WO2004035037A2

WO2004035037A2 - Derivatives of n-phenylanthranilic acid and 2-benzimidazolon as potassium channel and/or cortical neuron activity modulators

Info

Publication number: WO2004035037A2
Application number: PCT/IL2003/000855
Authority: WO
Inventors: Bernard Attali; Asher Peretz
Original assignee: Ramot At Tel Aviv University Ltd.
Priority date: 2002-10-21
Filing date: 2003-10-21
Publication date: 2004-04-29
Also published as: WO2004035037A3; AU2003272068A1; EP1553932A2; JP2006513154A; KR20050074493A; AU2003272068A8; CA2503075A1

Abstract

Compounds, compositions and methods are provided which are useful in the treatment of diseases through the modulation of potassium ion flux through voltage-dependent potassium channels and/or depressing cortical neuron activity. More particularly, the invention provides derivatives of N-phenylanthranilic acid or 2-benzimidazolone, compositions and methods that are useful in the treatment of central or peripheral nervous system disorders and as neuroprotective agents.

Description

DERIVATIVES OF N-PHENYLANTHRANILIC ACID AND

2-BENZIMIDAZOLONE AS POTASSIUM CHANNEL AND/OR CORTICAL

NEURON ACTIVITY MODULATORS

FIELD AND BACKGROUND OF THE INVENTION

The present invention relates to the field of pharmacology, and particularly to derivatives of N-phenylanthranilic acid and/or 2-benzimidazolone for the treatment of pathologies, especially pathologies related to potassium ion flux through voltage- dependent potassium channels and/or cortical neuron activity. Ion channels are cellular proteins that regulate the flow of ions, including calcium, potassium, sodium and chloride, into and out of cells. These channels are present in all animal cells and affect such processes as nerve transmission, muscle contraction and cellular secretion. Among the ion channels, potassium channels are the most ubiquitous and diverse, being found in a variety of animal cells such as nervous, muscular, glandular, immune, reproductive and epithelial tissue. These channels allow the flow of potassium in and/or out of the cell under certain conditions. For example, the outward flow of potassium ions upon opening of these channels makes the interior of the cell more negative, counteracting depolarizing voltages applied to the cell. These channels are regulated, e.g., by calcium sensitivity, voltage-gating, second messengers, extracellular ligands and ATP-sensitivity.

Potassium channels have now been associated with a number of physiological processes, including regulation of heartbeat, dilation of arteries, release of insulin, excitability of nerve cells, and regulation of renal electrolyte transport. Potassium channels are made by alpha subunits that fall into at least 8 families, based on predicted structural and functional similarities (Wei et al., Neuropharmacology 35(7): 805-829 (1997)). Three of these families (Kv, eag-related, and KQT) share a common motif of six transmembrane domains and are primarily gated by voltage. Two other families, CNG and SK/IK, also contain this motif but are gated by cyclic nucleotides and calcium, respectively. The three other families of potassium channel alpha subunits have distinct patterns of transmembrane domains. Slo family potassium channels, or BK channels have seven transmembrane domains (Meera et al., Proc. Natl. Acad. Sci. U.S.A. 94(25): 14066-71 (1997)) and are gated by both voltage and calcium or pH (Schreiber et al, J. Biol. Chem. 273: 3509-16 (1998)). Another family, the inward rectifier potassium channels (Kir), belongs to a structural family containing two transmembrane domains, and an eighth functionally diverse family (TP, or "two-pore") contains two tandem repeats of this inward rectifier motif.

Potassium channels are typically formed by four alpha subunits, and can be homomeric (made of identical alpha subunits) or heteromeric (made of two or more distinct types of alpha subunits). In addition, potassium channels made from Kv, KQT and Slo or BK subunits have often been found to contain additional, structurally distinct auxiliary, or beta, subunits. These subunits do not form potassium channels themselves, but instead they act as auxiliary subunits to modify the functional properties of channels formed by alpha subunits. For example, the Kv beta subunits are cytoplasmic and are known to increase the surface expression of Kv channels and/or modify inactivation kinetics of the channel (Heinemann et al, J. Physiol. 493: 625-633 (1996); Shi et al, Neuron 16(4): 843-852 (1996)). In another example, the KQT family beta subunit, minK, primarily changes activation kinetics (Sanguinetti et al, Nature 384: 80-83 (1996)). Slo or BK potassium channels are large conductance potassium channels found in a wide variety of tissues, both in the central nervous system and periphery. They play a key role in the regulation of processes such as neuronal integration, muscular contraction and hormone secretion. They may also be involved in processes such as lymphocyte differentiation and cell proliferation, spermatocyte differentiation and sperm motility. Three alpha subunits of the Slo family have been cloned, i.e., Slol, Slo2, and Slo3 (Butler et al, Science 261: 221-224 (1993); Schreiber et al, J. Biol. Chem., 273: 3509-16 (1998); and Joiner et al, Nature Neurosci. 1: 462-469 (1998)). These Slo family members have been shown to be voltage and/or calcium gated, and/or regulated by intracellular pH. Certain members of the Kv family of potassium channels were recently renamed (see, Biervert, et al, Science 279: 403-406 (1998)). KvLQTl was re-named KCNQ1, and the KvLQTl -related channels (KvLRl and KvLR2) were renamed KCNQ2 and KCNQ3, respectively. More recently, additional members of the KCNQ subfamily were identified. For example, KCNQ4 was identified as a channel expressed in sensory outer hair cells (Kubisch, et al, Cell 96(3): 437-446 (1999)). KCNQ5 (Kananura βt al, Neuroreport 11(9): 2063 (2000)), KCNQ2/3 (Main et al, Mol. Pharmacol. 58: 253-62 (2000), KCNQ3/5 (Wickenden et al, Br. J. Pharma 132: 381(2001)) and KCNQ6 have also recently been described. KCNQ2 and KCNQ3 have been shown to be nervous system-specific potassium channels associated with benign familial neonatal convulsions ("BFNC"), a class of idiopathic generalized epilepsy (see, Leppert, et al, Nature 337: 647-648

(1989)). These channels have been linked to M-current channels (see, Wang, et al, Science 282: 1890-1893 (1998)). The discovery and characterization of these channels and currents provides useful insights into how these voltage dependent (Kv) potassium channels function in different environments, and how they respond to various activation mechanisms. Such information has now led to the identification of modulators of KCNQ2 and KCNQ3 potassium channels or the M-current, and the use of such modulators as therapeutic agents.

A potassium channel opener that has gained much attention is retigabine (N-(2- amino-4-(4-fluorobenzylamino)-phenyl)carbamic acid ethyl ester). Retigabine was first described in European Patent No. 554,543. Compounds related to retigabine have also been proposed for use as potassium channel modulators, see for example U.S. Patent Application No. No. 10/022,579.

Retigabine is highly selective for potassium channels consisting of the subunits

KCNQ2 and KCNQ3. In addition, retigabine activates the homomultimerous channel, which contains only the subunit KCNQ2. Only marginal voltage-dependent currents are measurable in cells, which express only the homomeric channel from the KCNQ3 subunit, see U.S. 6,472,165.

U.S. Patent Application No. No. 10/075,521 teaches 2,4-disubstituted pyrimidine-5-carboxamide derivatives as KCNQ potassium channel modulators.

U.S. Patent Application No. 10/160,582 teaches cinnamide derivatives as KCNQ potassium channel modulators. U.S. Patent No. 5,565,483 and U.S. Patent Application Nos. 10/312,123,

10/075,703 and 10/075,522 teach 3-substituted oxindole derivatives as KCNQ potassium channel modulators.

U.S. Patent No. 5,384,330 teaches 1,2,4-triamino-benzene derivatives as KCNQ potassium channel modulators. U.S. Patent No. 6,593,349 teaches bisarylamines derivatives as KCNQ potassium channel modulators. The two aryl groups of the compounds taught in U.S. Patent No. 6,593,349 are a pyridine derivative and a five-membered heterocycle. A significant disadvantage of the KCNQ potassium channel modulators known in the art is that these are generally difficult to make, requiring complex multi-step syntheses and that in some cases these modulators are non-specific or even toxic.

There is, hence, a widely recognized need for, and it would be highly advantageous to have new and effective potassium channel modulators that are readily available and easy to synthesize.

SUMMARY OF THE INVENTION

The present invention provides compounds that are generally effective potassium channel modulators, especially voltage-dependent potassium channels such as KCNQ2 channel, KCNQ3 channels and KCNQ2/3 channels. Also, the present invention provides compounds that are generally effective at depressing cortical neuron activity. The compounds of the present invention are generally derivatives of N- phenylanthranilic acid or 2-benzimidazolone. Some of the compounds of the present invention are well known in the art and are readily available. Some of the compounds of the present invention are novel but are synthesized through a few (typically, one or two) high-yield steps from readily available starting materials.

According to one aspect of the present invention there is provided a method of modulating (preferably opening) a voltage-dependent potassium channel, the method comprising administering to the subject in need thereof a therapeutically effective amount of a compound selected from the group consisting of N-phenylanthranilic acid, a N-phenylanthranilic acid derivative, 2-benzimidazolone and a 2-benzimidazolone derivative, or a pharmaceutically acceptable salt thereof.

Preferably, the compound has a general Formula I or II (vide infra). Further preferably, the voltage-dependent potassium channels modulated are KCNQ2 channels, KCNQ3 channels and/or KCNQ2/3 channels.

According to another aspect of the present invention there is provided a method of depressing cortical neuron activity, the method comprising administering to the subject in need thereof a therapeutically effective amount of a compound selected from the group consisting of N-phenylanthranilic acid, a N-phenylanthranilic acid derivative, 2-benzimidazolone and a 2-benzimidazolone derivative, or a pharmaceutically acceptable salt thereof.

Preferably, the compound has a general Formula I or II (vide infra). According to yet another aspect of the present invention there is provided a pharmaceutical composition for the treatment or prevention of a condition or disorder, e.g., in the central or peripheral nervous system, in which modulating a voltage- dependent potassium chamiel and/or depressing a cortical neuron activity is beneficial, the pharmaceutical composition comprising, as an active ingredient, a compound selected from the group consisting of N-phenylanthranilic acid, a N-phenylanthranilic acid derivative, 2-benzimidazolone and a 2-benzimidazolone derivative, or a pharmaceutically acceptable salt thereof.

Preferably, the compound has the general Formula I or II (vide infra). Further preferably, the voltage-dependent potassium channels modulated are KCNQ2 channels, KCNQ3 channels and/or KCNQ2/3 channels.

According to the present invention, general Formulae I and II are:

Formula I Formula II

or a pharmaceutically acceptable salt thereof, wherein:

Z is an A-G(=K)-X-Y group, and wherein

A is alkyl or absent;

G is selected from the group consisting of carbon, sulfur and substituted or unsubstituted phosphor; K is selected from the group consisting of oxygen and sulfur;

X is selected from the group consisting of substituted or unsubstituted nitrogen, oxygen, sulfur or absent; and Y is selected from the group consisting of hydrogen, alkyl, hydroxyalkyl, cycloalkyl, aryl and a polyalkylene glycol residue, each of Q and W is independently selected from the group consisting of substituted or unsubstituted nitrogen, oxygen, sulfur and carbon; D is selected from the group consisting of oxygen and sulfur;

R¹ is selected from the group consisting of hydrogen, alkyl, cycloalkyl or aryl;

Each of R², R³, R⁴, R⁵, R⁶, R⁷, R⁸, R⁹, R¹⁰, R¹¹, R¹², R¹³ and R¹⁴ is independently selected from the group consisting of hydrogen, alkyl, hydroxyalkyl, trihaloalkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, heteroalicyclic, halo, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, sulfinyl, sulfonyl, cyano, nitro, amino and -NR¹⁵R¹⁶, or, alternatively, at least two of R², R³, R⁴, R⁵ and R⁶, of R⁷, R⁸, R⁹ and R¹⁰ and/or of R¹¹, R¹², R¹³ and R¹⁴ form a five- or six-membered aromatic, heteroaromatic, alicyclic or heteroalicyclic ring; and

R¹⁵ and R¹⁶ are each independently selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, carbonyl and sulfonyl, or, alternatively R¹⁵ and R¹⁶ form a five- or six-member heteroalicyclic ring; whereas if the phosphor and/or the nitrogen is substituted, the substituent is selected from the group consisting of alkyl, cycloalkyl and aryl.

According to one embodiment of the present invention, a compound of the present invention has the general Formula I. When a compound of the present invention is of the general Formula I, then Y is preferably selected from the group consisting of hydroxyalkyl and a polyalkylene glycol residue. Preferably, the polyalkylene glycol residue has a general formula III:

[O-(CH₂)m]n-OR¹⁷ Formula III

Wherein each of m and n is independently an integer of 1-10; and R¹⁷ is hydrogen, alkyl, cycloalkyl or aryl. According to a feature of the present invention, each of R², R³, R⁴, R⁵ and R⁶ is independently selected from the group consisting of hydrogen, alkyl, halo and trihaloalkyl and each of R⁷, R⁸, R⁹ and R¹⁰ is hydrogen. According to another embodiment of the present invention, a compound of the present invention has the general Formula II. When a compound of the present invention has the general Formula II, then preferably Q and W are each substituted or unsubstituted nitrogen; and D is oxygen, and even more preferably Q is a substituted nitrogen.

According to a preferred embodiment of the present invention, compounds are selected from the group consisting of compounds 1, 2, 3, 4, 5, 6, 7, 8, 9 and 10:

8

10

and pharmaceutically acceptable salts thereof. According to a feature of the present invention modulating of the voltage- dependent potassium channel and/or depressing the cortical neuron activity is for a treatment of a condition or disorder selected from the group of disorders consisting of epilepsy, ischemic stroke, migraine, ataxia, myokymia, neurogenic pain, Alzheimer's disease, Parkinson's disease, age-related memory loss, learning deficiencies, bipolar disorder, trigeminal neuralgia, spasticity, mood disorder, psychotic disorder, brain tumor, hearing and vision loss, anxiety and a motor neuron disease.

According to a feature of the present invention, administering of the a compound of the present invention is effected intranasally, subcutaneously, intravenously, intramuscularly, parenterally, orally, topically, intradermally, bronchially, buccally, sublingually, supositorially and mucosally.

According to a feature of the present invention, a compound of the present invention is part of a pharmaceutical composition, which further includes a pharmaceutically acceptable carrier.

According to a feature of the present invention, a pharmaceutical composition of the present invention further comprises an agent selected from the group consisting of an anti-bacterial agent, an antioxidant, a buffering agent, a bulking agent, an anti- inflammatory agent, an anti-viral agent, a chemotherapeutic agent and an anti- histamine.

According to a feature of the present invention, a pharmaceutically composition is packaged in a packaging material and identified in print, in or on the packaging material, for use in the treatment or prevention of a condition or disorder associated with altered activity of a voltage-dependent potassium channel. Preferably such a condition or disorder is selected from the group consisting of epilepsy, ischemic stroke, migraine, ataxia, myokymia, neurogenic pain, Alzheimer's disease, Parkinson's disease, age-related memory loss, learning deficiencies, bipolar disorder, trigeminal neuralgia, spasticity, mood disorder, psychotic disorder, brain tumor, hearing and vision loss, anxiety and a motor neuron disease.

