Treatment of tinnitus through modulation of chloride co-transporter nkcc1 in the auditory system

Abstract

The present invention relates to the treatment or prevention of tinnitus. More precisely, the present invention relates to a compound modulating chloride co-transporter NKCC1 (chloride co-transporter modulator) for use in the treatment of tinntitus. In addition, the present invention concerns pharmaceutical compositions comprising such an NKCC1 chloride co-transporter modulator as an active agent, a method for the treatment or prevention of tinnitus by administering such a chloride co-transporter modulator, and a screening method for the identification and characterization of compounds capable of modulating chloride co-transporter NKCC1.

1 . A compound modulating chloride co-transporter sodium potassium chloride co-transporter 1 (NKCC1) for use in the treatment or prevention of tinnitus. 
     
     
         2 . Compound for use according to  claim 1 , wherein the compound antagonizes NKCC1. 
     
     
         3 . Compound for use according to  claim 1  or  2 , wherein the compound antagonizes the function of NKCC1 by decreasing or inhibiting its activity and/or expression. 
     
     
         4 . Compound for use according to any of  claims 1  to  3 , wherein the activity and/or expression of KCC2 is increased by NKCC1 antagonism. 
     
     
         5 . Compound for use according to  claim 3 , wherein the compound decreases activity and/or expression of NKCC1 and increases activity and/or expression of KCC2. 
     
     
         6 . Compound for use according to any of  claims 1  to  5 , wherein the compound is selected from the group consisting of thiazides and sulfonamides. 
     
     
         7 . Compound for use according to any of  claims 1  to  6 , wherein the compound is a sulfonamide selected from the group consisting of acetazolamide, azosemide, bumetanide, chlorthalidone, clopamide, furosemide, hydrochlorothiazide (HCT, HCTZ, HZT), indapamide, mefruside, metolazone, piretanide, tripamide xipamide, dichlorphenamide (DCP), dorzolamide, ethoxzolamide, sultiame, or zonisamide or analogs thereof. 
     
     
         8 . Compound for use according to any of  claims 1  to  7 , wherein the sulfonamide compound is a diuretic compound, preferably selected from the group consisting of bumetanide, furosemide, piretanide, azosemide, and torsemide or analogs thereof. 
     
