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US 20100168030A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2010/0168030 A1 Zoltani et al. (43) Pub. Date: Jul. 1, 2010

(54) METHODS AND COMPOSITIONS FOR (52) U.S. c1...... 514/13 TREATMENT OF -CAUSED (57) ABSTRACT PATHOLOGY A method of treating an organophosphate toxin-caused car (76) Inventors: Csaba K. Zoltani, Lutherville, MD diac abnormality is described Which includes administering a (US); Gennady E. Platoff, pharmaceutical composition including a therapeutically Millersville, MD (US); Steven I. effective amount of a chloride current modulator to an indi Baskin, Bel Air, MD (US) vidual subject having a cardiac abnormality caused by intoxi cation With an organophosphate toxin. The chloride current Correspondence Address: modulator is effective to modulate a chloride conductance U S ARMY RESEARCH LABORATORY and thereby reduce a symptom or sign of an organophosphate ATTN: RDRL-LOC-I toxin-caused cardiac abnormality, thus treating the toxin 2800 POWDER MILL RD caused cardiac abnormality. Optionally, included is adminis ADELPHI, MD 20783-1197 (US) tering a therapeutic agent to inhibit an organophosphate toxin-caused distortion of the action potential of myocytes, (21) App1.No.: 11/288,269 support restoration of usual intracellular ionic concentra tions, and support an increase in cardiac contractility. A com (22) Filed: Nov. 29, 2005 position according to the invention is described Which includes a chloride current modulator; and a therapeutic agent Publication Classi?cation to inhibit an organophosphate toxin-induced distortion of the (51) Int. Cl. action potential of myocytes, support restoration of usual A61K 38/16 (2006.01) intracellular ionic concentrations, and support an increase in A61P 9/06 (2006.01) cardiac contractility.

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METHODS AND COMPOSITIONS FOR a simulated electrocardiogram (ECG or EKG) illustrating a TREATMENT OF control trace, A, representative of normal cardiac activity, and ORGANOPHOSPHATE-CAUSED a trace B illustrating differences due to organophosphate. A PATHOLOGY normal T-Wave is shoWn in the simulated control trace, A, at (a). Notable changes include a decrease in amplitude of the GOVERNMENT INTEREST QRS and the modulation of the T-Wave, the repolariZation of the ventricle. In particular, a T-Wave as a separate entity is no [0001] The invention described herein may be manufac tured, used, and licensed by or for the United States Govem longer in evidence, the shape of the QRS portion has been altered and the peak has been shortened. ment. [0009] Treatment of organophosphate intoxication has FIELD OF THE INVENTION focused predominantly on treatment of symptoms and clear ance of organophosphate compounds from the body. Particu [0002] This invention relates to compositions and methods lar treatments include administration of and ani for treatment of organopho sphate poisoning. In particular, the sodamine for treatment of symptoms such as gastrointestinal invention relates to compositions and methods for treatment and/or respiratory distress. Side effects may include arrhyth of cardiac arrhythmias and/or repolariZation abnormalities mia at higher doses. Further, particular including resulting from organophosphate poisoning. , and asoxime may be used in orga nophosphate-intoxicated patients to help restore acetylcho BACKGROUND OF THE INVENTION linesterase activity. [0003] are Widely used as [0010] Thus, there is a continuing need for neW treatments and as threat agents. Accidents in their use are for organophosphate poisoning and for treating conditions common in agriculture and treatment is still rudimentary. relating to organophosphate exposure. In particular, treat Organophosphate-caused ventricular ?brillation results in ments addressing cardiac sequelae of organophosphate expo heart failure and is recogniZed as the most common cause of sure and other types of toxin intoxication are especially mortality in cases of organophosphate intoxication. sought after. [0004] The ?rst organophosphate toxins Were developed early in the 20th century, primarily as pesticides. In the course SUMMARY OF THE INVENTION of research on their pesticidal properties, the potential of these agents for use in chemical Warfare and/or as threat [0011] A method of treating an organophosphate toxin agents Was discovered. induced cardiac abnormality in an individual subject is [0005] Organophosphate toxins bind to acetylcholinest described Which includes administering a pharmaceutical erase, preventing the hydrolysis of and leading composition including a therapeutically effective amount of a to its accumulation and consequent prolongation of its effects chloride current modulator to an individual subject having a at synapses. Symptoms of organophosphate toxin intoxica cardiac abnormality caused by intoxication With an organo tion include rhinorrhea, dyspnea, sWeating, miosis, bradycar phosphate toxin. The chloride current modulator is effective dia, tachycardia, loss of consciousness, convulsions, ?accid to modulate a chloride conductance and thereby reduce a paralysis, and apnea. Histological examination of organo symptom or sign of an organophosphate toxin-induced car phosphate-affected heart tissue shoWs cell sWelling, hemor diac abnormality, thus treating the toxin-induced cardiac rhaging and necrosis. abnormality. [0006] One of the most serious consequences of organo [0012] In a preferred option, the chloride current modulator phosphate intoxication is due to effects on the cardiovascular is a modulator of ICZ, We”. Modulators of chloride currents system. Organophosphate-caused heart failure is a syndrome include: a disulfonic stilbene; an arylaminobenZoate; a characterized primarily by left ventricular dysfunction, fenamate; an anthracene carboxylate; an indanylalkanoic reduced exercise tolerance, impaired quality of life and dra acid; a clo?bric acid; a clo?bric acid derivative; a sulfony matically shortened life expectancy. Decreased contractility lurea; a calixarene; suramin; and tamoxifen. Further pre of the left ventricle leads to reduced cardiac output With the ferred are modulators of ICZ, We” such as 4,4'-diisothiocyanos consequent systemic arterial and venous constriction. tilbene-2,2'-disulfonic acid (DIDS); 4,4'-dinitrostilbene-2,2' [0007] Cardiovascular effects of organophosphate intoxi disulfonic acid (DNDS); 4-acetamindo-4' cation are apparent When analyZed by electrocardiogram. The isothiocyanostilbene-2,2'-disulfonic acid (SITS); tamoxifen; electrocardiogram of an affected individual shoWs changes in 5-nitro-2-(3-phenylpropylamino)benZoic acid (NPPB); the P-Wave, the depolarization of the atria, and also prominent ni?umic acid (N FA); ?ufenamic acid; anthracene-9-carboxy modulation of the T-Wave, the repolariZation of the ventricles. late (9AC); diphenylaminecarboxylate (DPC); 2-(p-chlo After an initial bradycardia, Torsade de Pointes and tachycar rophenoxy)propionic acid (CPP); and indanyloxyacetic acid dia develop and the electrophysiology of the heart loses its (IAA-94). Mixtures of these modulators may also be admin predictability. In addition to these changes, the fast outWard istered. rectifying potassium current is blocked. The ensuing over [0013] Optionally, an inventive method further includes stimulation results in ion concentration imbalance and mem administering a therapeutic agent to inhibit organopho sphate brane current derangement. On the electrocardiogram, this is induced distortion of the action potential of myocytes, sup expressed by QT prolongation, ST and T Wave abnormalities. port restoration of usual intracellular ionic concentrations, Organophosphate-caused ventricular ?brillation culminates and support an increase in cardiac contractility. Such thera in heart failure and is recogniZed as the most common cause peutic agents include a second current modulator; an antiar of mortality in cases of organophosphate intoxication. rhythmic drug; a modulator of a mitochondrial membrane ion [0008] An example of effects of organophosphate com channel, ion pump and/or ion exchanger; inhibitor of protein pounds on cardiac tissue is shoWn in FIG. 1 Which illustrates kinase C; an anticonvulsant; an organophosphate clearing US 2010/0168030 A1 Jul. 1,2010

