ITMI20001667A1 - 2- OR 3- TENOIC ACID DERIVATIVES FOR ANTAGONIST ACTIVITY OF GLUTAMATE RECEPTORS. - Google Patents

Description

BACKGROUND OF THE INVENTION

Glutamate can activate both channel receptors and G protein-associated receptors. The former are commonly referred to as ionotropic receptors (iGlu) while the latter are called metabotropic receptors (mGlu). MGlu receptors are selectively stimulated by 1-amino-cyclopentan-1,3-di-carboxylic acid (1S, 3R-ACPD) and their structure is similar to that of other members of a complex family of G protein associated receptors which also includes receptors for GABAb, for pheromones and a sensor for Ca <2+>. 8 cDNAs (from mGluRl to mGluR8) have been identified that encode them and on the basis of their structure and their association with effectors are classified into three subgroups:

1 °) Group: It includes the mGlu receptors which activate the cycle of inositols (mGlul and mGlu5) and which are selectively stimulated by DHPG. Other agonists stimulate these receptors in the following order of potency: Quisqualate »1S, 3R-ACPD> CCG1> L-AP-4. A selective antagonist for these receptors has recently been described (see: Pellicciar! Et al. J. Med. Chem. 38: 3717-3719; 1995 and Moroni et al J. Pharmacol. Exp.Ther. 281: 721-729; 1997 ).

2 °) mGlu receptors which inhibit adenylate cyclase and are preferentially stimulated by DCG-IV and CCG1 (mGlu2, mGlu3). The order of power of the agonists is as follows: L-CCG1> 1 S, 3R-ACPD> QUIS ""> L-AP4. Good antagonists are now available for this subgroup (see: Pellicciari et al. J. Med. Chem. 39: 2259-2269; 1996 and Cozzi et al. Eur. J. Neurosci. 9: 1350-1355; 1997).

3 °) mGlu receptors stimulated preferentially by L-AP4 (mGlu4, mGluó, mGlu7, mGlu8) and also associated with adenyl cyclase inhibition but with this order of power: L-AP4 "1S, 3R-ACPD" " L-CCG1.

Their distribution in the CNS is now well characterized and it is known that multiple receptor subtypes can be expressed by the same neuron. The final effects of their activation depend on the types of receptors present and therefore, when they come into contact with the agonist, both inhibitory and excitatory effects can occur. For example, at the cerebellar level the stimulation of mGlul leads to activation of calcium-dependent potassium channels (the production of IP3 causes an increase in the intracellular concentration of calcium) and therefore to hyperpolarization; it also leads to phosphorylation of the NR2C subunit of the NMDA receptor and to a decreased function of this receptor-channel; at the hippocampal level the activation of mGlu 1 receptors increases neuronal excitability through inhibition of the voltage-operated potassium channels and in other structures of the central nervous system, such as the spinal cord, the activation of the mGlu receptors of the first group causes a considerable amplification of synaptic responses mediated by both NMDA and AMPA receptors. There are also mGlu receptors located at the presynaptic level capable of regulating the release of the transmitter with particularly interesting mechanisms. Thus, the stimulation of mGlu4 or mGlu7 can reduce the entry of Ca <2+> into the nerve terminals by directly inhibiting the voltage-dependent channels and thus reducing the synaptic release of the transmitter. A similar result can be obtained by stimulating the mGlu2 receptors which can also be located quite far from the active site of the synapse, but which, having a high affinity for glutamate, seem to be able to be activated, even in a tonic way, by the amino acid present in the extracellular spaces. . These receptors inhibit the formation of cAMP and somewhat reduce the effects of depolarization on transmitter release. On the contrary, the stimulation of other subtypes of mGlu receptors (perhaps mGlul) amplifies the depolarization-liberation coupling of the transmitter.