     
         9 . Compound for use according to any of  claims 1  to  8 , wherein diuretic compound is selected from the group consisting of
 (i) bumetanide, bumetanide aldehyde, bumetanide methyl ester, bumetanide cyanomethyl ester, bumetanide ethyl ester, bumetanide isoamyl ester, bumetanide octyl ester, bumetanide benzyl ester, bumetanide dibenzylamide, bumetanide diethylamide, bumetanide morpholinoethyl ester, bumetanide 3-(dimethylaminopropyl) ester, bumetanide N,N-diethylglycolamido ester, bumetanide N,N-dimethylglycolamido ester, bumetanide pivaxetil ester, bumetanide propaxetil ester, bumetanide methoxy(polyethyleneoxy) n-1 -ethyl ester, bumetanide benzyltrimethylammonium salt and bumetanide cetyltrimethylammonium salt, bumetanide [—(C═O)—SH] thioacid, bumetanide S-methyl thioester, bumetanide S-cyanotnethyl thioester, bumetanide S-ethyl thioester, bumetanide S-isoamyl thioester, bumetanide S-octyl thioester, bumetanide S-benzyl thioester, bumetanide S-(morpholinoethyl) thioester, bumetanide S—[3-(dimethylaminopropyl)] thioester, bumetanide S—(N,N-diethylglycolamido) thioester, bumetanide S—(N,N-dimethylglycolamido) thioester, bumetanide S-pivaxetil thioester, bumetanide S-propaxetil thioester, bumetanide S—[methoxy(polyethyleneoxy) n-1 -ethyl] thioester, bumetanide [(C═O)—S − ] benzyltrimethyl-ammonium thioacid salt and bumetanide [—(C═O)—S − ] cetyltrimethylammonium thioacid salt, metastable bumetanide [—(C═S)—OH] thioacid, bumetanide O-methyl thioester, bumetanide O-cyanomethyl thioester, bumetanide O-ethyl thioester, bumetanide O-isoamyl thioester, bumetanide O-octyl thioester, bumetanide O-benzyl thioester, bumetanide O-(morpholinoethyl) thioester, bumetanide O—[3-(dimethylaminopropyl)] thioester, bumetanide O—(N,N-diethylglycolamido) thioester, bumetanide, O—(N,N-dimethylglycolamido) thioester, bumetanide O-pivaxetil thioester, bumetanide O-propaxetil thioester, bumetanide O—[methoxy(poryethyleneoxy) n-1 -ethyl] thioester, bumetanide [—(C═S)—O − ] benzyltrimemyl-ammonium thioacid salt and bumetanide [—(C═S)—O − ] cetyltrimethylammonium thioacid salt, bumetanide thioaldehyde, bumetanide [—(C═S)—SH] dithioacid, bumetanide methyl dithioester, bumetanide cyanomethyl dithioester, bumetanide ethyl dithioester, bumetanide isoamyl dithioester, bumetanide octyl dithioester, bumetanide benzyl dithioester, bumetanide dibenzylthioamide, bumetanide diethylthioamide, bumetanide morpholinoethyl dithioester, bumetanide 3-(dimethylaminopropyl) dithioester, bumetanide N,N-diethylglycolamido dithioester, bumetanide N,N-dimethylglycolamido dithioester, bumetanide pivaxetil dithioester, bumetanide propaxetil dithioester, bumetanide methoxy(polyethyleneoxy) n-1 -ethyl dithioester, bumetanide benzyltrimethylammonium dithioacid salt and bumetanide cetyltrimethyl-ammonium dithioacid salt and 
 (ii) furosemide, furosemide aldehyde, furosemide methyl ester, furosemide cyanomethyl ester, furosemide ethyl ester, furosemide isoamyl ester, furosemide octyl ester, furosemide benzyl ester, furosemide morpholinoethyl ester, furosemide 3-(dimethylaminopropyl) ester, furosemide N,N-diethylglycolamido ester, furosemide N,N-dimethylglycolamido ester, furosemide pivaxetil ester, furosemide propaxetil ester, furosemide methoxy(polyethyleneoxy) n-1 -ethyl ester, furosemide benzyltrimethylammonium acid salt and furosemide cetyltrimethylammonium acid salt. In particular embodiments, the compound is not furosemide. In further embodiments of the present invention, the compound of formula IV can be furosemide [—(C═O)—SH] thioacid, furosemide S-methyl thioester, furosemide S-cyanomethyl thioester, furosemide S-ethyl thioester, furosemide S-isoamyl thioester, furosemide S-octyl thioester, furosemide S-benzyl thioester, furosemide S-(morpholinoethyl) thioester, furosemide S—[3-(dimethylaminopropyl)] thioester, furosemide S—(N,N-diethylglycolamido) thioester, furosemide S—(N,N-dimethylglycolamido) thioester, furosemide S-pivaxetil thioester, furosemide S-propaxetil thioester, furosemide S—[methoxy(poryethyleneoxy) n-1 -ethyl] thioester, furosemide [—(C═O)—S − ] benzyltrimethylammonium thioacid salt and furosemide [—(C═O)—S − ]cetyltrimethylammonium thioacid salt, metastable furosemide [—(C═S)—OH] thioacid, furosemide O-methyl thioester, furosemide O-cyanomethyl thioester, furosemide O-ethyl thioester, furosemide O-isoamyl thioester, furosemide O-octyl thioester, furosemide O-benzyl thioester, furosemide