WO2014044611A1

WO2014044611A1 - 1-aryl-2-heteroaryl benzimidazoles for the induction of neuronal regeneration

Info

Publication number: WO2014044611A1
Application number: PCT/EP2013/069022
Authority: WO
Inventors: Ursula MÖNNING; Andrea Thiele; Wolfgang Halfbrodt; Fiona Mcdonald
Original assignee: Bayer Pharma Aktiengesellschaft
Priority date: 2012-09-20
Filing date: 2013-09-13
Publication date: 2014-03-27

Abstract

The invention relates to the use of 1-aryl-2-heteroaryl benzimidazole derivatives, for the production of pharmaceutical agents for treating and preventing diseases which benefit from therapeutic promotion of regenerative processes, e.g. remyelination and neurogenesis.

Description

1-Aryl-2-Heteroaryl Benzimidazoles for the induction of neuronal regeneration

The present invention is directed to the use of 1 -aryl-2-heteroaryl benzimidazole derivatives, and their use for the production of pharmaceutical agents for treating and preventing diseases which benefit from therapeutic promotion of remyelination and neurogenesis which are induced as regenerative processes in the degenerated brain.

It is well described in the literature, that nearly all degenerative disorders of the central nervous system are associated with brain-specific inflammatory mechanisms leading finally to acute and chronic neurodegeneration (Endres and Dirnagl 2002, Martin 2001 , Dirnagl et al. 1999, Tomita and Fukuuchi 1996).

In the last decade it was discovered and extensively described that regenerative processes are induced in the injured brain. These processes can be driven by different mechanisms which might act synergistically. To the brains regenerative machinery belong neuronal sprouting, formation of new synapses and remyelination which lead to ensheetment of regenerating axonal processes. Another part of the brain's regenerative machinery is adult neurogenesis describing the development of mature glial cells and neurons out of precursor stem cells residing presumably in the subventricular zone of the lateral ventricle and the gyrus dentatus. Both proceses are activated upon injury and are aiming at limiting neuronal loss and restoring the brain's functional capacity. However, in contrast to the processes that take place during brain development in the adult degenerating brain these mechanisms are often incomplete or fail. An example for this failure is the observation that the myelin sheet produced during remyelination appears to be thinner as would be expected for an axon of a given diameter (Fancy SPJ et al. Annu Rev Neurosci 201 1 ; 34: 21 -43).

Critical factors for the remyelination process as well as for axonal sprouting are the clearance of myelin debris by phagocytic macrophages and the production of signalling factors like chemokines, cytokines and growth factors. These factors are necessary for successful generation of new axons and myelin sheets on regenerating axons as well as for oligodendrocyte precursor cell (OPC) proliferation. Activated microglia/ invading macrophages are important sources of such factors.

Therefore, therapeutic promotion of remyelination is one of the main aims in the treatment of neurodegenerative diseases (Fancy SPJ et al. 2010 Exp Neurol 225: 8-13; Fancy SPJ et al. Annu Rev Neurosci 201 1 ; 34: 21 -43). Until to date there is an unmet medical need for efficient therapy aiming at forcing remyelination and neurogenesis available for clinical use. Approaches using biologies to induce remyelination have been tested successfully in different laboratory animal disease models. However, nearly all of them failed in clinical studies.

Benzimidazoles that inhibit the activation of microglia after stimulation with the amyloid β-peptide are described in WO 01/51473 and WO03/68766. It is disclosed that benzimidazoles which inhibit the activation of microglia are used for the treatment of neuroinflammatory diseases, such as AIDS dementia, amyotrophic lateral sclerosis, Creutzfeldt-Jakob disease, Down's Syndrome, diffuse Lewy Body disease, Huntington's disease, leukencephalopathy, multiple sclerosis, Parkinson's disease, Pick's disease, Alzheimer's disease, stroke, temporary lobe epilepsy and as well for the treatment of tumors.

Compounds as disclosed in the afore mentioned patent applications are agents for interrupting the IL 12 and INFy production in cells of monocytic origin or T cells and NK cells.

It has now been discovered that 1-aryl-2-heteroaryl benzimidazoles of formula (I)

R¹ is a phenyl group that is optionally substituted in the 4-position with fluoro or methyl,

R² is a pyridin-3-yl group or a 3-thienyl group,

R³ is COOH or CONH₂,

n is 3, 4 or 5,

or a hydrate, a solvate, or a pharmaceutically applicable salt of said compounds, have beneficial effects on the induction of the production of new myelin sheaths

(remyelination) and axonal sprouting as well as formation of new synaptic terminals and are therefore useful for the induction of remyelination and neurogenesis in the degenerated brain..