According to still another aspect of the present invention there is provided a novel compound having a general Formula IV:

Formula IV or a pharmaceutically acceptable salt thereof, wherein:

Z is an A-G(=K)-X-Y group, and wherein: A is alkyl or absent;

G is selected from the group consisting of carbon, sulfur and substituted or unsubstituted phosphor;

K is selected from the group consisting of oxygen and sulfur; X is selected from the group consisting of substituted or unsubstituted nitrogen, oxygen, sulfur or absent; and

Y is selected from the group consisting of hydroxyalkyl and a polyalkylene glycol residue;

R¹ is selected from the group consisting of hydrogen, alkyl, cycloalkyl or aryl; Each of R², R³, R⁴, R⁵, R⁶, R⁷, R⁸, R⁹ and R¹⁰, is independently selected from the group consisting of hydrogen, alkyl, hydroxyalkyl, trihaloalkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, heteroalicyclic, halo, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, sulfinyl, sulfonyl, cyano, nitro, amino and - NR¹⁵R¹⁶, or, alternatively, at least two of R², R³, R⁴, R⁵ and/or R⁶, of R⁷, R⁸, R⁹ and R¹⁰ form a five- or six-membered aromatic, heteroaromatic, alicyclic or heteroalicyclic ring; and R¹⁵ and R¹⁶ are each independently selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, carbonyl and sulfonyl, or, alternatively R¹⁵ and R¹⁶ form a five- or six-member heteroalicyclic ring; whereas if the phosphor and/or the nitrogen is substituted, the substituent is selected from the group consisting of alkyl, cycloalkyl and aryl.

According to a feature of the present invention, the polyalkylene glycol residue of a novel compound of the present invention has a general formula V:

[O-(CH₂)m]n-OR¹⁷ Formula V wherein each of m and n is independently an integer of 1-10 and R¹⁷ is hydrogen, alkyl, cycloalkyl or aryl.

According to an additional feature, for a novel compound of the present invention, G is carbon, K is oxygen, each of R², R³, R⁴, R⁵ and R^δ is independently selected from the group consisting of hydrogen, alkyl, halo and trihaloalkyl; and each of R⁷, R⁸, R⁹ and R¹⁰ is hydrogen.

According to a preferred embodiment of the present invention, a novel compound of the present invention is selected from the group consisting of compounds 3, 4, 5, 6, 7, 8 and 9:

8

9 and pharmaceutically acceptable salts thereof.

According to an additional aspect of the present invention there is provided a pharmaceutical composition comprising, as an active ingredient, a compound of the present invention having a general formula IV. According to a feature of the present invention there is provided a pharmaceutical composition, having as an active ingredient, any one of the compounds 3, 4, 5, 6, 7, 8 and/or 9.

According to yet another aspect of the present invention there is provided a method for the synthesis of a compound of formula IV. The method comprises obtaining a N-phenylanthranilic acid or a derivative thereof; and reacting the N- phenylanthranilic acid or the derivative thereof with a hydroxyalkyl or a polyalkylene glycol terminating with a reactive group, which is capable of forming an ester bond with the N-phenylanthranilic acid or the derivative thereof.

The ester bond is preferably selected from the group consisting of a carboxylic ester bond, a carboxylic amide bond, a carboxylic thioester bond, a S-carboxy thioester bond and a S-carboxy amide bond, whereas the reactive group is preferably selected from the group consisting of hydroxy, amine and thiohydroxy.

The N-phenylanthranilic acid or the derivative thereof preferably has a general Formula VI:

Formula NI wherein,

A, G, K, R¹, R², R³, R⁴, R⁵, R⁶, R⁷, R⁸, R⁹ and R¹⁰ are as described hereinabove.

Preferably, G is carbon; K is oxygen; each of R², R³, R⁴, R⁵ and R⁶ is independently selected from the group consisting of hydrogen, alkyl, halo and trihaloalkyl; and each of R⁷, R⁸, R⁹ and R¹⁰ is hydrogen.

The polyalkylene glycol terminating with the reactive group preferably has a general formula NIL

V-[O-(CH₂)m]n-OR¹⁷

Formula Nil wherein:

V is hydroxy, amine or thiohydroxy; each of m and n is independently an integer of 1-10; and R¹⁷ is hydrogen, alkyl, cycloalkyl or aryl.

The present invention successfully addresses the shortcomings of the presently known configurations by providing compounds that act to modulate potassium channels and/or depress cortical activity, are generally available and/or are relatively easy to synthesize. Some of the compounds provided are already known in the art of pharmacology.

Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, suitable methods and materials are described below. In case of conflict, the patent specification, including definitions, will control. In addition, the materials, methods, and examples are illustrative only and not intended to be limiting. BRIEF DESCRIPTION OF THE DRAWINGS

The invention is herein described, by way of example only, with reference to the accompanying drawings. With specific reference now to the drawings in detail, it is stressed that the particulars shown are by way of example and for purposes of illustrative discussion of the embodiments of the present invention only, and are presented in the cause of providing what is believed to be the most useful and readily understood description of the principles and conceptual aspects of the invention. In this regard, no attempt is made to show details of the invention in more detail than is necessary for a fundamental understanding of the invention, the results taken with the drawings making apparent to those skilled in the art how the invention may be embodied in practice.

In the drawings:

FIGs. 1A-1D show results showing the leftward shift of the activation curve induced by meclofenamic acid (Compound 1) and 1-EBIO (Compound 10); FIGs. 2A-2C show results of the increase of KCNQ2/3 current induced by meclofenamic acid (Compound 1) and 1-EBIO (Compound 10);

FIGs. 3A-3E show results of the effect of meclofenamic acid (Compound 1) and 1-EBIO (Compound 10) on the deactivation process of KCNQ2/3 channels;

FIGs. 4A-4B show results demonstrating the depression of neuronal activity by meclofenamic acid (Compound 1);

FIG. 5 shows results demonstrating the neuroprotective effect of meclofenamic acid (Compound 1) from electroshock-induced seizures in adult mice;

FIGs. 6A-6C show the enhancement of the KCNQ2/3 current by diclofenac (Compound 2); FIGs. 7A-7C show the effects of Compound 6 on KCNQ2/3 currents;

FIGs. 8A-8B show the inhibition of evoked neuronal activity by compound 6; FIGs. 9A-9C show the inhibitory effect of different concentrations of Compound 6 on spontaneous neuronal activity;

FIGs. 10 A- 10C show the effects of Compound 5 on neuronal activity and on KCNQ2/3 current;

FIGs. 11 A-l IB show the increase in KCNQ2/3 current induced by the presence of Compound 3; FIGs. 12A-12C show the effects of Compound 4 on neuronal activity and on KCNQ2/3 current;

FIG. 13 shows the effect of Compound 9 on spontaneous neuronal activity; FIGs. 14A-14D show the effects of Compound 7 on KCNQ2/3 channels and neuronal activity;

FIGs. 15A-15B show the effect of Compound 8 on evoked and spontaneous neuronal activity;

FIGs. 16A-16D show the selectivity of meclofenamic acid (Compound 1) towards KCNQ2 and KCNQ3 homomeric channels, expressed in CHO cells; and FIG. 17 shows the chemical structures of Compounds 1-10.

DESCRIPTION OF THE PREFERRED EMBODIMENTS

The present invention provides compounds that, inter alia, are generally useful in the modulation of potassium ion flux through voltage-dependent potassium channels, specifically the KCNQ2, KCNQ3 and/or KCNQ2/3 channels and/or useful in depressing cortical neuron activity.

The principles and uses of the present invention may be better understood with reference to the Examples and accompanying descriptions. Before explaining at least one embodiment of the invention in detail, it is to be understood that the invention is not limited in its application to the details set forth in the following description or exemplified by the Examples. The invention is capable of other embodiments or of being practiced or carried out in various ways. Also, it is to be understood that the phraseology and terminology employed herein is for the purpose of description and should not be regarded as limiting.

According to one aspect of the present invention there is provided a method of modulating (preferably opening) a voltage-dependent potassium channel, the method comprising administering to the subject in need thereof a therapeutically effective amount of the compound N-phenylanthranilic acid, a N-phenylanthranilic acid derivative, 2-benzimidazolone, a 2-benzimidazolone derivative, or a pharmaceutically acceptable salt thereof. As used herein, the term "derivative" describes the result of a chemically altering, modifying or changing a molecule or a portion thereof, such that it maintains its original functionality in at least one respect. The compound preferably has a general Formula I or II. The voltage- dependent potassium channels modulated are preferably KCNQ2 channels, KCNQ3 channels and/or KCNQ2/3 channels.

According to another aspect of the present invention there is provided a method of depressing cortical neuron activity, the method comprising administering to the subject in need thereof a therapeutically effective amount of the compound N- phenylanthranilic acid, a N-phenylanthranilic acid derivative, 2-benzimidazolone, a 2- benzimidazolone derivative, or a pharmaceutically acceptable salt thereof.

Preferably, the compound has a general Formula I or II.

According to yet another aspect of the present invention there is provided a pharmaceutical composition for the treatment or prevention of conditions or disorders in which modulating a voltage-dependent potassium channel and/or depressing a cortical neuron activity is beneficial, the pharmaceutical composition comprising, as an active ingredient, the compound N-phenylanthranilic acid, a N-phenylanthranilic acid derivative, 2-benzimidazolone, a 2-benzimidazolone derivative, or a pharmaceutically acceptable salt thereof.

Preferably, the compound has the general Formula I or II. Further preferably, the voltage-dependent potassium channels modulated are KCNQ2 channels, KCNQ3 channels and/or KCNQ2/3 channels.

According to the present invention general Formulae I and II are:

Formula I Formula II

or a pharmaceutically acceptable salt thereof, wherein:

Z is an A-G(=K)-X-Y group, and wherein A is alkyl or absent;

Y is selected from the group consisting of hydrogen, alkyl, hydroxyalkyl, cycloalkyl, aryl and a polyalkylene glycol residue, each of Q and W is independently selected from the group consisting of substituted or unsubstituted nitrogen, oxygen, sulfur and carbon; D is selected from the group consisting of oxygen and sulfur;

R¹ is selected from the group consisting of hydrogen, alkyl, cycloalkyl or aryl; Each of R², R³, R⁴, R⁵, R⁶, R⁷, R⁸, R⁹, R¹⁰, R¹¹, R¹², R¹³ and R¹⁴ is independently selected from the group consisting of hydrogen, alkyl, hydroxyalkyl, trihaloalkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, heteroalicyclic, halo, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, sulfinyl, sulfonyl, cyano, nitro, amino and -NR¹⁵R¹⁶, or, alternatively, at least two of R², R³, R⁴, R⁵ and R⁶, of R⁷, R⁸, R⁹ and R¹⁰ and/or of R^π, R¹², R¹³ and R¹⁴ form a five- or six-membered aromatic, heteroaromatic, alicyclic or heteroalicyclic ring; and

R¹⁵ and R¹⁶ are each independently selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, carbonyl and sulfonyl, or, alternatively R¹⁵ and R¹⁶ form a five- or six-member heteroalicyclic ring; whereas if the phosphor and/or the nitrogen is substituted, the substituent is alkyl, cycloalkyl or aryl.

According to one embodiment of the present invention, a compound of the present invention has the general Formula I. When a compound of the present invention is of the general Formula I, then Y is preferably selected from the group consisting of hydroxyalkyl and a polyalkylene glycol residue.

Preferably, the polyalkylene glycol residue has a general formula III:

[O-(CH₂)m]n-OR¹⁷ Formula III wherein each of m and n is independently an integer of 1-10 and R¹⁷ is hydrogen, alkyl, cycloalkyl or aryl. According to a feature of the present invention, each of R², R³, R⁴, R⁵ and R⁶ is independently selected from the group consisting of hydrogen, alkyl, halo and trihaloalkyl and each of R⁷, R⁸, R⁹ and R¹⁰ is hydrogen.

According to another embodiment of the present invention, a compound of the present invention has the general Formula II. When a compound of the present invention has the general Formula II, then preferably Q and W are each substituted or unsubstituted nitrogen; and D is oxygen, and even more preferably Q is a substituted nitrogen.

According to a preferred embodiment of the present invention, compounds are selected from the group consisting of:

8

10

and pharmaceutically acceptable salts thereof.

The term "method" refers to manners, means, techniques and procedures for accomplishing a given task including, but not limited to, those manners, means, techniques and procedures either known to, or readily developed from known manners, means, techniques and procedures by practitioners of the chemical, pharmacological, biological, biochemical and medical arts.

By treating or preventing is meant that a compound of the present invention is used as a therapeutic, prophylactic or ameliorative agent, whether with respect to a pathology, condition or disorder, a symptom thereof or an effect thereof.

There are many pathologies, conditions and disorders with defective potassium channel functioning. Just as other potassium channel modulating compounds, the compounds of the present invention are for use within the framework of a treatment for pathologies, conditions and disorders associated with defective potassium modulation, so as to treat, ameliorate, prevent, inhibit, or limit the effects of the conditions and pathologies in animals including humans.

More particularly, the invention provides compounds, compositions and methods that are useful in the treatment of central or peripheral nervous system disorders (e.g., ischemic stroke, migraine, ataxia, Parkinson's disease, bipolar disorders, trigeminal neuralgia, spasticity, mood disorders, brain tumors, psychotic disorders, myokymia, neurogenic pain, seizures, epilepsy, hearing and vision loss, Alzheimer's disease, Parkinson's disease, age-related memory loss, learning deficiencies, anxiety and motor neuron diseases), and as neuroprotective agents (e.g., to prevent stroke and the like). Compounds of the invention have use as agents for treating convulsive states, for example that following grand mal, petit mal, psychomotor epilepsy or focal seizure. The compounds of the invention are also useful in treating disease states such as gastroesophogeal reflux disorder and gastrointestinal hypomotility disorders. Other pathologies and conditions that compounds of the present invention are useful in treating are listed in, for example, U.S. Patents 6,348,486; 6,117,900; 6,589,986 and 6,593,349 and U.S. Patent Applications 10/022,579; 10/075,703; 10/075,522; 10/114,148; 10/160,582 and 10/312,123, all of which are hereby incorporated by reference.

As voltage dependent potassium channels are found in all animal species, the compounds of the present invention are pharmaceutically effective when administered to subjects who are members of all animal species, including monkeys, dogs, cats, mice, rats, farm animals, livestock, fish and most importantly humans.

"Opening" and "activating" are used interchangeably herein to refer to the partial or full activation of a KCNQ channel by a compound, which leads to an increase in ion flux either into or out of a cell in which a KCNQ channel is found.

Techniques for formulation and administration of compounds as medicaments may be found in "Remington's Pharaiaceutical Sciences," Mack Publishing Co., Easton, PA, latest edition, which is incorporated herein by reference and are also discussed heremfurther.

The compounds of the present invention are configured to cross the blood brain barrier so as to allow many different types of dosage forms. Nevertheless, pharmaceutical compositions of the present invention may be provided to an individual in need of treatment (whether therapeutic, prophylactic or ameliorative) by a variety of preferred routes, such as subcutaneous, topical, oral, intraperitoneal, intradermal, intravenous, intranasal, bronchial, buccal, sublingual, suppository, intramuscular, oral, rectal, transmucosal, intestinal or parenteral delivery, including topical, intra-arterial, intramuscular, subcutaneous and intramedullary injections as well as infrathecal, direct intraventricular, intravenous, intraperitoneal, intranasal, or intraocular injections.

Alternately, one may administer a composition of the present invention in a local rather than systemic manner, for example, via injection of the composition directly into an organ often in a depot or slow release formulation, such as described below.

A therapeutically (or pharmaceutically) effective amount means an amount of active ingredient needed to achieve the desired outcome, which is generally to prevent, alleviate or ameliorate a condition or symptoms of the condition. Determination of a therapeutically effective amount is within the capability of those skilled in the art, especially in light of the detailed disclosure provided herein.

"Compound of the invention," as used herein refers to a compound according to Formula I or II or a combination thereof, and a pharmaceutically acceptable salt of a compound according to Formula I or II or a combination thereof and a solvated forms including hydrated forms such as monohydrate, dihydrate, trihydrate, hemihydrate, tetrahydrate and the like. The compounds may be true solvates or may merely retain adventitious solvent, or be a mixture of solvate and adventitious solvent.

