DE4220380A1 - Treatment of ulcerative colitis and Crohn's disease - by admin. of 9-amino-pyridazino:pyrrolo:isoquinoline derivs. which act as vaso-spasmolytic endothelin antagonists - Google Patents

Abstract

Treating ulcerative colitis and Crohn's disease comprises admin. of a 9-amino-pyridazino(4',5':3,4)pyrrolo(2,1-a) isoquinoline deriv. (I) or an addn., base or complex salt. R1 is H, 3-7C cycloalkyl, 2-5C alkenyl; PH1; propargyl; or 1-5C alkyl opt. subtd. by OH, 1-4C alkoxy, halo, NH2, mono- or di- 1-2C alkylamino, 2-4C acylamino, 3-7C cycloalkyl, Ph2, furyl, thienyl or a N-contg. 5- or 6-membered ring opt. contg. an O or S atom and opt. subtd. by 1-4C alkyl; R2 is R1 or, where R1 is H, R2 is NH2; di(1-2C alkylamino; acetonylamno; 2-3C acylamino, 1-3C alkylsulphonyl; 1-3C alkoxy-carbonyl; isopropylideneamino or a 5- or 6-membered N-contg. ring opt. contg. a further N atom or an O or S atom; or NR1R2 forms a 3-7 membered ring opt. contg. an O or S atom and opt. substd. by Ph1-1-4C alkyl; Ph1 is phenyl opt. mono- or di-substd. by halo or OMe; Ph2 is phenyl opt. substd. with 1 or more halo, 1-2C alkyl, 1-2C alkoxy, mono- or di- 1-2C alkylamino, NH2, 2-3C acyalmino or alkylsulphonylamino; R3-R5 are H or 1-4C alkyl; R6, R9 are H; OH; 1-4C alkoxy; 1-4C alkylthio; or -NR10R11; R7, R8 are OH; 1-4C alkoxy; or 1-4C alkylthio; R10 is H or 1-4C alkyl; R11 is H or 1-4C alkyl opt. subtd. by OH, OMe or furfuryl. Cpds. (I) are known from DE 3500941 and DE 3525048. They are also described in EP -252299. USE - Cpds. (I) act as vasospasmolyitc endothielin antagonists which block non-selective cation channels. Admin. of cpds. (I) is in conventional formulations at doses of 0.1-500 mg p.o. and 0.05-150 mg i.v.

Description

The invention relates to the use of 12-amino-pyridazino [4 ′, 5 ′: 3,4] pyrrolo [2,1-a] isoquinolines of the formula

as well as their physiologically compatible salts with Acids, bases and complexing agents for the production of Agents for the treatment of ulcerative colitis and Crohn's disease.

In formula I mean:
R ₁ and R ₂ , which may be the same or different, are hydrogen; C ₃ -C ₇ cycloalkyl; C ₂ -C ₅ alkenyl; Phenyl (the phenyl ring optionally being mono- or disubstituted by halogen or methoxy); Propargyl; a straight-chain or branched, saturated or unsaturated alkyl radical with 1-5 carbon atoms, which can be substituted by hydroxy, alkoxy with 1-4 carbon atoms, halogen, NH ₂ , NH-alkyl with 1-2 carbon atoms, N, N-dialkyl with 1 -2 carbon atoms, NH-acyl with 2-4 carbon atoms, cycloalkyl with 3-7 carbon atoms, phenyl (where the phenyl ring in turn can be substituted one or more times by halogen, alkyl with 1-2 carbon atoms, alkoxy with 1-2 carbon atoms, NH-alkyl with 1-2 carbon atoms, N, N-dialkyl with 1-2 carbon atoms, NH ₂ , N-acyl with 2-3 carbon atoms or alkylsulfonylamino), furyl, thienyl, a nitrogen-containing heterocyclic 5- or 6-ring which optionally may contain an oxygen or sulfur atom as a further heteroatom (the ring optionally being substituted by alkyl having 1-4 carbon atoms);
R ₁ and R ₂ together with the nitrogen atom form a 3- to 7-membered ring which may optionally contain an oxygen or nitrogen atom as a further heteroatom (the ring optionally being substituted by phenyl (C ₁ -C ₄ ) alkyl) (the phenyl ring in turn being substituted once or twice by halogen or methoxy));
R ₂ , if R _{1 is} hydrogen, an -NH ₂ ; Di (C ₁ -C ₂ ) alkylamino; Acetonylamino;
-NH (C ₂ -C ₃ ) acyl; Alkylsulfonyl or alkoxycarbonyl radical, each with 1-3 carbon atoms in the alkyl chain; the isopropylidene amino group

