EP1214291A1

EP1214291A1 - Biphenyl derivatives used as nhe-3 inhibitors

Info

Publication number: EP1214291A1
Application number: EP00965915A
Authority: EP
Inventors: Dieter Dorsch; Peter Raddatz; Norbert Beier; Claudia Wilm
Original assignee: Merck Patent GmbH
Current assignee: Merck Patent GmbH
Priority date: 1999-09-22
Filing date: 2000-09-04
Publication date: 2002-06-19
Also published as: DE19945302A1; CZ2002815A3; KR20020033818A; SK3342002A3; AU7649700A; CA2387529A1; JP2004500338A; NO20021407L; NO20021407D0; PL355097A1; HUP0202891A2; MXPA02003087A; WO2001021582A1; BR0014199A; HUP0202891A3; CN1555356A; ZA200203095B

Abstract

The invention relates to compounds of the formula (I), wherein X, R?1, R2, R3, R4 and R5¿ have the meaning indicated in claim 1. The inventive compounds represent inhibitors of the subtype 3 sodium/proton exchanger (NHE-3).

Description

Biphenyl derivatives as NHE-3 inhibitors

The invention relates to compounds of the formula I.

wherein

R ¹ , R ⁴ each independently of one another -C (= NH) -NH ₂ , which can also be obtained simply by -COA, -CO- [C (R ⁶ ) ₂ ] _n -Ar, -COOA,

-OH or can be substituted by a conventional amino protecting group,

NH-C (= NH) -NH ₂ , -CO-N = C (NH ₂ ) ₂ ,

R ² , R ³ , R ⁵ each independently of one another H, A, OR ⁶ , N (R ⁶ ) ₂ , NO ₂ , CN, Hai, NHCOA, NHCOAr, NHSO ₂ A, NHSO ₂ Ar, COOR ^e CON (R ⁶ ) ₂ , CONHAr, COR ⁶ , COAr, S (O) _n A, S (O) _n Ar,

-0- [C (R ⁶ ) ₂ ] _m -COOR ⁶ , - [C (R ⁶ ) ₂ ] _p -COOR ⁶ , -O- [C (R ⁶ ) ₂ ] _π C COONN ((RR ⁶⁶ )) ₂₂ ,, - [[CC ((RR ⁶⁶ )) ₂₂ ]] p _p --CCOONr (R ⁶ ) ₂ , -O- [C (R ⁶ ) ₂ ] _m -CONHAr, or - [C (R ⁶ ) ₂ ] _p -CONHAr,

X - [C (R ⁶ ) ₂ ] _n -, -CR ⁶ = CR ⁶ -, - [C (R ⁶ ) ₂ ] _n -O-, -O- [C (R ⁶ ) ₂ ] _n - -COO -, -OOC-, -CONR ⁶ - or -NR ⁶ CO-,

R ^c H, A or benzyl, A alkyl with 1-20 C atoms, in which one or two CH ₂ groups can be replaced by O or S atoms or by -CR ⁶ = CR ⁶ groups and / or 1-7 H atoms by F,

Ar unsubstituted or single, double or triple by A, Ar ',

OR ⁶ , OAr ^* , N (R ⁶ ) ₂ , NO ₂ , CN, shark, NHCOA, NHCOAr ',

NHS0 ₂ A, NHSO ₂ Ar ', COOR ⁶ , CON (R ⁶ ) ₂ , CONHAr', COR ⁶ , COAr ', S (O) _n A or S (O) _n Ar' substituted phenyl or 0 naphthyl.

Ar 'unsubstituted or single, double or triple by A, OR ⁶ ,

N (R ⁶ ) ₂ , NO ₂ , CN, shark, NHCOA, COOR ⁶ , CON (R ⁶ ) ₂ , COR ⁶ _. , - or S (O) _n A substituted phenyl or naphthyl,

Shark F, Cl, Br or I,

n 0, 1 or 2,

20

m 1 or 2,

p means 1 or 2,

25 and their salts and solvates as NHE-3 inhibitors

Other inhibitors of the Natnum / proton exchanger subtype 3 are described, for example, in EP 0 825 178

30

The invention was based on the task of finding new compounds with valuable properties, in particular those which can be used for the production of medicaments

35 DE 19819548 describes that the compounds of the formula I and their salts have factor Xa inhibitory properties and therefore for combating and preventing thromboembolic disorders such as thrombosis, myocardial infarction, arthroscopic sclerosis, inflammation, apoplexy, angina pectons, restenosis after angioplasty and Intermittent claudication can be used

Surprisingly, it was found that the compounds of the formula I and their salts inhibit the Natπum / Protoπen exchanger subtype 3 with good tolerability

The compounds of formula I can be used as active pharmaceutical ingredients in human and veterinary medicine

It is known that the Na ⁺ / H ⁺ exchanger is a family with at least 6 different isoforms (NHE-1 to NHE-6), all of which are now cloned, while the subtype NHE-1 is ubiquitous throughout the body in all Tissue is distributed, the other NHE subtypes are selectively expressed in specific organs such as in the kidney or in the lumen wall and contraluminal wall of the small intestine. This distribution reflects the specific functions that the various isoforms serve, namely the regulation of the intracellular pH and cell volume through the subtype NHE-1 and on the other hand Na ⁺ uptake and reuptake in the intestine and kidney through the isoforms NHE-2 and NHE-3. The isoform NHE-4 was mainly found in the stomach Expression of NHE-5 is restricted to the brain and neuronal tissue. NHE-6 is the isoform that forms the sodium proton exchanger in the mitochondrion

The isoform NHE-3 is expressed in particular in the apical membrane of the proximal kidney tubules. An NHE-3 inhibitor therefore exerts a protective effect on the kidneys

The therapeutic use of a selective inhibitor for NHE-3

Isoforming is versatile NHE-3 inhibitors inhibit or reduce tissue damage and cell necrosis after pathophysiological hypoxic and ischemic events that activate NHE activity such as this during renal ischemia or during kidney transplant removal, transport and reperfusion of a kidney

The compounds of formula I lower blood pressure and are suitable as active pharmaceutical ingredients for the treatment of hypertension. They are also suitable as diuretics

The compounds of the formula I, alone or in conjunction with NHE inhibitors of other subtype specificity, have an anti-ischemic effect and can be used for thrombosis, atherosclerosis, vascular spasms, for the protection of

Organs, e.g. kidney and liver, before and during operations, as well as for chronic or acute kidney failure

Furthermore, they can be used to treat stroke, the brain modem, ischemia of the nervous system, various forms of shock and to improve the respiratory drive in, for example, the following conditions, central sleep apnea, sudden child death, postoperative hypoxia and other respiratory disorders

The combination with a carbonic anhydrase inhibitor can further improve breathing activity. The compounds of the formula I have an inhibiting effect on the pro ferations of cells, for example fibroblast cell proliferation and the proliferation of smooth vascular muscle cells and can therefore be used to treat diseases can be used in which cell reproduction is a primary or secondary cause. The compounds of the formula I can be used against diabetic late complications, cancer, fibrotic diseases, endothelial dysfunction, organ hypertrophy and hyperplasia, in particular in the case of prostate hyperplasia or prostate hypertrophy. They are also suitable as diagnostics for determining and differentiating certain forms of hypertension, atherosclerosis, diabetes and proliferative diseases

Since the compounds of the formula I also have an advantageous effect on the level of the serum lipoproteins, they can be used alone or in combination with other medicaments for the treatment of an elevated blood lipid level The invention relates to the use of compounds of the formula I as claimed in claim 1 and their physiologically acceptable salts and / or solvates for the manufacture of a medicament for the treatment of thromboses, ischemic conditions of the heart, peripheral and central nervous system and stroke, ischemic conditions of peripheral organs and limbs and for the treatment of shock conditions.