The phrase "pharmaceutically acceptable salts" is meant to include salts of the active compounds which are prepared with relatively nontoxic acids or bases, depending on the particular substituents found on the compounds described herein. When compounds of the present invention contain relatively acidic functionalities, base addition salts can be obtained by contacting the neutral (i.e., non-ionized) form of such compounds with a sufficient amount of the desired base, either neat or in a suitable inert solvent. Examples of pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic a ino, or magnesium salt, or a similar salt. When compounds of the present invention contain relatively basic functionalities, acid addition salts can be obtained by contacting the neutral form of such compounds with a sufficient amount of the desired acid, either neat or in a suitable inert solvent. Examples of pharmaceutically acceptable acid addition salts include those derived from inorganic acids like hydrochloric, hydrobromic, nitric, carbonic, monohydrogencarbonic, phosphoric, monohydrogenphosphoric, dihydrogenphosphoric, sulfuric, monohydrogensulfuric, hydriodic, or phosphorous acids and the like, as well as the salts derived from relatively nontoxic organic acids like acetic, propionic, isobutyric, maleic, malonic, benzoic, succinic, suberic, fumaric, lactic, mandelic, phthalic, benzenesulfonic, p-tolylsulfonic, citric, tartaric, methanesulfonic, and the like. Also included are salts of amino acids such as arginate and the like, and salts of organic acids like glucuronic or galactunoric acids and the like (see, for example, Berge et al, "Pharmaceutical Salts", Journal of Pharmaceutical Science, 1977, 66, 1-19). Certain specific compounds of the present invention contain both basic and acidic functionalities that allow the compounds to be converted into either base or acid addition salts. The neutral forms of the compounds are preferably regenerated by contacting the salt with a base or acid and isolating the parent compound in a conventional manner. The parent form of the compound differs from the various salt forms in certain physical properties, such as solubility in polar solvents, but otherwise the salts are equivalent to the parent form of the compound for the purposes of the present invention.

In addition to salt forms, the present invention provides compounds, which are in a prodrug form. Prodrugs of the compounds described herein are those compounds that readily undergo chemical changes under physiological conditions in vivo to provide the compounds of the present invention. Additionally, prodrugs can be converted to the compounds of the present invention by chemical or biochemical methods in an ex vivo environment. For example, prodrugs can be slowly converted to the compounds of the present invention when placed in a transdermal patch reservoir with a suitable enzyme or chemical reagent.

Certain compounds of the present invention can exist in unsolvated forms as well as solvated forms, including hydrated forms. In general, the solvated forms are equivalent to unsolvated forms and are encompassed within the scope of the present invention. Certain compounds of the present invention may exist in multiple crystalline or amorphous forms. In general, all physical forms are equivalent for the uses contemplated by the present invention and are intended to be within the scope of the present invention.

Certain compounds of the present invention possess asymmetric carbon atoms (optical centers) or double bonds; the racemates, diastereomers, geometric isomers and individual isomers are encompassed within the scope of the present invention.

The compounds of the present invention may also contain unnatural proportions of atomic isotopes at one or more of the atoms that constitute such compounds. For example, the compounds may be radiolabeled with radioactive isotopes, such as for example tritium (³H), iodine-125 (¹²⁵I) or carbon-14 (¹⁴C). All isotopic variations of the compounds of the present invention, whether radioactive or not, are intended to be encompassed within the scope of the present invention.

As used herein in the specification and in the claims section that follows, the term "alkyl" refers to a saturated aliphatic hydrocarbon including straight chain and branched chain groups. Preferably, the alkyl group has 1 to 20 carbon atoms. Whenever a numerical range; e.g., "1-20", is stated herein, it means that the group, in this case the alkyl group, may contain 1 carbon atom, 2 carbon atoms, 3 carbon atoms, etc., up to and including 20 carbon atoms. More preferably, it is a medium size alkyl having 1 to 10 carbon atoms. Most preferably, it is a lower alkyl having 1 to 4 carbon atoms. The alkyl group may be substituted or unsubstituted. When substituted, the substituent group can be, for example, cycloalkyl, aryl, heteroaryl, heteroalicyclic, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, cyano, halo, carbonyl, thiocarbonyl, O-carbamyl, N-carbamyl, O-thiocarbamyl, N-thiocarbamyl, C-amido, N-amido, C-carboxy, O-carboxy, nitro, sulfonamido, trihalomethanesulfonamido, silyl, guanyl, guanidino, ureido, amino or NR_aRb, wherein R_a and Rtø are each independently hydrogen, alkyl, cycloalkyl, aryl, carbonyl, sulfonyl, trihalomethysulfonyl and, combined, a five- or six-member heteroalicyclic ring.

A "cycloalkyl" group refers to an all-carbon monocyclic or fused ring (i.e., rings which share an adjacent pair of carbon atoms) group wherein one of more of the rings does not have a completely conjugated pi-electron system. Examples, without limitation, of cycloalkyl groups are cyclopropane, cyclobutane, cyclopentane, cyclopentene, cyclohexane, cyclohexadiene, cycloheptane, cycloheptatriene, and adamantane. A cycloalkyl group may be substituted or unsubstituted. When substituted, the substituent group can be, for example, alkyl, aryl, heteroaryl, heteroalicyclic, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, cyano, halo, carbonyl, thiocarbonyl, C-carboxy, O-carboxy, O-carbamyl, N-carbamyl, C-amido, N-amido, nitro, amino and NR_a b as defined above.

An "alkenyl" group refers to an alkyl group which consists of at least two carbon atoms and at least one carbon-carbon double bond.

An "aryl" group refers to an all-carbon monocyclic or fused-ring polycyclic (i.e., rings which share adjacent pairs of carbon atoms) groups having a completely conjugated pi-electron system. Examples, without limitation, of aryl groups are phenyl, naphthalenyl and anthracenyl. The aryl group may be substituted or unsubstituted. When substituted, the substituent group can be, for example, halo, trihalomethyl, alkyl, hydroxy, alkoxy, aryloxy, thiohydroxy, thiocarbonyl, C-carboxy, O-carboxy, O-carbamyl, N-carbamyl, O-thiocarbamyl, N-thiocarbamyl, C-amido, N- amido, sulfinyl, sulfonyl, amino and NR_aRb as defined above.

A "heteroaryl" group refers to a monocyclic or fused ring (i.e., rings which share an adjacent pair of atoms) group having in the ring(s) one or more atoms, such as, for example, nitrogen, oxygen and sulfur and, in addition, having a completely conjugated pi-electron system. Examples, without limitation, of heteroaryl groups include pyrrole, furane, thiophene, imidazole, oxazole, thiazole, pyrazole, pyridine, pyrimidine, quinoline, isoquinoline and purine. The heteroaryl group may be substituted or unsubstituted. When substituted, the substituent group can be, for example, alkyl, cycloalkyl, halo, trihalomethyl, hydroxy, alkoxy, aryloxy, thiohydroxy, thiocarbonyl, sulfonamido, C-carboxy, O-carboxy, sulfinyl, sulfonyl, O- carbamyl, N-carbamyl, O-thiocarbamyl, N-thiocarbamyl, C-amido, N-amido, amino or NRgR^ as defined above.

A "heteroalicyclic" group refers to a monocyclic or fused ring group having in the ring(s) one or more atoms such as nitrogen, oxygen and sulfur. The rings may also have one or more double bonds. However, the rings do not have a completely conjugated pi-electron system. The heteroalicyclic may be substituted or unsubstituted. When substituted, the substituted group can be, for example, alkyl, cycloalkyl, aryl, heteroaryl, halo, trihalomethyl, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, cyano, nitro, carbonyl, thiocarbonyl, C-carboxy, O-carboxy, O-carbamyl, N-carbamyl, O-thiocarbamyl, N-thiocarbamyl, sulfinyl, sulfonyl, C-amido, N-amido, amino and NR_aR_D as defined above.

A "hydroxy" group refers to an -OH group. An "azido" group refers to a -N=N group.

An "alkoxy" group refers to both an -O-alkyl and an -O-cycloalkyl group, as defined herein.

An "aryloxy" group refers to both an -O-aryl and an -O-heteroaryl group, as defined herein. A "thiohydroxy" group refers to an -SH group.

A "thioalkoxy" group refers to both an -S-alkyl group, and an -S-cycloalkyl group, as defined herein.

An "thioaryloxy" group refers to both an -S-aryl and an -S-heteroaryl group, as defined herein.

A "carbonyl" group refers to a -C(=O)-R" group, where R" is hydrogen, alkyl, cycloalkyl, aryl, heteroaryl (bonded through a ring carbon) or heteroalicyclic (bonded through a ring carbon) as defined herein.

An "aldehyde" group refers to a carbonyl group, where R" is hydrogen. A "thiocarbonyl" group refers to a -C(=S)-R" group, where R" is as defined herein.

A "C-carboxy" group refers to a -C(=O)-0-R" groups, where R" is as defined herein.

An "O-carboxy" group refers to an R"C(=O)-O- group, where R" is as defined herein.

A "carboxylic acid" group refers to a C-carboxyl group in which R" is hydrogen.

A "halo" group refers to fluorine, chlorine, bromine or iodine. A "trihalomethyl" group refers to a -CX group wherein X is a halo group as defined herein.

A "trihalomethanesulfonyl" group refers to an X_βCS^O^- group wherein X is a halo group as defined herein.

A "sulfinyl" group refers to an -S(^:=O)-R" group, where R" is as defined herein. A "sulfonyl" group refers to an

group, where R" is as defined herein.

An "S-sulfonamido" group refers to a

group, with R_a and R_p as defined herein.

An "N-sulfonamido" group refers to an R_aS(=O)2-NR_D group, where R_a and Rb are as defined herein. A "trihalomethanesulfonamido" group refers to an

group, where R_a and X are as defined herein.

An "O-carbamyl" group refers to an -OC(=O)-NR_aRb group, where R_a and Rb are as defined herein. An "N-carbamyl" group refers to an RbOC(=O)-NR_a- group, where R_a and

Rtø are as defined herein.

An "O-thiocarbamyl" group refers to an -OC(=S)-NR_aR_D group, where R_a and Rfo are as defined herein.

An "N-thiocarbamyl" group refers to an

group, where R_a and Rtø are as defined herein.

An "Amino" group refers to an -NH2 group.

A "C-amido" group refers to a -C(=O)-NR_aR_D group, where R_a and Ri_j are as defined herein.

An "N-amido" group refers to an Rt,C(=O)-NR_a group, where R_a and Rfo are as defined herein.

A "quaternary ammonium" group refers to an -NHR_aRb group, wherein R_a and R_D are independently alkyl, cycloalkyl, aryl or heteroaryl.

An "ureido" group refers to an -NR_aC(=O)-NR^R_c group, where R_a and R^ are as defined herein and R_c is defined as either R_a or Rtø. A "guanidino" group refers to an -R_aNC(=N)-NRbR_c group, where R_a, R^ and R_c are as defined herein.

A "guanyl" group refers to an R_aR]_jNC(=N)- group, where R_a and R_D are as defined herein.

A "nitro" group refers to an -NO2 group. A "cyano" group refers to a -C=N group.

A "silyl" group refers to a -Si (R")3, where R" is as defined herein.

According to a preferred embodiment, the compound of the present invention has the general Formula I. In one preferred embodiment Y is selected from the group consisting of hydroxyalkyl and a polyalkylene glycol residue. Preferably such a polyalkylene glycol residue has a general formula III:

[O-(CH₂)m]n-OR¹⁷ Formula III wherein each of m and n is independently an integer of 1-10; and R is hydrogen, alkyl, cycloalkyl or aryl. Preferably, G is carbon, K is oxygen, each of R², R³, R⁴, R⁵ and R⁶ is independently selected from the group consisting of hydrogen, alkyl, halo and trihaloalkyl; and each of R⁷, R⁸, R⁹ and R¹⁰ is hydrogen.

According to a different preferred embodiment, the compound of the present invention has the general Foπnula II. When a compound of the present invention has the general Formula II, then preferably Q and W are each substituted or unsubstituted nitrogen; and D is oxygen, and even more preferably Q is a substituted nitrogen. Preferred compounds of the present invention include the compounds:

meclofenamic acid diclofenac

9 10

1-EBIO and pharmaceutically acceptable salts thereof.

According to another aspect of the present invention, there are provided novel compounds, which are useful in context of the present invention. These novel compounds are generally derivatives of N-phenylanthranilic acid having a hydroxyalkyl and a polyalkylene glycol residue covalently attached thereto. The hydroxyalkyl and a polyallcylene glycol residue generally increase the ability of a novel compound of the present invention to cross the blood brain barrier.

A preferred novel compound of the present invention is of the general Formula IV:

Formula IV

or a pharmaceutically acceptable salt thereof, wherein: Z is an A-G(=K)-X-Y group, and wherein: A is alkyl or absent;

R¹ is selected from the group consisting of hydrogen, alkyl, cycloalkyl or aryl; Each of R², R³, R⁴, R⁵, R⁶, R⁷, R⁸, R⁹ and R¹⁰, is independently selected from the group consisting of hydrogen, alkyl, hydroxyalkyl, trihaloalkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, heteroalicyclic, halo, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, sulfinyl, sulfonyl, cyano, nitro, amino and - NR¹⁵R¹⁶, or, alternatively, at least two of R², R³, R⁴, R⁵ and/or R⁶, of R⁷, R⁸, R⁹ and R ° form a five- or six-membered aromatic, heteroaromatic, alicyclic or heteroalicyclic ring; and

R ⁵ and R¹⁶ are each independently selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, carbonyl and sulfonyl, or, alternatively R¹⁵ and R¹⁶ form a five- or six-member heteroalicyclic ring. Whereas, if the phosphor and/or the nitrogen is substituted, the substituent is alkyl, cycloalkyl or aryl.

According to an additional feature, for a novel compound of the present invention, G is carbon, K is oxygen, each of R², R³, R⁴, R⁵ and R⁶ is independently selected from the group consisting of hydrogen, alkyl, halo and frihaloallcyl; and each of R⁷, R⁸, R⁹ and R¹⁰ is hydrogen. According to a preferred embodiment a novel compound of the present invention is selected from the group consisting of:

and pharmaceutically acceptable salts thereof. According to an additional aspect of the present invention there is provided a pharmaceutical composition comprising, as an active ingredient, a compound of the present invention having a general formula IV.

According to a feature of the present invention there is provided a pharmaceutical composition, having as an active ingredient, any one of the compounds 3, 4, 5, 6, 7, 8 and/or 9. According to yet another aspect of the present invention there is provided a method for the synthesis of the novel compounds described hereinabove. The method is effected by reacting a N-phenylanthranilic acid or a derivative thereof with a hydroxy alkyl or a polyallcylene glycol, which are terminated by a reactive group. The reactive group is selected such that it is capable of forming an ester bond with the N-phenylanthranilic acid or the derivative thereof.

As used herein, the phrase "an ester bond" describes a J(=L)-M bond, wherein J is carbon, sulfur or phosphor, preferably carbon, L is oxygen or sulfur and M is oxygen, sulfur or nitrogen (substituted or non-substituted, as is described hereinabove).

Preferred ester bonds, according to the present invention include a carboxylic ester bond [-C(=O)-O-], a carboxylic amide bond [-C(=O)-NR'-], a carboxylic thioester bond [-C(=O)-S-], a thiocarbonyl ester bond [-C(=S)-O-], a thiocarbonyl thioester bond [-C(=S)-S-] and a thiocarbonyl amide bond [-C(=S)-NR'-J.

Accordingly, the reactive group is preferably hydroxy, amino or thiohydroxy, as defined hereinabove, and the polyalkylene glycol terminating with the reactive group has a general formula VII:

V-[O-(CH₂)m]n-OR¹⁷

Formula Nil wherein, N is hydroxy, amine or thiohydroxy, each of m and n is independently an integer of 1-10; and R¹⁷ is hydrogen, alkyl, cycloalkyl or aryl.

