WO2001032176A1

WO2001032176A1 - Beta-carboline derivatives for use as analgesics

Info

Publication number: WO2001032176A1
Application number: PCT/GB2000/004163
Authority: WO
Inventors: Per Olaf Huusfeldt; Birgitte SØKILDE; Jane Marie Lundbeck
Original assignee: British Technology Group Inter-Corporate Licensing Limited
Priority date: 1999-11-01
Filing date: 2000-10-30
Publication date: 2001-05-10
Also published as: GB0026485D0; GB9925880D0; AU1044501A; GB2355659A

Abstract

Compounds of formula (2) wherein n is 2, 3 or 4; Z is an ester group of formula R3-O-CO- wherein R3 is C¿1-6? alkyl or C2-6 alkenyl; and R?1, R2 and R5¿ are independently selected from the group consisting of hydrogen, halogen, hydroxy, methoxy, nitro, and trifluoromethyl; R4 is hydrogen or C¿1-6? alkyl; in the form of the free base or an acid addition salt thereof are useful as GAT-3/4 type GABA uptake inhibitors, primarily to treat pain.

Description

BETA-CARBOLINE DERΓVATEVES FOR USE AS ANALGESICS

Background of the invention Field of the invention The invention relates to the use of β-carboline derivatives for analgesic and related purposes. Description of the related art

Nerve impulses pass along the specialised cells known as neurones. Neurones communicate with each other at synapses, mainly by chemical means. Neurotransmiters are small molecules which pass from vesicles within the neurone to the presynaptic membrane which is distanced from the postsynaptic membrane on a neighbouring neurone or an effector cell such as a muscle cell by the synaptic cleft. Neurotransmitters transmit a signal across the cleft.

Neurotransmitters can be inhibited by other molecules known as inhibitors, especially by garnma-aminobutyric acid (GAB A). The actions of GAB A at the synapse are terminated by re-uptake of GABA into the pre-synaptic neurones or into glial cells

The inhibition of this GABA uptake results in enhanced availability of this inhibitory neurotransmitter in the synaptic cleft and thus to increased "GABA-ergic" activity. Increased GABA-ergic activity can be useful in the treatment, for example of anxiety, pain and epilepsy, as well as muscular and movement disorders [see, for example, P. Krogsgaard-Larsen et al, Progress in Medicinal Chemistry, 22, 68-112 (1985)].

A well-known and potent inhibitor of GABA uptake from the synaptic cleft into presynaptic nerve terminals and glial cells is 3-piperidinecarboxylic acid

(nipecotic acid). However, being a relatively polar compound and therefore unable to cross the blood-brain barrier, 3-piperidinecarboxylic acid itself has found no practical utility as a drug.

During the 1990s a new class of proteins have been found, known as GABA transporters. They have been classified as GAT-1, GAT-2, GAT-3 and GAT-4. These proteins transport GABA from the synaptic cleft back into the neurone. Both GAT-1 and GAT-3/4 (GAT-3 is the rat and human homologue of GAT-4 in mice) are located exclusively within the central nervous system (CNS). Whilst GAT-1 is widely distributed, GAT-3/4 site expression is regionally specific. There are high levels of GAT-3/4 in the mid-brain, brain stem and spinal cord, key regions involved in pain control. Stimulation of peripheral sensory neurones known as C-fibres results in release of excitatory neurotransmitters in the spinal cord which result in the transmission and perception of pain. Pre-synaptic GABA receptors (GABA-B receptors) inhibit release of these neurotransmitters and reduce the perception of pain. Thus, activation of GABA receptors by GABA released from GABA-ergic neurones in the spinal cord and brainstem leads to a reduced release of neurotransmitters from these fibres and further ascending neurones and so to an analgesic effect.

The anti epileptic drug tiagabine is a selective inhibitor of GAT-1 and has analgesic properties, see M. D. Swedberg, J. Pharmacol. Exp. Ther. 269, 1021-1028 (1994). However, tiagabine potentiates GABA-ergic transmission in all regions of the CNS and can therefore have sedative side effects. Compounds capable of increasing GABA function in the spinal cord and brainstem would therefore have the potential to produce analgesic without sedation.