or a heterocyclic 5- or 6-ring containing a nitrogen atom and optionally an additional hetero atom containing an oxygen, nitrogen or sulfur atom;
R ₃ , R ₄ and R ₅ , which may be the same or different, are hydrogen or an alkyl radical having 1-4 carbon atoms;
R ₇ and R ₈ , which may be the same or different, hydroxy; Alkoxy of 1-4 carbon atoms; or alkylthio having 1-4 carbon atoms and R ₆ and R ₉ , which may be the same or different, is hydrogen; Hydroxy; Alkoxy of 1-4 carbon atoms; Alkylthio of 1-4 carbon atoms; or the rest

by doing
R _{10 is} hydrogen; or alkyl with 1-4 carbon atoms and
R _{11 is} hydrogen; or alkyl with 1-4 carbon atoms, where the alkyl radical can optionally be substituted by hydroxy, methoxy or furfuryl.

Preferred is the use of those compounds of the general formula I in which R ₁ and R ₂ , which may be the same or different, are for hydrogen; a straight-chain or branched alkyl group with 1-5 carbon atoms or R ₁ for hydrogen and R ₂ for amino; Methylamino; Dimethylamino; Isopropylidene amino; Dimethylamino-C ₁ -C ₄ alkyl; Methoxy-C ₁ -C ₄ alkyl; Cyclopropyl; Cyclopentyl; Cyclohexyl; Cyclohexylmethyl; Phenyl; Phenylethyl, where the phenyl ring is optionally mono- or disubstituted by methoxy or halogen; Pyrazolyl; Propargyl; [1-methylpyrrolidin-2-yl] ethyl; (Piperidin-1-yl) ethyl; Allyl; 4-benzyl-piperazin-1-yl; (Furan-2-yl) methyl; (Pyrrolidin-1-yl) ethyl; 2-hydroxyethyl; (Pyridin-4-yl) ethyl; Benzyl; (Thien-3-yl) ethyl; or R ₁ and R ₂ together with the nitrogen atom to which they are attached for pyrrolidine; Morpholine; Piperazine which is substituted by phenethyl or methoxyphenyl if desired; stand and
R ₇ and R _{8 are} independently hydrogen; Methyl; Methoxy; Hydroxy; or methylthio and
R ₃ , R ₄ , R ₅ , R ₆ and R _{9 represent} hydrogen.

The compounds of formula I are bases and can in the usual way with inorganic or organic Acids as well as salt and complexing agents in any transferred physiologically harmless adducts (salts) will.  

Acids suitable for salt formation are, for example Hydrochloric acid, hydrobromic acid, hydroiodic acid, Hydrofluoric acid, sulfuric acid, phosphoric acid, Nitric acid, acetic acid, propionic acid, butyric acid, Caproic acid, valeric acid, oxalic acid, malonic acid, Succinic acid, maleic acid, fumaric acid, lactic acid, Tartaric acid, citric acid, malic acid, benzoic acid, p-hydroxybenzoic acid, phthalic acid, cinnamic acid, Salicylic acid, ascorbic acid, methanesulfonic acid and the like.

Compounds of the general formula I are preferably used, in which R ₃ , R ₄ , R ₅ , R ₆ and R _{9 are} hydrogen and R ₇ and R _{8 are} methoxy, and / or
R _{1 is} hydrogen and R _{2 is} a group - (CH ₂ ) _0-3 -A, wherein A
Cyclopentyl, cyclohexyl, phenyl, mono- or dimethoxyphenyl

is
or R _{2 is} branched or unbranched alkyl of 4 or 5 carbon atoms; especially compounds wherein the group

For

where the
Phenyl group may be substituted by one or two methoxy groups; or R _{1 is} hydrogen and R _{2 is} one of the following groups
Cyclopentyl
Cyclohexyl
Phenyl

For

stands, in particular in which R _{1 is} hydrogen and R _{2 is} one of the following groups

stands.

Table 1 shows examples of compounds of the Formula I summarized:  

Table 1

From German patent applications DE 35 00 941.1 and DE 35 25 048.8 are cardiotonically active Compounds of the general formula I known become. From European patent application no. 252 299 (A) has become known that this Compounds cardio- and neuroprotective effects possess and in addition the tissue circulation and Tissue oxygenation in the central nervous system promote.

The present invention, however, is the Use of the compounds mentioned for the Manufacture of agents for the treatment of colitis Ulcerative and Crohn's disease.  