The invention further relates to the use of compounds of the formula I according to claim 1 and their physiologically acceptable salts and / or solvates for the production of a medicament for use in surgical operations and organ transplants and for the preservation and storage of transplants for surgical measures.

The invention also relates to the use of compounds of

Formula I according to claim 1 and its physiologically acceptable salts and / or solvates for the manufacture of a medicament for the treatment of diseases in which cell proliferation is a primary or secondary cause, for the treatment or prophylaxis of disorders of the fat metabolism or impaired respiratory drive.

The invention further relates to the use of compounds of the formula I according to claim 1 and their physiologically acceptable salts and / or solvates for the manufacture of a medicament for the treatment of ischemic kidney, ischemic bowel disease or for the prophylaxis of acute or chronic kidney disease.

Methods for identifying substances that inhibit the type 3 sodium / proton exchanger are e.g. in US 5,871,919.

For all radicals in the compounds of the formula I which occur more than once, such as R ⁶ , the meanings are independent of one another.

Hydrates and solvates are e.g. the hemi, mono- or dihydrates, among solvates e.g. Alcohol addition compounds such as With

Methanol or ethanol. In the above formulas, A means alkyl, is linear or branched, and has 1 to 20, preferably 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or

12 carbon atoms. A is preferably methyl, furthermore ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl or tert-butyl, further also pentyl, 1-, 2- or 3-methylbutyl, 1, 1-, 1, 2 - or 2,2-dimethylpropyl, 1-ethylpropyl, hexyl, 1-, 2-, 3- or 4-methylpentyl, 1, 1-, 1, 2-, 1, 3-, 2,2-, 2 , 3- or 3,3-dimethylbutyl, 1- or 2-ethylbutyl, 1-ethyl-1-methylpropyl, 1-ethyl-2-methylpropyl, 1, 1, 2- or 1, 2,2-trimethylpropyl, heptyl, Octyl, nonyl or decyl.

A also means e.g. Trifluoromethyl, pentafluoroethyl, allyl or crotyl.

COR ⁶ is acyl and preferably means formyl, acetyl, propionyl, but also butyryl, pentanoyl or hexanoyl. COOR ⁶ preferably means methoxycarbonyl, ethoxycarbonyl, propoxycarbonyl or butoxycarbonyl.

Shark preferably means F, Cl or Br, but also I.

R ² , R ³ and R ⁵ each represent, independently of one another, preferably H,

Fluorine, chlorine, bromine, iodine, hydroxy, methoxy, ethoxy, propoxy, nitro, amino, methylamino, dimethylamino, ethylamino, diethylamino, acetamido, sulfonamido, methylsulfonamido, phenyisulfonamido, methylthio, ethylthio, methylsulfinyl, ethylsulfinyl, ethylsulfonyl Phenylsulfinyl, phenylsulfonyl, cyan, carboxy, methoxycarbonyl, ethoxycarbonyl, carboxymethoxy, methoxycarbonylmethoxy, carboxymethyl, methoxycarbonylmethyl, aminocarbonylmethoxy, aminocarbonylmethyl, N-phenylaminocarbonylmethoxy, N-phenylaminocarbonylmethyl, also acyl.

9 c In particular, R, R mean H.

R ³ in particular denotes, for example, H, COOA or -OCH ₂ COOR ⁶ , where R ⁶ denotes H or alkyl having 1-4 C atoms.

R ^δ denotes H, A or benzyl, but especially H or alkyl with 1-4 C atoms.

X preferably denotes, for example, -CH ₂ -, -CH = CH-, -CH ₂ O-, -O-CH ₂ -, -COO-, -OOC-, -CONH- or -NHCO-; -CH ₂ O-, -O- CH ₂ - or -CH ₂ -CH ₂ - is very particularly preferred.

Ar preferably denotes unsubstituted phenyl or naphthyl, further preferably e.g. by A, fluorine, chlorine, bromine, iodine, hydroxy, methoxy, ethoxy, propoxy, butoxy, pentyloxy, hexyloxy, benzyloxy, phenethyloxy, methylthio, ethylthio, methylsulfinyl, ethylsulfinyl, methylsulfonyl, ethylsulfonyl, phenylsuifinyl, phenylsulfonyl, nitro, amino , Methylamino, ethylamino, dimethylamino, diethylamino, formamido, acetamido, propionylamino, butyrylamino, methylsulfonamido, ethylsulfonamido, propylsulfonamido, butylsulfonamido, phenylsulfonamido, (4-methylphenyl) sulfonethoxy, methoxy, carboxy - Thoxycarbonylethoxy, hydroxymethoxy, hydroxyethoxy, methoxyethoxy, carboxy, methoxycarbonyl, ethoxycarbonyl, cyano, phenylaminocarbonyl, acyl or benzoyl mono-, di- or tri-substituted phenyl or naphthyl, and also biphenyl.

Ar therefore preferably means, for example, o-, m- or p-tolyl, o-, m- or p-ethylphenyl, o-, m- or p-propylphenyl, o-, m- or p-isopropylphenyl, o-, m- or p-tert-butylphenyl, o-, m- or p-hydroxyphenyl, o-, m- or p-nitrophenyl, o-, m- or p-aminophenyl, o-, m- or p- (N-methylamino) phenyl, o-, m- or p-acetamidophenyl, o-, m- or p-methoxyphenyl, o-, m- or p-ethoxyphenyl, o-, m- or p-carboxyphenyl, o- , m- or p-methoxycarbonylphenyl, o-, m- or p- (N, N-dimethylamino) -phenyl, o-, m- or p- (N-ethylamino) -phenyl, o-, m- or p- (N, N-diethylamino) phenyl, o-, m- or p-acetylphenyl, o-, m- or p-formylphenyl, o-, m- or p-fluorophenyl, o-, m- or p- Bromophenyl, o-, m- or p- chlorophenyl, o-, m- or p-methylsulfonylphenyl, o-, m- or p- (phenylsulfonamido) phenyl, o-, m- or p- (methylsulfonamido) - phenyl, o-, m- or p-methylthiophenyl, more preferably 2,3-, 2,4-, 2,5-, 2,6-, 3,4- or 3,5-difluorophenyl, 2,3-, 2,4-, 2,5-, 2,6-, 3,4- or 3,5-dichlorophenyl, 2,3-, 2,4-, 2,5-, 2,6-, 3,4- or 3,5-dibromophenyl, 2,4- or 2,5-dinitrophenyl, 2,5- or 3,4-dimethoxyphenyl, 3-nitro-4 -chlorophenyl, 3-amino-4-chloro, 2-amino-3-chloro, 2-amino-4-chloro, 2-amino-5-chloro or 2-amino-6-chlorophenyl, 2-nitro -4-N, N-dimethylamino- or 3-nitro-4-N, N-dimethylaminophenyl, 2,3-diaminophenyl, 2,3,4-, 2,3,5-, 2,3,6- , 2,4,6- or 3,4,5-thchlorophenyl, 2,4,6-trimethoxy phenyl, 2-hydroxy-3,5-dichlorophenyl, p-iodophenyl, 3,6-dichloro-4-aminophenyl, 4-fluoro-3-chlorophenyl, 2-fluoro-4-bromophenyl, 2,5-difluoro 4-bromophenyl, 3-bromo-6-methoxyphenyl, 3-chloro-6-methoxyphenyl, 3-chloro-4-acetamidophenyl, 3-fluoro-4-methoxyphenyl, 3-amino-6-methylphenyl, 3-chloro 4-acetamidophenyl or 2,5-dimethyl-4-chlorophenyl.