The starting material, Ν-phenylanthranilic acid or the derivative thereof, preferably has a general formula VI:

Formula VI wherein,

A, G, K, R¹, R², R³, R⁴, R⁵, R⁶, R⁷, R⁸, R⁹ and R¹⁰, are as described hereinabove. Preferably, in Formula NI, G is carbon, K is oxygen, each of R², R³, R⁴, R⁵ and R⁶ is independently hydrogen, alkyl, halo and trihaloalkyl; and each of R⁷, R⁸, R⁹ and R¹⁰ is hydrogen.

In cases where the ester bond is an amide bond, the method according to this aspect of the present invention preferably further includes, prior to reacting with the hydroxyalkyl or the polyallcylene glycol, converting the Ν-phenylanthranilic acid or its derivative to a corresponding ester.

As is demonstrated in the Examples section that follows, the novel compounds of the present invention are easily and efficiently synthesized by the method of this aspect of the present invention.

Compounds of the present invention include known compounds such as meclofenamic acid (Compound 1), diclofenac (Compound 2) and 1-EBIO (Compound 10). 1-EBIO (Compound 10) has been found to increase the opening rate and hence the open probability Po of the channel in single-channel studies performed on intermediate (IK) and small conductance (SK) Ca2⁺-activated K+ channel, see Syme et al Am. J. Physiol. 278:C570-C581 (2000).

Compounds of the present invention also include novel compounds which are derivatives of Ν-phenylanthranilic acid, mainly derivatives of meclofenamic acid (Compound 1) and diclofenac (Compound 2). Exemplary novel compounds of the present invention are compounds 3, 4, 5, 6, 7, 8 and 9.

Herein it is demonstrated that these compounds are openers of KCΝQ2/3 channel complex heterologously expressed in CHO cells. These compounds are also shown herein to reduce both evoked and spontaneous action potentials in cortical neurons. The compounds of the present invention have two main effects: shifting of the voltage dependence of KCNQ2/3 channel activation to more hyperpolarized potentials and slow channel deactivation. Similar to the effect on recombinant KCNQ2/3 channels, compounds 4, 5, 6 and 7 induce an approximately 20 mV negative shift in the threshold of M-current activation in cortical neurons, from -50 mV to -70 mV. As a result of this leftward shift of the KCNQ2/3 threshold of activation, there is a progressive hyperpolarization of the resting membrane potential. Without being bound to any theory in particular, the data presented herein in the Examples section that follows suggests that the compounds of the present invention either destabilize a closed channel conformation or stabilize the KCNQ2/3 channel in an open state. Further, exposure of channels to the compounds described herein also leads to a slowing of deactivation that contributes to the stabilization of the KCNQ2/3 channel in the open state. Without being bound to any theory in particular, it is possible that the compounds of the present invention modify the channel gating by shifting the voltage dependence of the voltage sensor S4 movement in the hyperpolarizing direction.

From a functional point of view, the leftward shift in the voltage dependence and the slowing of deactivation, caused by the compounds of the present invention, leads to substantial M-current activation at normal resting and subthreshold potentials. The especially large activation of KCNQ2/3 chaimels, e.g., in the case of meclofenamic acid (Compound 1) at potentials around -60/-50 mV (more than 10-fold increase in KCNQ2/3 current amplitude), shows that the compounds of the present invention cause membrane hyperpolarization. In addition, since the M-current (KCNQ2/3) is non-inactivating, its marked activation by compounds of the present invention contributes to a significant steady-state potassium conductance at subthreshold and threshold potentials, acting as brake for neuronal firing. Indeed, it is also demonstrated that compounds of the present invention depress the evoked and spontaneous cortical neurons activity.

It is important to note that the voltage range through which compounds of the present invention activate KCNQ2/3 channels makes theses compounds exceptionally useful for the treatment of ischemic stroke.

There are similarities between the properties exhibited by the compounds of the present invention and retigabine.

First, both the compounds of the present invention and retigabine all shift the voltage dependence of KCNQ2/3 channel activation leftwards, decelerating deactivation kinetics and hyperpolarizing the resting membrane potential (for retigabine discussed, for example, in Tatulian et al J. Neurosci. 21: 5535-5545 (2001)).

Second, retigabine produces a secondary inhibitory action on KCNQ channels at positive potentials (above +20 mV, Tatulian et al. J. Neurosci. 21: 5535-5545 (2001)) as do the compounds of the present invention towards KCNQ2/3 channels.

An interesting difference between retigabine and the compounds of the present invention is related to the selectivity towards the KCNQ2 and KCNQ3 subunits. While retigabine exerts the strongest opener action on KCNQ3 homomeric channels (Tatulian et al J. Neurosci. 21: 5535-5545 (2001), it is shown herein that the compounds of the present invention are more potent on KCNQ2 homomeric channels

From a functional point of view, the leftward shift of the activation curve and the slowing of deactivation, produced by the compounds of the present invention, leads to substantial M-current activation at normal resting and subthreshold potentials. In addition, since the M-current (KCNQ2/3) is non-inactivating, activation by the compounds of the present invention is expected to contribute to a significant steady- state potassium conductance at subthreshold and threshold potentials, acting as a brake for neuronal firing. Indeed, compounds 4, 5, 6, 7 and 8 depress the evoked and spontaneous cortical neuron activity.

The voltage range through which the compounds of the present invention operate, indicates exceptionally suitability for the treatment of epilepsy, ischemic stroke and neuropathic pain.

As noted hereinabove, according to a feature of the present invention, a compound of the present invention forms a part of a pharmaceutical composition, which further includes a pharmaceutically acceptable carrier.

According to a further feature of the present invention, a pharmaceutical composition of the present invention is packaged in a packaging material and identified in print, in or on the packaging material, for use in the treatment or prevention of a condition or disorder, e.g., in the peripheral or central nervous system, associated with altered activity of a voltage-dependent potassium channel. The conditions or disorders thus identified are one or more of the conditions or disorders for which the specific pharmaceutical composition is suitable.

As used herein a "pharmaceutical composition" refers to a preparation of one or more of the compounds of the present invention (as active ingredient), or physiologically acceptable salts or prodrugs thereof, with other chemical components including but not limited to physiologically suitable carriers, excipients, lubricants, buffering agents, antibacterial agents, bulking agents (e.g. mannitol), antioxidants (e.g., ascorbic acid or sodium bisulfite), anti-inflammatory agents, anti-viral agents, chemotherapeutic agents, anti-histamines and the like. The purpose of a pharmaceutical composition is to facilitate administration of a compound to a subject. The term "active ingredient" refers to a compound, which is accountable for a biological effect. The terms "physiologically acceptable carrier" and "pharmaceutically acceptable carrier" which may be interchangeably used refer to a carrier or a diluent that does not cause significant irritation to an organism and does not abrogate the biological activity and properties of the administered compound. Herein the term "excipient" refers to an inert substance added to a pharmaceutical composition to further facilitate administration of a compound. Examples, without limitation, of excipients include calcium carbonate, calcium phosphate, various sugars and types of starch, cellulose derivatives, gelatin, vegetable oils and polyethylene glycols.

As stated above, techniques for formulation and administration of compounds as medicaments may be found in "Remington's Pharmaceutical Sciences," Mack Publishing Co., Easton, PA, latest edition, which is incorporated herein by reference.

Pharmaceutical compositions of the present invention may be manufactured by processes well known in the art, e.g., by means of conventional mixing, dissolving, granulating, dragee-making, levigating, emulsifying, encapsulating, entrapping or lyophilizing processes.

Pharmaceutical compositions for use in accordance with the present invention thus may be formulated in conventional manner using one or more physiologically acceptable carriers comprising excipients and auxiliaries, which facilitate processing of the active compounds into preparations which, can be used pharmaceutically. Proper formulation is dependent upon the route of administration chosen.

For injection, the compounds of the invention may be formulated in aqueous solutions, preferably in physiologically compatible buffers such as Hank's solution,

Ringer's solution, or physiological saline buffer. For transmucosal administration, penetrants appropriate to the barrier to be permeated are used in the formulation. Such penetrants are generally known in the art.

For oral administration, the compounds can be formulated readily by combining the active compounds with pharmaceutically acceptable carriers well known in the art. Such carriers enable the compounds of the invention to be formulated as tablets, pills, dragees, capsules, liquids, gels, syrups, slurries, suspensions, and the like, for oral ingestion by a patient. Pharmacological preparations for oral use can be made using a solid excipient, optionally grinding the resulting mixture, and processing the mixture of granules, after adding suitable auxiliaries if desired, to obtain tablets or dragee cores. Suitable excipients are, in particular, fillers such as sugars, including lactose, sucrose, mannitol, or sorbitol; cellulose preparations such as, for example, maize starch, wheat starch, rice starch, potato starch, gelatin, gum tragacanth, methyl cellulose, hydroxvpiOpylmethyl-cellulose, sodium carbomethylcellulose; and/or physiologically acceptable polymers such as polyvinylpyrrolidone (PVP). If desired, disintegrating agents may be added, such as cross-linked polyvinyl pyrrolidone, agar, or alginic acid or a salt thereof such as sodium alginate.

Dragee cores are provided with suitable coatings. For this purpose, concentrated sugar solutions may be used which may optionally contain gum arabic, talc, polyvinyl pyrrolidone, carbopol gel, polyethylene glycol, titanium dioxide, lacquer solutions and suitable organic solvents or solvent mixtures. Dyestuffs or pigments may be added to the tablets or dragee coatings for identification or to characterize different combinations of active compound doses.

Pharmaceutical compositions, which can be used orally, include push-fit capsules made of gelatin as well as soft, sealed capsules made of gelatin and a plasticizer, such as glycerol or sorbitol. The push-fit capsules may contain the active ingredients in admixture with filler such as lactose, binders such as starches, lubricants such as talc or magnesium stearate and, optionally, stabilizers. In soft capsules, the active compounds may be dissolved or suspended in suitable liquids, such as fatty oils, liquid paraffin, or liquid polyethylene glycols. In addition, stabilizers may be added. All formulations for oral administration should be in dosages suitable for the chosen route of administration.

For buccal administration, the compositions may take the form of tablets or lozenges formulated in conventional manner.

For administration by inhalation, the compounds for use according to the present invention are conveniently delivered in the form of an aerosol spray presentation from a pressurized pack or a nebulizer with the use of a suitable propellant, e.g., dichlorodifluoromethane, trichlorofluoromethane, dichloro- tetrafluoroethane or carbon dioxide. In the case of a pressurized aerosol, the dosage unit may be determined by providing a valve to deliver a metered amount. Capsules and cartridges of, e.g., gelatin for use in an inhaler or insufflator may be formulated containing a powder mix of the compound and a suitable powder base such as lactose or starch. The preparations described herein may be formulated for parenteral administration, e.g., by bolus injection or continuous infusion. Formulations for injection may be presented in unit dosage form, e.g., in ampoules or in multidose containers with optionally, an added preservative. The compositions may be suspensions, solutions or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilizing and/or dispersing agents.

Pharmaceutical compositions for parenteral administration include aqueous solutions of the active preparation in water-soluble form. Additionally, suspensions of the active compounds may be prepared as appropriate oily injection suspensions. Suitable lipophilic solvents or vehicles include fatty oils such as sesame oil, or synthetic fatty acids esters such as ethyl oleate, triglycerides or liposomes. Aqueous injection suspensions may contain substances, which increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol or dextran. Optionally, the suspension may also contain suitable stabilizers or agents which increase the solubility of the compounds to allow for the preparation of highly concentrated solutions.

Alternatively, the active ingredient may be in powder form for constitution with a suitable vehicle, e.g., sterile, pyrogen-free water, before use.

The preparation of the present invention may also be formulated in rectal compositions such as suppositories or retention enemas, using, e.g., conventional suppository bases such as cocoa butter or other glycerides.

In addition to the formulations described previously, a preparation of the present invention may also be formulated for local administration, such as a depot preparation. Such long acting formulations may be administered by implantation (for example subcutaneously or intramuscularly) or by intramuscular injection. Thus, for example, the preparation may be formulated with suitable polymeric or hydrophobic materials (for example, as an emulsion in an acceptable oil) or ion exchange resins, or as sparingly soluble derivatives such as sparingly soluble salts. Formulations for topical administration may include, but are not limited to, lotions, suspensions, ointments gels, creams, drops, liquids, sprays emulsions and powders.

Pharmaceutical compositions suitable for use in context of the present invention include compositions wherein the active ingredients are contained in an amount effective to achieve the intended purpose. More specifically, a therapeutically effective amount means an amount of active ingredient effective to prevent, alleviate or ameliorate a condition and/or symptoms thereof and/or effects thereof.

Determination of a therapeutically effective amount is within the capability of those skilled in the art, especially in light of the detailed disclosure provided herein. For any preparation used in the methods of the invention, the therapeutically effective amount or dose can be estimated initially from cell culture assays. For example, a dose can be formulated in animal models to achieve a circulating concentration range that includes the IC^Q as determined in cell culture. Such information can be used to more accurately determine useful doses in humans. Toxicity and therapeutic efficacy of the compounds described herein can be determined by standard pharmaceutical procedures in cell cultures or experimental animals, e.g., by determining the IC^Q and the LD50 (lethal dose causing death in 50 % of the tested animals) for a subject compound. The data obtained from these cell culture assays and animal studies can be used in formulating a range of dosage for use in human. The dosage may vary depending upon the dosage form employed and the route of administration utilized. The exact formulation, route of administration and dosage can be chosen by the individual physician in view of the patient's condition. (See e.g., Fingl, et al, 1975, in "The Pharmacological Basis of Therapeutics", Ch. 1 p.l). Dosage amount and interval may be adjusted individually to provide plasma levels of the active moiety which are sufficient to maintain the kinase modulating effects, termed the minimal effective concentration (MEC). The MEC will vary for each preparation, but can be estimated from in vitro data; e.g., the concentration necessary to achieve 50-90 % inhibition of a kinase may be ascertained using the assays described herein. Dosages necessary to achieve the MEC will depend on individual characteristics and route of administration. HPLC assays or bioassays can be used to determine plasma concentrations.

Dosage intervals can also be determined using the MEC value. Preparations should be administered using a regimen, which maintains plasma levels above the MEC for 10-90 % of the time, preferable between 30-90 % and most preferably 50-90 %. It is noted that, in the case of local administration or selective uptake, the effective local concentration of the drug may not be related to plasma concentration. In such cases, other procedures known in the art can be employed to determine the effective local concentration. Depending on the severity and responsiveness of the condition to be treated, dosing can also be a single administration of a slow release composition, with course of treatment lasting from several days to several weeks or until cure is effected or diminution of the disease state is achieved.

The amount of a composition to be administered will, of course, be dependent on the subject being treated, the severity of the affliction, the manner of administration, the judgment of the prescribing physician, etc.

Compositions of the present invention may, if desired, be presented in a pack or dispenser device, such as an FDA approved kit, which may contain one or more unit dosage forms containing the active ingredient. The pack may, for example, comprise metal or plastic foil, such as a blister pack. The pack or dispenser device may be accompanied by instructions for administration. The pack or dispenser may also be accompanied by a notice associated with the container in a form prescribed by a governmental agency regulating the manufacture, use or sale of pharmaceuticals, which notice is reflective of approval by the agency of the form of the compositions or human or veterinaiy administration. Such notice, for example, may be of labeling approved by the U.S. Food and Drug Administration for prescription drugs or of an approved product insert. Compositions comprising a preparation of the invention formulated in a compatible pharmaceutical carrier may also be prepared, placed in an appropriate container, and labeled for treatment of an indicated condition. The pharmaceutical composition of the present invention is used for the treatment, prevention or amelioration of conditions or disorders associated with altered activity of a voltage-dependent potassium channel or of a cortical neuron, comprising, as an active ingredient, a compound of the present invention which serve for modulating, generally by opening, a respective potassium channel or depressing cortical neuron activity. Generally, the potassium channel modulated is a KCNQ2 channel and/or a KCNQ3 channel and/or a KCNQ2/3 channel. Peripheral or central nervous system conditions or disorders associated with altered activity of a voltage- dependent potassium channel that are preferably treated or prevented by the pharmaceutical compositions of the present invention include, but are not limited to epilepsy, ischemic stroke, migraine, ataxia, myokymia, neurogenic pain, Alzheimer's disease, Parkinson's disease, age-related memory loss, learning deficiencies, bipolar disorder, trigeminal neuralgia, spasticity, mood disorder, psychotic disorder, brain tumor, hearing and vision loss, anxiety and a motor neuron disease. Preferably the composition is packaged in a packaging material and is identified in print, in or on the packaging material, for use in the treatment or prevention of a peripheral or central nervous system condition or disorder associated with altered activity of a voltage- dependent potassium channel.