As described by SH MURCH et al. (Lancet 339: 381-385, Feb. 15, 1992), endothelin I plays an important pathophysiological role in inflammatory bowel diseases such as ulcerative colitis and Crohn's disease. With the help of immunohistochonic methods, it was found that patients with Crohn's disease in the submucosa area and patients with ulcerative colitis in the area of the lamina propria of the large intestine epithelium have significantly and greatly increased endothelin I concentrations compared to normal healthy people. Local release of endothelin is believed to cause massive vasospasms with consecutive disseminated ischemia with micro-infarctions, which are believed to be the root cause of the diseases mentioned. The vasospasmogenic effectiveness of the endothelin is explained by a Ca ²⁺ overload of vascular myocytes. Endothelin primarily triggers an IP ₃ -mediated intracellular Ca ²⁺ release, which is followed by a massive transmembrane Ca ²⁺ influx through dihydropyridine-insensitive channels. (MS Simonson et al. Clin. Invest. Med. 14: 499-507, 1991; T. Masakai, J. Cardiovasc. Pharmacol. 13: Suppl. 5, S1-S4, 1989; DW Hay, RJ Pharmacol. 100: 383-392, 1990) These channels are unselective cation channels, the existence of which has also recently been described in cells of the colon mucosa. (Chr. Siemer and H. Gögelein, Europ. J. Physiol. 420: 319-328, 1992).

The endothelin-stimulated activation of Fura-22-loaded human leukemia cells (HL 60 cell) has proven to be a suitable screening model for finding functional endothelin antagonists. In line with G. GRYNKIEWICZ et al. (J. Iol. Chem. 260: 3440-3450, 1985), the intracellular Ca ²⁺ concentration in the cytoplasm of HL 60 cells (suspensions) can be monitored spectrofluorometrically and quantified as a measure of cell activation by endothelin. The stimulation was carried out by adding 0.1 μM endothelin; it could be inhibited in a dose-dependent manner by the substances according to the invention.

The functional endothelin antagonism of the substances according to the invention is blocked of unselective cations mediates channels. That's why proof of a functional is also suitable Thapsigargin antagonism on RBL-hm 1 cells as Screening method for functional Endothelin antagonists.

As a reliable screening method to find UKK blockers has been used to quantify the Kinetics of cytoplasmic Calcium ion concentration in Fura-2-loaded adhesive RBL-hm1 cells based on that of Grynkiewicz et al. (1985) proven fluorescence method.

The kinetics of the cytoplasmic Ca ⁺² concentration is continuously recorded as a concentration-time curve after the cell activation triggered by thapsigargin (0.1 µM). The curves of two activated cell cultures are compared in the presence and absence of 10 µM test substance. The area under the curves (AUC) is integrated and registered as a measure of cell activation. The inhibitory potency of the tested UKK blockers is determined using the following relationship:

% H = the percentage inhibition of calcium influx through unselective cation channels through 0.1 µM Thapsigargin stimulated and by 10 µM test substance is inhibited.

AUC _inh = area under the curve recorded in the presence of the stimulant plus 10 µM inhibitory test substance.

AUC contr. = Area under the curve that only after Addition of the stimulant is registered.

Literature:

GRYNKIEWICZ G., M. POENIE and RY TSIEN A new generation of Ca ²⁺ indicators with greatly improved fluorescence properties J. BIOL. CHEM. 260: 3440-3450, 1985
HIDE, M. and MA BEAVEN Calcium influx in a rat mast cell (RBL-2H3) line J. BIOL. CHEM. 266 15221-15229, 1991
KUDO, Y. and A. OGURA Glutamate-induced increase in intracellular Ca ²⁺ concentration in isolated hippocampal neurones BR. J. PHARMACOL. 89: 191-198, 1986
Execution of the investigation:
For screening purposes, Fura-2-loaded adhesive RBL-hm1 cells are stimulated with 0.1 μM thapsigargin in Ca ²⁺ -free incubation medium. After 4 minutes, extracellular Ca ^{2+ is} restored to 1.5 mM and the excessive increase in the cytoplasmic Ca ²⁺ concentration due to a massive transmembrane Ca ²⁺ influx through unselective cation channels is registered on the basis of the fura-2 fluorescence. This influx is to be inhibited exclusively and dose-dependently by unselective cation channel blockers. Neither classic calcium antagonists nor specific blockers of agonists that stimulate IP ₃ turnover can inhibit the transmembrane Ca ⁺² influx indirectly triggered by thapsigargin. The compounds of the present invention are characterized by an inhibition of UKK.

The results of the measurements are summarized in the following table. The percentage inhibition of UKK after thapsigargin stimulation (0.1 µM thapsigargin) in RBL-hm 1 cells is given. The uniform concentration of the test substances is 10 ^-5 mol).