Treatment of rats with test compound (example 1 ) (10 mg/kg) starting at day 1 after permanent middle cerebral artery occlusion (MCAO) followed by continuation of the treatment until day 14 after injury revealed the unexpected effects of example 1 . The rats were examined in behavioural tests, their brains underwent electronmicroscopical and immunohistochemical investigations. 4 weeks after injury not only behavioural improvements were observed, but also remyelinated axons were seen in the injured area as well as newly differentiated neurons (see figures 1 -6).

1 -aryl-2-heteroaryl benzimidazoles of formula (I) have beneficial effects on the induction of the production of new myelin sheaths (remyelination). Axonal sprouting and formation of new synaptic terminals as well as the generation of new neuronal cells in a rat model of stroke could be observed.

Therefore, the underlying problem of the present invention is solved by the provision of 1 - aryl-2-heteroaryl benzimidazoles for therapeutic promotion of remyelination and neurogenesis which are induced as regenerative processes in the degenerated brain.

A further aspect of the invention relates to compounds of formula (I)

wherein

Ri is 4-methylphenyl,

or a hydrate, a solvate, or a pharmaceutically applicable salt of said compounds.

A further aspect of the invention relates to compounds of formula (I)

wherein ¾2 is pyridin-3-yl,

A further aspect of the invention relates to compounds of formula (I)

wherein

R₃ is COOH,

or a hydrate, a solvate, or a pharmaceutically applicable salt of said compounds. A further aspect of the invention relates to compounds of formula (I) wherein n is 4,

A further aspect of the invention relates to the compound of formula (I)

wherein Ri is 4-methylphenyl,

f¾ is pyridin-3-yl,

and

n is 4,

or a hydrate, a solvate, or a pharmaceutically applicable salt of said compound.

A further aspect of the invention relates to compounds of formula (I) selected from the group consisting of:

5- {[1 -(4-methylphenyl)-2-(pyridin-3-yl)-1 H-benzimidazol-6-yl]oxy}pentanoic acid,

4-{[1 -(4-methylphenyl)-2-(pyridin-3-yl)-1 H-benzimidazol-6-yl]oxy}butanoic acid,

6- {[1 -(4-methylphenyl)-2-(pyridin-3-yl)-1 /-/-benzimidazol-6-yl]oxy}hexanoic acid,

6-{[1 -(4-methylphenyl)-2-(3-thienyl)-1 H-benzimidazol-6-yl]oxy}hexanoic acid,

4- {[1 -phenyl-2-(3-thienyl)-1 /-/-benzimidazol-6-yl]oxy}butanoic acid,

5- {[1 -(4-fluorophenyl)-2-(pyridin-3-yl)-1 H-benzimidazol-6-yl]oxy}pentanoic acid,

4-{[1 -phenyl-2-(3-thienyl)-1 /-/-benzimidazol-6-yl]oxy}butanamide,

For the demonstration of the in vivo regenerative activity of 1 -aryl-2-heteroaryl benzimidazoles, several animal models of neurodegeneration (transient and permanent middle cerebral artery occlusion (MCAO)) were employed. Summarizing the experimental data 1 -aryl-2-heteroaryl benzimidazoles of the present invention have a large potential for facilitation of regeneration after stroke. Possible mechanisms for the improvement of functional recovery after treatment with 1 -aryl-2-heteroaryl benzimidazoles, e.g. compound of example 1 , are the enhancement of neuronal (dendritic and axonal) sprouting and synapse formation, as well as remyelination in the diseased parts of the brain. The substructural data showing newly generated myelin sheets support the influence of example 1 on remyelination. Remyelination occurs in the adult CNS due to axonal regeneration only, this also hints towards an effect of 1 -aryl-2-heteroaryl benzimidazoles on axonal outgrowth and collateral sprouting. Additionally, immunohistochemical data support the influence of compounds of the present invention on neurogenesis, i.e. generation of new neurons within the lesioned area 4 weeks after permanent MCAO in rats. Four weeks after permanent MCAO; significant increased amounts of doublecortin positive neuronal precursor cells as well as NeuN positive newly generated neuronal cells were detected in the ipsilateral striatum in the example 1 treated groups. The newborn character of the NeuN positive neuronal cells was detected by their positivity for BrdU, which was integrated upon cellular proliferation. Thus, example 1 has a beneficial effect on neurogenesis in the lesioned area and supports differentiation of the newly generated cells.