Ar 'means in particular e.g. Phenyl or naphthyl, further preferably e.g. o-, m- or p-tolyl, o-, m- or p-ethylphenyl, o-, m- or p-propylphenyl, o-, m- or p-isopropylphenyl, o-, m- or p- tert-butylphenyl, o-, m- or p-hydroxyphenyl. o-, m- or p-nitrophenyl, o-, m- or p-aminophenyl, o-, m- or p- (N-methylamino) phenyl, o-, m- or p-acetamidophenyl, o-, m - or p-methoxyphenyl, o-, m- or p-ethoxyphenyl, o-, m- or p-carboxyphenyl, o-, m- or p-methoxycarbonylphenyl, o-, m- or p- (N, N-dimethylamino ) -phenyl, o-, m- or p- (N-ethylamino) -phenyl, o-, m- or p- (N, N-diethyiamino) -phenyl, o-, m- or p-acetylphenyl, o- , m- or p-formylphenyl, o-, m- or p-fluorophenyl, o-, m- or p-bromophenyl, o-, m- or p-chlorophenyl or o-, m- or p-methylsulfonylphenyl.

Accordingly, the invention relates in particular to the use of those compounds of the formula I in which at least one of the radicals mentioned has one of the preferred meanings indicated above. Some preferred groups of compounds can be expressed by the following sub-formulas la to li, which correspond to the formula I and in which the radicals not specified have the meaning given for the formula I, but in which

in la R ¹ , R ⁴ each independently of one another —C (= NH) —NH ₂ , which can also be simply substituted by OH or —CO — N = C (NH ₂ ) ₂ ; in Ib R ² , R ^{5 are} H; in Ic R ¹ , R ⁴ each independently of one another —C (= NH) —NH ₂ , which can also be simply substituted by OH or

-CO-N = C (NH ₂ ) ₂ , R ² , R ⁵ H and

R ^{3 is} H or COOR ⁶ , in Id R ¹ , R ⁴ each independently of one another —C (= NH) —NH ₂ , which can also be simply substituted by OH or

-CO-N = C (NH ₂ ) ₂ , R ² , R ⁵ H and

R ^{3 is} H, COOR ⁶ or -O- (CH ₂ ) COOR ⁶ , in le X is -CH ₂ -O- or -O-CH ₂ -, in If R ¹ , R ^{4 are} each independently of one another -C ( = NH) -NH ₂ , which can also be simply substituted by OH or -CO-N = C (NH ₂ ) ₂ , R ² , R ⁵ H,

R ³ H or COOR ⁶ and

X denotes -CH ₂ -0- or -O-CH ₂ -, in Ig R ¹ , R ⁴ each independently of one another -C (= NH) -NH ₂ , which can also be simply substituted by OH or -CO-N = C (NH ₂ ) ₂ , R ² , R ⁵ H,

R ³ H, COOR ⁶ or -O- (CH ₂ ) COOR ⁶ , and

-CH2-O-, -O-CH2- or -CH ₂ -CH ₂ - mean, in Ih R ¹ , R ⁴ each independently of one another -C (= NH) -NH ₂ , which can also be simply substituted by OH or -CO-N = C (NH ₂ ) ₂ , R ² , R ⁵ H,

R ³ H, COOR ⁶ , -O-CH ₂ -COOR ⁶ , CH ₂ -COOR ⁶ , -O-CH ₂ - CON (R ⁶ ) ₂ , CH ₂ -CON (R ⁶ ) ₂ , -O-CH ₂ -CONHAr or

CH ₂ -CONHAr, X -CH ₂ -O-, -O-CH2- or -CH ₂ -CH ₂ -,

R ⁶ H or A,

A alkyl with 1-4 C atoms, in Li R ¹ , R ⁴ each independently of one another —C can also be simply substituted by OH or

-CO-N = C (NH ₂ ) ₂ ,

R ² , R ⁵ H,

R ^J H, COOR ⁶ , -O-CH ₂ -COOR ⁶ , CH ₂ -COOR ⁶ , -O-CH ₂ -

CON (R ⁶ ) ₂ , or CH ₂ -CON (R ⁶ ) ₂ ,

X -CH ₂ -O-, -O-CH ₂ - or -CH ₂ -CH ₂ -,

R ⁶ H or A,

A is alkyl with 1-4 carbon atoms

The compounds of the formula I and also the starting materials for their preparation are otherwise prepared by methods known per se, as described in the literature (for example in the standard works such as Houben-Weyl, methods of organic chemistry, Georg-Thieme-Verlag, Stuttgart ) are described, namely under reaction conditions which are known and suitable for the reactions mentioned. Use can also be made of variants which are known per se and are not mentioned here

If desired, the starting materials can also be formed in situ, so that they are not isolated from the reaction mixture, but instead are immediately reacted further to give the compounds of the formula I.

Compounds of formula I can preferably be obtained by liberating compounds of formula I from one of their functional derivatives by treatment with a solvolysing or hydrogenolysing agent Preferred starting materials for solvolysis or hydrogenolysis are those which otherwise correspond to the formula I, but instead of one or more free amino and / or hydroxyl groups contain corresponding protected ammo- and / or hydroxyl groups, preferably those which instead of an H atom , which is connected to an N atom, carry an amino protective group, in particular those which carry an R'-N group instead of an HN group, in which R 'denotes an amino protective group, and / or those which replace the H atom a hydroxyl group carry a hydroxyl protective group, for example those which correspond to the formula I, but instead of a group -COOH carry a group -COOR ", in which R" denotes a hydroxyl protective group

Preferred starting materials are also the oxadiazoldenvate, which can be converted into the corresponding amidino compounds

The oxadiazole group can be introduced, for example, by implementing the

Cyan compounds with hydroxylamine and reaction with phosgene, dialkyl carbonate, chloroformate, N, N'-carbonyldιιmιdazol or Acetanhydnd

There may also be several - identical or different - protected amino and / or hydroxyl groups in the molecule of the starting material. If the protective groups present are different from one another, they can be split off selectively in many cases

The term "amino protecting group" is generally known and refers to groups which are suitable for protecting (blocking) an amino group from chemical reactions, but which are easily removable after the desired chemical reaction has been carried out elsewhere in the molecule unsubstituted or substituted acyl, aryl, aralkoxymethyl or aralkyl groups are particularly suitable for such groups. Since the amino protective groups are removed after the desired reaction (or reaction sequence), their type and size are otherwise not critical, but those with 1 are preferred -20, in particular 1-8 carbon atoms The term "acyl group" is to be understood in the broadest sense in connection with the present process. It encompasses ahphatic, araliphatic, aromatic or hetero- cyc's carboxylic acids or suifonic acids derived acyl groups and in particular alkoxycarbonyl, aryloxycarbonyl and especially aralkoxycarbonyl groups. Examples of such acyl groups are alkanoyl such as acetyl, propionyl, butyryl, aralkanoyl such as phenylacetyl, aroyl such as benzoyl or tolanyloxycarbonyl, aryloxy, such as aryloxy such as methoxycarbonyl, ethoxycarbonyl, 2,2,2-trichloroethoxycarbonyl, BOC (tert-butyloxycarbonyl), 2-iodoethoxycarbonyl, aralkyloxycarbonyl such as CBZ ("carbobenzoxy"), 4-methoxybenzyloxycarbonyl, FMOC, arylsulfonyl such as Mtr. Preferred amino protective groups are BOC and Mtr, also CBZ, Fmoc, Benzyl and Acetyl

The term "hydroxyl protecting group" is also generally known and refers to groups which are suitable for protecting a hydroxyl group against chemical reactions, but which are easily removable after the desired chemical reaction has been carried out elsewhere in the molecule such groups are the unsubstituted or substituted aryl, aralkyl or acyl groups mentioned above, and also alkyl groups. The nature and size of the hydroxyl-protecting groups is not critical since they are preferred after the desired chemical reaction or reaction sequence

Groups with 1-20, in particular 1-10, carbon atoms. Examples of hydroxyl protecting groups include benzyl, p-nitrobenzoyl, p-toluenesulfonyl, tert-butyl and acetyl, with benzyl and tert-butyl being particularly preferred

Depending on the protective group used, the compounds of formula I can be liberated from their functional derivatives, for example with strong acids, expediently with TFA or perchloric acid, but also with other strong inorganic acids such as hydrochloric acid or strong organic strong sulfuric acid Carboxylic acids such as tchloric acetic acid or sulfonic acids such as benzene or p-toluenesulfonic acid. The presence of an additional inert solvent is possible, but not always necessary. Organic solvents, for example carboxylic acids such as acetic acid, ethers such as tetrahydrofuran or dioxane, amides such as DMF, are preferably suitable as inert solvents. halogenated hydrocarbons such as dichloromethane, also alcohols such as methanol, ethanol or isopropanol, and water

Mixtures of the abovementioned solvents are also suitable for TFA is preferably used in excess without the addition of another solvent, perchloric acid in the form of a mixture of acetic acid and 70% perchloric acid in a ratio of 9: 1. The reaction temperatures for the cleavage are advantageously between about 0 and about 50 °, preferably between 15 and 30 ° (room temperature).