agent; an inhibitor of a muscarinic potassium channel; and an rophenyl)ethenyl diethyl phosphate), acetylcholine receptor antagonist; and combinations thereof. (2-chloro- l -(2,4-dichlorophenyl)ethenyl diethyl phosphate), [0014] A composition according to the invention is crotoxyphos (l -phenylethyl3 -(dimethoxypho sphinoyloxy) described Which includes a chloride current modulator; and a isocrotonate), (2,2-dichlorovinyl dimethyl phos therapeutic agent to inhibit organopho sphate-induced distor phate), ((E)-2-dimethylcarbamoyl-l-methylvi tion of the action potential of myocytes, support restoration of nyl dimethyl phosphate or 3-dimethoxyphosphinoyloxy-N, usual intracellular ionic concentrations, and support an N-dimethylisocrotonamide), dimethylvinphos ((Z)-2 increase in cardiac contractility. In a preferred option, the chloro-l-(2,4-dichlorophenyl)vinyl dimethyl phosphate), chloride current modulator is a modulator of ICZ, We”. Further fospirate (dimethyl 3,5,6-trichloro-2-pyridyl phosphate), optionally, the therapeutic agent included in an inventive heptenophos(7-chlorobicyclo [3 .2 .0] hepta-2,6-dien-6-yl composition is selected from the group consisting of: a sec dimethyl phosphate), methocrotophos ((E)-2-(N-methoxy ond current modulator, an antiarrhythmic drug, an acetylcho N-methylcarbamoyl)-l-methylvinyl dimethyl phosphate) or line receptor antagonist, an anticonvulsant, an organophos (3-dimethoxyphosphinoyloxy-N-methoxy-N-methylisocro phate clearing agent, an inhibitor of protein kinase C, a tonamide), ((EZ)-2-methoxycarbonyl-l-meth modulator of a mitochondrial membrane moiety, and a com ylvinyl dimethyl phosphate), (dimethyl (E) bination thereof. l-methyl-2-(methylcarbamoyl)vinyl phosphate), naftalofos (diethyl naphthalimido-oxyphosphonate), BRIEF DESCRIPTION OF THE DRAWINGS ((EZ)-2-chloro-2-diethylcarbamoyl- l -methylvinyl dimethyl [0015] FIG. 1 shoWs a simulated electrocardiogram (ECG phosphate), propaphos (4-(methylthio)phenyl dipropyl phos or EKG) illustrating a control trace, A, representative of nor phate), TEPP (), and tetrachlorvin mal cardiac activity, and a trace B illustrating differences due phos ((Z)-2-chloro-l-(2,4,5-trichlorophenyl)vinyl dimethyl to organophosphate; and phosphate); pesticides include: dioxa [0016] FIG. 2 is a graph shoWing simulated action poten benZofos ((RS)-2-methoxy-4H-l,3,27t5-benZodioxaphos tials and changes thereof in various states relating to organo phinine 2-sul?de), fosmethilan (Si[N-(2-chlorophenyl)bu phosphate intoxication. tyramidomethyl]0,0-dimethyl phosphorodithioate), and (S-ot-ethoxycarbonylbenzyl 0,0-dimethyl phos DETAILED DESCRIPTION OF THE PREFERRED phorodithioate); aliphatic organothiophosphate pesticides EMBODIMENTS such as: acethion (S-(ethoxycarbonylmethyl) 0,0-diethyl phosphorodithioate), amiton (S-2-diethylaminoethyl 0,0-di [0017] A method of treating a toxin-induced cardiac abnor ethyl phosphorothioate), cadusafos (S,S-di-sec-butyl O-ethyl mality in an individual subject is provided Which includes phosphorodithioate), (0,0-diethyl(RS)iO administering a pharmaceutical composition containing a (l,2,2,2-tetrachloroethyl)phosphorothioate), chlormephos therapeutically effective amount of an ion channel modulator (S-chloromethyl 0,0-diethyl phosphorodithioate), deme to an individual subject having a cardiac abnormality caused phion (reaction mixture of 0,0-dimethyl O-2-methylthioet by intoxication With a toxin. The ion channel modulator is hyl phosphorothioate and 0,0-dimethyl S-2-methylthioethyl effective to modulate an ion channel conductance, thereby phosphorothioate), (reaction mixture of 0,0-di reducing a symptom or sign of a toxin-induced cardiac abnor ethyl O-2-ethylthioethyl phosphorothioate and 0,0-diethyl mality and treating the toxin-induced cardiac abnormality. S-2-ethylthioethyl phosphorothioate), (0,0-di [0018] In a particular embodiment, an inventive method is ethyl S-2-ethyl thioethyl phosphorodithioate), (0,0, a method of treating an organophosphate toxin induced car O',O'-tetraethyl S,S'-methylene bis(phosphorodithioate)), diac abnormality. An organophosphate toxin is an organo (O-ethyl S,S-dipropyl phosphorodithioate), phosphate compound Which is a pesticide and/or chemical IPSP (S-ethylsul?nylmethyl 0,0-di-isopropyl phospho Warfare agent having inhibitor activity. rodithioate), isothioate (S-2-isopropylthioethyl 0,0-dim Acetylcholinesterase inhibitor activity may be assayed by any ethyl phosphorodithioate), (diethyl (dimethox of various methods knoWn in the art. For example, an Ellman ythiophosphorylthio)succinate), methacrifos (methyl (E)-3 assay may be used to detect acetylcholinesterase inhibitor (dimethoxyphosphinothioyloxy)-2-methylacrylate), activity of an organophosphate pesticide or toxin. oxydemeton (methyl (S-2-ethylsul?nylethyl 0,0-dimethyl Brie?y described such an assay involves addition of human phosphorothioate), oxydeprofos ((RS)-S-2-ethylsul?nyl-l acetylcholinesterase in an appropriate buffer, such as 100 mM methylethyl 0,0-dimethyl phosphorothioate), oxydisulfoton sodium phosphate buffer, pH 7.4, to an organophosphate (0,0-diethyl S-2-ethylsul?nylethyl phosphorodithioate), pesticide or nerve agent toxin of interest. A substrate for (0,0-diethyl S-ethylthiomethyl phosphorodithio acetylcholinesterase such as acetylthiocholine is added to the ate), (0,0,0',O'-tetraethyl dithiopyrophosphate), mixture along With Ellman’s reagent 5'5-dithio-bis-(2-ni (S-tert-butylthiomethyl 0,0-diethyl phospho trobenZoate). After a brief incubation, optical density is mea rodithioate), and thiometon, (S-2-ethylthioethyl 0,0-dim sured at 415 nm Where optical density is inversely propor ethyl phosphorodithioate); aliphatic amide organothiophos tional to the inhibiting activity of the organophosphate phate pesticides including: amidithion (S-2 pesticide or nerve agent toxin (Ellman, G. L. et al., Biochemi methoxyethylcarbamoylmethyl 0,0-dimethyl cal , 7: 88-95, 1961 and Wilson, B. W. et al., pho sphorodithioate), cyanthoate (Si[N-(l -cyano- l -meth International Journal of Toxicology, 21: 385-388, 2002). ylethyl)carbamoylmethyl]0,0-diethyl phosphorothioate), [0019] Many organophosphate toxins are knoWn and are (0,0-dimethyl S-methylcarbamoylmethyl phos typically esters, amides, or thiol derivatives of phosphoric, phorodithioate), ethoate-methyl (S-ethylcarbamoylmethyl phosphonic, phosphorothioic, or phosphonothioic acids. 0,0-dimethyl phosphorodithioate), (S-formyl [0020] Organophosphate toxins Which are pesticides illus (methyl)carbamoylmethyl]0,0-dimethyl phosphorodithio tratively include bromfenvinfos (2-bromo-l-(2,4-dichlo ate), mecarbam (S-(N-ethoxycarbonyl-N-methylcarbamoyl US 2010/0168030 A1 Jul. 1,2010

methyl) 0,0-diethyl phosphorodithioate), (0,0 methylpyrimidin-4-yl 0,0-dimethyl phosphorothioate), dimethyl S-methylcarbamoylmethyl phosphorothioate), primidophos (0,0-diethyl O-2-N-ethylacetamido-6-meth , 0,0-diethyl S-isopropylcarbamoylmethyl phos ylpyrimidin-4-yl phosphorothioate), pyrimitate (O-Z-dim phorodithioate), sophamide, S-methoxymethylcarbamoylm ethylamino-6-methylpyrimidin-4-yl 0,0-diethyl phospho ethyl 0,0-dimethyl phosphorodithioate), and Vamidothion rothioate), and ((RS)iO-(2-tert (0,0-dimethyl Si(RS)-2-(l-methylcarbamoylethylthio) butylpyrimidin-S-yl) O-ethyl O-isopropyl ethyl phosphorothioate); organothiophosphate pesti phosphorothioate); quinoxaline organothiophosphate pesti cides illustratively include: chlorphoxim (2-(2-chlorophe cides illustratively including: (0,0-diethyl nyl)-2-(diethoxyphosphinothioyloxyimino)acetonitrile, O-quinoXalin-Z-yl phosphorothioate) and quinalphos-methyl (0,0-diethyl 0t-cyanobenZylideneamino-oXy (0,0-dimethyl O-quinoXalin-Z-yl phosphorothioate); thia phosphonothioate), and phoXim-methyl (0,0-dimethyl diaZole organothiophosphate pesticides such as: athidathion 0t-cyanobenZylideneamino-oXyphosphonothioate); hetero (0,0-diethyl S-2,3 -dihydro-5 -methoxy-2-oxo-1,3,4-thiadia cyclic organothiophosphate pesticides such as: Zol-3-ylmethyl phosphorodithioate), lythidathion (S-5 (S-6-chloro-2,3-dihydro-2-oxo-l,3-oXaZolo[4,5-b]pyridin ethoXy-2,3-dihydro-2-oxo- l ,3,4-thiadiaZol-3 -yhnethyl 0,0 3-ylmethyl 0,0-dimethyl phosphorothioate), dimethyl phosphorodithioate), (S-2,3-dihydro (O-3-chloro-4-methyl-2-oxo-2H-chromen-7-yl 0,0-diethyl 5 -methoxy-2-oXo-l ,3 ,4-thiadiaZol-3-ylmethyl 0,0 phosphorothioate), coumithoate (0,0-diethyl O-(7,8,9,l0 dimethyl phosphorodithioate) or tetrahydro-6-oxo-6H-benZo[c]chromen-3-yl)phospho (3 -dimethoxypho sphinothioylthiomethyl -5 -methoxy- l ,3 , 4 rothioate), (S,S'-(l,4-dioXane-2,3-diyl) 0,0,0', thiadiaZol-2(3H)-one), and prothidathion (S-2,3-dihydro-5 O'-tetraethyl bis(phosphorodithioate)), (S-5 isopropoxy-2-oxo-1,3,4-thiadiazol-3 -ylmethyl 0,0-diethyl methoxy-4-oxo-4H-pyran-2-ylmethyl 0,0-dimethyl phosphorodithioate); triaZole organothiophosphate pesti phosphorothioate), menaZon (S-4,6-diamino- l ,3,5-triaZin-2 cides such as: isaZofos (O-S-chloro-l-isopropyl-lH-l,2,4 ylmethyl 0,0-dimethyl phosphorodithioate), morphothion triaZol-3-yl 0,0-diethyl phosphorothioate) and triaZophos (0,0-dimethyl S-morpholinocarbonylmethyl phospho (0,0-diethyl O-l-phenyl-lH-l,2,4-triaZol-3-yl phospho rodithioate), (S-6-chloro-2,3-dihydro-2-oxoben rothioate); phenyl organothiophosphate pesticides including: ZoXaZol-3-ylmethyl 0,0-diethyl phosphorodithioate), pyra aZothoate (O-4-(4-chlorophenylaZo)phenyl 0,0-dimethyl clofos ((RS)-O-l-(4-chlorophenyl)pyraZol-4-yl O-ethyl phosphorothioate), bromophos (O-4-bromo-2,5-dichlo S-propyl phosphorothioate), pyridaphenthion (O-(l,6-dihy rophenyl 0,0-dimethyl pho sphorothioate), bromophos-ethyl dro-6-oxo-l-phenylpyridaZin-3-yl) 