Therefore, 1 -aryl-2-heteroaryl benzimidazoles of the present invention show a surprising effect on the overall regeneration in the lesioned area of the brain. Treatment with test compound (example 1 ) increased neurogenesis with the preferential induction of neuronal precursor cells and neuronal differentiation and accelerated the regeneration by the generation of new myelin sheets. These processes built up the basis for the observed functional improvements.

1. Examples

Examples 1 -6 were prepared as disclosed in the application WO2003/68766

(corresponding example in brackets).

Example 1 : 5-{[1 -(4-methylphenyl)-2-(pyridin-3-yl)-1 H-benzimidazol-6-yl]oxy}pentanoic acid (example 2) Example 2: 4-{[1 -(4-methylphenyl)-2-(pyridin-3-yl)-1 /-/-benzimidazol-6-yl]oxy}butanoic acid (example 3)

Example 3: 6-{[1 -(4-methylphenyl)-2-(pyridin-3-yl)-1 /-/-benzimidazol-6-yl]oxy}hexanoic acid (example 1 )

Example 4: 6-{[1 -(4-methylphenyl)-2-(3-thienyl)-1 H-benzimidazol-6-yl]oxy}hexanoic acid (example 5)

Example 5: 4-{[1 -phenyl-2-(3-thienyl)-1 /-/-benzimidazol-6-yl]oxy}butanoic acid (example 9)

Example 6: 5-{[1 -(4-fluorophenyl)-2-(pyridin-3-yl)-1 H-benzimidazol-6-yl]oxy}pentanoic acid (example 14) Example 7: 4-{[1 -phenyl-2-(3-thienyl)-1 /-/-benzimidazol-6-yl]oxy}butanamide

55 mg of ammonium chloride were stirred in 2 ml of toluene. 0.51 ml of aluminium trimethyl (2 M solution in toluene) were added and the resulting mixture was stirred for 30 minutes at room temperature. A solution of 200 mg of methyl 4-{[1 -phenyl-2-(3- thienyl)-1 /-/-benzimidazol-6-yl]oxy}butanoate (example 9a, WO2003/68766) in 2 ml of toluene was added and the reaction mixture was stirred for 5 hours at reflux temperature. The reaction mixture was concentrated in vacuo. After flash chromatography of the residue on silica gel 20 mg of the product were obtained.

H-NMR (De-DMSO): δ (ppm) = 1 .87 tt (2H); 2.19 t (2H); 3.88 t (2H); 6.52 d (1 H); 6.75 s (br) (1 H), 6.92 dd (1 H); 7.19 dd (1 H); 7.25-7.28 m (1 H), 7.28 s (br) (1 H), 7.42-7.51 m (2H). 7.54-7.58 m (1 H), 7.60-7.72 m (4H). Biological examples:

1. Remyelination in the rat permanent CAO model (stroke model)

Adult male Fischer F344 rats weighing 250-300 g (Harlan Winkelmann Laboratories, Netherlands, and Charles River Laboratory, Germany) were used. Permanent MCAO (pMCAO) was induced under anesthesia with halothane by electro-coagulation of the proximal portion of the right middle cerebral artery according to a modified method of Tamura et al. (1981 ). Permanent MCAO resulted in an unilateral focal cerebral ischemia.

Test compound (example 1 ) (10 mg/kg) was administered intraperitoneally in a volume of 2 ml/kg body weight (BW). Administration was performed once daily starting one day after pMCAO and continued for 13 days. The control group received vehicle solution (33% propylene glycol/ saline) under the same administration protocol as the example 1 treated animals. Animals were sacrificed at day 28 after injury and perfusion-fixed with 2.5% glutaraldehyde. Glutaraldehyde-perfused brains were subjected to ultrastructural analysis using conventional electron microscopy.

Test compound (example 1 ) delivered i.p. for 2 weeks starting one day after ischemia resulting in significantly improved functional outcome of the animals compared to the vehicle treated control group at 14 days after injury. It could be demonstrated by conventional electron microscopy that the enhancement of recovery was accompanied at the cellular level by production of new myelin sheaths. This production was induced by test compound (example 1 ) recognizable by thin myelin layers (fig. 1 ), and either axonal sprouting and/ or the formation of new synaptic terminals 28 days after injury (fig. 2). A high number of synaptic terminals clustered in certain parts in the lesioned side hints to the latter process (fig. 2).