The groups BOC, OBut and Mtr can e.g. B. preferably with TFA in dichloromethane or with about 3 to 5N HCl in dioxane at 15-30 °, the FMOC group with an about 5 to 50% solution of dimethylamine, diethylamine or piperidine in DMF at 15 -30 °.

Protective groups which can be removed hydrogenolytically (for example CBZ, benzyl or the release of the amidino group from their oxadiazole derivative) can, for. B. by treatment with hydrogen in the presence of a catalyst (z. B. a noble metal catalyst such as palladium, conveniently on a support such as coal or like wet Raney nickel with the addition of e.g. acetic acid). Suitable solvents are the above, especially z. B. alcohols such as methanol or ethanol or amides such as DMF. The hydrogenolysis is generally carried out at temperatures between about 0 and 100 ° and pressures between about 1 and 200 bar, preferably at 20-30 ° and 1-10 bar. Hydrogenolysis of the CBZ group succeeds e.g. B. good on 5 to 10% Pd / C in methanol or with ammonium formate (instead of hydrogen) on Pd / C in methanol / DMF at 20-30 °.

Compounds of the formula I in which R ¹ and R ^{4 are} -C (= NH) -NH ₂ can preferably be obtained from the corresponding cyano compound. The conversion of a cyano group into an amidino group takes place by reaction with, for example, hydroxylamine and subsequent reduction of the N-

Hydroxyamidine with hydrogen in the presence of a catalyst such as e.g. Pd / C or Raney nickel.

To produce an amidine of the formula I (R ¹ = -C (= NH) -NH ₂ ), ammonia can also be added to a nitrile of the formula I (R ¹ = CN). The addition is preferably carried out in several stages by, in a manner known per se, a) converting the nitrile with H ₂ S into a thioamide which is mixed with an alkyl agent, for example CH ₃ I, is converted into the corresponding S-alkyl imidothioester, which in turn reacts with NH ₃ to form the amidine, b) converts the nitrile with an alcohol, for example ethanol in the presence of HCl, into the corresponding imido ester and this with ammonia treated, or c) reacting the nitrile with lithium bis (trimethylsilyl) amide and then hydrolyzing the product.

The cyan compounds are prepared by methods known per se.

Compounds of the formula I where n are R ¹ and R ^{4 are} -CON (= NH) -NH ₂ can preferably be obtained from the corresponding alkoxycarbonyl compounds by reacting with guanidine.

It is also possible to convert a compound of the formula I into another compound of the formula I by one or more radicals, R ¹ , R ² , R ³ , R ⁴ and / or R ⁵ into one or more radicals (e) converting R ¹ , R ² , R ³ , R ⁴ and / or R ⁵ , for example by acylating an amino group or nitro groups (for example by hydrogenation on Raney nickel or Pd carbon in an inert solvent such as methanol or ethanol) to

Amino groups reduced.

Esters can e.g. are saponified with acetic acid or with NaOH or KOH in water, water-THF or water-dioxane at temperatures between 0 and 100 °.

Furthermore, free amino groups can be acylated in the usual way with an acid chloride or anhydride or alkylated with an unsubstituted or substituted alkyl halide, advantageously in an inert solvent such as dichloromethane or THF and / or in the presence of a base such as triethylamine or pyridine at temperatures between -60 and + 30 °.

The reaction is usually carried out in an inert solvent, in the presence of an acid-binding agent, preferably an alkali metal or alkaline earth metal hydroxide, carbonate or bicarbonate or another Salt of a weak acid of the alkali or alkaline earth metals, preferably of potassium, sodium, calcium or cesium. The addition of an organic base such as methylamine, dimethylaniline, pyπdin or quinoline or an excess of the amine component of the formula II or the alkylation derivative of the formula III can also be advantageous Depending on the conditions used, the reaction time is between a few minutes and 14 days, the reaction temperature is between about 0 ° and 150 °, normally between 20 ° and 130 °

Suitable inert solvents are, for example, hydrocarbons such as hexane, petroleum ether, benzene, toluene or xylene, chlorinated hydrocarbons such as trichlorethylene, 1,2-dichloroethane, carbon tetrachloride, chloroform or dichloromethane, alcohols such as methanol, ethanol, isopropanol, n-propanol, n- Butanol or tert-butanol, ethers such as diethyl ether, dnsopropyl ether, tetrahydrofuran (THF) or dioxane, glycol ethers such as ethylene glycol monomethyl or monoethyl ether (methyl glycol or ethyl glycol), ethylene glycol dimethyl ether (diglyme), ketones such as acetone or butanone, amides such as acetamide, dimethylacetamide, N-methylpyrrolidone (NMP) or dimethylformamide (DMF), nitls such as acetonitrile, suifoxides such as dimethyl sulfoxide (DMSO) carbon disulphide, carboxylic acids such as formic acid or acetic acid nitro compounds such as nitromethane or nitro benzol or nitro benzol

A base of the formula I can be converted into the associated acid addition salt using an acid, for example by reacting equivalent amounts of the base and the acid in an inert solvent such as ethanol and subsequent evaporation. Acids, the physiologically acceptable salts, are particularly suitable for this reaction In this way, inorganic acids can be used, for example sulfuric acid, nitric acid, hydrohalic acids such as hydrochloric acid or hydrobromic acid, phosphoric acids such as orthophosphoric acid, sulfamic acid, and also organic acids, in particular aliphatic, alicyclic, araliphatic, aromatic or heterocyclic mono- or polybasic carbon-, sulfonic - or sulfuric acids, e.g. formic acid, acetic acid, propionic acid, pivalic acid, diethyl acetic acid,

Malonic acid, succinic acid, pimeline acid, fumaric acid, maleic acid, Lactic acid, tartaric acid, malic acid, citric acid, gluconic acid, ascorbic acid, nicotinic acid, isonicotinic acid, methane or ethanesulphonic acid, ethanedisulphonic acid, 2-hydroxyethanesulphonic acid, benzenesulphonic acid, p-toluenesulphonic sulphonic acid, naphthenesulphonic acid mono-, sulfuric acid -sulphonic acid -sulphonic acid -sulphonic acid -sulphonic acid -sulphonic acid -sulphonic acid -sulphonic acid- harmless acids, for example picrates, can be used to isolate and / or purify the compounds of the formula I.