O-(morpholinoethyl) thioester, furosemide O—[3-(dimethylaminopropyl)] thioester, furosemide O—(N,N-diethylglycolamido) thioester, furosemide O—(N,N-dimethylglycolamido) thioester, furosemide O-pivaxetil thioester, furosemide O-propaxetil thioester, furosemide O—[methoxy(polyethyleneoxy) n-1 -ethyl] thioester, furosemide [—(C═S)—O − ] benzyltrimethyl-ammonium thioacid salt and furosemide cetyltrimethylammonium thioacid salt, furosemide thioaldehyde, furosemide [—(C═S)—SH] dithioacid, furosemide methyl dithioester, furosemide cyanomethyl dithioester, furosemide ethyl dithioester, furosemide isoamyl dithioester, furosemide octyl dithioester, furosemide benzyl dithioester, furosemide dibenzylthioamide, furosemide diethylthioamide, furosemide morpholinoethyl dithioester, furosemide 3-(dimethylamino[rho]ropyl) dithioester, furosemide N,N-diethylglycolamido dithioester, furosemide N,N-dimethylglycolamido dithioester, furosemide pivaxetil dithioester, furosemide propaxetil dithioester, furosemide methoxy(polyethyleneoxy) n-1 -ethyl dithioester, furosemide benzyltrimethylammonium dithioacid salt and furosemide cetyltrimethylammonium dithioacid salt and 
 (iii) piretanide, piretanide aldehyde piretanide methyl ester, piretanide cyanomethyl ester, piretanide ethyl ester, piretanide isoarmyl ester, piretanide octyl ester, piretanide benzyl ester, piretanide dibenzylamide, piretanide diethylamide, piretanide morpholinoethyl ester, piretanide 3-(dimethylaminopropyl) ester, piretanide N,N-diethylglycolamide ester, piretanide dimethylglycolamide ester, piretanide pivaxetil ester, piretanide propaxetil ester, piretanide methoxy(polyethyleneoxy) n-1 -ethyl ester, piretanide benzyltrimethylammonium salt and piretanide cetyltrimethylammonium salt. In particular embodiments, the compound is not piretinide, piretanide [—(C═O)—SH] thioacid, piretanide S-methyl thioester, piretanide S-cyanomethyl thioester, piretanide S-ethyl thioester, piretanide S-isoamyl thioester, piretanide S-octyl thioester, piretanide S-benzyl thioester, piretanide S-(morpholinoethyl) thioester, piretanide S—[3-(dimethylaminopropyl)] thioester, piretanide S—(N,N-diethylglycolamido) thioester, piretanide S—(N,N-dimethylglycolamido) thioester, piretanide S-pivaxetil thioester, piretanide S-propaxetil thioester, piretanide S—[methoxy(polyethyleneoxy) n-1 -ethyl] thioester, piretanide [-(C═O)—S − ] benzyltrimethylammonium thioacid salt and piretanide [—(C═O)—S − ] cetyltrimethylammonium thioacid salt, metastable piretanide [—(C═S)—OH] thioacid, piretanide O-methyl thioester, piretanide O-cyanomethyl thioester, piretanide O-ethyl thioester, piretanide O-isoamyl thioester, piretanide O-octyl thioester, piretanide O-benzyl thioester, piretanide O-(morpholinoethyl) thioester, piretanide O—[3-(dimethylaminopropyl)] thioester, piretanide O—(N,N-diethylglycolamido) thioester, piretanide, O—(N,N-dimethylglycolamido) thioester, piretanide O-pivaxetil thioester, piretanide O-propaxetil thioester, piretanide O—[methoxy(polyethyleneoxy) n-1 -ethyl] thioester, piretanide [—(C═S)—O − ] benzyltrimethylammonium thioacid salt and piretanide [—(C═S)—O − ] cetyltrimethylammonium thioacid salt, piretanide thioaldehyde, piretanide [—(C═S)—SH] dithioacid, piretanide methyl dithioester, piretanide cyanomethyl dithioester, piretanide ethyl dithioester, piretanide isoamyl dithioester, piretanide octyl dithioester, piretanide benzyl dithioester, piretanide dibenzylthioamide, piretanide diethylthioamide, piretanide morpholinoethyl dithioester, piretanide 3-(dimethylaminopropyl) dithioester, piretanide N,N-diethylglycolamido dithioester, piretanide N,N-dimethylglycolamido dithioester, piretanide pivaxetil dithioester, piretanide propaxetil dithioester, piretanide methoxy(polyethyleneoxy) n-1 -ethyl dithioester, piretanide benzyltrimethylammoniurn dithioacid salt and piretanide cetyltrimethylarnmoniurn dithioacid salt and 
 (iv) tetrazolyl-substituted azosemides, in particular methoxymethyl tetra-zolyl-substituted azosemides, methylthiornethyl tetrazolyl-substituted azosemides and N-mPEG350-tetrazolyl-substituted azosemides), azosemide benzyltrimethylammonium salt and/or azosemide cetyltrimethylammonium salt and 
 (v) pyridine-substituted torsemide quaternary ammonium salts or the corresponding inner salts (zwitterions), in particular methoxymethyl pyridinium torsemide salts, methylthiomethyl pyridinium torsemide salts and N-mPEG350-pyridinium torsemide salts. 
 