0,0-diethyl phospho (O-4-bromo-2,5-dichlorophenyl 0,0-diethyl phosphorothio rothioate), and quinothion (0,0-diethyl O-Z-methylquinol ate), (S-4-chlorophenylthiomethyl 0,0-di 4-yl phosphorothioate); benZothiopyran ethyl phosphorodithioate), chlorthiophos (isomeric reaction organothiophosphate pesticides include: dithicrofos (S-(6 mixture in Which O-2,5-dichlorophenyl-4-methylthiophenyl chloro -3 ,4-dihydro -2H-l -benZothi-in-4 -yl) 0,0-diethyl 0,0-diethyl phosphorothioate predominates), phosphorodithioate) and thicrofos (S-(6-chloro-3,4-dihydro (O-4-cyanophenyl 0,0-dimethyl phosphorothioate), cythio 2H-l -benZothi-in-4-yl) 0,0-diethyl phosphorothioate); ben ate (0,0-dimethyl O-4-sulfamoylphenyl phosphorothioate), ZotriaZine organothiophosphate pesticides such as: aZinphos dicapthon (O-2-chloro-4-nitrophenyl 0,0-dimethyl phos ethyl (S-(3 ,4-dihydro -4 -oXobenZo [d] - [l ,2,3] -triaZin-3 - phorothioate), dichlofenthion, O-2,4-dichlorophenyl 0,0-di ylmethyl) 0,0-diethyl phosphorodithioate) and aZinphos ethyl phosphorothioate), etaphos ((RS)4O-2,4-dichlorophe methyl (S-(3 ,4-dihydro -4 -oXobenZo [d] - [l ,2,3] -triaZin-3 - nyl O-ethyl S-propyl phosphorothioate), famphur (O-4 ylmethyl) 0,0-dimethyl phosphorodithioate); isoindole dimethylsulfamoylphenyl 0,0-dimethyl phosphorothioate), organothiophosphate pesticides include: dialifos, ((RS)iS fenchlorphos (0,0-dimethyl O-2,4,5-trichlorophenyl phos 2-chloro-l-phthalimidoethyl 0,0-diethyl phosphorodithio phorothioate), (0,0-dimethyl O-4-nitro-m-tolyl ate) and (0,0-dimethyl S-phthalimidomethyl phos phosphorothioate), fensulfothion (0,0-diethyl O-4-methyl phorodithioate); isoXaZole organothiophosphate pesticides sul?nylphenyl phosphorothioate), (0,0-dimethyl such as: (0,0-diethyl O-5-phenyl-1,2-oxazol-3-yl O-4-methylthio-m-tolyl phosphorothioate), fenthion-ethyl phosphorothioate) and Zolaprofos ((RS)-O-ethyl S-3 -methyl (0,0-diethyl O-4-methylthio-m-tolyl phosphorothioate), 1,2-oxazol-5-ylmethyl S-propyl phosphorodithioate); pyra heterophos ((RS)iO-ethyl O-phenyl S-propyl phospho Zolopyrimidine organothiophosphate pesticides such as: rothioate), jodfenphos (O-2,5-dichloro-4-iodophenyl 0,0 chlorpraZophos (O-(3-chloro-7-methylpyraZolo[ l , 5-a]pyri dimethyl phosphorothioate), mesulfenfos (0,0-dimethyl midin-2-yl) 0,0-diethyl phosphorothioate) and pyraZophos O-4-methylsul?nyl-m-tolyl phosphorothioate), (ethyl Z-diethoxypho sphinothioyloxy-S -methylpyraZolo [l , (0,0-diethyl O-4-nitrophenyl phosphorothioate), parathion 5-a]pyrimidine-6-carboxylate); pyridine organothiophos methyl (0,0-dimethyl O-4-nitrophenyl phosphorothioate), phate pesticides including: (0,0-diethyl O-3,5, phenkapton (S-2,5-dichlorophenylthiomethyl 0,0-diethyl 6-trichloro-2-pyridyl phosphorothioate) and chlorpyrifos phosphorodithioate), phosnichlor (O-4-chloro-3 -nitrophenyl methyl (0,0-dimethyl O-3,5,6-trichloro-2-pyridyl 0,0-dimethyl phosphorothioate), ((RS)iO-4 phosphorothioate); pyrimidine organothiophosphate pesti bromo-2-chlorophenyl O-ethyl S-propyl phosphorothioate), cides such as: butathiofos (O-2-tert-butylpyrimidin-5-yl prothiofos ((RS)4O-2,4-dichlorophenyl O-ethyl S-propyl 0,0-diethyl phosphorothioate), (0,0-diethyl O-2 phosphorodithioate), sulprofos ((RS)4O-ethyl O-4-(meth isopropyl-6-methylpyrimidin-4-yl phosphorothioate), etrim ylthio)phenyl S-propyl phosphorodithioate), temephos (0,0, fos (O-6-ethoxy-2-ethylpyrimidin-4-yl 0,0-dimethyl phos O',O'-tetramethyl 0,0'-thiodi-p-phenylene bis(phospho phorothioate), lirimfos (O-6-ethoxy-2-isopropylpyrimidin rothioate)), trichlormetaphos-3 ((RS)4O-ethyl O-methyl 4-yl 0,0-dimethyl phosphorothioate), pirimiphos-ethyl O-2,4,5-trichlorophenyl phosphorothioate), and trifenofos (O-2-diethylamino-6-methylpyrimidin-4-yl 0,0-diethyl ((RS)iO-ethyl S-propyl O-2,4,6-trichlorophenyl phospho phosphorothioate), pirimiphos-methyl (O-2-diethylamino-6 rothioate); phosphonate pesticides such as: butonate ((RS)-2, US 2010/0168030 A1 Jul. 1,2010

2,2-trichloro-l-(dimethoxyphosphinoyl)ethyl butyrate) and Torsade de Pointes and tachycardia. Such signs and symp trichlorfon (dimethyl (RS)-2,2,2-trichloro-l-hydroxyeth toms are monitored by methods typically used to monitor ylphosphonate); phosphonothioate pesticides such as: mecar cardiac parameters, such as ECG. Further symptoms and phon (methyl (RS) [methoxy(methyl) phosphinothioylthio] signs are described herein. In some cases, symptoms and acetyl (methyl) ); phenyl ethylphosphonothioate signs associated With intoxication are not speci?cally associ pesticides including: ((RS)iO-ethyl S-phenyl eth ated With toxin exposure. In such cases, intoxication may be ylphosphonodithioate) and trichloronat ((RS)4O-ethyl O-2, con?rmed by tests for presence of the toxin, a metabolite of 4,5-trichlorophenyl ethylphosphonothioate); phenyl phe the toxin, a degradation product of the toxin and/ or a personal nylphosphonothioate pesticides such as: cyanofenphos history indicative of likelihood of exposure to the toxin. ((RS)iO-4-cyanophenyl O-ethyl phenylphosphonothioate), [0023] An inventive method includes treatment of disor EPN ((RS)4O-ethyl O-4-nitrophenyl phenylphosphono ders associated With organophosphate toxicity, particularly thioate) and ((RS)4O-4-bromo-2,5-dichlorophe nyl O-methyl phenylphosphonothioate); phosphoramidate cardiac abnormalities such as heart failure caused by ven pesticides including: crufomate ((RS)-4-tert-butyl-2-chlo tricular ?brillation due to organophosphate-toxicity and car rophenyl methyl methylphosphoramidate), diogenic shock due to organophosphate-toxicity. ((RS)-ethyl 4-methylthio-m-tolyl isopropylphosphorami [0024] One characteristic of organophosphate intoxication date), fosthietan (diethyl l,3-dithietan-2-ylidenephosphora is distortion of ion gradients across excitable membrane cell midate), mephosfolan (diethyl 4-methyl-l,3-dithiolan-2 membranes, particularly cardiac cell membranes. Thus, an ylidenephosphoramidate), (diethyl 1,3-dithiolan inventive method includes administering a compound to 2-ylidenephosphoramidate), and pirimetaphos ((RS)-2 modulate organophosphate-induced activation and/or inhibi diethylamino-6-methylpyrimidin-4-yl methyl tion of one or more cardiac cell transmembrane ion currents, methylphosphoramidate); phosphoramidothioate pesticides so as to inhibit organophosphate-induced distortion of the such as: ((RS)4O,S-dimethyl acetylphosphorami action potential of myocytes, support restoration of usual dothioate), isocarbophos ((RS)4O-2-isopropoxycarbon intracellular ionic concentrations, and support an increase in ylphenyl O-methyl phosphoramidothioate), isofenphos cardiac contractility. ((RS)iO-ethyl O-2-isopropoxycarbonylphenyl isopropy [0025] The present invention provides methods for modu lphosphoramidothioate), ((RS)4O,S-dim lation of selected membrane currents to aid in inhibiting ethyl phosphoramidothioate), and propetamphos ((RS)-(E) organopho sphate-induced distortion of the action potential of O-2-isopropoxycarbonyl- l -methylvinyl O-methyl myocytes, support restoration of usual intracellular ionic con ethylphosphoramidothioate); phosphorodiamide pesticides centrations, and support an increase in cardiac contractility. It including: (tetramethylphosphorodiamidic ?uo has been found by the present inventors that modeling inhi ride), , N,N'-di-isopropylphosphorodiamidic ?uo bition of an organophosphate-induced activation of a mem ride), and (octamethylpyropho sphoric tetra-amide). brane current Which is typically sWelling-activated has such [0021] Chemical Warfare organophosphates are commonly bene?cial effects on modeled cell parameters. called “organophosphate nerve agents” and include those [0026] In a particular example, modeled inhibition of a exempli?ed in Ellison, D. H., Handbook of Chemical and transmembrane chloride current activated in an organophos Biological Warfare Agents, 1999, CRC Press. Speci?c phate exposed cell or tissue aids in reestablishing character examples of organophosphate nerve agents include phos istics of normal cell and tissue functionality, for instance by phoramidocyanidic acid, dimethyl-, ethyl ester also knoWn as inhibiting organophosphate-induced distortion of the action GA and ; phosphono?uoridic acid, methyl-, l-methyl potential of myocytes, supporting restoration of usual intra ethyl ester also knoWn as GB and ; phosphono?uoridic cellular ionic concentrations, and supporting an increase in acid, methyl-, 1,2,2-trimethylpropyl ester also knoWn as GD cardiac contractility. FIG. 2 illustrates a model of organo and ; phosphono?uoridic acid, ethyl-, l-methylethyl phosphate intoxication of cardiac cells and the effect of ter ester also knoWn as GE; phosphono?uoridic acid, methyl-, minating the cell membrane chloride current. A normal action cyclohexyl ester also knoWn as GF and ; phos potential typical of cardiac tissue unaffected by organophos phoramido?uoridic acid, dimethyl-, 2-(dimethylamino)ethyl phate is shoWn in the rightmost trace (A) representing a ester also knoWn as