In this light, the regulation of the functioning of neuronal circuits becomes particularly interesting. In the hippocampus, stimulation of the mGlu4 and mGlu7 receptors reduces the transmission at the level of the glutamatergic synapses, while the stimulation of mGlu5 can increase the excitability of the circuit, perhaps also because it amplifies the ionotropic responses. The result of the reduction in transmission and the increase in excitability is that low intensity stimuli are blocked, while powerful stimuli, capable of overcoming the presynaptic inhibition, are amplified. In this way, the strategic localization of mGlu receptors can lead to the formation of filtering systems capable of increasing the signal / noise ratio of the stimuli that converge on this neuronal circuit. These systems, in which other types of receptors also come into play, seem to operate at the level of different sensory signals and could operate in the regulation of painful afferents at both the spinal and thalamic levels. The pharmacology of mGlu receptors appears to promise broad therapeutic fields of application. The fact that stimulation of mGlu receptors can cause an increase in the sensitivity of ionotropic receptors to the same transmitter, makes these receptors an ideal target for. modulation of excitatory synaptic function.

Numerous molecules are available capable of modifying glutamate excitatory neurotransmission (Lubeluzole, Ketanina, Cerestat, Eliprodil, Aniracetam, LY-3 14582, ACEA 1021, GV-150526A) and of which a clinical use has been proposed. Unfortunately, the clinical experimentation of many of the ligands of the ionotropic glutamate receptors proposed as antiepileptic molecules or capable of reducing ischemic brain damage, has shown that the relationship between therapeutic results and side effects is not favorable. Numerous research groups are attempting to identify molecules that have acceptable side effects and that are endowed with acceptable side effects and that are selectively active on NMDA or AMPA receptor subtypes. Their identification could change the prognosis of diseases in which modern medicine still lacks acceptable therapies (e.g. stroke, CNS degenerative diseases).

DESCRIPTION OF THE INVENTION

Compounds characterized by the presence of a carboxylate thiophene ring have now been found which exhibit selective antagonist activity against the mGlu 1 and mGIu5 receptors (Group 1).

The copeposites of the invention have the following formula I.

in which:

R, is hydrogen, methyl, cyclopropyl or a group of formula

R2 is hydrogen, methyl, methoxy, hydroxy, halogen, cyano;

one of R3 or R4 is carboxy while the other has the same meanings as R2.

The invention also relates to the salts of compounds I with pharmaceutically acceptable acids or bases, the physiologically equivalent derivatives such as esters or amides and the individual enantiomers of compounds I.

R, is preferably hydrogen, R2 is preferably hydrogen or methyl, one of R3 or R4 is preferably carboxy while the other is hydrogen or methyl.

The configuration of the asymmetric carbon atom is preferably R. The compounds of formula I can be prepared by reaction of a compound of formula II

where R2 is as defined above while one of R'3 or R'4 is a protected carboxy group, for example as an alkyl ester, and the other has the same meanings as R2, with alpha-phenylglycinol, preferably with (R) - alpha-phenylglycinol, in the presence of trimethylsilylcyanide, followed by hydrolysis of the obtained N-substituted alpha-amino-nitriles, for example by means of lead tetraacetate and removal of any protective groups on the carboxyl.

The reaction with alpha-phenylglycinol is stereoselective and is carried out in the presence of anhydrous polar solvents such as alcohols, ethers, esters, ketones at temperatures between -10 and 10 ° C.

Hydrolysis with lead tetraacetate is usually carried out in the presence of anhydrous solvents, such as alcohols, ketones, esters, halogenated hydrocarbons or their mixtures at a temperature of about 0 ° C. The final hydrolysis in acids provides the desired compounds.

The compounds of formula II are known or can be prepared with known methods.

For example, 2-formyl-thiophenes can be transformed into the corresponding protected imidazolidine derivatives by reaction with Ν, Ν-dimethylethylenediamine to be then subjected to carboxylation reactions in position 5 (in the presence of strong bases and C02), or by halogenation followed by carboxylation.

By way of example, some synthesis schemes that can be used for the preparation of the compounds of formula I are reported

Scheme 1

Scheme 2

Scheme 3

Scheme 4

The compounds of formula I have antagonist activity of glutamate receptors and can be used both for scientific purposes (for the characterization of glutamate receptors) and for therapeutic purposes for the treatment of diseases in which there is an excessive stimulation of the mGlu receptors. Among the clinical situations in which this excessive stimulation has been demonstrated are to be reported: Focal (stroke) and global (cardiac arrest or fibrillation) cerebral ischemia, CNS trauma, cerebral and subarachnoid hemorrhage, m. Parkinson's, m. Alzheimer's disease, Huntington's chorea, amyotrophic lateral sclerosis, AIDS dementia, seizure disorders, pain and hyperalgesic syndromes, muscle spasms and myoclonus, schizophrenia, anxious and depressive psychiatric syndromes, drug addiction, vomiting.