2. Induction of Neuorogenesis in the rat permanent MCAO model (stroke model)

Focal cerebral ischemia was induced by permanent occlusion of the right middle cerebral artery (pMCAO). The rats were left untreated or treated with test compound (example 1 ) (10 mg/kg/day) for two weeks after occlusion. During the first 5 days after occlusion rats received daily injections of BrdU (50 mg/kg/day). BrdU is a thymidine analog that incorporates into the DNA in proliferating cells. Four weeks after the ischemic insult rats were sacrificed and subjected to immunohistological investigations regarding neurogenesis. The brain slices were stained for BrdU, NeuN, a nuclear marker of adult neuronal cells, and S100p a cytoplasmic marker of adult astroglial cells. The ipsilateral Caudate putamen was investigated using confocal laser scanning microscopy. The results document that the number of proliferating astrocytes does not differ between the groups (fig. 3b). However, an increased amount of NeuN+/ BrdU+ cells within the striatum of treated animals 4 weeks after pMCAO (fig. 3a, c) (p<0.01 ; t- Test) (fig. 3) could be observed. Test compound (example 1 ) induced a significant increase in the number of doublecortin (DCX) positive cells, representing immature neuronal cells, within the lesioned striatum of pMCAO treated rats. Immunoreactive immature neuronal cells were detected by immunohistochemistry in the ipsilateral lesioned DCX. Immunohistochemistry was performed on paraformaldehyde fixed free floating sections using the anti-doublecortin antibody C-18 (Santa Cruz) and a routine peroxidase protocol with the ABC vectastain kit as detection method. The number of DCX immunoreactive cells was counted in 6 sections/ animal (n=6 animals in the untreated group and 5 in the treated one) which were 480 μιη distant from each other covering the whole lesioned striatum. For cell counting a stereological procedure based on the MicroBrightfieid software was used (Gunderson et a/. ). With the same software the volume of the striatum was determined using the Cavalieri Method. The number of DCX immunoreactive cells in the lesioned striatum of example 1 treated rats was significantly increased and about 2.5 times higher as in the untreated animals (p = 0,0043: ANOVA) (Fig. 2). The volume of the investigated area was not different between treated and untreated animals.

3. Behavior testing in rat permanent CAO (pMCAO) model

Adult male Fischer F344 rats weighing 250-300 g (Harlan Winkelmann Laboratories, Netherlands, and Charles River Laboratory, Germany) were used. Permanent MCA occlusion was induced under anesthesia with halothane by electro-coagulation of the proximal portion of the right middle cerebral artery according to a modified method of Tamura et al. (1981 ). Permanent MCAO resulted in a unilateral focal cerebral ischemia. Administration of the test compound (example 1 ) was performed by i.p administration of 10 mg/kg once daily beginning 1 day after MCAO and continuing for 13 days. The start of the administration was delayed for one day in order to study the drug effects only on chronic microglia activation that occurs during chronic phase of ischemia.

All animals were subjected to a neurological assessment to measure the functional outcome (1 day before occlusion, 2 days and 14 days after occlusion). The behavioral tests reflect spontaneous motor activity, motor function of the forelimbs as well as coordination. These functions were impaired after the damage of the dorsolateral cortex. To some extent, impairment in these tests may also reflect damage of the striatum, which is affected by the cerebral infarct, too. A ranking score was employed to assess post-ischemic motor deficits.

All behavioral tests were conducted by a "blinded" investigator according to modified versions of tests described by De Ryck et al. (1989), Garcia et al. (1995), and Roof et al.

(2001 ).

3.1. Spontaneous motor function

Examination of spontaneous behavior in a Perspex box [score 0-6; maximal score 6= no impairment].

The animals were observed for 3 minutes in a Perspex box. The rat's activity was assessed by its ability to approach the walls of the cage. Scores indicate the following: 4 rat moved around, explored the environment, and approached all walls of the cage 3 slightly affected rat moved around in the cage but approached only three walls 2 slightly affected rat moved around the cage but raised up to only two walls

1 severely affected rat raised up to only 1 wall and barely moved in the cage

0 rat did not move at all.