Additional objects, advantages, and novel features of the present invention will become apparent to one ordinarily skilled in the art upon examination of the following examples, which are not intended to be limiting. Additionally, each of the various embodiments and aspects of the present invention as delineated hereinabove and as claimed in the claims section below finds experimental support in the following examples.

EXAMPLES

Reference is now made to the following examples, which together with the above descriptions, illustrate the invention in a non-limiting fashion.

MA TERIALS AND EXPERIMENTAL METHODS Chemical syntheses:

Meclofenamic acid (Compound 1), diclofenac (Compound 2) and 1 -ethyl benzimidazolone (1-EBIO, Compound 10) are commercially available and were purchased from Sigma-Aldrich (St. Louis, MO, USA).

The chemical structures of Compounds 1 - 10 of the present invention are presented in Figure 17.

Synthesis of Compound 7: Diclofenac (50 mg, 0.17 mmol) was dissolved in dry dichlromethane (DCM) and a catalytic amount of N-dimethylaminopyridine (DMAP) and diethylene glycol

(0.08 ml, 0.85 mmol) were added. The mixture was cooled to 0 °C while stirring, and a solution of dicyclohexyl carbodiimide (DCC, 52.6 mg, 0.255 mmol) in DCM was added dropwise. The resulting suspension was then stirred at 0 °C for 30 minutes, while being monitored by TLC (using a mixture of 1:1 EtOAc:Hexane as eluent). The solid was thereafter removed by filtration and was washed with DCM. The filtrate was concentrated under reduced pressure and chromato graphed on silica gel to afford the pure product (43 mg, 66 %).

Η-NMR (200 MHz, CDC1₃): δ = 7.35 (2H, d, J=8); 7.19-7.06 (2H, m); 6.99(1H, d, J=8); 6.96 (IH, d, J=8.); 6.56 (IH, d, J=8); 4.35 (2H, m); 3.8 (2H, s); 3.65-3.75(4H, m); 3.53-3.57(2H, m) ppm. Synthesis of Compound 8:

Compound 8 was prepared according to the procedure described above for Compound 7, using di ethyl eneglycol methyl ether instead of diethylene glycol. The product was obtained in 53 % yield.

1H-NMR (200 MHz, CDC1₃): δ = 7.35 (2H, d, J=8); 7.19-7.06 (2H, m); 6.99(1H, d, J=8); 6.96 (IH, d, J=8.); 6.56 (IH, d, J=8); 4.34-4.29 (2H, m); 3.85 (2H, s); 3.75- 3.70(2H, m); 3.61-3.59 (2H, m); 3.58-3.50 (2H, m) ppm. Synthesis of Compounds 3, 4, 5 ,6 and 9 - general procedure:

To the corresponding acid (meclofenamic acid, diclofenac or a derivative thereof) (0.506 mmol), dissolved in dichloromethane, N-hydroxy succinimide (0.76 mmol) and DCC (0.76 mmol) were added. The mixture was stirred for 1 hour while being monitored by TLC (using a mixture of 1:1 EtOAc:Hexane as eluent). After completion of the reaction the mixture was filtered and the solvent was evaporated. The crude product was separated by column chromato graphy to give the pure N- hydroxy succinimide ester intermediate of Compounds 3, 4, 5, 6 and 9, as follows:

Intermediate of Compound 3: Yield 90 %. 1H-NMR (200 MHz, CDC1₃): δ = 8.69 (IH, s); 8.09-8.14 (IH, dd, J=1.7, J=8.6); 7.32-7.37 (IH, dt, J=1.8, J=8.5); 7.14- 7.28 (3H, m); 6.72-6.80 (2H, m); 2.9 (4H, s); 2.29 (3H, s) ppm.

Intermediate of Compound 4: Yield 90 %. 1H-NMR (200 MHz, CDC1₃): δ = 8.91 (IH, s); 8.10-8.15 (IH, dd, J=1.6, J=8.18); 7.33-7.48 (5H, m); 7.24 (IH, d, J=7.8); 6.80-6.88 (IH, dt, J=1.06, J=7.1); 2.9 (4H, s) ppm.

Intermediate of Compound 5: Yield 100 %. Η-NMR (200 MHz, CDC1₃): δ = 8.6 (IH, s); 8.12 (IH, dd, J=1.53, J=7.4); 7.32(1H, ); 7.06-7.13 (3H, m); 6.66- 6.74 (2H, m); 2.9 (4H, s); 2.32 (3H, s); 2.14(3H, s) ppm. Intermediate of Compound 6 and 9: Yield 89 %. 1H-NMR (200MHz, CDC1₃): δ = 7.35 (3H, d, J=8); 7.18-7.26 (IH, m); 6.93-7.07 (2H, m); 6.6-6.63 (IH, d, J=7.9); 6.2 (IH, s); 4.13 (2H, s); 2.84 (4H, s) ppm.

The corresponding intermediate was dissolved in 1 ml of dimethylformamide (DMF), diethyleneglycolamine, or (2-hydroxyethyl)methylamine for Compound 9, (1 equivalent), was added and the mixture was stirred for 30 minutes, while being monitored by TLC (using EtOAc as eluent). After completion of the reaction the solvent was removed under reduced pressure. The product was purified by column chromatography to give Compounds 3, 4, 5 , 6 and 9, as follows: Compound 3: Yield 55 %. 1H-NMR (200 MHz, CDC1₃): δ = 9.31 (IH, s);

7.45-7.50 (IH, dd, J=0.6, J=8); 6.99-7.22 (5H, m); 6.67-6.94 (IH, dt, J=1.8, J=6.5); 3.54-3.74 (8H, m); 2.33 (3H, s) ppm.

Compound 4: Yield 70 %. 1H-NMR (200 MHz, CDC1₃): δ = 9.44 (IH, s); 7.46-7.50 (IH, dd, J=0.6, J-8); 7.4 (2H, d, J=9); 7.16-7.32 (5H, m); 7.09 (IH, bs); 6.79-6.84 (IH, dt, J=1.8, J=6.5); 3.54-3.73(8H, m) ppm.

Compound 5: Yield 88 %. 1H-NMR (200 MHz, CDC1₃): δ = 9.19 (IH, s); 7.41-7.45 (IH, dd, J=1.53, J=7.4); 7.14-7.21(2H, m); 7.06-7.13 (IH, t, J=); 6.93-6.96 (2H, m); 6.65-6.69 (2H, m); 3.6-3.8 (8H, m); 2.32(3H, s); 2.20 (3H, s) ppm.

Compound 6: Yield 72 %. 1H-NMR (200MHz, CDCI₃): δ = 7.35 (2H, d, J=8); 7.19-7.06 (2H, m); 6.99(1H, d, J=8); 6.96 (IH, d, J=8.); 6.56 (IH, d, J-8); 3.69 (2H, s); 3.55-3.49(4H, m) ppm.

Compound 9: Yield 63%. 1H-NMR (200 MHz, CDC1₃): δ = 7.35 (2H, d,

J=8); 7.19-7.06 (2H, m); 6.99(1H, d, J=8); 6.96 (IH, d, J=8.); 6.56 (IH, d, J=8); 3.78-

3.74 (2H, m); 3.64-3.57 (2H, m); 3.23 (2H, s); 2.95 (3H, s) ppm.

Biology CHO cell culture and transfection:

CHO (Chinese Hamster Ovary) cells were grown in Dulbecco's modified

Eagle's medium supplemented with 2 mM glutamine, 10% fetal calf serum and antibiotics. Briefly, 40,000 cells seeded on poly-D-lysine-coated glass coverslips (13 mm diameter) in a 24-multiwell plate were transfected with pIRES-CD8 (0.5 μg) as a marker for transfection and with KCNQ2 (0.5 μg) and/or KCNQ3 (0.5μg). For electrophysiology, transfected cells were visualized approximately 40 hours following transfection, using the anti-CD8 antibody-coated beads method (Jurman et al, 1994). Transfection was performed using 3.5 μl of lipofectamine (Gibco-BRL) according to the manufacturer's protocol.

Neuronal cortical culture:

Sprague Dawley rat embryos (El 8) were removed by caesarian section and their cortices were dissected out. The tissue was digested with papain (100 U; Sigma, St. Louis, MO) for 20 min, triturated to a single-cell suspension, and plated at a density of 40,000 cells per ml on a substrate of bovine collagen type IV and 100 μg/ml poly-L- lysine in 13 mm diameter glass coverslip of a 24-multiwell plate. The culture medium consisted of Modified Eagle's Medium containing 5% horse serum (Biological Industries, Beit HaEmek, Israel), B-27 neuronal supplement (Invitrogen, Carlsbad, CA), 100 U/ml penicillin, 100 μg/ml streptomycin, and 2 mM glutamine. D-Glucose was supplemented to a final concentration of 6 g/1. Cytosine-1-D-arabinofuranoside (5 μM) was added after 5 days to arrest glial cell proliferation. All cultures were maintained at 37°C in humidified air containing 5% CO₂. Electrophysiology: For current measurements in CHO cells, recordings were performed 40 h following transfection, using the whole-cell configuration of the patch-clamp technique (Hamill et al, Nature 294: 462-464 (1981)). Signals were amplified using an Axopatch 200B patch-clamp amplifier (Axon Instruments, Foster City, California, USA), sampled at 2 kHz and filtered at 800 Hz via a 4-pole Bessel low pass filter. Data were acquired using pClamp 8.1 software (Axon Instruments, Foster City, California, USA) and an Elonex Pentium III computer in conjunction with a DigiData 1322 A interface (Axon Instruments, Foster City, California, USA). The patch pipettes were pulled from borosilicate glass (Warner Instrument. Corp., Hamden, Connecticut, USA) with a resistance of 2-5 MΩ and were filled with (in mM): 130 KC1, 1 MgCl₂, 5 K₂ATP, 5 EGTA, 10 HEPES, adjusted with KOH at pH 7.4 (290 mOsm). The external solution contained (in [mM] ): 140 NaCl, 4 KC1, 1.8 CaCl₂, 1.2 MgCl₂, 11 glucose, 5.5 HEPES, adjusted with NaOH at pH 7.4 (310 mOsm). Series resistances (3-13 MΩ) were compensated (75-90%) and periodically monitored. For current-clamp measurements of rat cortical neurons, recordings were performed 10-14 days after plating. The patch pipettes were filled with (in [mM] ): 135 KCl, 1 K₂ATP, 1 MgATP, 2 EGTA, 1.1 CaCl₂, 5 glucose, 10 HEPES, adjusted with KOH at pH 7.4 (315 mOsm). The external solution contained (in [mM] ): 140 NaCl, 4 KCl, 2 CaCl₂, 2 MgCl₂, 5 glucose, 10 HEPES, adjusted with NaOH at pH 7.4 (325 mOsm). For evoking the neuronal action potentials, 50-300 pA currents were injected into the cells for 800 ms (square pulse). Recordings were sampled at 5 kHz and filtered at 2 KHz via a 4-pole Bessel low pass filter. For voltage-clamp measurements of rat cortical neurons, the patch pipettes were filled with (in mM): 90 K-acetate, 40 KCl, 3 MgCl₂, 2 K₂ATP, 20 HEPES, adjusted with KOH at pH 7.4 (310-315 mOsm). The external solution contained (in mM): 120 NaCl, 23 NaHCO₃, 3 KCl, 2.5 CaCl₂, 1.2 MgCl₂, 11 glucose, 0.0005 tetrodotoxin (TTX), 5 HEPES, adjusted with NaOH at pH 7.4 (325 mOsm.

Current measurements in Xenopus oocytes were performed as described in the art (Peretz et al. J. Physiol 545:751-766 (2002)). Briefly, two-electrode voltage-clamp measurements were performed 3-5 days following cRNA micro injection into oocytes. Oocytes were bathed in a modified ND96 solution containing (in mM: 96 NaCl, 2 KCl, 1 MgCl₂, 0.1 CaCl₂ and 5 HEPES titrated to pH = 7.4 with NaOH). Whole-cell currents were recorded at room temperature (20°C-22°C) using a GeneClamp 500 amplifier (Axon Instruments, Foster City, California, USA). Glass microelectrodes (A-M Systems, Inc., Carlsborg, Wisconsin, USA) were filled with 3M KCl and had tip resistances of 0.5-1.5 MΩ. Stimulation of the preparation, data acquisition and analyses were performed using the pCLAMP 6.02 software (Axon Instruments, Foster City, California, USA) and a 586 personal computer Pentium 4 interfaced with a Digidata 1200 interface (Axon Instruments, Foster City, California, USA). Current signals were filtered at 0.5 kHz and digitized at 2 kHz. Data analyses:

Data analysis was performed using the Clampfit program (pClamp 8.1, Axon Instruments, Foster City, California, USA), Microsoft Excel 98 (Microsoft Corp., Redmond, Washington, USA), Axograph 4.6 (Axon Instruments, Foster City, California, USA) and Prism 2.0 (GraphPad, San Diego, California, USA). Leak subtraction was performed off-line, using the Clampfit program of the pClamp 8.1 software. To analyze the KCNQ2/3 channel deactivation, a single exponential fit was applied to the tail currents. Chord conductance (G) was calculated by using the equation: G = I / (V - V_rev )

where I corresponds to the current amplitude measured at the end of the pulse and V_rev, the calculated reversal potential assumed to be -90 mV in CHO cells and -98 mV in Xenopus oocytes. G was estimated at various test voltages V and then, nomialized to a maximal conductance value, G_max, calculated at +40 mV. Activation curves were fitted by a Boltzmann distribution:

G/G_max = l / { l + exp[ ( V₅₀ - V) / s ] }

where V₅o is the voltage at which the current is half-activated and s is the slope factor. All data were expressed as mean ± SEM. Statistically significant differences were assessed by Student's -test.

EXPERIMENTAL RESULTS

Assays conducted with meclofenamic acid (Compound 1), diclofenac (Compound 2) and 1-EBIO (Compound 10)

EXAMPLE 1

The effect of Meclofenamic acid (Compound 1) and 1-EBIO (Compound 10) on the

KCNQ2/3 current

The leftwards-shift of the voltage dependence of activation of the KCNQ2/3 current induced by Meclofenamic acid (1) and 1-EBIO (10) is discussed with reference to Figures 1 A- ID.

When KCNQ2 and KCNQ3 subunits are expressed separately as homomeric channels in various expression systems, they give rise to relatively small potassium currents, especially for KCNQ3 (Wang et al. Science 282: 1890-1893 (1998) and Yang et al. J. Biol. Chem. 273: 19419-19423 (1998)). However, KCNQ2 co- expressed with KCNQ3 produces a current whose amplitude is about 10 times that of the sum of the two homomeric channels and whose biophysical and pharmacological properties are very similar to those of the native M-current (Main et al. Mol. Pharmacol. 58: 253-262 (2000), Wang et al. Science 282: 1890-1893 (1998) and Yang et al J. Biol. Chem. 273: 19419-19423 (1998)).

CHO cells were co-transfected with the two corresponding cDNAs of KCNQ2 and KCNQ3 at an equimolar ratio and exposed to meclofenamic acid (1) and 1-EBIO (10) so as to identify the effect of these compounds on M-current.