On the other hand, compounds of the formula I with bases (for example sodium or potassium hydroxide or carbonate) can be converted into the corresponding metal, in particular alkali metal or alkaline earth metal, or into the corresponding ammonium salts

Physiologically acceptable organic bases, such as ethanolamine, can also be used

The invention furthermore relates to the use of the compounds of the formula I as NHE-3 inhibitors and / or their physiologically acceptable salts for the production of pharmaceutical preparations, in particular by a non-chemical route. Together with at least one solid, liquid and / or semi-liquid carrier or auxiliary and optionally in combination with one or more other active ingredients in a suitable dosage form

The invention further relates to pharmaceutical preparations containing at least one NHE-3 inhibitor of the formula I and / or one of its physiologically acceptable salts and solvates

These preparations can be used as pharmaceuticals in human or veterinary medicine. Suitable carrier substances are organic or inorganic substances which are suitable for enteral (for example oral), parenteral or topical application and do not react with the new compounds, for example water, vegetable oils, benzyl alcohols, alkylene glycols, polyethylene glycols, glycene triacetate, gelatin, carbohydrates such as lactose or starch, magnesium stearate, talc, petroleum jelly, tablets, pills, dragees, capsules, powder granules, syrups, juices or drops are used for oral use, for rectal use On- Use Suppositoπen, for parenteral application solutions, preferably ohmic or aqueous solutions, further suspensions, emulsions or implants, for topical application ointments, creams or powders, or transdermally in patches. The new compounds can also be lyophilized and the lyophilized products obtained, for example Production of injection preparations can be used. The specified preparations can be sterilized and / or auxiliaries such as lubricants, preservatives, stabilizers and / or wetting agents, emulsifiers, salts for influencing the osmotic pressure, buffer substances, coloring, taste and / or contain several other active ingredients, for example one or more vitamins

Suitable pharmaceutical preparations for administration in the form of aerosols or sprays are, for example, solutions, suspensions or emulsions of the active ingredient of the formula I in a pharmaceutically acceptable solvent

The compounds of formula I and their physiologically acceptable salts and solvates can be used for the treatment and / or prophylaxis of the diseases or conditions described above

The substances according to the invention are generally preferably administered in doses between about 0.1 and 100 mg, in particular between 1 and 10 mg, per dosage unit. The daily dosage is preferably between about 0.001 and 10 mg / kg body weight. The specific dose for each patient depends However, it depends on a variety of factors, for example on the effectiveness of the special compound used, on the age, body weight, general health, gender, on the diet, on the time and route of administration, on the rate of excretion, combination of drugs and the severity of the respective disease, which the therapy applies the oral

Application is preferred

Above and below, all temperatures are given in ° C. In the examples below, "customary working up" means adding water if necessary, adjusting, if necessary, to pH values between 2 and 10, depending on the constitution of the end product, extracted with ethylamine acetate or dichloromethane, separates, dries the organic phase over sodium sulfate, evaporates and purifies by chromatography on silica gel and / or by crystallization

Mass spectrometer (MS) El (electron impact ionization) M ⁺ FAB (Fast Atom Bombardment) (M + H) ⁺

example 1

A solution of 2.06 g of 3-bromobenzone and 1.50 g of 3-tolylboronic acid in 50 ml of dimethoxyethane is mixed with 247 mg of palladium (II) acetate, 335 mg of Tπ-o-tolylphosphine, 20 ml of water and 954 mg of anhydrous sodium carbonate added and heated with stirring for 18 hours at 100 ° C. The mixture is worked up in the customary manner, the residue is chromatographed on a silica gel column using petroleum ether / ethyl acetate 9 1 and 3'-methylbiphenyl-3-carbonιtrιl is obtained as a colorless solid ("A"), El 193 A solution of 1.17 g "A" in 10 ml carbon tetrachloride is made with 1.09 g

N-bromosuccinic (NBS) and 60 mg of azobisisobutyronitrile are added and the mixture is heated at 70 ° C. for 18 hours. The mixture is worked up in the customary manner, the residue is chromatographed on a column of silica gel with petroleum ether / ethyl acetate 9 l and 3'-bromomethylbiphenyl-3-carbontril is obtained as a colorless solid ("B") A solution of 500 mg "B" and 238 mg 3-Hydroxybenzonιtrιl in 10 ml

Acetonitnl is mixed with 652 mg casium carbonate and stirred for 40 hours at room temperature. After working up, the residue is chromatographed on a reversed-phase column with acetonitrile / water 65 35. A 3 '- (3-cyanophenoxymethyl) - bιphenyl-3- is obtained as a colorless solid. carbonιtrιl ("C"), FAB 311

A solution of 90 mg of "C" and 125 mg of hydroxylammonium chloride in 10 ml of ethanol is mixed with 1.2 g of polymer-bound dimethylaminopyridine (DMAP) and stirred for 18 hours at room temperature. The solvent is filtered off and N-hydroxy-3 '- [ 3- (N-hydroxy-carbamimidoyl) phenoxymethyl] biphenyl-3-carboxamid ("D") as a colorless solid, FAB 377

A solution of 76 mg "D" in 10 ml of methanol is mixed with 100 mg of water-moist Raney nickel and 30 mg of acetic acid and hydrogenated for 18 hours at room temperature and normal pressure. The mixture is filtered off, the solvent is removed and 3 '- (3-carbamimidoyl- phenoxymethyl) - bιphenyl-3-carboxamιdιn, acetate, El 327 (M ^τ - NH ₃ ), 310 (M ⁺ - 2 NH ₃ )

The connections are obtained analogously

3 '- (3-carbamimidoyl-phenoxymethyl) biphenyl-4-carboxamidine, D acetate, FAB 345;

3 '- (4-carbamimidoyl-phenoxymethyl) biphenyl-4-carboxamidine, D acetate, FAB 345;

3 '- (4-carbamimidoyl-phenoxymethyl) biphenyl-3-carboxamidine, D acetate, FAB 345;

4 '- (4-carbamimidoyl-phenoxymethyl) biphenyl-4-carboxamidine,

4 '- (4-carbamimidoyl-phenoxymethyl) biphenyl-3-carboxamidine,

4 '- (3-carbamimidoyl-phenoxymethyl) biphenyl-3-carboxamidine and

4 '- (3-carbamimidoyl-phenoxymethyl) biphenyl-4-carboxamidine.

Example 2

Analogously to Example 1, 3-bromobenzonitrile is also used to react

3-methoxyphenylboronic acid the compound 3'-methoxybiphenyl-3-carbonitrile.

Subsequent ether cleavage with aluminum triiodide in acetonitrile and

Reaction with 3-bromomethyl-benzonithl gives 3 '- (3-cyano-benzyloxy) -biphenyl-3-carbonithl.

By reaction with hydroxylamine and reduction with hydrogen under

Ra-Ni catalysis gives 3 '- (3-carbamimidoyl-benzyloxy) biphenyl-3-carboxamidine

You get analog 4 '- (4-Carbamιmιdoyl-benzyloxy) -bιphenyl-4-carboxamιdιn, diacetate, FAB

345,

4 '- (3-Carbamιmιdoyl-benzyloxy) -bιphenyl-4-carboxamιdιn, diacetate, FAB

345

Example 3

Analogously to Example 1, reaction of 3-cyanophenylboronic acid with 3-bromo-5-methyl-benzoic acid methyl ester gives the compound 3'-cyan-5-methyl-biphenyl-3-carboxylic acid methyl ester by bromination with

NBS and reaction with 3-Hydroxybenzonιtrιl gives 3'-cyano-5- (3-cyanophenoxymethyl) -bιphenyl-3-carboxylic acid methyl ester reaction with hydroxylamine and reduction with H ₂ / Ra-Ni gives the compound 3'-carbamιmιdoyl-5 - (3-carbamιmιdoyl-phenoxymethyl) -bιphenyi-3-carboxylic acid methyl ester

Saponification of the ester with aqueous NaOH gives 3'-

Carbamιmιdoyl-5- (3-carbamιmιdoyl-phenoxymethyl) -bιphenyl-3-carboxylic acid

Chromatography on a reversed-phase column with an acetonitrile / water / TFA mixture gives 3'-carbamimidoyl-5- (3-carbamimidoyl-phenoxymethyl) bisphenyl-3-carboxylic acid, bisfluoroacetate

The connections are obtained analogously

4'-Carbamιmιdoyl-4- (4-carbamιmιdoyl-phenoxymethyl) -bιphenyl-3-carboxylic acid methyl ester, FAB 403,

4'-Carbamιmιdoyl-4- (3-carbamιmιdoyl-phenoxymethyl) -bιphenyl-2-carboxylic acid methyl ester, FAB 403, 3'-Carbamimidoyl-4- (4-carbamimidoyl-phenoxymethyl) biphenyl-2-carboxylic acid methyl ester, FAB 403;

3'-Carbamimidoyl-4- (3-carbamimidoyl-phenoxymethyl) biphenyl-2-carboxylic acid methyl ester, FAB 403;

4'-Carbamimidoyl-5- (3-carbamimidoyl-phenoxymethyl) biphenyl-4-carboxylic acid methyl ester, FAB 403;

3'-Carbamimidoyl-5- (3-carbamimidoyl-phenoxymethyl) biphenyl-4-carboxylic acid methyl ester, FAB 403.