Furthermore, the forelimbs were observed during rising up to the wall. Scores indicate the following:

2 both forelimbs were used symmetrically

1 only 1 forelimb was used correctly

0 rat did not rise up at all.

3.2. Sensorimotoric function

Examination of different limb placing tests (forelimb placement on tabletop in response to a stimuli) [score 0-6; score 6= no impairment].

3.2.1 . Fore paw reaching

The rat was held in the air by the tail 10 cm above a table and the stretch of the forelimbs towards the table surface was ranked. Scores indicate the following:

2 both forelimbs were extended symmetrically

1 rat reacted by turning around at the tail

0 no reaction of one forelimb was observed. 3.2.2 Left forepaw placement

The rat was placed along the table edge to check the lateral placement of the left forelimb. The forelimb was gently pulled down and retrieval and placement were verified. Scores indicate the following:

2 rat placed immediately the left forelimb on the table

1 rat placed the forelimb on the table but with a delay

0 rat did not respond.

3.2.3 Right forepaw placement

(Identical to 3.2.2.. but with the right forepaw)

3.3. Coordinated motor function

Examination of the behavior on an inclined platform [score 0-3 (2); score 3 (2)= no impairment].

Performance was rated as follows:

3 rat climbed normally and gripped tightly to the grid

2 rat climbed on the inclined platform but with some foot slips of the forelimbs

1 rat climbed on the inclined platform but with many wrong gripps of the forelimbs and the hind limbs

0 rat did not climb on the grid.

To evaluate baseline performance, all animals were tested on day 1 before and day 2 post ischemia. Animals reached maximum or sub maximum scores. The same procedure was used post-operatively to assess ischemia-induced motor deficits. Animals were tested once per day on day 2 and day 14 post ischemia. On day 2 post ischemia, significant impairments were observed for all tasks. Animals without impairment in the forepaw placing test were excluded from the study.

As shown in fig. 4, 14 days after occlusion the behavior was significantly improved by drug treatment with a recovery of 64%, whereas in the vehicle group a spontaneous absolute recovery of 22.5 % was observed.

Moreover, peripheral application of test compound (example 1 -7), delivered from day 1 to day 14 post ischemia, also enhanced functional recovery (table 1 ). Treatments were not associated with body weight loss or other apparent adverse systemical or neurological consequences. 4. Behavior testing in rat transient CAO (tMCAO) model

A tMCAO model (Male Sprague-Dawley rats) was employed according to the method described by Koizumi et al. (1986) with minor modifications. Surgery was carried out under halothane anesthesia. Briefly, the left common carotid artery, external carotid artery, and pterygopalatine artery were exposed and ligated. The middle cerebral artery was occluded by inserting the suture into the internal carotid artery and advancing it to the origin of the MCA. The carotid artery was ligated tightly around the inserted filament to secure it at its place. After 75 minutes of MCAO, the suture was removed under halothane anesthesia, and the blood flow was restored. Body temperature was maintained at 37.0 ± 0.5^lC.

I. p. application of 10 mg/kg test compound (example 1 ) starting 3 days after tMCAO and continued for further 12 days once daily significantly enhanced striatal recovery measured by methamphetamine-induced rotation. Methamphetamine-induced rotation is a measure for striatal dysfunction. It directly reflects the degree of asymmetry between the left (ipsilateral, lesioned) and the right (contralateral, unlesioned) striatum since metamphetamine induces dopamine release that leads to rotational behaviour if it is not balanced on both striata of the brain.

MCAO rats were selected based on infarct volume assessed with magnet resonance imaging (MRI) at one day after occlusion (infarction volumes over 150 mm³ were selected). The initial infarct volume of the control (n=12), 3 mg/kg (n=1 1 ), and 10 mg/kg group (n=1 1 ) were 245.8, 248.6, and 244.7 mm³, respectively. Brains of rats which died or which had subarachnoid hemorrhage were excluded from the analysis. Sham-operated rats (n=10) were also prepared for evaluation in the behavioral test. Test compound (example 1 . 3 or 10 mg/kg) was administered intraperitoneally in a volume of 2 ml/kg BW. Administration was performed once daily starting 3 days after tMCAO and continued for 12 days. The control group received the vehicle solution (33% propylene glycol/ saline) under the same administration protocol as the test compound (example 1 ) treated animals. Sham-operated animals received neither treatment nor vehicle.