Certain β-carboline derivatives which are potent inhibitors of GAT-3/4 are disclosed in PCT Patent Application WO 96/34865 (filed by Novo Nordisk A/S and assigned to the present applicant). These are principally compounds of formula (1):

wherein n is 1 to 5,

R¹ and R² independently are principally hydrogen, halogen, hydroxy, nitro, -NR⁹R¹⁰, -SONRⁿR¹², -COOR¹³, -CONR¹⁴R¹⁵, Cι-₆-alkyl, C,-₆-alkoxy, trifluoromethyl or trifluoromethoxy wherein R⁹, R¹⁰, R¹¹, R¹², R¹³, R¹⁴ and R¹⁵ independently are hydrogen, Cι-₆-alkyl, C₂-₆-alkenyl or C₂-₆ alkynyl; and

R⁴ is hydrogen or Cι-₆-alkyl; and

R⁵ and R⁶ independently are hydrogen, halogen, hydroxy, nitro, -NR¹⁶R¹⁷, -COOR¹⁸, Cι-₆-alkyl, Cι-₆-alkoxy, trifluoromethyl or trifluoromethoxy wherein R¹⁶, R¹⁷ and R¹⁸ independently are hydrogen, Cι-₆-alkyl, C₂.₆-alkenyl or C₂-₆-alkynyl; and Z is any of several defined 5-membered ring heterocyclic groups, preferably 3- substituted- 1 ,2,4-oxadiazol-5-yl groups; in the form of a free base or a pharmaceutically acceptable salt thereof.

The problem of the present invention is to find further potent GAT-3/4 inhibitors, preferably with a high selectivity for GAT-3/4 inhibition relative to GAT-1 inhibition.

Summary of the invention

It has now been found that compounds of formula (2):

wherein: n is 2, 3 or 4;

Z is an ester group of formula R³-O-CO- wherein R³ is C]-₆ alkyl or C₂-₆ alkenyl; and

R¹, R² and R⁵ are independently selected from the group consisting of hydrogen, halogen, hydroxy, methoxy, nitro, and trifluoromethyl; and R⁴ is hydrogen or Cι-₆ alkyl; in the form of the free base or an acid addition salt thereof are useful for any novel purpose involving the inhibition of GABA uptake via a GAT-3 or GAT-4 subtype carrier, preferably as analgesics, as well as for treatment of trauma, movement disorders (other than resulting from epilepsy) and spasticity.

These compounds are known from US Patent 4,539,407 to have anticonvulsant activity and thus to be useful in the treatment of epilepsy. However, no other effect is disclosed in that patent.

The invention extends to the further medical use of the compounds of formula (2), in any terms permitted by patent law of individual countries or regions. In particular it includes the use of compounds of formula (2) in the preparation of a medicament for the purposes described above and particularly as an analgesic.

The compounds of formula (2) differ from those of WO 96/34865 in that the 3 -position substituent in the β-carboline ring system is an ester group instead of a heterocyclic group.

Description of the preferred embodiments

The compounds of formula (2) may exist as geometric and optical isomers and all isomers and mixtures thereof are included herein. Isomers may be separated by means of standard methods such as chromatographic techniques or fractional crystallisation of suitable salts.

Examples of pharmaceutically acceptable acid addition salts include inorganic and organic acid addition salts such as hydrochloride, hydrobromide, sulphate, phosphate, acetate, phthalate, fumarate, maleate, citrate, lactate, tartrate, oxalate, or similar pharmaceutically acceptable inorganic or organic acid addition salts. R¹ and R² are preferably hydrogen.

R⁴ is preferably hydrogen or methyl.

R⁵ is preferably hydrogen or a 3- or 4- chloro, bromo or trifluoromethyl substituent. Within the definition of the ester group, Z, the alkyl and alkenyl groups R³ may have straight or branched chains; ethyl and n-propyl are currently preferred, but chain branching at the α-carbon atom is specifically included.