GV and GP; phosphonothioic acid, ethyl-, control. The leftmost trace (B) shoWs a model of an action S-[2-(diethylamino)ethyl]O-ethyl ester also knoWn as VE; potential of an organophosphate affected cardiac ventricular phosphorothioic acid, S-[2-(diethylamino)ethyl]0,0-diethyl cell. Note the shortened cycle length, the raised equilibrium ester also knoWn as VG and Amiton; phosphonothioic acid, potential and the reduced amplitude. Restoring the usual methyl-, S-[2-(diethylamino)ethyl]O-ethyl ester also knoWn potassium ion concentrations across the cell membrane helps, as VM; phosphonothioic acid, methyl-, S-[2-(diethylamino) as shoWn in (C), but the cycle length, a measure of the tachy ethyl]O-(2-methylpropyl) ester also knoWn as VRm RVX, cardia, is still less than half of the control condition. Antago and Russian VX; phosphonothioic acid, ethyl-, S-[2-[bis(l niZing the organophosphate-caused chloride membrane cur methylethyl)amino]ethyl]O-ethyl ester also knoWn as VS; rent, as shoWn in the trace (D) restores the normal action and phosphonothioic acid, methyl-, S-[2-[bis(l -methylethyl) potential, indicating restored membrane potential and con amino]ethyl]O-ethyl ester also knoWn as VX. tractility. [0022] A therapeutically effective amount is that amount [0027] Thus, compounds administered in a method accord Which decreases a symptom or sign of organophosphate ing to the invention include those that inhibit usually inactive induced cardiac abnormalities. Exemplary symptoms and membrane currents activated folloWing exposure of tissue to signs include arrhythmia, ?brillation, abnormal ECG readout organophosphate. such as changes in the P-Wave, the depolarization of the atria, [0028] In one embodiment, an inventive method of treating and prominent modulation of the T-Wave, rising resting organophosphate poisoning in an individual subject includes potential, Brugada-like symptoms, an initial bradycardia, administering a pharmaceutical composition Which contains US 2010/0168030 A1 Jul. 1,2010

a therapeutically effective amount of a chloride channel Mechanism of action of novel cardiotonic agents, J. Cardio modulator to an individual subject in need thereof. vascular Pharmacology, 2002, 40:323-338. [0029] In a particular embodiment, the chloride channel [0036] Organophosphate poisoning is a toxicity caused by modulator is an antagonist of a chloride current activated in a multiple effects of the organophosphate compounds on meta cell exposed to organopho sphate. In a preferred embodiment, bolic processes. Thus, cardiac cells are affected by organo the chloride current modulator is a modulator of a chloride phosphate via several pathWays and organophosphate-related current knoWn as “ICLSWEH.” ICLSWEH, also knoWn as “ICLVOZ,” is cardiac pathology appears to be the result of such multiple a cell volume regulated chloride current present in cardiac effects. Some of the toxic effects of organophosphate occur cells. See, for example, Hume, J. R. et al., Physiol. RevieWs, by mechanisms such as the af?nity of organophosphate com 80:31-81, 2000 and Lang, F. et al., Physiol. RevieWs, 78:247 pounds for acetylcholinesterase. Effects of acetylcholinest 306, 1998. erase inhibition include stimulation of the aforementioned [0030] Chloride current antagonists illustratively include “ICLSWZH” chloride conductance. In addition, organophos disulfonic stilbenes, arylaminobenZoates, fenamates, phate toxins block the fast outWard rectifying potassium cur anthracene carboxylates, indanylalkanoic acids, clo?bric rent With the subsidiary effect upon energy production at the acid, clo?bric acid derivatives, sulfonylureas, calixarenes, cellular level. suramin, and tamoxifen. [0037] A conductance modulator is optionally delivered in [0031] Particular chloride conductance modulators admin conjunction With a therapeutic agent in addition to a conduc istered in an embodiment of an inventive method include tance modulator in one embodiment. A therapeutic agent may inhibitors of an “ICLSWEH” chloride current. For example, a be included in a pharmaceutical composition With a conduc preferred disulfonic stilbene inhibitor of an “ICLSWZH” chlo tance modulator and/ or administered separately. ride current included in a method and composition according [0038] A composition including a chloride current modu to the invention is 4,4'-diisothiocyanostilbene-2,2'-disulfonic lator, particularly an inhibitor of ICLSWE,ZZ and a therapeutic acid (DIDS). In another embodiment, disulfonic stilbenes agent, is administered to inhibit organophosphate-induced included in a method and composition according to the inven distortion of the action potential of myocytes, support resto tion illustratively include 4,4'-dinitrostilbene-2,2'-disulfonic ration of usual intracellular ionic concentrations, and support acid (DNDS) and 4-acetamindo-4'-isothiocyanostilbene-2, an increase in cardiac contractility. Exemplary therapeutic 2'-disulfonic acid (SITS). agents illustratively include a second current modulator, an [0032] In another embodiment, an “ICLSWZH” chloride con antiarrhythmic drug, an acetylcholine receptor antagonist, an ductance inhibitor included in an inventive method and com anticonvulsant, an organopho sphate clearing agent, an inhibi position is tamoxifen. In further embodiments, an “ICLSWZH” tor of protein kinase C, a modulator of a mitochondrial mem chloride conductance inhibitor included in an inventive brane moiety, and a combination thereof. Combinations of a method and composition is illustratively 5-nitro-2-(3 -phenyl chloride current modulator, particularly an inhibitor of ICZ, propylamino) benZoic acid (NPPB); ni?umic acid (NFA); swell and a therapeutic agent may have a synergistic effect in ?ufenamic acid; anthracene-9-carboxylate (9AC); dipheny inhibiting organophosphate-induced distortion of the action laminecarboxylate (DPC); 2-(p-chlorophenoxy)propionic potential of myocytes, supporting restoration of usual intra acid (CPPia clo?bric acid derivative); and indanyloxyacetic cellular ionic concentrations, and supporting an increase in acid (IAA-94). cardiac contractility. In particular, lesser amounts of a chlo [0033] An inhibitor of “ICLSWEH” chloride current may be ride current modulator may be necessary to achieve reduction identi?ed using methods knoWn in the art. For example, one in symptoms and signs of cardiac abnormality When the or more cells placed in a recording chamber may be exposed modulator is included in a composition With a therapeutic to a stimulus causing a change in cell volume. A chloride agent. Such a composition may therefore provide bene?ts of current activated in response to the change in cell volume may treatment of toxin-induced cardiac abnormality as Well as be detected by methods such as Whole cell or patch clamp cost savings and reduction in side effects. Further, lesser techniques. An inhibitor of “ICLSWZH” chloride current may be amounts of a therapeutic agent may be administered When identi?ed as an agent that causes a speci?c decrease in “ICZ, included in an inventive composition. Swen” chloride current. [0039] An exemplary therapeutic agent suitable in this [0034] A chloride conductance inhibitor included in a com regard is an organophosphate clearing agent. The term “orga position and method according to the invention preferably nophosphate clearing agent” as used herein is intended to produces at least about a 50% decrease in an organophosphate mean a compound having a bene?cial effect on an organo exposure-induced membrane chloride current. Further, a phosphate-poisoned individual, particularly an effect of chloride conductance inhibitor included in a composition and inhibiting the action of organophosphate in the affected indi method according to the invention preferably induces at least vidual such as by stimulating removal, sequestration, or about a 50% increase in cardiac action potential duration in an of the organophosphate and/ or regenerating ace organophosphate-intoxicated substrate, that is, the tissue tylcholinesterase activity such that the toxic effects are inhib being acted on. ited. [0035] One consequence of organophosphate intoxication [0040] An organophosphate clearing agent includes an is deleterious accumulation of intracellular calcium. A chlo oxime agent such as obidoxime, pralidoxime and asoxime, ride conductance inhibitor included in a composition and and salts or other derivatives thereof exempli?ed by asoxime method according to the invention causes at least about a chloride, pralidoxime chloride, pralidoxime methane 1-12% decrease in intracellular calcium concentration and sulfonate, knoWn as the mesilate, and pralidoxime methyl preferably causes at least about a 2-6% decrease in. intracel sulphate. lular calcium concentration. Such a decrease in intracellular [0041] Exemplary dosing and administration of organo calcium concentration may be measured in a standard in vitro phosphate clearing agents includes intravenous administra calcium sensitiZing assay, such as that detailed in M. Endoh, tion of pralidoxime mesilate in amounts of about 20-50 US 2010/0168030 A1 Jul. 1,2010