Example 4

Analogously to Example 1, 3'-methylbiphenyl-3-carboxylic acid methyl ester is obtained by reacting methyl 3-bromobenzoate with 3-tolylboronic acid. By bromination with NBS and reaction with 3-hydroxybenzoic acid methyl ester, 3 '- (3-methoxycarbonyl-phenoxymethyl) biphenyl-3-carboxylic acid methyl ester is obtained therefrom. By reaction with guanidine hydrochloride in methanolic sodium methaolate solution, N- [3 '- (3-guanidinocarbonylphenoxymethyl) biphenyl-3-carbonyl] guanidine is obtained therefrom

The compound N- [4 '- (4-guanidinocarbonylphenoxymethyl) biphenyl-4-carbonyl] guanidine is obtained analogously. Example 5

A solution of 7.0 g of 3-bromo-5-methylphenol and 5.97 g of methyl bromoacetate and 13 g of casium carbonate in 100 ml of acetonitrile is stirred at room temperature overnight. After the usual work-up, 9.70 g (3-bromo 5-methylphenoxy) -essιgsauremethylester ("AB") A suspension of 2.0 g "AB", 100 mg tetrakιs (trιphenylphosphιn) - palladium and 0.85 g of sodium carbonate in 50 ml of toluene is heated to boiling. Then a solution of 2.94 g of 3-cyanophenylboronic acid in 30 ml of methanol was added dropwise and the mixture was heated under reflux for 14 hours. Working up as usual gives 2.17 g of (3'-cyano-5-methyl-biphenyl-3-yloxy) methyl acetate ("AC ")

A solution of 1.2 g of "AC" and 0.765 g of NBS in 50 ml of carbon tetrachloride is irradiated with UV light at room temperature. After conventional work-up, 1.54 g (3'-cyano-5-bromomethyl-biphenyl-3 -yloxy) - methyl acetate ("AD")

A solution of 185 mg of "AD", 63.1 mg of 4-hydroxybenzontrium and 172.7 mg of casium carbonate in 10 ml of acetonitrile is stirred at room temperature for 4 days. After the usual work-up, [3'-cyan-5- (4-cyanophenoxymethyl) is obtained. -bιphenyl-3-yloxy] -essιgsauremethylester ("AE")

A solution of 60 mg of "AE", 69.5 mg of hydroxylammonium chloride and 101 mg of triethylamine in 10 ml of methanol is refluxed for 14 hours. After the solvent has been removed, the mixture is taken up in water. The mixture is separated off and 70 mg of [3'-N-hydroxyamido] are obtained -5- (4-N-hydroxyamιdιno- phenoxymethyl) -bιphenyl-3-yloxy] -essιgsauremethylester ("AF") By

Reduction with H ₂ / Raney nickel is obtained therefrom [3'-amino-5- (4-amino-phenophenoxymethyl) -phenyl-3-yloxy] -essιgsauremethylester, FAB 433

The connections are obtained analogously

[4'-Amιdιno-5- (4-amιdιnophenoxymethyl) -bιphenyl-3-yloxy] - methyl acetate, FAB 433

[3'-Amιdιno-5- (3-amιdιnophenoxymethyl) -bιphenyl-3-yloxy] - methyl acetate, FAB 433

[4'-Amιdιno-5- (3-amιdιnophenoxymethyl) -bιphenyl-3-yloxy] - methyl acetate, FAB 433

If in the first stage methyl bromoacetate is replaced by tert-butyl bromoacetate, the tert-butyl esters obtained in the last stage can be cleaved with trifluoroacetic acid and the corresponding carboxylic acids are obtained

[3'-Amιdιno-5- (4-amιdιnophenoxymethyl) -bιphenyl-3-yloxy] - acetic acid, bistrifluoroacetate, FAB 419,

[4'-amino-5- (4-amino-phenoxymethyl) -phenyl-3-yloxy] - acetic acid, [3'-amino-5- (3-amino-phenoxymethyl) -phenyl-3-yloxy] - acetic acid,

[4'-Amιdιno-5- (3-amιdιnophenoxymethyl) -bιphenyl-3-yloxy] - acetic acid

Example 6

A solution of 5.0 g of 3'-bromomethyl-bιphenyl-3-carbonιtrιl and 5 ml of Tn-ethylphosphite are added together and slowly heated to 150 °

The mixture is subsequently stirred at 150 ° for 6 h and, after customary working up, 6.05 g of (3'-cyano-biphenyl-3-ylmethyl) -phosphonic acid ethyl ester ("BA") are obtained. A solution of 1.0 g of "BA" and 3- Cyanobenzaldehyde in 20 ml of ethylene glycol dimethyl ether is added under ice cooling and nitrogen to 150 mg of sodium hydride. Stirring is continued for 4 hours, working as usual and maintaining

0.93 g of 3 '- [2- (3-cyanophenyl) -vιnyl] -bιphenyl-3-carbonιtrιl ("BB") After hydrogenation of 360 mg "BB" with Pd-C-5% in methanol, 360 mg are obtained 3 '- [2- (3-cyanophenyl) ethyl] bιphenyl-3-carbonιtrιl ("BC") After reaction with hydroxylammmonium chloride and hydrogenation with Raney nickel, the compound 3' - [2- (3 -Amιdιno- phenyl) -ethyl] -bιphenyl-3-carboxamιdιn, FAB 343

The compound 3 '- [2- (4-amidophenyl) ethyl] -biphenyl-3-carboxamid, FAB 343 is obtained analogously Example 7

The compound N- [3 '- (4-guanidinocarbonyl-benzyloxy) biphenyl-3-carbonylj-guanidine, dihydrochloride, FAB 431

is obtained, for example, according to the following reaction scheme:

1-chloro-1-ylpyπdιnιumιodιd N- ethylpyrro don N-ethyldnsopropylamiπ (Mukaiyama)

The following connections are obtained analogously

N- [3 '- (3-guanidocarbonylbenzyloxy) biphenyl-3-carbonyl] guanidine, dihydrochloride, FAB 431,

N- [3 '- (4-guanidocarbonyl-benzyloxy) biphenyl-4-carbonyl] guanidine, dihydrochloride, FAB 431,

N- [3 '- (3-Guanιdιnocarbonyl-benzyloxy) -bιphenyl-4-carbonyl] -guanιdιn,

Dihydrochloride, FAB 431

Pharmacological tests

The methodology which was used to characterize the compounds of the formula I as NHE-3 inhibitors is shown below