A propylene bridge is preferred, i.e. n is preferably 3.

It has been demonstrated that the novel compounds of formula (2) which selectively inhibit the uptake of GABA, via the GAT-3/4 subtype carrier, from the synaptic cleft, possess useful pharmacological properties in the central nervous system, in that they cause a selective enhancement of GABA-ergic activity. They are primarily useful to treat pain.

The compounds of formula (2) may be prepared as follows: Method A:

A compound of formula (3) wherein R¹, R² and Z are as defined above, may be reacted with an azaheterocyclic compound of formula (4) wherein n, R⁴ and R⁵ are defined above and X is a suitable leaving group such as halogen, p-toluene sulphonate or mesylate. This alkylation reaction may be carried out in a suitable solvent such as acetone, dibutylether, 2-butanone, tetrahydrofuran, methylisobutylketone, methyhsopropylketone, toluene, benzene or DMF in the presence of a base e.g. potassium carbonate, sodium hydride or potassium tert.-butoxide at a temperature up to reflux for the solvent used, for e.g. 1 to 200 h. Method B:

A compound of formula (5) wherein R¹, R², n, Z and X are as defined above may be reacted with an azaheterocyclic compound of formula (6) wherein R⁴ and R⁵ are as defined above. This reaction may be carried out in a suitable solvent as defined above in the presence of a base as defined above and possibly a catalyst, e.g. an alkali metal iodide, at a temperature up to reflux for the solvent used, for e.g. 1 to 200 h. Method C:

An azaheterocyclic ketone of formula (7) wherein R¹, R², n, R and Z are as defined above may be reacted with a Grignard reagent of formula (8) wherein R⁵ is as defined above and Y is chloro, bromo or iodo. This reaction may be carried out in a suitable solvent such as diethyl ether, THF, toluene or benzene at a suitable temperature up to reflux temperature for the solvent used, for e.g. 1 to 5 h. Pharmacological Data

³H-GABA uptake was measured by a modification of the method of Fjalland et al., (1978). A crude membrane preparation was prepared from selected brain areas of male Wistar rats. To test for GAT-1 inhibition, whole forebrain was used. To test for GAT-4 inhibition the fresh inferior colliculus was used, this being an area of the brain associated with GAT-4 activity. The tissue was homogenised in 20 ml of ice- cold 0.32 M sucrose with a hand driven teflon/glass Potter-Elvehjem homogeniser. The homogenate was centrifuged at 4°C for 10 min. at 1,000 x g, and the pellet was discarded. The supernatant was recentrifuged at 4°C for 20 min. at 10,000 x g. The pellet was then homogenised in 50 volumes 0.32 M sucrose. To 300 μl (GAT-1) or 200 μl (GAT-4) of uptake-buffer (200 nM NaCl, 15.3 mM KC1, 6.67 mM MgSO₄, 3.83 mM CaCl₂, 16.67 mM glucose, 66.67 mM Tris, pH 7.5 at 30°C) was added 100 μl test substance and 50 μl tissue suspension. In the GAT-4 inhibition test 100 μl of l-(2-(((diphenylmethylene)amino)oxy)ethyl)-l,2,5,6-tetrahydro-3-pyridinecarboxylic acid (1 nM final concentration) was also added. The samples were mixed and incubated at 30°C for 8 min. Then Η-GABA (0.9 nM final concentration) and unlabelled GABA (25 nM final concentration) was added, and the incubation was continued for an additional 8 min. The reaction was terminated by rapid filtration through Whatman GF/F glass fiber filters under vacuum. The filters were then washed twice in 10 ml of ice-cold isotonic (0.9%) saline, and the tritium trapped on the filters was quantified by conventional scintillation spectroscopy. Non-specific binding was determined in duplicate using nipecotic acid (500 μM final concentration). Total binding minus non-specific binding gives the specific binding.