mg/kg body Weight every 4-6 hours, usually for about 3 days bition of organophosphate-induced distortion of the action or less. Pralidoxime chloride may be administered intrave potential of myocytes, restoration of usual intracellular ionic nously in amounts of about 1 g every 4-6 hours. Effective concentrations, and an increase in cardiac contractility. Anti levels in blood serum may be in the range of about 4-15 arrhythmic drugs illustratively include class I antiarrhyth micrograms/milliliter. Pralidoxime iodide may be adminis mics Which are sodium channel blockers such as class IA tered intravenously in amounts of about 20-50 milligrams/kg antiarrhythmics illustratively including quinidine (Quini body Weight intermittently to alleviate symptoms. Obi dex), procainamide (Pronestyl) and disopyramide (Norpace); doxime may be given in amounts ranging from about 2-4 class IB antiarrhythmics illustratively including lidocaine mg/kg body Weight intravenously. may be @(ylocaine), tocainide (Tonocard), and mexiletine (Mexitil); given as about 250-1000 milligrams diluted in 1-5 milliliters class IC antiarrhythmics illustratively including encainide of distilled Water intramuscularly 4 times/day for 2-7 days. (Enkaid), and ?ecainide (Tambocor); class II antiarrhythmics One of skill in the art Will knoW hoW to adjust such doses in Which are beta blockers illustratively including propranolol order to meet the demands of a particular therapeutic situa (Inderal), acebutolol (Sectral), esmolol (Brevibloc), meto tion. prolol, and atenolol; class III antiarrhythmics Which are [0042] In a further example of a therapeutic agent, a second potassium channel blockers illustratively including sotalol conductance modulator is optionally administered in order to (Betapace), dofetilide and amiodarone (Cordarone) and class inhibit organophosphate-induced distortion of the action IV antiarrhythmics Which are calcium channel blockers illus potential of myocytes, support restoration of usual intracel tratively including verapamil (Calan, Isoptin), diltiaZem lular ionic concentrations, and support an increase in cardiac (CardiZem) and mebefradil (Posicor). In addition, some anti contractility. For example, more than one modulator of a arrhythmics such as alinidine may act as chloride channel chloride current may be administered. The multiple chloride blockers, see Millar J S and Williams E M., Pacemaker selec current modulators optionally modulate the same or different tivity: in?uence on rabbit atria of ionic environment and of chloride channels. In another example, such a compound is a alinidine, a possible anion antagonist. Cardiovasc. Res. 1981, sodium channel opener such as aconitine, and/or an agent 15(6):335-50. Digoxin is a further antiarrhythmic optionally such as veratridine Which sloWs the sodium current inactiva administered. Dosage of such agents is knoWn in the art and is tion, (Wang, G K and Wang S Y, Veratridine block of rat illustrated in standard texts such as R. N. Fogoros, Antiar skeletal muscle Nav1.4 sodium channels in the inner vesti rhythmic Drugs: A Practical Guide, BlackWell Publishers, bule, J Physiol., 2003, 548:667-675.), and/or a calcium chan 1997, and Mosby’s Drug Consult, 2005, Mosby Inc., nel opener such as BAY K 8644 and/or ?ecanide. Elsevier, St. Louis Mo., ISBN 0-323-03393-8. For example, [0043] In a further embodiment, a second conductance quinidine gluconate is typically administered in doses rang modulator is administered to address the effects of organo ing from about 5-10 mg/kg (Mosby’s Drug Consult p.II phosphate toxins in blocking the fast outWard rectifying 2466-2475); procainamide is generally given in amounts of potassium current. For example, an inhibitor of an inWard up to 50 mg/kg/day (Mosby’s Drug Consult p.II-2409-2412); recti?er may be administered. An inhibitor of an inWard rec disopyramide is administered in amounts ranging from about ti?er inhibits KAsh, a muscarinic potassium channel found in 400-1200 mg/day (Mosby’s Drug Consult p.II-852-855); atrial myocytes, for example. Such an inhibitor is illustra lidocaine is generally given in amounts in the range of 50-100 tively tertiapin, a peptide originally isolated from Apis mel mg administered intravenously (Mosby’s Drug Consult p.II lifera honeybee venom. Tertiapin-Q is a synthetic version of 1744-1751); tocainide is usually given in amounts ranging the peptide in Which a is replaced by a glutamine from 1200-1800 mg/day (Mosby’s Drug Consult p.II-2833); to yield the peptide having the sequence: ALCNCNRII mexiletine is generally given in amounts of about 600-1200 IPHQCWKKCGKK. See Drici, M. D. et al., Br J Pharrnacol. mg/day (Mosby’s Drug Consult p.II-1951-1953); ?ecainide 2000, 131(3):569-77, for discussion of tertiapin effects on is usually administered in amount ranging from about 100 potassium channels in mammalian heart. 300 mg/day (Mosby’s Drug Consult p.II-1 189-1 1 90); propra [0044] Another example of a therapeutic agent included in nolol is typically given in amounts in the range of about a method of treatment and/ or a pharmaceutical composition 80-640 mg/day (Mosby’s Drug Consult p.II-2445-2451); according to the invention is an acetylcholine receptor acebutolol is generally given in amounts in the range of about antagonist. An acetylcholine receptor antagonist may be an 200-1200 mg/day (Mosby’s Drug Consult p.II-13-14); antagonist of a nicotinic and/ or muscarinic receptor. Musca esmolol is typically administered in amounts ranging from rinic antagonists are preferred in one embodiment. Exem about 50-500 micrograms/kg/min (Mosby’s Drug Consult plary muscarinic acetylcholine receptor antagonists include p.II-1019-1022); sotalol is generally given in amounts rang scopolamine, ipratropium, methantheline, propantheline, ing from 160-640 mg/day (Mosby’s Drug Consult p.II-2642 tolterodine (tartrate), anisotropine, clidinium, dicyclomine, 2650); dofetilide is generally given in amounts ranging from glycopyrrolate, homatropine, hyoscyamine, mepenZolate, 250-1000 micrograms/day (Mosby’s Drug Consult p.II-872 methscopolamine, and pirenZepine. Atropine is a preferred 876); amidarone is usually administered in an amount in the muscarinic acetylcholine receptor antagonist. range of about 400-1600 mg/ day in oral form (Mosby’s Drug [0045] An anticonvulsant may be administered in an inven Consult p.II-121-130); verapamil is typically given in an tive method and/ or as part of a pharmaceutical composition of amount in the range of about 120-480 mg/ day (Mosby’s Drug the invention. Preferred anticonvulsants include benZodiaZ Consult p.II-2977-2983); digoxin is administered as epines such as , loraZepam, and midaZolam, each described in Mosby’s Drug Consult p.II-810-819; and dilt given in amounts in the range of about 0.05-5 mg/kg body iaZem is generally administered in amounts in the range of Weight, preferably in the range of about 0.1-1 mg/kg body about 180-360 mg/day in oral formulation (Mosby’s Drug Weight. Consult p.II-827-839). [0046] Another example of a therapeutic agent is an anti [0047] In one embodiment, a therapeutic agent is adminis arrhythmic drug administered in order to further support inhi tered Which addresses effects of organophosphate intoxica US 2010/0168030 A1 Jul. 1,2010

tion on mitochondrial function. As noted above, one of the knoWn for terrorist activity being an indicator making expo effects of organophosphate intoxication is inhibition of oxy sure to organophosphate more likely. gen metabolism. Consequences of such inhibition include [0053] Organophosphate poisoning may occur by any of distortion of usual ion concentrations across the mitochon various routes illustratively including ingestion, inhalation, drial membrane. For example, a particular problem is accu skin exposure, mucosal exposure, and parenteral exposure. mulation of calcium in mitochondria, due in part to insu?i Organophosphate exposure leading to toxic effects may be in cient production of ATP necessary to sustain activity of any of various forms such as exposure to organophosphate mechanisms for maintenance of normal calcium concentra containing gases or liquids. tions, such as the Na"-Ca++ exchanger. Further, sodium and [0054] Symptoms and signs of organophosphate poisoning potassium concentrations in mitochondria are distorted due, include early symptoms such as anxiety, headache, vertigo, at least in part, to the decreased production of ATP and con hyperpnea, dyspnea, hypertension, bradycardia and cardiac sequent inhibition of a Na"-K+ pump. A further deleterious arrhythmias such as sinus or AV nodal arrhythmias. Further effect of organophosphate intoxication on cells and mito symptoms include loss of consciousness, convulsions and chondria is increased generation of reactive oxygen species. cardiac arrest. Of particular