Table 2

The vasospasmolytic effectiveness of the above functional endothelin or Thapsigargin antagonists have been isolated on Vascular preparation confirmed: on retrograde perfused spontaneously beating LANDENDORFF hearts from rats was the coronary perfusion continuously using electromagnetic flux measurement (equipment from Hugo Sachs Elektronik, MARCH), quantified. With this Measurement arrangement can be of extent, duration and Course shape of vascular spasms with great accuracy to register. One perfuses with 100 nM Endothelin concentration, so becomes the coronary Perfusion flow reduced from 11 to 5 ml / min. Restriction of perfusion can be caused by the substances according to the invention are canceled. The Potency of the compounds of the invention regarding thapsigargin inhibition on Fura-2 loaded RBL-hm1 cells or the effectiveness of Endothelin inhibition on Fura-2 loaded HL 60 cells correlated clearly with that on the Langendorff preparation proven vasospasmolytic effectiveness of investigated substances. It can be concluded from this be that the vasospasmolytic Endothelin antagonism of the investigated substances there is a blockage of the unselective cation channels.  

The compounds can be both enteral and administered parenterally. As a dose for oral 0.1 to 500 mg of active ingredient per dose, for IV use from 0.05 to 150 mg per dose suggested. The desired therapeutic dose is depending on the indication and pharmaceutical form and can be determined experimentally.

Suitable forms of use are, for example Tablets, capsules, suppositories, solutions, juices, Emulsions, aerosols or dispersible powders. Appropriate tablets can, for example, by Mixing the active ingredient (s) with known ones Excipients, for example inert Diluents, such as calcium carbonate, Calcium phosphate or milk sugar, disintegrants, such as Corn starch or alginic acid, binders such as starch or gelatin, lubricants such as magnesium stearate or talk, and / or means to achieve the Depot effects, such as carboxypolymethylene, Carboxymethyl cellulose, cellulose acetate phthalate, or Polyvinyl acetate can be obtained. The tablets can also consist of several layers.

Correspondingly, coated tablets can be coated by similar to the tablets produced with cores agents usually used in coated tablets, for example Kollidon or shellac, rubber arabic, talc, titanium dioxide or sugar will. To achieve a deposit effect or The core can also avoid incompatibilities consist of several layers. The same can also the coated tablet to achieve a deposit effect consist of several layers, the above in the Tablets mentioned excipients can be used can.  

Juices of the active substances according to the invention or Active ingredient combinations can also be a Sweeteners such as saccharin, cyclamate, glycerin or Sugar as well as a taste-improving agent, e.g. B. Flavorings, such as vanillin or orange extract, contain. You can also use suspension aids or thickeners, such as Sodium carboxymethyl cellulose, wetting agent, for example condensation products of fatty alcohols with ethylene oxide, or protective substances, such as p-hydroxybenzoates.

Injection solutions are made in the usual way, e.g. B. with the addition of preservatives, such as p-hydroxybenzoates, or stabilizers, such as Alkali salts of ethylenediaminetetraacetic acid, made and in injection bottles or ampoules bottled.

The one or more active ingredients or Capsules containing active ingredient combinations can for example, by making the Active ingredients with inert carriers, such as milk sugar or Sorbitol, mixes and encapsulates in gelatin capsules.

Suitable suppositories can be, for example Mix with the appropriate carriers, such as Neutral greases or polyethylene glycol or its Manufacture derivatives.

Process for the preparation of the compounds of the formula I are in the German patent applications DE-35 00 941.1 and DE 35 25 048.8 described on the reference is hereby made.

Claims (7)