For the foreseen therapeutic uses, compounds I will be formulated in suitable pharmaceutical compositions, using conventional techniques and excipients.

Compounds I can be administered orally, parenterally, rectally or transdermally, at dosages that will depend on several factors (weight, sex, age of the patient, severity and type of pathology) and on the pharmacokinetic and toxicological characteristics of each single compound. Examples of formulations include capsules, tablets, ampoules, granules containing single doses of 10 to 500 mg of compounds I.

The following examples illustrate the invention in greater detail.

Example 1

a) 1,3-Dimethyl-2- (3-methyl-2-thienyl) imidazolidine (2)

N, N'-dimethylethylenediamine (17.5 g, 198.5 mmol). The resulting mixture is reacted under reflux for 12 hours, occasionally removing the water that forms from the reaction using the Dean-Stark apparatus. The solvent is evaporated under vacuum and the residue is distilled under vacuum to obtain derivative 2 (33.8 g, 172.2 mmoles, 87% yield) as oil (eg 75 ° C at 0.37 mmHg).

'H-NMR (CDC13) d \ 2.20 (9H, bs, NCH3 and CH3-Ar), 2.49 (2H, m, CH2), 3.30 (2H, m, CH2), 3.74 (IH, s, CH), 6.68 (1H, d, J = 5.48, H Ar), 7.16 (IH, d, J = 5.48, H Ar).

b) 5-formyl-4-methyl-thiophene-2-carboxylic acid (3)

A solution of 1,3-dimethyl-2- (3-methyl-2-thienyl) imidazolidine (2) (1 1.30 g, 57.7 mmol) in anhydrous THF (610 ml) maintained under magnetic stirring in an inert atmosphere it is cooled to -78 ° C and treated with TMEDA (9.52 ml, 63.05 mmol) and nBuLi (42 ml, 1.5 M solution in hexane). The mixture is reacted at -78 ° C for 2 hours. The reaction mixture is then poured into a flask containing a pasty mixture of diethyl ether (200 ml) and solid C02 (enough to have a partial solidification of the mixture). The mixture thus obtained is allowed to return to room temperature under stirring overnight. The mixture is evaporated to dryness under vacuum and treated under stirring at room temperature with an aqueous solution of H2SO4 at 10% (250 ml). The mixture is extracted with ethyl acetate (2 x 200 ml) and the organic phase is extracted with a saturated solution of NaHC03 (200 ml). The aqueous phase is acidified with HCl (3N) and extracted with ethyl acetate (4 x 100 ml). The organic phases are combined, dried over anhydrous Na2S04) and evaporated under vacuum to obtain acid 3 (8.3 g, 48.7 mmoles, 84% yield) as a gray solid, (mp 177-1 80 ° C)

’H-NMR (CDC13) d: 2.53 (3H, s, CH3), 7.54 (IH, s, Ar), 10.02 (IH, s, CHO).

c) Methyl 5-formyl-4-methyl-thiophene-2-carboxylic ester (4)

A solution of 5-formyl-4-methyl-thiophene-2-carboxylic acid (3) (8.20 g, 48.2 mmol) in methanol (200 ml) is bubbled several times with gaseous HCl and the mixture is made react at room temperature for 36 hours. The solvent is removed under vacuum and the residue is divided between water (200 ml) and ethyl acetate (200 ml), the organic phases are washed with a saturated solution of NaHC03 (2 x 100 ml), dried over anhydrous Na2SO4 and evaporated under vacuum obtaining ester 4 (7.1 g, 38.6 mmoles, 80% yield) as a white solid (mp 75-77 ° C), which is used as such for the subsequent reaction.