Values for non-GABA uptake carrier-mediated uptake of ³H-GABA for some representative compounds of the present invention, along with the best results from the compound of PCT Application WO 96/34865, are shown below. Comparing the compound of present Example 1 with the best compounds of WO 96/34865, which are those of Examples 4 and 7 thereof, it will be seen that the compound of the invention shows greater GAT-4 inhibition. Substitution of the rings is generally less favourable, although the 3-chloro substituent in the right-hand benzene ring gave a particularly good result. Replacement of the ester group Z by a -COOH group was unfavourable, as shown by the comparison included ("Comp"). In several instances the compounds of the invention display high ratios of IC₅₀, GAT-l:GAT-4. Thus, for the compound of Example 1 of the present invention, the ratio is about 41 compared with 14 for the best compound (in that respect) of WO 96/34865, the compound of Example 28.

PCT Application WO 96/34865

Z = oxadiazolyl

Present invention

Z = ester group (carboxylic acid group for comparison)

The compounds of formula (2) are preferably formulated with a pharmaceutical carrier or diluent. The compositions containing the compounds of formula (2) may be prepared by conventional techniques and in conventional forms, for example capsules, tablets, solutions or suspensions.

The pharmaceutical carrier employed may be a conventional solid or liquid carrier. Examples of solid carriers are lactose, terra alba, sucrose, talc, gelatin, agar, pectin, acacia, magnesium stearate and stearic acid. Examples of liquid carriers are syrup, peanut oil, olive oil and water.

Similarly, the carrier or diluent may include any time delay material known to the art, such as glyceryl monostearate or glyceryl distearate, alone or mixed with a wax. If a solid carrier for oral administration is used, the preparation can be tabletted, placed in a hard gelatin capsule in powder or pellet form or it can be in the form of a troche or lozenge. The amount of solid carrier will vary widely, but will usually be from about 25 mg to about 1 g. If a liquid carrier is used, the preparation may be in the form of a syrup, emulsion, soft gelatin capsule or sterile injectable liquid such as an aqueous or non-aqueous liquid suspension or solution.

Generally, the compounds of formula (2) are dispended in unit dosage form comprising 50-200 mg of active ingredient in or together with a pharmaceutically acceptable carrier per unit dosage. The dosage of the compounds of formula (2) is 1-500 mg/day, e.g. about 100 mg per dose, when administered to patients, e.g. humans, as a drug.

A typical tablet, which may be prepared by conventional tabletting techniques contains: Core:

Active compound (as free compound or salt thereof) 100 mg

Colloidal silicon dioxide ("Aerosil", TM) 1.5 mg

Cellulose, microcryst. ("Avicel", TM) 70 mg

Modified cellulose gum ("Ac-Di-Sol", TM) 7.5 mg

Magnesium stearate

Coating:

HPMC Approx. 9 mg

*Mywacett, TM 9-40 T Approx. 0.9 mg

*Acylated monoglyceride used as plasticizer for film coating. The route of administration may be any route, which effectively transports the active compound to the appropriate or desired site of action, such as oral or parenteral e.g. rectal, transdermal, subcutaneous, intravenous, intraurethral, intramuscular, topical, intransal, ophthalmic solution or an ointment, the oral route being preferred.

The following Examples illustrate the preparation of the compounds used in the invention. Preparative Examples

Example 1

Propyl 9-(3-(4-Phenyl-4-hydroxypiperidino)propyl)-β -carboline-3-carboxylate.

Propyl-9H-pyrido[3,4-b]indolecarboxylate 2.5g (10 mmol) was dissolved in N,N-dimethylformamide (125 mL) and 0.55g (14 mmol) NaH was added under nitrogen, the mixture was stirred lh at room temperature. 4.72 g (30 mmol) 2-bromo- 1-chloropropane was added and the reaction mixture was stirred for 20 h. at room temperature. The mixture was evaporated in vacuo and redissolved in CH₂C1₂ (125ml). 2N HC1 (100 mL) was added and the mixture was filtered. The organic phase was washed with 5% NaHCO₃(aq) (100 mL) and NaCl (brine) (25 mL). It was then dried with MgSO , filtered and evaporated in vacuo to give the crude propyl (9H-l-(3-chloro/bromopropyl)pyrido[3,4-b]indolecarboxylate. Recrystallisation from 2-propanol gave 3.19g (98% in yield).