1. Use of compounds of the formula (I) in the
R ₁ and R ₂ , which may be the same or different, are hydrogen; C ₃ -C ₇ cycloalkyl; C ₂ -C ₅ alkenyl; Phenyl (the phenyl ring optionally being mono- or disubstituted by halogen or methoxy); Propargyl; a straight-chain or branched, saturated or unsaturated alkyl radical with 1-5 carbon atoms, which can be substituted by hydroxy, alkoxy with 1-4 carbon atoms, halogen, NH ₂ , NH-alkyl with 1-2 carbon atoms, N, N-Di (C ₁ -C ₂ ) alkylamino, NH-acyl with 2-4 carbon atoms, cycloalkyl with 3-7 carbon atoms, phenyl (where the phenyl ring can in turn be substituted, one or more times by halogen, alkyl with 1-2 carbon atoms, alkoxy with 1 -2 carbon atoms, NH-alkyl with 1-2 carbon atoms, N, N-dialkyl with 1-2 carbon atoms, NH ₂ , N-acyl with 2-3 carbon atoms or alkylsulfonylamino), furyl, thienyl, a nitrogen-containing heterocyclic 5- or 6 Ring which may optionally contain an oxygen or sulfur atom as a further heteroatom (the ring optionally being substituted by alkyl having 1-4 carbon atoms); mean; or R ₁ and R ₂ together with the nitrogen atom form a 3- to 7-membered ring which may optionally contain an oxygen or nitrogen atom as a further hetero atom (the ring optionally being substituted by phenyl (C ₁ -C ₄ ) alkyl ( the phenyl ring in turn being mono- or disubstituted by halogen or methoxy)); mean; or
R ₂ , if R _{1 is} hydrogen, also -NH ₂ ; Di (C ₁ -C ₂ ) alkylamino; Acetonylamino; -NH (C ₂ -C ₃ ) acyl; an alkylsulfonyl or alkoxycarbonyl radical each having 1-3 carbon atoms in the alkyl chain; the isopropylidene amino group or can be a heterocyclic 5- or 6-ring containing a nitrogen atom and optionally an additional hetero atom containing an oxygen, nitrogen or sulfur atom;
R ₃ , R ₄ and R ₅ , which may be the same or different, represent hydrogen or an alkyl radical having 1-4 carbon atoms;
R ₇ and R ₈ , which may be the same or different, hydroxy; Alkoxy of 1-4 carbon atoms; or alkylthio with 1-4 carbon atoms and R ₆ and R ₉ , which may be the same or different, is hydrogen; Hydroxy; Alkoxy of 1-4 carbon atoms; Alkylthio of 1-4 carbon atoms; or the rest mean in the
R _{10 is} hydrogen; or alkyl of 1-4 carbon atoms and
R _{11 is} hydrogen; or alkyl with 1-4 carbon atoms, where the alkyl radical can optionally be substituted by hydroxy, methoxy or furfuryl; as well as their physiologically tolerable salts with acids, bases or complexing agents for the preparation of agents for the treatment of ulcerative colitis and Crohn's disease. 2. Use of compounds of formula I according to claim 1 in which R ₁ and R ₂ , which may be the same or different, for hydrogen; or a straight-chain or branched alkyl group with 1-5 carbon atoms or R ₁ for hydrogen and R ₂ for amino; Methylamino; Dimethylamino; Isopropylidene amino; Dimethylamino-C ₁ -C ₄ alkyl; Methoxy-C ₁ -C ₄ alkyl; Cyclopropyl; Cyclopentyl; Cyclohexyl; Cyclohexylmethyl; Phenyl; Phenylethyl, where the phenyl ring is optionally mono- or disubstituted by methoxy or halogen; Pyrazolyl; Propargyl; [1-methylpyrrolidin-2-yl] ethyl; (Piperidin-1-yl) ethyl; Allyl; 4-benzyl-piperazin-1-yl; (Furan-2-yl) methyl; (Pyrrolidin-1-yl) ethyl; 2-hydroxyethyl; (Pyridin-4-yl) ethyl; Benzyl; (Thiophene-3-yl) ethyl; or R ₁ and R ₂ together with the nitrogen atom to which they are attached for pyrrolidine; Morpholine; Piperazine which is substituted by phenethyl or methoxyphenyl if desired; stand and
R ₇ and R _{8 are} independently hydrogen; Methyl; Methoxy; Hydroxy; or methylthio and
R ₃ , R ₄ , R ₅ , R ₆ and R _{9 represent} hydrogen. 3. Use of a compound of formula I according to claim 1 or 2, wherein R ₃ , R ₄ , R ₅ , R ₆ and R _{9 are} hydrogen and R ₇ and R _{8 are} methoxy. 4. Use of a compound of formula I according to any one of claims 1 to 3, wherein R _{1 is} hydrogen and R _{2 is} a group - (CH ₂ ) _0-3 -A, wherein A
Cyclopentyl
Cyclohexyl
Phenyl
Mono- or dimethoxylhenyl is
or R _{2 is} branched or unbranched alkyl having 4 or 5 carbon atoms. 5. Use of a compound of formula I according to any one of claims 1 to 3, wherein the group For stands in which the phenyl group may be substituted by one or two methoxy groups. 6. Use of a compound of formula I according to any one of claims 1-3, wherein R _{1 is} hydrogen and R _{2 is} one of the following groups
Cyclopentyl
Cyclohexyl
Phenyl stands. 7. Use of a compound of formula I according to claim 6, wherein R _{1 is} hydrogen and R _{2 is} one of the following groups stands.