Turning to Figures 1A and IB, representative traces were recorded from the same cell before (left panel) and after (right panel) external application of 100 μM meclofenamic acid (Compound 1, Figure 1A) and 100 μM 1-EBIO (Compound 10, Figure IB). The membrane potential was stepped from -70 mV to +40 mV for 1.5 second pulse duration, in 10 mV increments, followed by 0.75 second pulse to -60 mV, producing the tail current. The holding potential of all experiments was -85 mV.

The normalized conductance (G/G_max) was plotted as a function of the voltage steps, for the control (open squares) and meclofenamic acid (Compound 1, Figure 1C) or 1-EBIO (Compound 10, Figure ID) treated cells (solid square). The activation curves were fitted using the Boltzmann distribution.

Figure 1A (left panel) shows representative traces of the KCNQ2/3 current activated by step depolarization above a voltage threshold of about -50 mV. Addition of 100 μM meclofenamic acid externally shown in Figure 1A (right panel) produced a pronounced leftward shift of 22.7 mV in the voltage-dependence of KCNQ2/3 current activation, from V₅₀ = -19.65 ± 1.85 V (n = 24) to V₅₀ = -42.34 ± 2.08 mV (n = 14). The slope parameter of the Boltzmann fitting curve did not change significantly with s = -9.46 ± 0.41 mV/e fold and s = -10.50 ± 0.93 mV/e fold, for control and meclofenamic acid (Compound 1), respectively.

In Figure IB (right panel) the leftward shift caused by 100 μM 1-EBIO (Compound 10) of 7.9 mV (p<0.005) in the voltage-dependence of KCNQ2/3 current activation, from V₅₀ = -22.9 ± 1.8 mV to V₅₀ = -30.8 ± 2.8 mV (n = 4). The slope parameters of the Boltzmann fitting curve did not change significantly and were s = - 10.2 ± 1.1 mV/e-fold and s = -9.5 ± 0.7 mV/e-fold, for control and 1-EBIO (Compound 10), respectively. Consequently, upon exposure to meclofenamic acid (Compound 1) or 1-EBIO

(Compound 10) the KCNQ2/3 current activated at more hyperpolarized potentials above a voltage threshold of about -60 mV versus -50 mV for control (Figures 1C and ID). EXAMPLE 2

Augmentation of the KCNQ2/3 current amplitude by Meclofenamic acid

(Compound 1) and 1-EBIO (Compound 10)

The augmentation of the KCNQ2/3 current amplitude by meclofenamic acid (Compound 1) and 1-EBIO (Compound 10) is discussed with reference to Figures 1 A-1D and Figures 2A-2C.

As is shown in Figure 2A, the KCNQ2/3 current increases in the presence of meclofenamic acid (Compound 1) and 1-EBIO (Compound 10). Traces were recorded in the absence (control) and presence of meclofenamic acid (Compound 1, left panel) or 1-EBIO (Compound 10, right panel). The cells were stepped to -20 mV for 1.5 second pulse duration. In this train protocol, the interval between the pulses was 30 seconds.

Figure 2B shows the percentages of the current measured in the presence (+) or absence (-) of meclofenamic acid (Compound 1, left panel) or 1-EBIO (Compound 10, right panel), where the control is 100 %.

In Figure 2C, the current amplitude (nA) was plotted against the step voltage, in the hyperpolarized potential range (-70 mV to -20 mV) to emphasize the negative shift of the threshold for channel activation, in response to meclofenamic acid (Compound 1, left panel) or 1-EBIO (Compound 10, right panel) application, Within about one minute of external application of meclofenamic acid

(Compound 1) a large increase in KCNQ2/3 current amplitude across a range of test potentials between -50 to 0 mV was observed (Figure 1C and Figure 2A, left panel). The effect of meclofenamic acid (Compound 1) was fully reversible (data not shown). In the train protocol, when the cells were stepped to -20 mV the application of meclofenamic acid (Compound 1) induced an increase of the current amplitude by up to 72 %, from 844 ± 130 pA to 1451 ± 164 pA (n = 15), for control and meclofenamic acid (Compound 1), respectively (Figure 2B, left panel). From the normalized conductance-voltage relation (G/G_max) and the normalized current-voltage relation (I/Imax) presented in Figures 1C and 2C, respectively, one can see that meclofenamic acid (Compound 1) increased KCNQ2/3 potassium current primarily via a leftward shift in the voltage-dependence of channel activation. As the test potentials became more positive and approached saturation values of the activation curve (i.e., +20 mV), the effects of meclofenamic acid (Compound 1) on KCNQ2/3 current amplitude became very small. Clearly, the most pronounced action of meclofenamic acid (Compound 1) was exerted at negative physiologically relevant potentials. At -50 mV, -40 mV and -30 mV, meclofenamic acid (Compound 1) increased KCNQ2/3 current amplitude by more than 10-fold, 5-fold and 2.5-fold, respectively (Figures 1A, 1C and 2C, left panels).

Similarly, addition of 100 μM 1-EBIO (Compound 10) quickly led to an increase in KCNQ2/3 current amplitude across a range of test potentials between -50 to -10 mV (Figure IB, Figure 2A and Figure 2B, right panels), although the effect was less pronounced than the effect of meclofenamic acid (Compound 1). The effect of 1- EBIO (Compound 10) was fully reversible (data not shown). In the train protocol, when the cells were stepped to -30 mV the application of 1-EBIO (Compound 10) induced an increase of the current amplitude by up to 57 %, from 945 ± 135 pA (n = 14) to 1483 ± 194 (n = 14), for control and 1-EBIO (Compound 10), respectively. From the normalized conductance-voltage relation (G/G_maχ) and the normalized current- voltage relation (I/I_max) presented in figures ID and 2C, right panel, respectively, one can see that 1 -EBIO (Compound 10) increased KCNQ2/3 potassium current primarily via a leftward shift in the voltage-dependence of channel activation. As the test potentials became more positive and approached the saturation values of the activation curve, the effects of 1-EBIO (Compound 10) on KCNQ2/3 current amplitude became weaker. Figure ID shows the leftward shift of the threshold for channel activation of about 10 mV. In addition, Figure 2C, right panel shows that the most pronounced action of 1-EBIO (Compound 10) was exerted at negative physiologically relevant potentials. At -50 mV and -40 mV, 1-EBIO (Compound 10) increases the KCNQ2/3 current amplitude by more than 10-fold and 3-fold, respectively.

EXAMPLE 3 The effect of meclofenamic acid (Compound 1) and 1-EBIO (Compound 10) on

KCNQ2/3 deactivation kinetics The slowing down of KCNQ2/3 deactivation kinetics caused by meclofenamic acid (Compound 1) and 1-EBIO (Compound 10) is discussed with reference to Figures 3A-3E. In Figure 3A the tail current of a cell before (control) and following application of meclofenamic acid (Compound 1) is shown. The prepulse was -20 mV while the tail potential was -60 mV.

In Figure 3B the time constant resulting from the exponential fit of the tail current decay (τ_deact)_. in the absence (-) and the presence of meclofenamic acid (Compound 1) (+) is depicted.

Although meclofenamic acid (Compound 1) and 1-EBIO (Compound 10) did not affect the KCNQ2/3 activation kinetics, they markedly slowed down the deactivation process. The decay of the tail current or deactivation reflects the transition of the channel from the open state to the close state. As seen in Figure 3 A, the cells were depolarized to -20 mV and then repolarized to -60 mV. The decay of the tail current was fitted using one exponential function. In response to the addition of 100 μM meclofenamic acid (Compound 1), the time constant for deactivation increased by about 2-fold, from τ_deact ⁼ 79.6 ± 4.5 msec to τ_de ct = 167.5 ± 11.6 msec (n = 10). The results are highly significant, as shown in Figure 3B and Figure 3C.

As noted above, 1-EBIO (Compound 10) did affect significantly (p<0.001) the KCNQ2/3 deactivation kinetics. The decay of the tail current was fitted using one exponential function. In response to the addition of 100 μM 1-EBIO (Compound 10), the time constant for deactivation increased from τ_deact ⁼ 91.2 ± 8.4 msec to Xd_eact ⁼ 110.1 ± 9.5 msec (n = 14, Figure 3E). Figure 3D shows that 1-EBIO (Compound 10) slows down the deactivation kinetics of KCNQ2/3 channels.

EXAMPLE 4 Inhibition of evoked and spontaneous neuronal activity by meclofenamic acid (Compound 1) and 1-EBIO (Compound 10)

The inhibition of evoked and spontaneous neuronal activity by meclofenamic acid (Compound 1) and 1-EBIO (Compound 10) is discussed with reference to Figures 4A-4B.

In Figures 4A-4B neuronal activity depression by meclofenamic acid (Compound 1) is shown. In Figure 4A, evoked rat cortical neuronal activity before

(first row) during (second and third rows) and after (fourth and fifth rows) application of different meclofenamic acid (Compound 1) concentrations. Each column corresponds to a different neuron.

In Figure 4B, spontaneous activity recorded before, during and after 10 μM meclofenamic acid (Compound 1) application is depicted. As discussed hereinabove, one of the main functions of the M-current is monitoring the excitability of neurons in the brain. From the result above, it is clear that meclofenamic acid (Compound 1) and 1-EBIO (Compound 10) enhance the heterologously expressed M-current. The question remains how meclofenamic acid (Compound 1) and 1-EBIO (Compound 10) modulate the excitability of neurons expressing the M-current. To answer this question, a primary culture of rat cortical neurons and the current clamp configuration of the patch clamp technique were used.

First, how meclofenamic acid (Compound 1) and 1-EBIO (Compound 10) affect the evoked action potential activity of the rat cortical neurons was studied. In Figure 4A it is seen that the evoked potentials are reversibly inhibited by meclofenamic acid (Compound 1), in the range of 5-20 μM. Each lane of Figure 4A is recorded from a different neuron, using the current-clamp configuration of the patch- clamp technique.

Second, it is seen that the spontaneous activity of the rat cortical neurons is completely but reversibly inhibited by 10 μM meclofenamic acid (Compound 1), Figure 4B.

EXAMPLE 5 Protection of mice from seizures produced by electroshock using meclofenamic acid

(Compound 1) The effect of meclofenamic acid (Compound 1) in protecting mice from electroshock induced seizures is discussed with reference to Figure 5.

Five groups of 10 ICR mice each received intraperitoneally saline or meclofenamic acid (Compound 1) at 25 mg/lcg, 50 mg/kg, 100 mg/kg and 150 mg/lcg and were subjected 30 minutes later to an electric shock (50 mA, 0.2 second duration, 60 Hz). The relative fraction of mice that did not produce seizures was plotted for each dose in Figure 5.

In view of the strong depressing activity of meclofenamic acid (Compound 1) on cortical neurons, the anticonvulsant activity in mice subjected to seizures produced by electroshock was examined. Meclofenamic acid (Compound 1) dissolved in saline was injected intraperitoneally (in a volume of 10 ml/kg) at doses ranging from 25 mg/lcg to 150 mg/lcg to ICR adult mice and its anticonvulsant activity was compared with saline. Thirty minutes after drug administration, seizures were produced by electroshock (50 mA, 0.2 second duration, 60 Hz). Figure 5 shows that 50 mg/lcg meclofenamic acid (Compound 1) significantly protected 50% of the mice from electroshock and at 100 mg/kg fully prevented seizures. At 150 mg/kg meclofenamic acid (Compound 1) led to sedation of the mice.

EXAMPLE 6

The effect of diclofenac (Compound 2) on the voltage dependence activation of the KCNQ2/3 current and the deactivation ofKCNQ2/3 current

The leftwards-shift of the voltage dependence of activation of the KCNQ2/3 current induced by diclofenac (Compound 2) is discussed with reference to Figures 6A-6C.

In Figures 6A-6C the enhancement of the KCNQ2/3 current caused by diclofenac (Compound 2) is shown. In Figure 6A whole cell currents of KCNQ2/3 heterogously expressed in CHO cells recorded before and after perfusion of 50 μM diclofenac are shown. In Figure 6A the cell membrane was stepped from -90 mV to - 50 mV (1 second) followed by tail step to -60 mV (0.75 second). Recordings were taken every 30 seconds. In Figure 6B the percentage of the current presented in the presence (+) or absence (-) of diclofenac, where the control is 100%, taken from the experiment presented in Figure 6A is shown. In Figure 6C the normalized conductance (G/G_max) is plotted as a function of the voltage steps, for the control (open squares) and diclofenac (closed squares), for KCNQ2/3 current.

In Figure 6C, the addition of 50 μM diclofenac (Compound 2) induced a leftward shift of -14.5 mV in the voltage-dependence of KCNQ2/3 activation, from V₅₀ = -30.9 ± 4.1 mV to V₅₀ = -45.4 ± 2.7 mV (n = 1, p <0.01).

In Figure 6C, it is seen that as with meclofenamic acid (Compound 1), treatment of CHO cells with diclofenac (Compound 2) slowed down the deactivation kinetics of KCNQ2/3 channels

In Figures 6A and 6B it is seen that the KCNQ2/3 current amplitude is increased by diclofenac (2) at physiologically relevant potentials. In a train protocol, when the cells were stepped from -85 mV to -50 mV the application of diclofenac (Compound 2) induced an increase of the current amplitude by up to 262 ±26 % (n = 6).

The effects of diclofenac (Compound 2) were fully reversible (data not shown). In general, these effects of both meclofenamic acid (Compound 1) and diclofenac (Compound 2) appear to result from a stronger impact of the openers on KCNQ2 than on KCNQ3 channel subunits, as is demonstrated in Example 7 below.

EXAMPLE 7 Selectivity of meclofenamic acid (Compound 1) action

The opener properties of meclofenamic acid (Compound 1) and diclofenac on heteromeric KCNQ2/Q3 channels raise the question of whether these compounds act equally well or more selectively on either subunit. To address this problem, the effect of 50 μM meclofenamic acid on homomeric KCNQ2 channels and homomeric KCNQ3 expressed separately in CHO cells was tested. The results are presented in Figures 16A-D.

As is shown in Figure 16C, it was found that meclofenamic acid generally exerted a stronger action on KCNQ2 than on KCNQ3 channels as it produced a substantial leftward shift of -26.9 mV in the activation curve of KCNQ2 channels, from V₅₀ = -23.6 ± 2.2 mV (n = 8) to V₅₀ = -50.5 ± 1.4 mV (n = 5) in control and meclofenamic acid-treated cells, respectively (Figure 16C left panel, p < 0.01). The leftward shift produced by meclofenamic acid on the activation curve of KCNQ3 channels was weaker (-15 mV) from V₅o = -39.0 ± 3.5 mV (n = 11) to V₅o = -54.0 ± 2.0 mV (n = 6) in control and meclofenamic acid-treated cells, respectively (Figure 16C right panel, p < 0.01).

Meclofenamic acid (Compound 1) significantly reduced the speed of KCNQ2 channel closure with the time constant of deactivation increasing from τ eac_t = 92.6 ± 3.9 msec to τ_deact = 152.7 ± 4.9 msec (Figures 16A and 16D; n = 8, p O.001). In contrast, it did not affect the deactivation kinetics of KCNQ 3 channels (τ _eact = 319.6 ± 40.8 msec and τdeact ⁼ 17.2 ± 30.2 msec for control and meclofenamic acid-treated cells, respectively, n = 8). Reflecting the stronger effect of the opener on KCNQ2 versus KCNQ3 channels, the application of 50 μM meclofenamic acid produced an increase of 240 ± 26 % and 120 ± 4 % of the KCNQ2 and KCNQ3 current amplitudes, respectively, when the cells were stepped from -85 mV to -40 mV (Figures 16A and B; n = 8, pθ.001).

Interestingly, homomeric KCNQ1 and heteromeric KCNQ1/KCNE1 currents were not enhanced by 50 μM diclofenac or 50μM meclofenamic acid across a range of test potentials between -50 to 0 mV (data not shown). Instead, both openers reduced the current amplitude at positive potentials (from 0 mV to 40 mV).