     
     
         10 . Compound for use according to any of  claims 1  to  6 , wherein the thiazide compound is selected from the group consisting of bendroflumethiazide, benzthiazide, chlorothiazide, hydrochlorothiazide, hydroflumethiazide, methylclothiazide, polythiazide, trichlor-methiazide, chlorthalidone, indapamide, metolazone or quinethazone or analogs thereof. 
     
     
         11 . Compound for use according to any of  claims 1  to  10 , wherein the compound is used in combination with one or more carbonic anhydrase inhibitors, in particular acetazomalide, dichlorphenamide, dorzolamide, brinzolamide and/or methazolamide. 
     
     
         12 . Compound for use according to any of  claims 1  to  5 , wherein the compound is an antibody, an antibody fragment or a humanized antibody antagonizing NKCC1 activity or an RNA selected from the group consisting of an siRNA, shRNA and anti-sense RNA, reducing NKCC1 expression. 
     
     
         13 . Compound for use according to any of  claims 1  to  5 , wherein the compound is selected from CCC interacting protein (CIP1) or a functional peptide derived therefrom, N-ethylmaleimide (NEM) or staurosporin. 
     
     
         14 . Compound for use according to any of  claims 1  to  13 , wherein the compound is used in combination with one or more GABAergic agonists and/or one or more glycine agonists, in a co-treatment protocol either simultaneously or in a staggered regimen protocol. 
     
     
         15 . A pharmaceutical composition comprising a compound modulating chloride co-transporter NKCC1 as defined in any  claims 1  to  14 . 
     
     
         16 . Pharmaceutical composition according to  claim 15 , further comprising one or more carbonic anhydrase inhibitor and/or GABAergic agonists and/or glycine agonists. 
     
     
         17 . Pharmaceutical composition according to  claim 15  or  16 , wherein the pharmaceutical composition is provided in a liquid, semi-liquid or viscous form, preferably in a gel-like form. 
     
     
         18 . Pharmaceutical composition according to any of  claims 15  to 17, wherein the composition contains a, preferably biodegradable polymer, preferably selected from the group consisting of hyaluronic acid resp. hyaluronates, lecithin gels, (poly)alanine derivatives, pluronics, poly(ethyleneglycol), poloxamers, chitosans, xyloglucans, collagens, fibrins, polyesters, poly(lactides), poly(glycolide) or their co-polymers PLGA, sucrose acetate isobutyrate, and glycerol monooleate. 
     
     
         19 . Pharmaceutical composition according to any of  claims 15  to  18  for use in the treatment or prevention of tinnitus. 
     
     
         20 . Pharmaceutical composition according to  claim 19  for use in the treatment or prevention of tinnitus, whereby the pharmaceutical composition is administered locally. 
     
     
         21 . Pharmaceutical composition according to  claim 20  for use in the treatment or prevention of tinnitus, whereby the pharmaceutical composition is administered to the middle/inner ear interface, preferably to the round and/or oval window membrane. 
     
     
         22 . A method for treatment tinnitus comprising administering a therapeutically effective amount of a compound modulating one or more chloride co-transporters as defined in  claims 1  to  14  or a pharmaceutical composition according to  claims 15  and  21 . 
     
     
         23 . Method for treatment tinnitus according to  claim 22 , further comprising co-administering of a carbonic anhydrase inhibitor and/or a GABAergic agonist and/or a glycine agonist. 
     
     
         24 . Method for treatment according to  claim 22  or  23 , wherein administration is performed systemically or locally, preferably locally on or in the ear. 
     
     
         25 . A screening method for the identification and characterization of compounds capable of modulating one or more chloride transporters, by (a) providing cells stably expressing the one or more chloride transporters, (b) adding a test compound to the cells, (c) adding a transporter cation and (d) measuring the cation transport across the cell membrane. 
     
     
         26 . A screening method according to  claim 25 , wherein one or more of the following steps are carried out: (i) provision of cells with heterologous expression of one or more of the transporters, (ii) additionally adding a fluorescence dye binding to the transporter cation, and (iii) measuring the initial cation transport rate and (iv) blocking interfering transporter activity of another transporter. 
     
     
         27 . A screening method according to  claim 25  or  26 , wherein the one or more chloride transporters are selected from NKCC1 and KCC2.