1H-NMR (CDCI3) d: 2.48 (3H, s, CH3), 3.81 (3H, s, C02CH3), 7.51 (IH, s, Ar), 9.96 (IH, s, CHO).

d) (2R) -N - [(R) -a-PhenylglycinesI] -2- (5'-carbomethoxy-3'methic) -2'-thienyl) "glycinonitrile and (12) (2S) -N - [( R) -a-Phenylglycinyl] -2- (5'-carbomethoxy-S'methyl-l-thienity-glycinonitrile (13)

To a solution of methyl 5-formyl-4-methyl-thiophene-2-carboxylic ester (4) (1.98 g, 10.8 mmoles) in anhydrous methanol (100 ml) maintained under magnetic stirring, in an inert atmosphere and at room temperature (R) -a-phenylglycinol (2.17 g, 15.8 mmoles) is added and the mixture is left to react for 2 hours. The mixture is cooled to 0 ° C (ice bath), TMSCN (2.9 ml, 21.44 mmoles) is added and it is allowed to react for 12 hours removing the ice bath. The solvent is evaporated under vacuum and the residue is purified by flash chromatography, using petroleum ether / ethyl acetate as eluent mixture (gradient 90/10 up to 75/25). The two amino nitriles 5 (1.88 g, 5.7 mmoles, 53% yield) and 6 (0.39 g, 1.18 mmoles, 11% yield) are obtained as amorphous solids.

5: 'H-NMR (CDCI3) 5: 2.15 (3H, s, CH3), 3.65 (1H, dd, J = 10.8, 9.4, £ H2OH), 3.72-3.90 (4H, m, CH ^ OH and C02CH3) , 4.20 (1H, dd, J = 9.4, 4.0, CH-F), 4.51 (IH, s, CHCN), 7.20-7.40 (5H, m, Ar), 7.45 (IH, s, H-Ar).

, 3C-NMR (CDC13) 6: 14.49, 21.48, 46.34, 52.72, 60.88, 63.58, 67.43, 68.14, 117.67, 127.75, 128.13, 129.11, 129.48, 131.92, 136.68, 137.16, 137.97, 138.70, 164.57, 162.81.

6: 'H-NMR (CDC13) 5: 2.11 (3H, s, CH3), 3.65-4.00 (5H, m, CH2OH and C02CH3), 4.45 (1H, dd, J = 9.4, 4.0, CH-Ar), 4.87 (1H, s, CHCN), 7.20-7.40 (5H, m, Ar), 7.49 (1H, s, H-Ar).

I3C-NMR (CDC13) d: 14.49, 45.60, 52.76, 62.46, 67.30, 76.63, 118.10, 127.94, 128.95, 129.05, 131.60, 136.77, 137.61, 138.05, 142.23, 163.03.

e) (2S) -2- (5'-Carboxy-3'methyl-2'-thienI) -glycine (7) (3-MATIDA) To a solution of (2R) -N - [(R) -a- phenylglycinyl] -2- (5'-carbomethoxy-3'methyl-25-thienyl) -glycinonitrile (5) (0.90 g, 2.72 mmol) in a mixture (1: 1) of anhydrous methanol (30 ml) and anhydrous dichloromethane (30 ml) cooled to 0 ° C, placed under magnetic stirring and in an inert atmosphere, lead tetraacetate (2.54 g, 5.52 mmoles) is added. The mixture obtained is left to react at 0 ° C for a further 30 minutes and subsequently treated with 50 ml of water, leaving the reaction mixture to return to room temperature. The mixture is filtered on celite and the recovered solvent is evaporated under vacuum. The residue obtained is dissolved in HCl (6N, 40 ml) and reacted under reflux for 16 hours. The mixture is left to return to room temperature, washed with dichloromethane (3 x 10 ml) and the aqueous phase evaporated under vacuum. The residue obtained is purified by ion exchange chromatography with Dowex 50WX2-200 resin, eluting with 10% pyridine. The final Γα-amino acid (7) is obtained as a gray solid (0.32 g, 1.49 mmoles, 55% yield) (mp 245-247 ° C).

’H-NMR (D20) d \ 2.15 (3H, s, CH3), 5.01 (IH, s, CH), 7.35 (IH, s, H-Ar).

nC-NMR (D20 + Pir d6) δ \ 16.83, 55.72, 136.47, 137.37, 142.13, 144.81, 171.47, 174.99.

Examples 2-3

Similarly to example 1, the following compounds were obtained: (2S) N-2- (5'carboxy-4'-methyl-2'-thienyl) -glycine (4-MATIDA)

'H-NMR (D20) d: 2.42 (3H, s, CH3), 5.01 (1H, s, CH), 7.06 (IH, s, Th). ! 3C-NMR ((D20 + Py d6) d: 14.63, 53.60, 132.27, 136.20, 137.00, 141.10, 168.90, 171.40.