The infarct volume was estimated using T2-weighted Magnet Resonance Imaging (MRI ). T2 weighted MRI highlights areas with increased amounts of water/ fluids seen by the high signal intensity on the image when compared to the normal brain tissue. Thus, T2 weighted MRI enables the diagnosis of the location and the volume of the infarcted brain area. MRIs were taken at 1 and 15 days after tMCAO. The infarct volume was assessed by T2-weighted RI at 2 weeks after tMCAO. The infarct volume of the compound treated group at a dose of 10 mg/kg had a tendency to decrease by 24.2 ± 9.3% compared with the vehicle treated control group. However, the mean volumes of infarction among each group did not differ significantly (fig. 5).

Methamphetamine-induced rotation behavior was tested 14 days after tMCAO. Methamphetamine stimulates the release of dopamine from intact dopaminergic neurons. In unilateral lesioned animals this causes them to turn toward the lesioned side. The rate at which the animals rotate can be used as an indication of the severity of the dopaminergic denervation. Rats received methamphetamine (4 mg/kg. i.p.) and beginning 10 minutes after administration, ipsilateral and contralateral rotation was counted individually for four times 2 minutes at intervals of 10 minutes (total counting time of 8 minutes).

Methamphetamine-induced ipsilateral rotation was dramatically increased in tMCAO rats. Treatment with test compound (example 1 ) decreased rotation behavior in a dose dependent manner. Whereas in the 3 mg/kg group the decrease in the number of rotations did not reach significance was the number of rotations significantly decreased by 51 .9 ± 14.6% (p<0.05) in the10 mg/kg treatment group, compared with the vehicle treated control group (fig. 6). These data demonstrate indirectly that treatment with the compounds of the present invention resulted in a dose dependent improvement of striatal function the basis of which could only be regeneration.

Possible mechanisms for the improvement of functional recovery after treatment with the compounds of the present invention are the enhancement of neuronal (dendritic and axonal) sprouting and synapse formation as well as remyelination and/ or prevention of ongoing degeneration in the diseased parts of the brain.

Figures:

Figure 1 : New myelin sheaths detected by electron microscopy in the cortex.

In the untreated brain degenerated myelin is detected in the cortex near the lesion site. The degenerated myelin is recognized by swollen myelin fibres (arrows) in (a).

(b) represents an enlarged view of the depicted region in (a) with degenerating myelin (arrows) surrounding dark colored degenerated axons.

(c) test compound (example 1 ) initiates the production of new myelin sheaths within the cortical area near the lesion site (c).

(d) represents an enlarged view of the depicted region in (c) showing new delicate intact myelin sheaths (arrows).

(e) the depicted region (e) represents either sprouting axons or new synaptic terminals (arrow). Figure 2: Induction of doublecortin positive cells by test compound (example 1 ).

Test compound (example 1 ) induced a significant increase in the number of doublecortin (DCX) positive cells, representing immature neuronal cells, within the lesioned striatum of pMCAO treated rats.

(a) immunoreactive immature neuronal cells (arrows) were detected in the ipsilateral lesioned DCX by immunohistochemistry using DAB as a substrate. The sections were counterstained with hematoxylin.

(b) the number of DCX immunoreactive cells in the lesioned striatum of test compound (example 1 ) treated rats was significantly increased and about 2.5 times higher as in the same area of the untreated striatum (p = 0,0043; AN OVA). Cpu - Caudate putamen; LV - left ventricle; arrows point to DCX positive cells.

Figure 3: Influence of test compound (example 1 ) on generation of new neurons and proliferation of astrocytes in the lesioned striatum 28 days after pMCAO.

Neurogenesis in rats treated with test compound (example 1 ) after pMCAO.

(A) the amount of NeuN/ BrdU positive cells within the striatum of treated animals 4 weeks after pMCAO (p<0.01 ; t-Test) was increased.

(B) in contrast, there was no difference in number of proliferating astrocytes between the groups.

(C) represents an example of a newly generated neuron as seen by BrdU and NeuN immunoreactivity with confocal microscopy. Below and on the right hand side of the photographs a section through the slice is seen showing that the immunoreactivities belong to this single neuron.

Figure 4: Effect of test compound (example 1 ) after MCAO on behavior.