The compounds of the formula I were characterized with regard to their selectivity with respect to the isoforms NHE-1 to NHE-3. The three isoforms were stably exposed in mouse fibroblast cell lines. The inhibitory effect of the compounds was determined by determining the ElPA-sensitive ²² Na ⁺ uptake into cells after intracellular acidosis. To characterize the Na ⁺ / H ⁺ exchange inhibitors in terms of their isoform selectivity, we examined the compounds for their inhibition of the NHE isoforms NHE-1, -2 and -3, which were stable in a mouse fibroblast cell line were exposed (see procedure), in that the ElPA-sensitive ²² Na ⁺ uptake into the cells was determined after intracellular acidosis

material and methods

LAP1 cells that express the different NHE isoforms

The LAP1 cells that express the isoforms NHE-1, -2 and -3 (a

Mouse fibroblast cell line), were developed by Prof J Pouyssegur (Nice, Frank- The transfections were carried out according to the method of Franchi et al (1986). The cells were cultured in Dulbecco's modified Eagle's medium (DMEM) with 10% inactivated fetal calf serum (FCS). The cells used to select the NHE were used So-called "acid killing method" by Sardet et al (1989) used. The cells were first removed for 30 minutes in a NHCl-containing bicarbonate- and sodium-free buffer by washing with a bicarbonate-, NH CI- and sodium-free buffer and it was removed with a Bicarbonate-free NaCI-containing buffer incubated Only those cells that functionally express NHE were able to survive in the intracellular acidification to which they were exposed

Characterization of NHE inhibitors in relation to their isoform selectivity

Using the mouse fibroblast cell lines mentioned above, which express the isoforms NHE-1, NHE-2 and NHE-3, compounds were tested for selectivity towards the isoforms according to the procedure described by Counillon et al (1993) and Scholz et al (1995) Cells were acidified intracellularly using the NH CI prepulse method and then by incubation in a bicarbonate-free ²² Na ⁺ -containing buffer. Due to the intracellular acidification, NHE was activated and sodium was taken up in the cells. The effect of the test compound was inhibited by EIPA (ethyl -isopropylamilond) - sensitive ²² Na ⁺ uptake printed out

The cells which expressed NHE-1, NHE-2 and NHE-3 were inoculated with a density of 5-7.5 × 10 ⁴ cells / well in microtiter plates with 24 wells and grown to confluence for 24 to 48 hours. The medium was aspirated and the cells were incubated for 60 minutes at 37 ° C in the NH ₄ CI buffer (50 inM NH ₄ CI, 70 mM Chohnchlond, 15 mM MOPS, pH 7.0). Then the buffer was removed and the cells were rapidly twice with the Chohnchlond wash buffer (120mM Chohnchlond, 15mM PIPES / Tris, 0.1mM Ouabain, 1mM MgCl ₂ , 2mM CaCl ₂ , pH 7.4), the cells were incubated in this buffer for 6 minutes Incubation time, the incubation buffer was aspirated

The cells were scanned four times to remove extracellular radioactivity quickly washed with ice cold phosphate buffered saline (PBS). The cells were then solubilized by adding 0.3 ml of 0.1 N NaOH per well. The cell fragment-containing solutions were transferred to scintillation tubes. Each well was washed twice more with 0.3 ml of 0.1 N NaOH and the washing solutions were also added to the corresponding scintillation tubes. Scintillation cocktail was added to the tubes containing the cell lysate and the radioactivity absorbed into the cells was determined by determining the β radiation.

Inhibition of Na ⁺ uptake in rabbit erythrocytes

The Na ⁺ / H ⁺ exchange activity was also determined by observing the uptake of ²² Na ⁺ ions in acidified rabbit erythrocytes. Rabbit erythrocytes have found widespread use in studies of Na ⁺ / H ⁺ exchange activity (Escobales & Fugueroa, 1991; Morgan & Canessa, 1990). The ElPA-sensitive portion of the ²² Na ⁺ uptake in acidified erythrocytes was regarded as Na ⁺ / H ⁺ -dependent ²² Na ⁺ uptake.

cell preparation

The preparation of the red blood cells and the internal acidification of the red blood cells were carried out based on the methods of Morgan and Canessa (1990).

The blood was obtained from rabbits (e.g. New Zealand White). It was collected in 50 ml Falcon zentfuge tubes containing 5 ml sodium heparin solution (250 U / ml). The blood and heparin solution were mixed well. The red blood cells were obtained by centrifugation at 2000 xg at 4 ° C; Plasma and white blood cell cuff were removed. The remaining solution was filtered through 200 µm gauze. The filtrate was resuspended to the original volume with washing buffer (140 mM KCl, 0.15 mM MgCl ₂ , 10 mM TRIS / MOPS, pH 7.4). The red blood cells were again obtained by centrifugation (2000 × g, 4 ° C.). The washing process was repeated twice. Intracellular acidification

For intracellular acidification, 5 ml of the deposited collected red blood cells were again suspended with 45 ml acidifying buffer (170 mM KCl, 0.15 mM MgCl ₂ , 0.1 mM ouabain, 10 mM glucose, 10 mM

Sucrose, 20 mM Tris / Mes, pH 6.2). The red blood cell suspension was incubated for 10 minutes at 37 ° C (with occasional mixing). To fix the internal pH, up to 200 μM and 1 mM DIDS or DIAMOX (acetazolamide) are added. The mixture was incubated for a further 30 minutes at 37 ° C.

The red blood cells were then obtained by centrifugation (4 minutes at 2000 × g, 4 ° C.); they were again suspended with ice-cold unbuffered washing solution (170 mM KCl, 40 mM sucrose, 0.15 mM MgCl ₂ ) and thus washed four times.

Incubation and measurement of the ²² Na ⁺ uptake

The incubation was carried out in Macrowell tube strips in a format of 8 x 12. The incubation was started by adding 200 μl incubation buffer (160 mM KCl, ²² NaCl (37 MBq / well), 10 mM

NaCI, 0.15 mM MgCl ₂ , 0.1 mM ouabain, 10 mM glucose, 40 mM sucrose, 10 mM Tris / MOPS, pH 8.0, 0.5 mM Diamox, 1% DMSO) with 20 μl of the (preheated ) acidified solution of the red blood cells. The test substances were first dissolved with 100% DMSO and the solution was then diluted to the appropriate concentrations with incubation buffer. Incubation was carried out for 5 minutes at 37 ° C. (In preliminary experiments it was shown that under these incubation conditions the ²² Na uptake rate was linear over the 5-minute incubation period). The incubation was stopped by adding 800 µl ice cold stop solution (112 mM MgCl ₂ , 0.1 mM ouabain). The tubes were kept on ice for a short time. The tubes were then covered with parafilm and the red blood cells were obtained by centrifugation at 2000 × g and 4 ° C. for 7 minutes. The supernatant was aspirated with a self-made suction device, with which one could aspirate 4 adjacent tubes at the same time; Spacer rings at the further ends of the tips prevented the tips from being too deep immerse the tubes and aspirate the pellet of red blood cells. All ²² Na-containing supernatants and washing solutions were kept and disposed of as radioactive waste. The red blood cells were washed three times with 900 μl of ice-cold stop solution, by repeating the suspension / decentration step described above. After the last washing, 200 μl of water were added to the red blood cell pellet. The tubes were then treated with ultrasound for 2 x 30 minutes. The Macrowell tube strips were then taken apart and each tube was placed upside down in its own scintillation tube; the hemolyzed solution of the red blood cells emptied into the scintillation vial by gentle shaking. 3 ml of Aquasafe 300 PS scintillation fluid was added to each vial, and the vials were capped and mixed well.

The radioactivity taken up in the red blood cells was determined in a scintillation counter by monitoring the ß-decay. The determination was carried out in triplicate for each substance concentration. Each value of the average of the Zählungsbe- was subtracted atmosphere in the presence of 10 uM EIPA to the non-Na ⁺ / H ⁺ - Na ⁺ dependent ²² -Aufπahme in the erythrocytes to incorporate. The mean of the remaining counts in the absence of a substance was used as a 100% control; the mean values in the presence of the test compounds were expressed as a percentage of this control value. The percentages of the recording data were recorded semi-logarithmically;

IC50 values were obtained by fitting the values to a nonlinear curve using the equation f (x) = 100 / (1 + (IC50 / x) ^** n).