This compound was a bromo/chloro mixture and was used below without any further purification.

2.0g (6 mmol) of propyl 9H-l-(3-chloro/bromopropyl)pyrido[3,4- b]indolecarboxylate prepared above was dissolved in N,N-dimethylformamide (100 mL). 4-Phenyl-4-hydroxypiperidine 1.07g (6 mmol) and triethylamine (1 mL) were added. The reaction mixture was stirred under nitrogen at 60°C for 12 h. The reaction mixture was evaporated in vacuo and redissolved in CH C1₂ (100 mL), washed with 5% NaHCO₃ (aq) (50 mL) and NaCl (brine) (50 mL) and then dried with magnesium sulfate. Evaporation in vacuo gave the crude product as an brown oil. Further purification was performed on a silica gel column (150g; CH₂Cl /EtOH/ NH₄OH(25%aq); 185:14:1 as eluant). 2.48g was isolated as the free amine. To 700mg of the free amine was added HC1 (2.3M in ethanol), the solvent was evaporated and the product (as hydrochloride salt) recrystallised from ethanol and diethyl ether, yield 752 mg. For physical data, see list after Examples 3-10. Example 2

Ethyl 9-(3-(4-Phenyl-4-hydroxypiperidino)-propyl)-β-carboline-3-carboxylate.

Ethyl 9H-l-(3-chloro/bromopropyl)pyrido[3,4-6]indole carboxylate was synthesised in the same manner as in Example 1. The title compound (hydrochloride salt) was then synthesised in the same manner as in Example 1, to give 1 g white crystals, yield 54%. Examples 3-10

The other compounds of the Table shown above were synthesised in the same manner as Example 1. Physical data

Compound 1 : Hydrochloride.

Mp: 168-170°C.

Sample for NMR contains ethanol and 2HC1, Mp:179-180°C.

Η-NMR: (D6-DMSO 200MHz) 1.05(d,t;3H); 1.7(d;2H); 1.8(q;lH); 2.4(m;2H); 2.55(d,d;lH); 3.2(m;4H); 4.4(t;2H); 4.35(t;2H); 5.4(broad s;3H);

7.25(t;lH); 7.35(t;2H); 7.45(d,t;2H); 7.8(t;lH); 8.1(d,lH); 8.6(d;lH); 9.2(s;lH); 9.6(s;lH); 11.5(s;lH).

Compound 2: Hydrochloride.

Mp: amorphous, about 113°C. Contains ethanol.

Η-NMR: (CDCl₃;200MHz) 1.25(t;3H); 1.5(t;3H); 1.65(s;lH); 2.05- 2.2(m;4H); 3.45(d,t;4H); 3.7(d;2H); 3.75(q;l,5H); 4.6(m;4H); 7.3(d;l,5H); 7.4(d,t;2,5H); 7.5(d;2H); 7.65(d,d;2H); 8.2(d;lH); 8.9(s;lH); 9.05(s;lH).

Compound 3: Hydrochloride. 3HC1 salt,

Mp: 182-185°C.

FAB MS 502.2 (M⁺ + 1). The sample crystallised was reckoned to contain 0.38 mole H₂O as water of crystallisation.

Sample for NMR contains ethanol. Η-NMR: (CD₃OD; 200MHz) 1.5(t;3H); 1.9(d;2H); 2.4-2.7(m;4H); 3.6(m;6H); 4.65(q;2H); 5.0(t;2H); 7.2-7.4(m;3H); 7.5(d,2H); 8.3(d,lH); 8.7(d;lH); 9.5(d;2H); 9.9(s;lH). Compound 4: Liquid chromatography mass spectrometry (LC-MS): M+l=475; Retention time (Rt)=3.00 min; area; 82%.