Behavioral scores of test compound in MCAO rats after i.p. injection. Animals were tested 2 days before [-2d], 2 days [+2d] and 14 days [+14d] after the MCAO. Test compound) was given daily from day 1 to day 14 [10 mg/kg/d], controls received vehicle only. The vehicle was 33% propylene glycol/saline solution [2 ml/kg]. Results show absolute behavioral scores determined at post ischemia day 2 [+2d] and day 14 [+14d] and are expressed as means ± S.D. Treatments are shown at the base of each set of columns. (n=animal number/group). *p<0.05; -test

Figure 5: Infarct volume 2 weeks after tMCAO.

The infarct volume was estimated using T2-weighted MRI . MRI were taken at 1 and 15 days after tMCAO. The infarct volume of the test compound (example 1 ) treated group at a dose of 10 mg/kg had a tendency to decrease by 24.2 ± 9.3% compared with the vehicle treated control group (control). However, the mean volumes of infarction among each group did not differ significantly. Figure 6: Methamphetamine induced rotation behavior.

The numbers of rotations within 8 minutes of observation are shown in

(a) test compound (example 1 ) in a dose of 10 mg/kg BW significantly improved striatal function.

(b) demonstrates the rotation behavior as observed during 2 minutes in four 10 minutes intervals. The highest dose of test compound (example 1 ) significantly reduced the number of methamphetamine induced rotations. (Sham: untreated and without lesion; Control: untreated with tMCAO; example 1 (3mg/kg) and example 1 (10 mg/kg) - tMCAO, treated with respective concentrations of test compound (example 1 ). BW= body weight; *p <0.05 (-test). Table:

Table 1 : Effect of examples 1 -7 on functional recovery after MCAO Functional recoveries in MCAO rats after treatment with test compounds (examples 1 -7). Animals were tested regarding their behavior 2 days before, as well as 2 days and 14 days after the permanent MCAO. Test compounds were given i.p. daily from day 1 to day 14 [10 mg/kg], controls received vehicle only [33% propylene glycol /saline solution]. Functional recovery was calculated as the difference in the behavioral scores at post ischemia day 2 [2d] and day 14 [+14d] and are expressed as % of maximal possible improvement according to basic values. Recovery observed after vehicle treatment in the respective studies were termed as "spontaneous recovery"

Claims

Claims:

1. Use of 1-aryl-2-heteroaryl benzimidazoles derivatives according to formula (I)

(I)

wherein

R² is a pyridin-3-yl group or a 3-thienyl group,

R³ is COOH or CONH₂,

n is 3, 4 or 5,

or a hydrate, a solvate, or a pharmaceutically applicable salt thereof, for the induction of remyelination and neurogenesis in the degenerated brain.

2. Use according to claim 1 , wherein

Ri is 4-methylphenyl.

or a hydrate, a solvate, or a pharmaceutically applicable salt thereof.

3. Use according to claim 1-2, whereby wherein

R2 is pyridin-3-yl,

or a hydrate, a solvate, or a pharmaceutically applicable salt thereof.

4. Use according to claim 1-3, whereby

R₃ is COOH,

or a hydrate, a solvate, or a pharmaceutically applicable salt thereof.

5. Use according to claim 1-4, whereby

n is 4,

or a hydrate, a solvate, or a pharmaceutically applicable salt thereof.

6. Use according to claim 1 , whereby is 4-methylphenyl,

is pyridine-3-yl,

is COOH, is 4,

hydrate, a solvate, or a pharmaceutically applicable salt thereof.

7. Use according to claims 1-6, whereby the compounds of formula (I) are selected from the group consisting of:

5- {[1-(4-methylphenyl)-2-(pyridin-3-yl)-1 /-/-benzimidazol-6-yl]oxy}pentanoic acid, 4-{[1-(4-methylphenyl)-2-(pyridin-3-yl)-1 /-/-benzimidazol-6-yl]oxy}butanoic acid,

6- {[1-(4-methylphenyl)-2-(pyridin-3-yl)-1 /-/-benzimidazol-6-yl]oxy}hexanoic acid,

6-{[1-(4-methylpheny!)-2-(3-thienyl)-1 H-benzimidazol-6-yl]oxy}hexanoic acid,

4- {[1-phenyl-2-(3-thienyl)-1 /-/-benzimidazol-6-yl]oxy}butanoic acid,

5- {[1-(4-fluorophenyl)-2-(pyridin-3-yl)-1 H-benzimidazol-6-yl]oxy}pentanoic acid, 4-{[1-phenyl-2-(3-thienyl)-1 /-/-benzimidazol-6-yl]oxy}butanamide,

or or a a hydrate, a solvate, or a pharmaceutically applicable salt thereof.