Literature:

Counillon et al. (1993) Mol. Pharmacol. 44: 1041-1045 Escobales and Figueroa (1991) J. Membrane Biol. 120, 41-49 Franchi et al. (1986) Proc. Natl. Acad. Be. USA 83: 9388-9392 Morgan and Canessa (1990) J. Membrane Biol. 118, 193-214 Sardet et al. (1989) Cell 56: 271-280

Scholz et al. (1995) Cardiovasc. Res. 29: 260-268 test results

IC50 (NHE-3) = 1-2 µM

diacetate;

Code EMD 221963 IC50 (NHE-3) = 1 μM

diacetate;

Code EMD 246326 IC50 (NHE-3) = 3 μM ,

diacetate;

Code EMD 246327 IC50 (NHE-3) = 3-4 μM.

The following examples relate to pharmaceutical preparations

Example A: Injection glasses

A solution of 100 g of an NHE-3 inhibitor of the formula I and 5 g of diminium hydrogenphosphate is adjusted to pH 6.5 in 3 l of double-distilled water with 2N hydrochloric acid, sterile filtered, filled into injection glasses, lyophilized under sterile conditions and sterile sealed Each injection glass contains 5 mg of active ingredient

Example B: Suppositories

A mixture of 20 g of an NHE-3 inhibitor of the formula I is melted with 100 g of soy lecithin and 1400 g of cocoa butter, poured into molds and allowed to cool. Each suppository contains 20 mg of active ingredient

Example C: solution

A solution is prepared from 1 g of an NHE-3 inhibitor of the formula I, 9.38 g of NaH ₂ PO ₄ H ₂ O, 28.48 g of Na ₂ HPO ₄ 12 H ₂ O and 0.1 g of benzalkonium chloride in 940 ml of double distilled water. Adjust to pH 6.8, make up to 1 l and sterilize by irradiation. This solution can be used in the form of eye drops

Example D: ointment

500 mg of an NHE-3 inhibitor of the formula I are mixed with 99.5 g of petroleum jelly under aseptic conditions

Example E: tablets

A mixture of 1 kg of an NHE-3-Inhιbιtors of formula I, 4 kg of lactose, 1, 2 kg of potato starch, 0.2 kg of talc and 0.1 kg of magnesium stearate is compressed into tablets in a conventional manner such that each tablet 10th mg contains active ingredient Example F: coated tablets

Analogously to Example E, tablets are pressed, which are then coated in a conventional manner with a coating of sucrose, starchy potatoes, talc, tragacanth and colorant

Example G: Cape

2 kg of an NHE-3 inhibitor of the formula I are filled in hard gelate capsules in a customary manner, so that each capsule contains 20 mg of the active ingredient

Example H: ampoules

A solution of 1 kg of NHE-3 inhibitor of the formula I in 60 l of double-distilled water is sterile filtered, filled into ampoules, lyophilized under sterile conditions and sealed sterile. Each ampoule contains 10 mg of active ingredient

Claims

claims

Compounds of formula I.

wherein

R ¹ , R ⁴ each independently of one another -C (= NH) -NH ₂ , which is also simply by -COA, -CO- [C (R ⁶ ) ₂ ] _n -Ar, -COOA, -OH or by a conventional amino protecting group can be substituted, NH-C (= NH) -NH ₂ , -CO-N = C (NH ₂ ),

R ² , R ³ , R ⁵ each independently of one another H, A, OR ⁶ , N (R ⁶ ) ₂ ,

N0 ₂ , CN, shark, NHCOA, NHCOAr, NHS0 ₂ A, NHSO ₂ Ar, COOR ⁶ , CON (R ⁶ ) ₂ , CONHAr, COR ⁶ , COAr, S (O) _n AS (O) _n Ar, -O - [C (R ⁶ ) ₂ ] _m -COOR ⁶ , - [C (R ⁶ ) ₂ ] _p -COOR ⁶ , -O- [C (R ⁶ ) ₂ ] m-CON (R ⁶ ) ₂ , - [ C (R ⁶ ) ₂ ] _p -CON (R ⁶ ) ₂ , -O- [C (R ⁶ ) ₂ ] _m -CONHAr or - [C (R ⁶ ) ₂ ] _P -CONHAr,

X - [C (R ⁶ ) ₂ ] _n -, -CR ° = CR ⁶ -, - [C (R ⁶ ) ₂ ] _n -O-, -0- [C (R ⁶ ) ₂ ] _n -, - COO-, -OOC-, -CONR ⁶ - or -NR ⁶ CO-, R ⁶ H, A or benzyl,

A alkyl with 1 -20 C atoms, in which one or two CH ₂ -

Groups by O or S atoms or by -CR ⁶ = CR ⁶ -

Groups and / or 1 -7 H atoms can be replaced by F,

Ar unsubstituted or single, double or triple by A, Ar '. OR ⁶ , OAr ', N (R ⁶ ) ₂ . NO ₂ , CN, shark, NHCOA,

NHCOAr ', NHSO ₂ A, NHSO ₂ Ar', COOR ⁶ , CON (R ⁶ ) ₂ , CONHAr ', COR ⁶ , COAr', S (O) _n A or S (O) _π Ar 'substituted phenyl or naphthyl,

Ar 'unsubstituted or single, double or triple by A,

OR ⁶ , N (R ⁶ ) ₂ , N0 ₂ , CN, shark, NHCOA, COOR ⁶ , CON (R ⁶ ) ₂ , COR ⁶ or S (O) _n A substituted phenyl or naphthyl,

Shark F, Cl, Br or I,

n 0, 1 or 2,

m 1 or 2,

p means 1 or 2,

as well as their salts and solvates as NHE-3 inhibitors

Compounds according to claim 1

a) 3 '- (3-Carbamιmιdoyl-phenoxymethyl) -bιphenyl-3-carboxamidine, b) 3' - (3-Carbamιmιdoyl-benzyloxy) -bιphenyl-3-carboxamidi, c) 3'-carbamimidoyl-5- (3-carbamimidoyl-phenoxymethyl) biphenyl-3-carboxylic acid; d) N- [3 '- (3-guanidinocarbonylphenoxymethyl) biphenyl-3-carbonylj-guanidine; e) [3'-Amidino-5- (4-amidinophenoxymethyl) biphenyl-3-yloxy] methyl acetate; f) [3'-Amidino-5- (4-amidinophenoxymethyl) biphenyl-3-yloxy] acetic acid

and their salts and solvates as NHE-3 inhibitors.

3. Use of compounds of formula I according to claim 1 and their physiologically acceptable salts and / or solvates for the manufacture of a medicament for the treatment of thromboses, ischemic conditions of the heart, the peripheral and central

Nervous system and stroke, ischemic conditions of peripheral organs and limbs and for the treatment of shock conditions.

4. Use of compounds of formula I according to claim 1 and their physiologically acceptable salts and / or solvates for the manufacture of a medicament for use in surgical operations and organ transplants and for the preservation and storage of grafts for surgical measures.

5. Use of compounds of formula I according to claim 1 and their physiologically acceptable salts and / or solvates for the manufacture of a medicament for the treatment of diseases in which cell proliferation is a primary or secondary cause, for the treatment or prophylaxis of disorders of the fat metabolism or disturbed breathing drive.

6. Use of compounds of the formula I according to claim 1 and their physiologically acceptable salts and / or solvates for the production of a medicament for the treatment of ischemic kidney, ischemic bowel diseases or for the prophylaxis of acute or chronic kidney diseases

Pharmaceutical preparation, characterized by a content of at least one NHE-3 inhibitor according to claim 1 and / or a shrer physiologically acceptable salts and / or solvates