Compound 5: Hydrochloride. Contains ethanol. Mp: 201-205°C. Η-NMR: (D6-DMSO 200MHZ) 1.3(t;3H); 1.6(d;2H); 2.0-2.4(m;4H); 2.6-

3.2(broad m;6H); 4.4(q;2H); 4.7(m;2H); 5.5(broad s; IH); 7.3(d;lH); 7.4(t;4H); 7.6(t;lH); 7.8(d;lH); 8.4(d;lH); 8.9(s;lH); 9.2(S;1H). Comparative compound (acid): Hydrochloride. Mp>250°C. Η-NMR: (D6-DMSO 200MHz) 1.7(d;2H); 2.35(m;4H); 3.1-3.5(m; 7H);

4.7(t;2H); 5.6(s;lH); 7.4(m;5H); 7.7(t;lH); 7.9 (d;lH); 8.5(d;lH); 9.0(s;lH); 9.3(s;lH); 10.6(s;lH).

Compound 6: Hydrochloride. Mp: amorphous. Contains ethanol. Η-NMR: (D6-DMSO 200MHz) 1.4(d;6H); 1.7(d;2H); 2.45(m;2H); 3.1-

3.3(m;4H); 3.45(d;2H); 4.7(t;2H); 5.2(h;lH); 7.35(d;2H); 7.45(t;3H); 7.8(t;lH); 8.05(d;lH); 8.6(d;lH); 9.2(s;lH); 9.5(s;lH); 11.3(s;lH). Compound 7: Free amine. Mp: amorphous. LC-MS: M+l=538; Rt-2.88 min; area 96%.

Compound 8: Hydrochloride. Mp: amorphous.

LC-MS: M+l-526; Rt=3.50 min; area 97%. Compound 9: Free amine. LC-MS: M+l=488; Rt=2.92 min; area: 72%. Compound 10: Free amine. Mp: amorphous. LC-MS: M+l=493; Rt-3.33 min; area: 100%. Compound 11 : Free amine. Mp: amorphous. LC-MS: M+l=526; Rt=3.50 min; area: 99%.

Claims

1. A method of treating a patient suffering from pain, trauma, a movement disorder, other than resulting from epilepsy, or spasticity, which comprises administering to said patient an effective amount of a compound of formula

(2):

wherein: n is 2, 3 or 4;

Z is an ester group of formula R³-O-CO- wherein R³ is Ci-₆ alkyl or C - β alkenyl; and

R¹, R² and R⁵ are independently selected from the group consisting of hydrogen, halogen, hydroxy, methoxy, nitro, and trifluoromethyl; and R⁴ is hydrogen or Cι-₆ alkyl; in the form of the free base or an acid addition salt thereof.

2. The method of Claim 1 , wherein R¹ and R² are both hydrogen atoms.

3. The method of Claim 2, wherein n is 3.

4. The method of Claim 3, wherein R³ is ethyl or n-propyl.

5. The method of Claim 1, wherein Z is C₂H₅-O-CO- or n-C₃H₇-O-CO-, n is 3 and R¹ and R² are both hydrogen atoms.

6. The method of Claim 5, wherein R and R are hydrogen atoms.

7. Use of a compound of formula (2):

wherein: n is 2, 3 or 4;

Z is an ester group of formula R³-O-CO- wherein R³ is Cι-₆ alkyl or C - β alkenyl; and

R¹, R² and R⁵ are independently selected from the group consisting of hydrogen, halogen, hydroxy, methoxy, nitro, and trifluoromethyl; and

R⁴ is hydrogen or Cι-₆ alkyl; in the preparation of a formulation for treating a patient suffering from pain, trauma, a movement disorder, other than resulting from epilepsy, or spasticity.

8. Use according to Claim 7, wherein the compound of formula (2) is as further defined in any one of Claims 2 to 6.