Assays conducted with Compounds 3-9

Novel Compounds 3, 4, 5, 6, 7, 8 and 9 were tested for KCNQ2/3 opening activity as is described below. It is important to note that compounds 3-9 were also tested and found to have no effect on KCNQ1/KCNE1 cardiac channels and displayed a selective brain specificity. These results are not shown.

EXAMPLE 8 The effect of Compound 6 on KCNQ2/3 channels and on rat cortical neurons

The effect of Compound 6 on recombinant KCNQ2/3 potassium channels heterogously expressed in CHO cells and on rat cortical neurons is discussed with reference to Figures 7A-7C, Figures 8A-8B and Figures 9A-9C.

In Figures 7A-7C the effects of Compound 6 on KCNQ2/3 currents are shown. In Figure 7A whole-cell currents recorded before and after perfusion of 25 μM Compound 6 are shown. In Figure 7B the percentage of the current presented in the presence (+) or absence (-) of Compound 6, where the control is 100%, taken from the experiment presented in Figure 7A. In Figure 7C the normalized conductance (G/G_maχ) was plotted as a function of the voltage steps, for the control (open squares) and Compound 6 (closed squares), for KCNQ2/3 current are shown.

In Figures 8A-8B inhibition of the evoked neuronal activity by the Compound 6 is shown. In Figure 8 A neuronal activity, as evoked by square depolarizing current, inhibited by lOμM Compound 6 and recovered after wash is shown. In Figure 8 A the depolarizing current was 50 pA for 800 msec. In Figure 8B the evoked neuronal activity, using the ramp protocol, recorded before, after external perfusion of Compound 6 and recovered after wash is shown. In the ramp protocol depicted in Figure 8B, the depolarizing current was ramped from 0 pA to 300 pA within 800 msec.

In Figures 9A-9C inhibition of the spontaneous neuronal activity by different concentration of Compound 6 is shown. Shown is spontaneous activity recorded before, after addition and after wash of Compound 6, for 20μM in Figure 9A, for lOμM in Figure 9B and 5μM in Figure 9C.

Figure 7C shows the effects of 25 μM Compound 6 on recombinant KCNQ2/3 channels expressed in CHO cells. As with meclofenamic acid (Compound 1) and diclofenac (2), when externally applied Compound 6 produced a significant leftward shift of ~13 mV in the voltage-dependence of KCNQ2/3 activation, from V₅o = -30.9 ±

4.1 mV to V₅₀ = -43.45 ± 2.3 mV (n = 7, pθ.01).

In Figure 7A it is seen that Compound 6 also slowed down the deactivation kinetics of KCNQ2/3 channels. In Figures 7A and 7B it is seen that the KCNQ2/3 current amplitude was increased by Compound 6 at physiologically relevant potentials. In a train protocol, when the cells were stepped from -85 mV to -50 mV the application of 25 μM

Compound 6 induced an increase of the current amplitude by up to 220%> (n = 10).

The effect of Compound 6 was reversible (data not shown). As one of the main functions of the M-current is to dampen the neuronal spiking discharges, what the effect of Compound 6 on the evoked and spontaneous action potential activity of cultured rat cortical neurons was examined. Using the current-clamp configuration of the patch clamp technique, the effects of Compound 6 on neuronal action potentials evoked either by a squared pulse (Figure 8 A; 50-300 pA, 800 msec) or a ramp (Figure 8B; 0-300 pA in 800 msec) of depolarizing current were examined. The resting membrane potential was close to -60 mV and, when needed, was maintained at this level by injecting DC current. Superfusion of 10 μM Compound

6 reduced drastically and reversibly the number of evoked action potentials in cortical neurons. Within less than one minute following external superfusion of 10 μM Compound 6, the cortical neurons fired action potentials with a widening of interspike interval (Figure 8 A, 2nd row). After 2 minutes of opener exposure, only one spike could be evoked by the same depolarizing current (Figure 8A, 3rd row). Thus,

Compound 6 consistently reduced the number of evoked action potentials. Upon washout of the Compound for less than 2 minutes, neurons recovered their initial spiking activity (Figure 8 A, 5th row).

Similar results were obtained with ramp currents depicted in Figure 8B.

Using higher density cultures of rat cortical neurons, spontaneous spiking activity was recorded (Figures 9). Compound 6 dose-dependently (5 μM - 20 μM) produced within less than 2 minutes a profound depression of spontaneous action potentials. The depressing action of Compound 6 could be reversed by washout of the

Compound 6 Compound for all three concentrations.

EXAMPLE 9

The effect of Compound 5 on KCNQ2/3 channels and on rat cortical neurons

The effect of Compound 5 on recombinant KCNQ2/3 potassium channels heterogously expressed in CHO cells and on rat cortical neurons is discussed with reference to Figures lOA-lOC. In Figures 10A-10C the effects of Compound 5 on the neuronal activity and

KCNQ2/3 current is shown. In Figure 10A, evoked rat cortical neuronal activity recorded before, after application of 25 μM Compound 5, and after washing is shown.

In Figure 10B KCNQ2/3 currents recorded before (upper panel) and after (lower panel) application of 25μM Compound 5. In Figure 10B, the cell membrane was stepped from -80 mV to -40 mV in 10 mV increments (holding potential = -90 mV).

In Figure 10C the normalized conductance (G/G_max) was plotted as a function of the voltage steps, for the control (open squares) and Compound 5 (closed squares).

From Figures 10 A- 10C it is seen that Compound 5 has KCNQ2/3 potassium channels opening properties similar to those of Compound 6. In Figure 10C it is seen that 25 μM Compound 5 produced a significant leftward shift of ~8 mV in the voltage-dependence of KCNQ2/3 activation, from V₅₀ =

-30.9 ± 4.1 mV to V₅₀ = -38.3 ± 1.9 mV (n = 5, p<0.05).

In Figure 10B it is seen that Compound 5 slowed down the deactivation kinetics of KCNQ2/3 channels. It is also seen that 15 μM Compound 5 enhances the current amplitude at the physiologically relevant potentials of -40 and -50 mV.

In Figure 10A it is also seen that superfusion of 25 μM Compound 5 reversibly inhibited the number of evoked action potentials in cortical neurons. EXAMPLE 10

The effect of Compound 3 on KCNQ2/3 channels

The effect of Compound 3 on recombinant KCNQ2/3 potassium channels heterogously expressed in CHO cells is discussed with reference to Figures 11 A-l IB. In Figures 11 A-l IB the KCNQ2/3 current increase in the presence of

Compound 3 is shown. In Figure 11 A are shown currents in the absence (control) and presence of 25 μM Compound 3. In Figure 11 A, the cells were stepped to -50 mV for 1.5 second pulse duration and the interval between the pulses was 30 second. In Figure 1 IB the percentage of the current presented in the presence (+) or absence (-) of Compound 3, where the control is 100%>, is shown.

In Figures 11A-1 IB it is seen that Compound 3 has potent KCNQ2/3 potassium channel opening properties at 25 μM. In a train protocol, when the cells were stepped from -85 mV to -50 mV the application of 25 μM Compound 3 induced an increase of the current amplitude by up to 299 ± 47 % (n = 6; p < 0.002). The effect of Compound 3 was fully reversible (data not shown). .

EXAMPLE 11

The effect of Compound 4 on KCNQ2/3 channels and on rat cortical neurons

The effect of Compound 4 on recombinant KCNQ2/3 potassium channels heterologously expressed in CHO cells and on rat cortical neurons is discussed with reference to Figures 12A-12C.

In Figures 12A-12C, the effects of Compound 4 on neuronal activity and

KCNQ2/3 current are shown. In Figure 12A KCNQ2/3 currents recorded before (left panel) and after (right panel) application of 50μM Compound 4 are shown. The cell membrane was stepped from -80 mV to -40 mV in 10 mV increments (holding potential = -90 mV). In Figure 12B spontaneous cortical neuron activity recorded before, after addition of 20 μM Compound 4 and after a wash is shown. In Figure

12D the normalized conductance (G/G_maχ) was plotted as a function of the voltage steps, for the control (open squares) and Compound 4 (closed squares). In Figures 12A-12C it is evident that Compound 4 is a potent KCNQ2/3 channel opener. As seen in Figure 12A when CHO cells were stepped from -85 mV to

-50 mV and -40 mV, the application of 50 μM Compound 4 induced an increase of the current amplitude by more than 4-fold and 1.5-fold, respectively, as discussed hereinabove for Compound 3. This increase in KCNQ2/3 current amplitude results from the leftward shift produced by Compound 4 on the voltage-dependent curve of activation, Figure 12C.

In Figure 12B it is seen that when applied to rat cortical neurons, 20 μM Compound 4 markedly depressed the spontaneous spiking activity. As seen in Figure 12B, the effect of Compound 4 was fully reversible.

EXAMPLE 12 The effect of Compound 9 on KCNQ2/3 channels and on rat cortical neurons The effect of Compound 9 on rat cortical neurons is discussed with reference to

Figure 13. In Figure 13 is shown spontaneous neuronal activity (action potentials) as modulated by 20 μM Compound 9.

As seen in Figure 13, and in contrast to other molecules of the present invention, 20 μM of Compound 9 exhibit an inhibitory activity on evoked and spontaneous spiking activity of cortical neurons. However, in contrast to other molecules of the present invention, Compound 9 displayed only a weak opener action on recombinant KCNQ2/3 channels heterologously expressed in CHO cells. This result suggests that Compound 9 exerts neuronal depressant activity via mechanisms that do not involve KCNQ2/3 channels.

EXAMPLE 13 The effect of Compound 7 on KCNQ2/3 channels and on rat cortical neurons The effect of Compound 7 on recombinant KCNQ2/3 potassium channels heterologously expressed in CHO cells and on rat cortical neurons is discussed with reference to Figures 14A-14D.

In Figures 14A-14D the effects of 20 μM Compound 7 on KCNQ2/3 channels and neuronal activity are shown. In Figure 14A the KCNQ2/3 whole cell currents recorded before and after perfusion of 20 μM Compound 7 are shown. In Figure 14B the percentage of the current presented in the presence (+) or absence (-) of Compound 7, where the control is 100%, taken from the experiment presented in Figure 14 A. In Figure 14C, the normalized conductance of the KCNQ2/3 current (G/G_max) is plotted as a function of the voltage steps, for the control (open squares) and Compound 7 (closed squares). In Figure 14D is shown the modulation of spontaneous neuronal activity (action potentials) by 20 μM Compound 7.

Figure 14 shows the effects of Compound 7 on KCNQ2/3 channels. As with compounds 6 and 5 (discussed hereinabove), Compound 7 is a potent KCNQ2/3 channel opener with a marked leftward shift in the voltage dependent activation curve (Figure 14C). This effect led to a potent increase of the channel amplitude as determined by a train protocol (Figures 14A and 14B). Likewise, 20 μM Compound 7 exhibited a very potent inhibitory activity on evoked and spontaneous spiking activity of cortical neurons (Figure 14D). This effect was fully reversible.

EXAMPLE 14 The effect of Compound 8 on KCNQ2/3 channels and on rat cortical neurons The effect of Compound 8 on recombinant KCNQ2/3 potassium channels heterologously expressed in CHO cells and on rat cortical neurons is discussed with reference to Figures 15 A-l 5B.

In Figures 15A-15B, the evoked and spontaneous neuronal activity as modulated by Compound 8 is shown. In Figure 15 A, is shown evoked rat cortical neuronal activity recorded before, during and after application of lOμM Compound 8.

In Figure 15B the spontaneous neuronal activity (action potentials) as modulated by 5 μM Compound 8 is shown.

Figures 15A-B show that low concentrations of Compound 8 produced a potent inhibitory activity on evoked and spontaneous spiking activity of cortical neurons. However, like Compound 9 discussed hereinabove, Compound 8 displayed only a weak opener action of recombinant KCNQ2/3 channels heterologously expressed in CHO cells. This result suggests that Compound 8 exerts neuronal depressant activity via mechanisms that do not involve KCNQ2/3 channels.

It is appreciated that certain features of the invention, which are, for clarity, described in the context of separate embodiments, may also be provided in combination in a single embodiment. Conversely, various features of the invention, which are, for brevity, described in the context of a single embodiment, may also be provided separately or in any suitable subcombination. Although the invention has been described in conjunction with specific examples thereof, it is evident that many alternatives, modifications and variations will be apparent to those skilled in the art. Accordingly, it is intended to embrace all such alternatives, modifications and variations that fall within the spirit and broad scope of the appended claims. All publications, patents and patent applications mentioned in this specification are herein incorporated in their entirety by reference into the specification, to the same extent as if each individual publication, patent or patent application was specifically and individually indicated to be incorporated herein by reference. In addition, citation or identification of any reference in this application shall not be construed as an admission that such reference is available as prior art to the present invention.

Claims

WHAT IS CLAIMED IS:

1. A method of modulating a voltage-dependent potassium channel, the method comprising administering to the subject in need thereof a therapeutically effective amount of a compound selected from the group consisting of N- phenylanthranilic acid, a N-phenylanthranilic acid derivative, 2-benzimidazolone and a 2-benzimidazolone derivative, or a pharmaceutically acceptable salt thereof.

2. The method of claim 1, wherein said compound has a general Formula I or II:

Formula I Formula II

or a pharmaceutically acceptable salt thereof, wherein:

Z is an A-G(=K)-X-Y group, and wherein

A is alkyl or absent;

K is selected from the group consisting of oxygen and sulfur;

X is selected from the group consisting of substituted or unsubstituted nitrogen, oxygen, sulfur or absent; and

Y is selected from the group consisting of hydrogen, alkyl, hydroxyalkyl, cycloalkyl, aryl and a polyallcylene glycol residue, each of Q and W is independently selected from the group consisting of substituted or unsubstituted nitrogen, oxygen, sulfur and carbon;

D is selected from the group consisting of oxygen and sulfur;

Each of R², R³, R⁴, R⁵, R⁶, R⁷, R⁸, R⁹, R¹⁰, Rⁿ, R¹², R¹³ and R¹⁴ is independently selected from the group consisting of hydrogen, alkyl, hydroxyalkyl, trihaloalkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, heteroalicyclic, halo, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, sulfinyl, sulfonyl, cyano, nitro, amino and -NR¹⁵R¹⁶, or, alternatively, at least two of R², R³, R⁴, R⁵ and R⁶, of R⁷, R⁸, R⁹ and R¹⁰ and/or of R¹¹, R¹², R¹³ and R¹⁴ form a five- or six-membered aromatic, heteroaromatic, alicyclic or heteroalicyclic ring; and

R¹⁵ and R¹⁶ are each independently selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, carbonyl and sulfonyl, or, altematively R¹⁵ and R¹⁶ form a five- or six-member heteroalicyclic ring; whereas if said phosphor and/or said nitrogen is substituted, the substituent is selected from the group consisting of alkyl, cycloalkyl and aryl.

3. The method of claim 2, wherein said compound has the general Formula I.

4. The method of claim 3, wherein Y is selected from the group consisting of hydroxyalkyl and a polyalkylene glycol residue.

5. The method of claim 4, wherein said polyalkylene glycol residue has a general formula III:

[O-(CH₂)m]n-OR¹⁷

Formula III wherein: each of m and n is independently an integer of 1-10; and

R¹⁷ is hydrogen, alkyl, cycloalkyl or aryl.

6. The method of claim 3, wherein: G is carbon; K is oxygen; each of R², R³, R⁴, R⁵ and R⁶ is independently selected from the group consisting of hydrogen, alkyl, halo and trihaloalkyl; and each of R⁷, R⁸, R⁹ and R¹⁰ is hydrogen.

7. The method of claim 2, wherein said compound has the general formula II.

8. The method of claim 7, wherein:

Q and W are each substituted or unsubstituted nitrogen; and D is oxygen.