(2S) -N-2- (4'-carboxy-5'-methyl-2'-thienyl) -glycine (5-MATIDA)

'H-NMR (D20) d: 2.50 (3H, s, CH3), 4.85 (1H, s, CH), 7.26 (1H, s, Th). I3C-NMR ((D20 + Py d6) d: 14.09, 48.37, 130.08, 136.20, 137.00, 145.41, 168.90, 170.56.

(2S) -N-2- (5'-carboxy-2'-thienyl) -glycine (ATIDA)

* H-NMR (D20) d: 5.45 (IH, s, CH), 7.34 (IH, d, J = 3.8, 4-H Th), 7.77 (IH, d, J = 3.8, 3-H Th).

, 3C-NMR ((DzO + Py d6) d: 53.84, 127.81, 128.90, 137.61, 141.45, 166: 82, 171.58.

Example 4

The molecules under study were evaluated on the following experimental tests: 1) Antagonism of the stimulating action of phospholipase C and formation of inositol phosphates by 1 S, 3R-ACPD (300 μΜ) in slices of rat bark. Molecules active in this test are considered to be mGlul or mGlu5 antagonists (Group 1).

2) Antagonism of the action of L-CCG1 (3 μΜ) on the formation of cAMP induced by forskolin (30 μΜ) on slices of rat striatum. Molecules active on this test are to be considered mGlu2 or mGlu3 antagonists (Group 2)

3) Antagonism of the action of L-AP4 (10 μΜ) on the formation of cAMP induced by forskolin (30 μΜ) in rat cerebellar slices. Molecules active on this test are to be considered antagonists mGlu4, mGlu7 and mGlu8 (Group 3).

The molecules active on the mGlu of the 1st group were then tested on the enhancement of the release of transmitter from rat cortical slices and the molecules active on the mGlu of the 2nd group were tested on the inhibition of the release of the transmitter from striatal slices. All the molecules studied were also tested on mouse cortical preparations (cortical wedges: see Mannaioni et al. Br. J. Pharmacol. 118: 1530-1536; 1996) to evaluate their eventual selectivity and their action on ionotropic receptors. The methods used for the above experiments are described in: Lombardi et al British J. Pharmacol. 110: 1407-14121993 ,; and Lombardi et al. British J. Pharmacol.

117: 189-195,1996 and Moroni et al. Eur. J. Pharmacol. 347: 189-195; 1998. _____ Results

Among the molecules tested, the compound of Example 1 (UFP 713) appears to be particularly interesting, a selective antagonist of the mGluRs receptors of the first group. This molecule antagonizes the effect of 1S, 3R-ACPD (300 μΜ) on the formation of inositol phosphates in slices of rat cortex with an IC50 of 10 μΜ. It has no effect up to 300 μΜ on the other subgroups of mGlu receptors or on the ionotropic receptors for glutamate. An interesting action of UPF 713 is its ability to inhibit neuronal death in cell cultures subjected to oxygen and glucose deprivation as previously observed for other mGlu 1 antagonists (see: Pellegrini-Giampietro Neuropharmacology 38: 1607-1619; 1999) _

Claims (6)

CLAIMS 1. Compounds of formula I in which; R, is hydrogen, methyl, cyclopropyl or a group of formula R2 is hydrogen, methyl, methoxy, hydroxy, halogen, cyano; one of R3 or R4 is carboxy while the other has the same meanings as R2, their salts with pharmaceutically acceptable acids or bases, physiologically equivalent derivatives and single enantiomers. 2. Compounds according to claim 1 wherein R is hydrogen, R2 is hydrogen or methyl, one of R3 or R4 is carboxy while the other is hydrogen or methyl. 3. Compounds according to claim 1 or 2 in which the configuration of the asymmetric carbon atom is R. 4. Compound according to claim 1, (2S) -2- (5'-Carboxy-3'methyl-2'-thienyl) -glycine. 5. Pharmaceutical compositions containing as an active principle a compound of claims 1-4 in admixture with suitable excipients. 6. Compounds of claims 1-4 as glutamate receptor antagonists.