interest in the context of an embodiment of an inventive method are cardiovascular symp [0048] A therapeutic agent Which addresses effects of orga toms including electrocardiography changes such as atria nophosphate intoxication on mitochondrial function includes ?brillation, entopic ventricular heartbeats, abnormal QRS an agonist or antagonist of a mitochondrial membrane chan complex and sinus bradycardia. Elevated blood organophos nel ion exchanger and/or pump effective to normaliZe dis phate concentration and decreased acetylcholinesterase torted ion concentrations across the mitochondrial mem activity can con?rm exposure to organopho sphate. HoWever, brane. The mitochondrial membrane moiety may be selected Where likelihood of organophosphate exposure is present, from the group consisting of: an ion channel, an ion pump, an rapid treatment is required since death can occur in less than ion exchanger. Such compounds may be included to improve 10 minutes in severe cases. Thus, in one embodiment, an mitochondrial energy production, dampen mitochondrial cal individual selected for treatment With an inventive composi cium accumulation and reactive oxygen species production. tion and/or method is suffering from organophosphate-in For example, since calcium concentrations inside the mito duced heart-failure and/ or an organophosphate-induced con chondria are typically increased folloWing organophosphate tractility de?cit. Such an individual may have a history or exposure, an agent may be administered Which is effective to likelihood of organophosphate exposure and symptoms of activate or enhance an outWard movement of calcium from a organophosphate-induced heart failure and/or an organo mitochondrion. Exemplary agents effective to stimulate cal phosphate-induced contractility de?cit. Optionally, organo cium e?llux include menadione (Henry T R et al., J. Toxicol. phosphate exposure is con?rmed by assay for organophos Environ. Health, 45(4):489-504, 1995). phate, an organophosphate metabolite, a clinical and/or [0049] In other examples, an agonist or antagonist of a pathophysiological ?nding consistent With organophosphate mitochondrial membrane channel and/ or pump stimulates exposure, especially by acetylcholinesterase activity assay. activity of a mitochondrial membrane potassium channel, [0055] The methods of the invention include both acute and such as mitoKATP and/ or mitoKCa. A particular compound chronic therapies. Relatively long-term administration of a Which stimulates mitoKATP is diaZoxide. therapeutic agent also Will be bene?cial after a patient has [0050] Optionally further included is administration of a suffered from chronic organophosphate-caused heart failure therapeutic agent Which is a protein kinase C (PKC) inhibitor. to provide increased exercise tolerance and functional capac In the presence of organophosphate, rising inorganic phos ity. For example, a chloride conductance inhibitior, such as phate (Pi) and declining phosphocreatine (PCr) are present in DIDS, can be administered to a patient after having suffered cells, affecting the electrophysiology of the tissue. An exem heart failure due to organophosphate caused toxicity to pro plary PKC inhibitor Which may be used is H-7 (1-(5-iso mote enhanced functional capacity. quinolinesulfonyl)-2-methylpiperaZine, an isoquinoline [0056] For example, DIDS can be immediately adminis sulfonamide), a pharmacological inhibitor of PKC Which tered to a patient, e.g. intravenously or intraperitonially, that prevents and/or inhibits the changes in the PCr and Pi. has suffered or is suffering from congestive heart failure or [0051] Preferably, a patient is identi?ed and selected for cardiogenic shock. Such immediate administration prefer treatment according to an inventive method. For example, a ably Would entail administration of a therapeutic agent Within patient in need of treatment may be an individual that is minutes after a subject exhibits a symptom and/or sign of suffering from organophosphate intoxication such that an organophosphate-caused ventricular ?brillation or cardio increase in myocardial contractility With reduced energy genic shock. In one embodiment, an oral dosage is preferred. requirements is an intended therapy and Where inhibition of [0057] A patient in need of treatment for organopho sphate organophosphate-induced distortion of the action potential of poisoning is discussed herein in general terms relating to myocytes and support of restoration of usual intracellular human individuals. HoWever, inventive methods and compo ionic concentrations is desirable. sitions for individuals of other species, particularly mamma [0052] As noted above, organophosphate toxins are in com lian species such as non-human primates, sheep, dogs, cats, mon use in agricultural settings, as Well as being a haZard cattle, pigs, horses and the like, are considered to be Within the encountered on the battle?eld or in terrorist situations. Thus, scope of the invention. an individual in need of treatment of organophosphate poi [0058] While compositions and methods according to the soning may be characterized by an occupation likely to bring invention are described as relating to treatment of organo the individual in contact With organopho sphate and/ or history phosphate-induced cardiac symptoms and signs, it is appre of organophosphate exposure. Often of interest in identifying ciated that other cells and tissues are affected by organophos an individual in need of treatment is a history of the individu phate and that compositions and methods described herein al’s recent activity or location, presence in a War Zone or area may be applicable to treatment of those other cells and tis US 2010/0168030 A1 Jul. 1,2010

sues. In particular, excitable membranes of cells such as neu example, parabens, chlorobutanol, phenol, sorbic acid, and rons may have membrane potentials and ion concentrations the like. It may also be desirable to include isotonic agents, for across those membranes are distorted as a result of organo example, sugars, sodium chloride, and substances similar in phosphate intoxication and it is appreciated that inventive nature. Prolonged delivery of an injectable pharmaceutical methods and compositions may be used to treat such sequelae form can be brought about by the use of agents delaying of organophosphate exposure. absorption, for example, aluminum mono stearate and gelatin. [0059] An inventive pharmaceutical composition includes [0065] Solid dosage forms for oral administration include a modulator of a biological cell membrane conductance as capsules, tablets, pills, poWders, and granules. In such solid described herein and a pharmaceutically acceptable carrier. dosage forms, a conductance modulator is admixed With at The term “pharmaceutically acceptable carrier” as used least one inert customary excipient (or carrier) such as sodium herein is intended to refer to a carrier or diluent that is gen citrate or dicalcium phosphate or (a) ?llers or extenders, as for erally non-toxic to an intended recipient and Which does not example, starches, lactose, sucrose, glucose, mannitol, and signi?cantly inhibit activity of an active agent included in the silicic acid, (b) binders, as for example, carboxymethylcellu composition. lose, alignates, gelatin, polyvinylpyrrolidone, sucrose, and [0060] An inventive composition is suitable for administra acacia, (c) humectants, as for example, glycerol, (d) disinte tion to patients by a variety of systemic and/or local routes grating agents, as for example, agar-agar, calcium carbonate, illustratively including intravenous, oral, parenteral, intrath plant starches such as potato or tapioca starch, al ginic acid, ecal, intraventricular, intracardiac, pericardiac, and mucosal. certain complex silicates, and sodium carbonate, (e) solution [0061] An inventive composition may be administered retarders, as for example, para?in, (f) absorption accelerators, acutely or chronically. For example, in an emergency situa as for example, quaternary ammonium compounds, (g) Wet tion, a conductance modulator included in a composition as ting agents, as for example, cetyl alcohol, glycerol described herein may be administered as a unitary dose or in monostearate, and glycols (h) adsorbents, as for example, multiple doses over a relatively limited period of time, such as kaolin and bentonite, and (i) lubricants, as for example, talc, seconds -hours. In a further embodiment, administration may calcium stearate, magnesium stearate, solidpolyethylene gly include multiple doses administered over a period of days cols, sodium lauryl sulfate, or mixtures thereof. In the case of years, such as for chronic treatment of long-lasting sequelae capsules, tablets, and pills, the dosage forms may also com of organophosphate poisoning. prise buffering agents. [0062] A therapeutically effective amount of a current [0066] Solid compositions of a similar type may also be modulator