9. The method of claim 8 wherein Q is a substituted nitrogen.

10. The method of claim 2, wherein said compound is selected from the group consisting of:

10

and pharmaceutically acceptable salts thereof.

11. The method of claim 1, wherein said voltage-dependent potassium channel comprises a KCNQ2 channel and/or a KCNQ3 channel and/or a KCNQ2/3 channel.

12. The method of claim 1, wherein said modulating of said voltage- dependent potassium channel is for a treatment of a condition or disorder selected from the group consisting of epilepsy, ischemic stroke, migraine, ataxia, myokymia, neurogenic pain, Alzheimer's disease, Parkinson's disease, age-related memory loss, learning deficiencies, bipolar disorder, trigeminal neuralgia, spasticity, mood disorder, psychotic disorder, brain tumor, hearing and vision loss, anxiety and a motor neuron disease.

13. The method of claim 1, wherein said administering is effected intranasally, subcutaneously, intravenously, intramuscularly, parenterally, orally, topically, intradermally, bronchially, buccally, sublingually, supositorially and mucosally.

14. The method of claim 1, wherein said compound forms a part of a pharmaceutical composition which further includes a pharmaceutically acceptable carrier.

15. The method of claim 14, wherein said pharmaceutical composition further comprises an agent selected from the group consisting of an anti-bacterial agent, an antioxidant, a buffering agent, a bulking agent, an anti-inflammatory agent, an anti- viral agent, a chemotherapeutic agent and an anti-histamine.

16. A method of depressing cortical neuron activity, the method comprising administering to the subject in need thereof a therapeutically effective amount of a compound selected from the group consisting of N-phenylanthranilic acid, a N- phenylanthranilic acid derivative, 2-benzimidazolone and 2-benzimidazolone derivative.

17. The method of claim 16, wherein said compound has a general Formula

I or II:

Formula I Formula II

or a pharmaceutically acceptable salt thereof, wherein:

Z is an A-G(=K)-X-Y group, and wherein:

A is alkyl or absent; G is selected from the group consisting of carbon, sulfur and substituted or unsubstituted phosphor;

K is selected from the group consisting of oxygen and sulfur;

D is selected from the group consisting of oxygen and sulfur;

Each of R², R³, R⁴, R⁵, R⁶, R⁷, R⁸, R⁹, R¹⁰, R¹¹, R¹², R¹³ and R¹⁴ is independently selected from the group consisting of hydrogen, alkyl, hydroxyalkyl, trihaloalkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, heteroalicyclic, halo, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, sulfinyl, sulfonyl, cyano, nitro, amino and -NR¹⁵R¹⁶, or, alternatively, at least two of R², R³, R⁴, R⁵ and R⁶, of R⁷, R⁸, R⁹ and R¹⁰ and/or of Rⁿ, R¹², R¹³ and R¹⁴ form a five- or six-membered aromatic, heteroaromatic, alicyclic or heteroalicyclic ring; and

R¹⁵ and R^lδ are each independently selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, carbonyl and sulfonyl, or, alternatively R¹⁵ and R¹⁶ form a five- or six-member heteroalicyclic ring; whereas if said phosphor and/or said nitrogen is substituted, the substituent is selected from the group consisting of alkyl, cycloalkyl and aryl.

18. The method of claim 17, wherein said compound has the general Formula I.

19. The method of claim 18, wherein Y is selected from the group consisting of hydroxyalkyl and a polyalkylene glycol residue.

20. The method of claim 19, wherein said polyalkylene glycol residue has a general formula III: [O-(CH₂)m]n-OR¹⁷

Formula III wherein: each of m and n is independently an integer of 1-10; and R¹⁷ is hydrogen, alkyl, cycloalkyl or aryl.

21. The method of claim 18, wherein: G is carbon;

K is oxygen; each of R², R³, R⁴, R⁵ and R⁶ is independently selected from the group consisting of hydrogen, alkyl, halo and trihaloalkyl; and each of R⁷, R⁸, R⁹ and R¹⁰ is hydrogen.

22. The method of claim 17, wherein said compound has the general formula II.

23. The method of claim 22, wherein:

Q and W are each substituted or unsubstituted nitrogen; and D is oxygen.

24. The method of claim 22, wherein said Q is a substituted nitrogen.

25. The method of claim 17, wherein said compound is selected from the group consisting of:

10

and pharmaceutically acceptable salts thereof.

26. The method of claim 16, wherein said depressing said cortical neuron activity is for a treatment of a condition or disorder selected from the group consisting of epilepsy, ischemic stroke, migraine, ataxia, myokymia, neurogenic pain, Alzheimer's disease, Parkinson's disease, age-related memory loss, learning deficiencies, bipolar disorder, trigeminal neuralgia, spasticity, mood disorder, psychotic disorder, brain tumor, hearing and vision loss, anxiety and a motor neuron disease.

27. The method of claim 16, wherein said administering is effected intranasally, subcutaneously, intravenously, intramuscularly, parenterally, orally, topically, intradermally, bronchially, buccally, sublingually, supositorially and mucosally.

28. The method of claim 16, wherein said compound forms a part of a pharmaceutical composition which further includes a pharmaceutically acceptable earner.

29. The method of claim 28, wherein said pharmaceutical composition further comprises an agent selected from the group consisting of an anti-bacterial agent, an antioxidant, a buffering agent, a bulking agent, an anti-inflammatory agent, an anti-viral agent, a chemotherapeutic agent and an anti-histamine.

30. A pharmaceutical composition for the treatment or prevention of a condition or disorder in which modulating a voltage-dependent potassium channel and/or depressing a cortical neuron activity is beneficial, the pharmaceutical composition comprising, as an active ingredient, a compound selected from the group consisting of N-phenylanthranilic acid, a N-phenylanthranilic acid derivative, 2- benzimidazolone and 2-benzimidazolone derivative, and a pharmaceutically acceptable carrier.

31. The pharmaceutical composition of claim 30, wherein said compound has the general Formula I or II:

Formula I Formula II

or a pharmaceutically acceptable salt thereof , wherein:

Z is an A-G(=K)-X-Y group, and wherein:

A is alkyl or absent;

K is selected from the group consisting of oxygen and sulfur;

Y is selected from the group consisting of hydrogen, alkyl, hydroxyalkyl, cycloalkyl, aryl and a polyalkylene glycol residue, each of Q and W is independently selected from the group consisting of substituted or unsubstituted nitrogen, oxygen, sulfur and carbon;

D is selected from the group consisting of oxygen and sulfur;

Each of R², R³, R⁴, R⁵, R⁶, R⁷, R⁸, R⁹, R¹⁰, Rⁿ, R¹², R¹³ and R¹⁴ is independently selected from the group consisting of hydrogen, alkyl, hydroxyalkyl, trihaloalkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, heteroalicyclic, halo, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, sulfinyl, sulfonyl, cyano, nitro, amino and -NR¹⁵R¹⁶, or, alternatively, at least two of R², R³, R⁴, R⁵ and R⁶, of R⁷, R⁸, R⁹ and R¹⁰ and/or of R¹¹, R¹², R¹³ and R¹⁴ form a five- or six-membered aromatic, heteroaromatic, alicyclic or heteroalicyclic ring; and R¹⁵ and R¹⁶ are each independently selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, carbonyl and sulfonyl, or, alternatively R¹⁵ and R¹⁶ form a five- or six-member heteroalicyclic ring; whereas if said phosphor and/or said nitrogen is substituted, the substituent is selected from the group consisting of alkyl, cycloalkyl and aryl, the pharmaceutical composition further comprising: a pharmaceutically acceptable carrier.

32. The pharmaceutical composition of claim 31, wherein said compound has said general Formula I.

33. The pharmaceutical composition of claim 32, wherein Y is selected from the group consisting of hydroxyalkyl and a polyalkylene glycol residue.

34. The pharmaceutical composition of claim 33, wherein said polyallcylene glycol residue has a general formula III:

[O-(CH₂)m]n-OR¹⁷ Formula III wherein: each of m and n is independently an integer of 1-10; and R¹⁷ is hydrogen, alkyl, cycloalkyl or aryl.

35. The pharmaceutical composition of claim 32, wherein: G is carbon;

K is oxygen; each of R², R³, R⁴, R⁵ and R⁶ is independently selected from the group consisting of hydrogen, alkyl, halo and trihaloalkyl; and each of R⁷, R⁸, R⁹ and R¹⁰ is hydrogen

36. The pharmaceutical composition of claim 31, wherein said compound has said general formula II.

37. The pharmaceutical composition of claim 36, wherein: Q and W are each substituted or unsubstituted nitrogen; and D is oxygen.

38. The pharmaceutical composition of claim 37, wherein said Q is a substituted nitrogen.

39. The pharmaceutical composition of claim 31, wherein said compound is selected from the group consisting of:

9 10 and pharmaceutically acceptable salts thereof.

40. The pharmaceutical composition of claim 30, wherein said voltage- dependent potassium channel comprises a KCNQ2 channel and/or a KCNQ3 channel and/or a KCNQ2/3 channel.

41. The pharmaceutical composition of claim 30, wherein said condition or disorder is selected from the group consisting of epilepsy, ischemic stroke, migraine, ataxia, myokymia, neurogenic pain, Alzheimer's disease, Parkinson's disease, age- related memory loss, learning deficiencies, bipolar disorder, trigeminal neuralgia, spasticity, mood disorder, psychotic disorder, brain tumor, hearing and vision loss, anxiety and a motor neuron disease.

42. The pharmaceutically composition of claim 30, packaged in a packaging material and identified in print, in or on said packaging material, for use in the treatment or prevention of a condition or disorder associated with altered activity of a voltage-dependent potassium channel and/or a cortical neuron.

43. The pharmaceutical composition of claim 42, wherein said condition or disorder is selected from the group consisting of epilepsy, ischemic stroke, migraine, ataxia, myokymia, neurogenic pain, Alzheimer's disease, Parkinson's disease, age- related memory loss, learning deficiencies, bipolar disorder, trigeminal neuralgia, spasticity, mood disorder, psychotic disorder, brain tumor, hearing and vision loss, anxiety and a motor neuron disease.

44. The pharmaceutical composition of claim 43, further comprising an agent selected from the group consisting of an anti-bacterial agent, an antioxidant, a buffering agent, a bulking agent, an anti-inflammatory agent, an anti-viral agent, a chemotherapeutic agent and an anti-histamine.

45. A compound comprising N-phenylanthranilic acid or a derivative thereof and a hydroxyalkyl residue or a polyalkylene glycol residue covalently attached thereto, or a pharmaceutically acceptable salt thereof.

46. The compound of claim 45, having a general Formula IV:

Formula IV

or a pharmaceutically acceptable salt thereof, wherein:

Z is an A-G(=K)-X-Y group, and wherein:

A is alkyl or absent;

K is selected from the group consisting of oxygen and sulfur;

Y is selected from the group consisting of hydroxyalkyl and a polyallcylene glycol residue;

Each of R², R³, R⁴, R⁵, R⁶, R⁷, R⁸, R⁹ and R¹⁰, is independently selected from the group consisting of hydrogen, alkyl, hydroxyalkyl, trihaloalkyl, cycloalkyl, alkenyl, allcynyl, aryl, heteroaryl, heteroalicyclic, halo, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, sulfinyl, sulfonyl, cyano, nitro, amino and - NR¹⁵R¹⁶, or, alternatively, at least two of R², R³, R⁴, R⁵ and/or R⁶, of R⁷, R⁸, R⁹ and R form a five- or six-membered aromatic, heteroaromatic, alicyclic or heteroalicyclic ring; and

R¹⁵ and R¹⁶ are each independently selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, carbonyl and sulfonyl, or, alternatively R and R form a five- or six-member heteroalicyclic ring; whereas if said phosphor and/or said nitrogen is substituted, the substituent is selected from the group consisting of alkyl, cycloalkyl and aryl.

47. The compound of claim 46, wherein said polyalkylene glycol residue has a general formula V:

[O~(CH₂)m]n-OR¹⁷ Formula V wherein: each of m and n is independently an integer of 1-10; and

17

R is hydrogen, alkyl, cycloalkyl or aryl.

48. The compound of claim 47, wherein: G is carbon;

49. A compound selected from the group consisting of:

4

and pharmaceutically acceptable salts thereof.

50. A pharmaceutical composition comprising, as an active ingredient, the compound of claim 45.

51. A pharmaceutical composition comprising, as an active ingredient, the compound of claim 46.

52. The pharmaceutical composition of claim 51, comprising, as an active ingredient, a compound 3 having the structure:

or a pharmaceutically acceptable salt thereof.

53. The pharmaceutical composition of claim 51, comprising, as an active ingredient, a compound 4 having the stmcture:

or a pharmaceutically acceptable salt thereof.

54. The pharmaceutical composition of claim 51, comprising, as an active ingredient, a compound 5 having the structure:

or a pharmaceutically acceptable salt thereof.

55. The pharmaceutical composition of claim 51, comprising, as an active ingredient, a compound 6 having the stmcture:

or a pharmaceutically acceptable salt thereof.

56. The pharmaceutical composition of claim 51, comprising, as an active ingredient, a compound 7 having the stmcture:

or a pharmaceutically acceptable salt thereof.

57. The pharmaceutical composition of claim 51, comprising, as an active ingredient, a compound 8 having the structure:

or a pharmaceutically acceptable salt thereof.

58. The pharmaceutical composition of claim 51, comprising, as an active ingredient, a compound 9 having the stmcture:

or a pharmaceutically acceptable salt thereof.

59. A method of synthesizing the compound of claim 45, the method comprising: obtaining a N-phenylanthranilic acid or a derivative thereof; reacting said N-phenylanthranilic acid or said derivative thereof with a hydroxyalkyl or a polyallcylene glycol terminating with a reactive group, said reactive group is capable of forming an ester bond with said N-phenylanthranilic acid or said derivative thereof.

60. The method of claim 59, wherein said ester bond is selected from the group consisting of a carboxylic ester bond, a carboxylic amide bond, a carboxylic thioester bond, a S-carboxy ester bond, a S-carboxy thioester bond and a S-carboxy amide bond.

61. The method of claim 59, wherein said reactive group is selected from the group consisting of hydroxy, amine and thiohydroxy.

62. The method of claim 59, wherein said N-phenylanthranilic acid or said derivative thereof has a general Formula VI:

Formula VI wherein,

A is alkyl or absent;

G is selected from the group consisting of carbon, sulfur and substituted or unsubstituted phosphor, whereas if said phosphor is substituted, the substituent is selected from the group consisting of alkyl, cycloalkyl and aryl;

K is selected from the group consisting of oxygen and sulfur;

Each of R², R³, R⁴, R⁵, R⁶, R⁷, R⁸, R⁹ and R¹⁰, is independently selected from the group consisting of hydrogen, alkyl, hydroxyalkyl, trihaloalkyl, cycloalkyl, alkenyl, allcynyl, aryl, heteroaryl, heteroalicyclic, halo, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, sulfinyl, sulfonyl, cyano, nitro, amino and - NR¹⁵R¹⁶, or, alternatively, at least two of R², R³, R⁴, R⁵ and/or R⁶, of R⁷, R⁸, R⁹ and R fomi a five- or six-membered aromatic, heteroaromatic, alicyclic or heteroalicyclic ring; and

R¹⁵ and R¹⁶ are each independently selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, carbonyl and sulfonyl, or, alternatively R¹⁵ and R¹⁶ form a five- or six-member heteroalicyclic ring.

63. The method of claim 62, wherein: G is carbon;

K is oxygen; each of R², R³, R⁴, R⁵ and R^δ is independently selected from the group consisting of hydrogen, alkyl, halo and trihaloalkyl; and each of R⁷, R⁸, R⁹ and R¹⁰ is hydrogen.

64. The method of claim 59, wherein said polyalkylene glycol terminating with said reactive group has a general Formula NIL

N-[O-(CH₂)m]n-OR¹⁷

Foπnula VII wherein: