-3,3-dimethylbutanamide; The method of claim 1, wherein the human has a seizure disorder.

6. 6. The agent for use according to claim 4 or claim 5, wherein the Kv7 potassium channel is one or more of Kv7.2, Kv7.3, Kv7.4 or Kv7.5, and the use may be selective for enhancing the opening of one or more of Kv7.2, Kv7.3, Kv7.4 or Kv7.5 over Kv7.

1.

7. 6. An agent for use according to claim 4 or claim 5, which comprises the opening of Kv7.2 / Kv7.3 (KCNQ2 / 3) potassium channels.

8. 8. The agent for use according to any one of claims 1 to 7, wherein the ASM is a benzodiazepine, carbamazepine, cenobamate, felbamate, gabapentin, lacosamide, lamotrigine, levetiracetam, oxcarbazepine, phenobarbital, phenytoin, pregabalin, rufinamide, tiagabine, topiramate, valproic acid, vigabatrin, zonisamide, or a combination thereof, and the ASM may be valproic acid, levetiracetam, phenytoin, lacosamide, or cenobamate. (i) the ASM does not enhance the opening of Kv7 potassium channels in the human; (ii) the ASM reduces neuronal excitability in the human; (a) the ASM may reduce neuronal excitability in the human by blocking sodium channels; (b) the ASM may reduce neuronal excitability in the human by blocking calcium channels; and / or (c) the ASM may reduce neuronal excitability in the human by binding to synaptic vesicle glycoprotein 2A (SV2A); and / or (iii) the ASM increases neuronal inhibition in the human Agent for use according to any one of claims 1 to 8.

10. 10. The method of claim 1, wherein the ASM is (a) a glutamatergic agent and / or (b) a GABAergic agent.

11. The seizure disorder is (i) associated with Kv7 potassium channel dysfunction; and / or (ii) focal onset epilepsy, photosensitive epilepsy, self-induced syncope, refractory epilepsy, Angelman syndrome, benign rolandic epilepsy, CDKL5 disorder, childhood absence epilepsy and juvenile absence epilepsy, Dravet syndrome, frontal lobe epilepsy, Glut1 deficiency, hypothalamic hamartoma, infantile spasms / West syndrome, juvenile myoclonic epilepsy, Landau-Kleffner syndrome, Lennox-Gastaut syndrome (LGS), myoclonic absence epilepsy, Otahara syndrome, Panaetopoulos syndrome, PCDH 11. The agent for use according to any one of claims 1 to 10, wherein the seizure disorder is selected from the group consisting of epilepsy-19, progressive myoclonic epilepsy, Rasmussen syndrome, ring chromosome 20 syndrome, reflex epilepsy, temporal lobe epilepsy, Lafora progressive myoclonic epilepsy, neurocutaneous syndrome, tuberous sclerosis, early infantile epileptic encephalopathy, early onset epileptic encephalopathy, generalized epilepsy febrile seizures plus, Rett syndrome, multiple sclerosis, or paroxysmal dyskinesia, and the seizure disorder may be focal onset epilepsy. (i) the agent is orally administered to the human; and / or (ii) the ASM is orally administered to the human Agent for use according to any one of claims 1 to 11.

13. (i) The agent is administered to the human at a dose of 2 to 100 mg, and the agent comprises (a) administering to the human 5 to 50 mg; (b) administering to the human 10 to 30 mg, and / or (c) administering 5, 10, 15, 20, 25, or 30 mg to the human may be administered at a dose of (ii) the agent is administered to the human at a dose of 2 to 100 mg per day, the agent comprising: (a) administering to the human 5 to 50 mg per day; (b) administering to the human 10 to 30 mg per day; or (c) administering 10, 15, 20, 25, or 30 mg per day to the human and / or (iii) the agent is administered to the human at a dose of 0.01 to 2.0 mg / kg, the agent comprising: (a) administering to the human a dose of 0.03 to 1.0 mg / kg; or (b) administering to the human a dose of 0.05 to 0.5 mg / kg. may be administered at a dose of Agent for use according to any one of claims 1 to 12.

14. An agent for use as described in any one of claims 1 to 13, wherein the agent is orally administered to the human between about 30 minutes prior to and about 2 hours after ingestion of a meal, and the agent may be orally administered to the human during a meal or within 15 minutes after ingestion of a meal.

15. The ASM is valproic acid, which is (i) 2 to 16 mg / kg to the human; or (ii) administering to the human a dose of 4 to 12 mg / kg.

15. The agent for use according to any one of claims 1 to 14, which may be administered in a dose of

16. The ASM is phenytoin, which is (i) 0.05 to 5 mg / kg to the human; or (ii) administering to the human a dose of 0.1 to 1 mg / kg 15. The agent for use according to any one of claims 1 to 14, which may be administered in a dose of

17. The ASM is lacosamide, which is (i) 0.1 to 5 mg / kg to the human; or (ii) administering to the human a dose of 0.5 to 1 mg / kg 15. The agent for use according to any one of claims 1 to 14, which may be administered in a dose of

18. The ASM is a cenobamate, and the cenobamate is (i) 0.05 to 5 mg / kg to the human; or (ii) administering to the human a dose of 0.1 to 1 mg / kg 15. The agent for use according to any one of claims 1 to 14, which may be administered in a dose of

19. An agent for use according to any one of claims 1 to 18, wherein combined administration of the agent and the ASM provides improved efficacy compared to individual administration of the agent alone or the ASM.

20. A pharmaceutical composition comprising an agent, an anticonvulsant (ASM), and a pharma- ceutical acceptable carrier, The pharmaceutical composition, wherein the agent is N-[4-(6-fluoro-3,4-dihydro-1H-isoquinolin-2-yl)-2,6-dimethylphenyl]-3,3-dimethylbutanamide.

21. The ASM is a benzodiazepine, carbamazepine, cenobamate, felbamate, gabapentin, lacosamide, lamotrigine, levetiracetam, oxcarbazepine, phenobarbital, phenytoin, pregabalin, rufinamide, tiagabine, topiramate, valproic acid, vigabatrin, zonisamide, or a combination thereof, and the ASM is (i) valproic acid, phenytoin, levetiracetam, lacosamide, or cenobamate, (ii) valproic acid, (iii) phenytoin, (iv) levetiracetam, (v) lacosamide, or (vi) cenobamate The pharmaceutical composition according to claim 20, which may be

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