and of a therapeutic agent described herein Will employed as ?llers in soft and hard-?lled gelatin capsules vary independently depending on the particular compound using such excipients as lactose or milk sugar as Well as high and on the particular agent used, the severity of the toxin molecular Weight polyethyleneglycols, and the like. exposure including the route of toxin exposure and the iden [0067] Solid dosage forms such as tablets, dragees, cap tity of the toxin, the length of time since toxin exposure and sules, pills, and granules can be prepared With coatings and the general physical characteristics of the individual to be shells, such as enteric coatings and others Well knoWn in the treated. One of skill in the art could determine a therapeuti art. They may contain opacifying agents, and can also be of cally effective amount in vieW of these and other consider such composition that they release the active compound or ations typical in medical practice. In general it is contem compounds in a certain part of the intestinal tract in a delayed plated that a therapeutically effective amount Would be in the manner. Examples of embedding compositions Which can be range of about 0.001 mg/kg-l00 mg/kg body Weight, more used are polymeric substances and Waxes. The active com preferably in the range of about 0.01-10 mg/kg, and further pounds can also be in micro-encapsulated form, if appropri preferably in the range of about 0.1-5 mg/kg. Further, dosage ate, With one or more of the above-mentioned excipients. may be adjusted depending on Whether treatment is to be [0068] The enteric coating is typically a polymeric mate acute or continuing. rial. Preferred enteric coating materials have the characteris [0063] Compositions suitable for delivery may be formu tics of being bioerodible, gradually hydrolyZable and/or lated in various forms illustratively including physiologically gradually Water-soluble polymers. The amount of coating acceptable sterile aqueous or nonaqueous solutions, disper material applied to a solid dosage generally dictates the time sions, suspensions or emulsions, and sterile poWders for interval betWeen ingestion and drug release. A coating is reconstitution into sterile injectable solutions or dispersions. applied With to a thickness such that the entire coating does Examples of suitable aqueous and nonaqueous carriers, dilu not dissolve in the gastrointestinal ?uids at pH beloW 3 asso ents, solvents, and vehicles include Water, , polyols ciated With stomach acids, yet dissolves above pH 3 in the such as propylene glycol, polyethylene glycol, glycerol, and small intestine environment. It is expected that any anionic the like, suitable mixtures thereof; vegetable oils such as olive polymer exhibiting a pH-dependent solubility pro?le is oil; and injectable organic esters such as ethyloleate. Proper readily used as an enteric coating in the practice of the present ?uidity can be maintained, for example, by the use of a invention to achieve delivery of the active to the loWer gas coating such as lecithin, by the maintenance of the required trointestinal tract. The selection of the speci?c enteric coating particle siZe in the case of dispersions, and by the use of material depends on properties such as resistance to disinte surfactants, such as sodium lauryl sulfate. Such formulations gration in the stomach; imper'meability to gastric ?uids and are administered by a suitable route including parenteral and active agent diffusion While in the stomach; ability to dissi oral administration. Administration may include systemic or pate at the target intestine site; physical and chemical stability local injection, and particularly intravenous injection. during storage; non-toxicity; and ease of application. [0064] These compositions may also contain adjuvants [0069] Suitable enteric coating materials illustratively such as preserving, Wetting, emulsifying, and dispensing include cellulosic polymers such as hydroxypropyl cellulose, agents. Prevention of the action of microorganisms can be hydroxyethyl cellulose, hydroxypropyl methyl cellulose, ensured by various antibacterial and antifungal agents, for methyl cellulose, ethyl cellulose, cellulose acetate, cellulose US 2010/0168030 A1 Jul. 1,2010

acetate phthalate, cellulose acetate trimellitate, hydroxypro sion, solution, suspension, syrup, or elixir. In addition to the pylmethyl cellulose phthalate, hydroxypropylmethyl cellu active compounds, the liquid dosage forms may contain inert lose succinate and carboxymethylcellulose sodium; acrylic diluents commonly used in the art, such as Water or other acid polymers and copolymers, preferably formed from solvents, solubiliZing agents and emulsi?ers, as for example, acrylic acid, methacrylic acid, methyl acrylate, ammonium ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl methylacrylate, ethyl acrylate, methyl methacrylate and/or acetate, benZyl alcohol, benZyl benZoate, propyleneglycol, ethyl; vinyl polymers and copolymers such as polyvinyl pyr 1,3-butyleneglycol, dimethylformamide, oils, in particular, rolidone, polyvinyl acetate, polyvinylacetate phthalate, viny cottonseed oil, groundnut oil, corn germ oil, olive oil, castor lacetate crotonic acid copolymer, and ethylene-vinyl acetate oil and sesame oil, glycerol, tetrahydrofurfuryl alcohol, poly copolymers; shellac; and combinations thereof. A particu ethyleneglycols and fatty acid esters of sorbitan or mixtures larly preferred enteric coating material for use herein is those acrylic acid polymers and copolymers available under the of these substances, and the like. trade name EUDRAGIT®, Roehm Pharma (Germany). The [0074] Besides such inert diluents, the composition can EUDRAGIT® series L, L-30D S copolymers, and cross also include adjuvants, such as Wetting agents, emulsifying linkedpolymers, see for example US. Pat. No. 6,136,345, are and suspending agents, sWeetening, ?avoring, and perfuming most preferred since these are insoluble in stomach and dis agents. solve in the intestine. [0075] Suspensions, in addition to an inventive conjugate, [0070] The enteric coating provides for controlled release may contain suspending agents, as for example, ethoxylated of the active agent, such that release is accomplished at a isostearyl , polyoxyethylene sorbitol and sorbitol predictable location in the loWer intestinal tract beloW the esters, microcrystalline cellulose, aluminum metahydroxide, point at Which drug release Would occur absent the enteric bentonite, agar-agar or tragacanth, or mixtures of these sub coating. The enteric coating also prevents exposure of the stances, and the like. active agent and carrier to the epithelial and mucosal tissue of [0076] An ion channel modulator and/ or therapeutic agent the buccal cavity, pharynx, esophagus, and stomach, and to administered in an inventive method is de?ned herein as an the associated With these tissues. The enteric coating ion channel modulator and/ or therapeutic agent and pharma therefore helps to protect the active agent and a patient’s ceutically acceptable salts, derivatives, oxides and hydrates internal tissue from any adverse event prior to drug release at thereof. The term “pharmaceutically acceptable salt, deriva the desired site of delivery. Furthermore, the coated solid tive, oxide and hydrate” refers to a formulation that is sub dosages of the present invention alloW optimiZation of drug stantially non-toxic to the individual being treated and Which absorption, active agent protection, and safety. Multiple does not substantially inhibit the activity of an active agent enteric coatings targeted to release the active agent at various being administered. The term “derivative” as used herein regions in the loWer gastrointestinal tract Would enable even refers to a channel modulator and/or a therapeutic agent more effective and sustained improved delivery throughout Which is chemically modi?ed to include a nitrogen, oxygen, the loWer gastrointestinal tract. carbon, , halogen or phosphorus containing moiety [0071] The enteric coating optionally contains a plasticiZer illustratively including a C l-C4 substituted or unsubstituted, to prevent the formation of pores and cracks that alloW the straight chain or branched alkyl, an amine, a sulfhydryl and an penetration of the gastric ?uids into the solid dosage. Suitable oxide. The term “oxide” as used herein refers to an oxygen plasticizers illustratively include, triethyl citrate (Citro?ex 2), containing derivative of a channel modulator and/or a thera triacetin (glyceryl triacetate), acetyl triethyl citrate (Citro?ec peutic agent. Exemplary oxygen containing derivatives A2), CarboWax 400 (polyethylene glycol 400), diethyl phtha include an oxygen containing moiety such as a carboxyl, a late, tributyl citrate, acetylated monoglycerides, glycerol, carbonyl, a sulfonyl, a sulfoxy, a hydroxyl, a nitro, a phos fatty acid esters, propylene glycol, and dibutyl phthalate. In phate, and a C l-C4 substituted or unsubstituted, straight chain particular, a coating composed of an anionic carboxylic or branched alkyl linked to the channel modulator and/or a acrylic polymer typically contains approximately 10% to therapeutic agent by ester or ether linkage. 25% by Weight of a plasticiZer, particularly dibutyl phthalate, [0077] Further examples and details of pharmacological polyethylene glycol, triethyl citrate and triacetin. The coating formulations and ingredients are found in standard references can also contain other coating excipients such as detacki?ers, such as: A. R. Gennaro, Remington: The Science and Practice antifoaming agents, lubricants (e.g., magnesium stearate), of Pharmacy, Lippincott Williams & Wilkins, 20th ed. and stabiliZers (e.g., hydroxypropylcellulose, acids and (2003); L. V. Allen, Jr. et al., Ansel’s Pharmaceutical Dosage bases) to solubiliZe or disperse the coating material, and to Forms and Drug Delivery Systems, 8th Ed. (Philadelphia, improve coating performance and the coated product. Pa.: Lippincott, Williams & Wilkins, 2004); J. G. Hardman et [0072] The enteric coating is applied to a solid dosage using al., Goodman & Gilman’s The Pharmacological Basis of conventional coating methods and equipment. For example, Therapeutics, McGraW-Hill Professional, 10th ed. (2001). an enteric coating can be applied to a solid dosage using a [0078] While compositions and methods described herein coating pan, an airless spray technique, ?uidized bed coating are primarily described in the context of organophosphate equipment, or the like. Detailed information concerning intoxication, it is appreciated that compositions and methods materials, equipment and processes for preparing coated dos as described may be applicable to treatment of other types of age forrns may be found in Pharmaceutical Dosage Forms: toxic agent exposure in Which a dormant ion current is acti Tablets, eds. Lieberman et al. (New York: Marcel Dekker, vated folloWing exposure such that cell membrane potentials Inc., 1989), and in L. V. Allen, Jr. et al., Ansel’s Pharmaceu and ion concentrations across those membranes are distorted, tical Dosage Forms and Drug Delivery Systems, 8th Ed. symptoms and signs indicative of pathological excitable cell (Philadelphia, Pa.: Lippincott, Williams & Wilkins, 2004). function. Further, an individual may be exposed to combina [0073] Liquid dosage forms for oral administration include tions of toxic agents for Which inventive methods and com a pharmaceutically acceptable carrier formulated as an emul positions are applicable. US 2010/0168030 A1 Jul. 1,2010

[0079] Embodiments of inventive compositions and meth modulate a chloride conductance, thereby reducing a ods are illustrated in the following examples. These examples symptom or sign of a toxin-caused cardiac abnormality are provided for illustrative purposes and are not considered and treating the toxin-induced cardiac abnormality. limitations on the scope of inventive compositions and meth 2. The method of claim 1 Wherein the chloride current ods. modulator is a modulator of lCl, sWell. 3. The method of claim 1 Wherein the chloride current EXAMPLE 1 modulator is selected from the group consisting of: a disul [0080] An individual having symptoms of organophos fonic stilbene, an arylaminobenZoate, a fenamate, an phate-induced cardiac arrhythmia is injected intravenously anthracene carboxylate, an indanylalkanoic acid, clo?bric With 10 mg/kg 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid, a clo?bric acid derivative, a sulfonylurea, a calixarene, acid (DIDS) dissolved in normal saline. The effect of the suramin, and tamoxifen. treatment is monitored by ECG. 4. The method of claim 2 Wherein the modulator of lCl, sWell is selected from the group consisting of: 4,4'-diisothio EXAMPLE 2 cyanostilbene-2,2'-disulfonic acid (DIDS); 4,4'-dinitrostil bene-2,2'-disulfonic acid (DNDS); 4-acetamindo-4'-isothio [0081] An individual presents With. symptoms of organo cyanostilbene-2,2'-disulfonic acid (SITS); tamoxifen; phosphate-induced cardiac arrhythmia. A solution of 20 5-nitro-2-(3 -phenylpropylamino) benZoic acid (NPPB); mg/kg of indanyloxyacetic acid (lAA-94) is prepared by dis ni?umic acid (N FA); ?ufenamic acid; anthracene-9-carboxy solving the lAA-94 in ethanol and then diluting the material late (9AC); diphenylaminecarboxylate (DPC); 2-(p-chlo to the ?nal concentration in normal saline. The organophos rophenoxy)propionic acid (CPP); and indanyloxyacetic acid phate affected individual is injected intravenously With the (lAA-94). lAA-94 preparation. The effect of the treatment may be moni 5. The method of claim 1 further comprising administering tored by ECG. a therapeutic agent to inhibit organophosphate-induced dis tortion of the action potential of myocytes, support restora EXAMPLE 3 tion of usual intracellular ionic concentrations, and support an [0082] An individual having symptoms of organophos increase in cardiac contractility. phate-induced cardiac arrhythmia is injected intravenously 6. The method of claim 5 Wherein the therapeutic agent is With 0.5 mg/kg of tamoxifen dissolved in ethanol and then a second current modulator. diluted to ?nal concentration in normal saline. The effect of 7. The method of claim 5 Wherein the therapeutic agent is the treatment may be monitored by ECG. an antiarrhythmic drug. 8. The method of claim 5 Wherein the therapeutic agent is EXAMPLE 4 a modulator of a mitochondrial membrane moiety selected from the group consisting of: an ion channel, an ion pump, an [0083] An individual having symptoms of organophos ion exchanger, and a combination thereof. phate-induced cardiac arrhythmia is injected intravenously 9. The method of claim 5 Wherein the therapeutic agent is With 1 mg/kg of 5-nitro-2-(3-phenylpropylamino)benZoic an inhibitor of protein kinase C. acid (NPPB) dissolved in ethanol and then diluted to ?nal 10. The method of claim 5 Wherein the therapeutic agent is concentration in normal saline. The effect of the treatment an anticonvulsant. may be monitored by ECG. 11. The method of claim 5 Wherein the therapeutic agent is an organophosphate clearing agent. EXAMPLE 5 12. The method of claim 11 Wherein the organophosphate [0084] An individual having a history of organophosphate clearing agent is selected from the group consisting of: obi exposure and symptoms of cardiac rhythm abnormalities doxime, asoxime, pralidoxime, a salt thereof; and a combi ingests 25 mg/kg of 4,4'-diisothiocyanostilbene-2,2'-disul nation thereof. fonic acid (DIDS) in tablet form. The effect of the treatment 13. The method of claim 1 Wherein the cardiac abnormality may be monitored by ECG. is ventricular ?brillation. [0085] Any patents or publications mentioned in this speci 14. The method of claim 1 Wherein the cardiac abnormality ?cation are incorporated herein by reference to the same is cardiogenic shock. extent as if each individual publication is speci?cally and 15. The method of claim 6 Wherein the second current individually indicated to be incorporated by reference. modulator is an inhibitor of a muscarinic potassium channel. [0086] The compositions and methods described herein are 16. The method of claim 15 Wherein the inhibitor of a presently representative of preferred embodiments, exem muscarinic potassium channel is selected from the group plary, and not intended as limitations on the scope of the consisting of: tertiapin, tertiapin-Q, and a combination invention. Changes therein and other uses Will occur to those thereof. skilled in the art. Such changes and other uses can be made 17. The method of claim 5 Wherein the therapeutic agent is Without departing from the scope of the invention as set forth an acetylcholine receptor antagonist. in the claims. 18. A pharmaceutically effective amount of a composition 1. A method of treating an organophosphate toxin-caused consisting of: cardiac abnormality in an individual subject, comprising: a chloride current modulator of lCl sWell; and administering a pharmaceutical composition comprising a a combination therapeutic agent limited to the class of therapeutically effective amount of a chloride current agents that in a single dose inhibit organophosphate modulator to an individual subject having a cardiac caused distortion of the action potential of myocytes, abnormality caused by intoxication With an organophos support restoration of usual intracellular ionic concen phate toxin, the chloride current modulator effective to trations, and support an increase in cardiac contractility. US 2010/0168030 A1 Jul. 1, 2010 1 1

19. (canceled) clearing agent, an inhibitor of protein kinase C, a modulator 20. The composition of claim 18 Wherein the therapeutic of a mitochondrial membrane moiety, and a combination agent is selected from the group consisting of: a second cur- thereof rent modulator, an antiarrhythmic drug, an acetylcholine receptor antagonist, an anticonvulsant, an organophosphate * * * * *