WO2009104027A1

WO2009104027A1 - Therapeutic application of triciclic aromatic and saturated benzo(4,5)thieno-(2,3-d)pyrimidine derivates, as well as their therapeutically acceptable salts

Info

Publication number: WO2009104027A1
Application number: PCT/HU2008/000017
Authority: WO
Inventors: Péter BÁNHEGYI; György Kéri; László ÖRFI; Zsolt SZÉKELYHIDI; Frigyes Wáczek
Original assignee: Vichem Chemie Kutato Kft
Current assignee: Vichem Chemie Kutato Kft
Priority date: 2008-02-19
Filing date: 2008-02-19
Publication date: 2009-08-27
Anticipated expiration: 2010-08-19

Abstract

The subject of the invention is therapeutic application of tricyclic aromatic and saturated benzo[4,5]thieno[2,3-d]pyrimidine derivatives and their therapeutically acceptable salts. The compounds according to the invention is used particularly as active agent of therapeutic preparations of tyrosine-kinase inhibiting action.

Description

Therapeutic application of tricyclic aromatic and saturated benzo[4,5]thieno- [2,3-d]pyrimidine derivatives, as well as their therapeutically acceptable salts.

The invention relates to therapeutic application of tricyclic aromatic and saturated benzo[4,5]thieno-[2,3-d]pyrimidine derivatives, as well as to that of their therapeutically acceptable salts.

The compounds applied in accordance with the invention can be characterized by the formula (I):

( I )

wherein Rl is hydrogen, phenyl, mono-, di-. tri-, tetra- or penta-substituted phenyl, five- or six-membered substituted or unsubstituted heterocyclic group having one or more heteroatoms, advantageously pyrazole, imidazole, isoxazole furane. pyrrole, thiophene, thiazole, isothiazole, triazole, pyrane, pyridine, pyrimidine, dioxane. morpholine, thiomorpholine, pyridazine, pyrazine or piperazine group; R2 is hydrogen, linear or branched alkyl, cycloalkyl, unsubstituted or substituted phenyl group; A is methylene (CH2) or methylidene (CH).

The invention covers also the above mentioned application of therapeutically acceptable salts of the compounds of the formula (I).

The compounds of the formula (I) are known but their therapeutic application is new. The compounds of the formula (I) can be advantageously applied as active substances of pharmaceutical preparations of tyrosine-kinaze inhibiting effect. These preparations have advantageously anti-proliferative. anti-tumor or anti-viral effect, so they can be applied for treatment of tumorous, hyper-proliferative. viral infection diseases, as well as those pertaining to inflammatory processes. The preparations can be formulated with the usual vehicles, media and auxiliary materials in any used form of pharmaceutics.

The EGFR-PTK (Epidermal Growth Factor Receptor Protein Tyrosine Kinase) is an enzyme belonging to the group of transmembrane (acting through the cell membrane) receptor tyrosine kinase enzymes fulfilling receptor function on the cell membrane having the task to maintain communication between the cells (as well as division and differentiation of the cells)

(Axel Ullrich, Waun Ki Hong; The role of EGFR in solid tumors and implications for therapy, Oncology Biotherapeutics, 2000, VoI 1 , Number 1 )

It accomplishes its function in pairs (with another kinase enzyme) on the cell membrane.

It can be divided into three main parts of its structure:

1.) extracellular binding region (acting outside the cell);

2.) transmembrane region (acting through the membrane);

3.) intracellular binding region (acting inside the cell).

During the functioning of the enzyme the molecule EGF arriving to the cell is bound on a specific binding location outside the cell resulting in that the EGFR-PTK becomes linked to a neighboring kinase, if the latter is also EGFR-PTK a homo- dimer, if it is kinase enzyme of another type then a hetero-dimer is generated, then the intracellular part of the enzyme gains kinase catalytic activity, as result thereof it catalyzes transfer of a phosphate group from the adenosine-triphosphate (ATP) molecule being present in the cell to a tyrosinic hydroxil group of an intracellular protein.

This process can be delineated as follows: T7HU2008/000017

Adenosine-triphosphate (ATP) Adenosine-diphosphate (ADP)

The signal arriving to the cell from outside is transmitted to the inside of the cell as result of the binding of phosphoryl radical.

There are four subtypes of the EGFR-PTK: EGFR, HER2 (Human EGF-Related Receptor), HER3, HER4.

In a number of diseases (primarily tumorous, hyperproliferative ones or viral infections) the abnormal (usually increased) function of the inter-cell communication route mediated by EGFR-PTK is either attested or its likelihood can be demonstrated.

The results of increased function of cancerous cells are as follows: - proliferation of cells; - invasiveness (the ability to invade into other tissues);

- suspension of the apoptosis (spontaneous necrocytosis);

- metastasis (generation of remote metastases);

- angiogenesis (growth of veins in the tumor).

Consequently, the inhibition of the function of enzyme system EGFR-PTK is investigated for a long time as a possible therapy of the above mentioned diseases.

The possible methods of inhibition of the enzyme EGFR-PTK are as follows:

L) - A monoclonal antibody inhibiting binding of the molecule EGF and activation of the intracellular catalytic domain by binding itself to the extracellular EGF receptor.

2.) - A molecule binding itself steadily to the intracellular kinase binding location and inhibiting the binding of the protein to be provided with phosphoryl radical to the catalytic domain.

3.) - A molecule binding itself steadily to the intracellular kinase binding location of the ATP binding region, which inhibits binding of ATP necessary for the phosphoryl radical binding to the catalytic domain.

There are publications in the professional literature about the EGFR-PTK inhibiting effect of the 4-phenylamino-pyrrholo-pyrimidine. (Peter M. Traxler et al, J. Med.

Chem, 1996, 39, 2285-2292)

In accordance with the predictions of the molecule modeling program of our own development work, the efficiency and the selectivity can be improved if the nitrogen atom of the pyrrole ring is changed with sulfur and/or the tetrahydro-benzene ring is aromatized.

The invention relates to the therapeutic application of tricyclic aromatic and saturated benzo[4,5]thieno-[2,3-d]pyrimidine compounds where Rl is hydrogen, phenyl, mono-, di-, tri-, tetra- or penta-substituted phenyl, five- or six-membered substituted or unsubstituted heterocyclic group containing one or more heteroatoms, advantageously imidazole, isoxazole, furane, pyrrhole, thiophene, thiazole, isothiazole, triazole, pyrane, pyridine, pyrimidine, dioxane, morpholine, thiomorpholine, pyridazine, pyrazine, piperazine group; R2 is hydrogen, linear or branched alkyl, cycloalkyl, unsubstituted and substituted phenyl group; A is methylene (CH2), methylidene (CH) and their therapeutically acceptable salts. The subject of the invention also the use of tricyclic aromatic and saturated benzo[4,5]thieno-[2,3-d]pyrimidine compounds of the formula (I) as active agent of medicinal preparations of thyrosine-kinase inhibiting effect.

The compounds of the formula (I) where Rl, R2 and A are as mentioned above, applied according to the invention, as well as their therapeutically acceptable salts are produced according to the process shown below,

1-12

According to the process - as shown in the above figure - an oxo- compound of the formula (II) containing a six-membered ring is condensed with cyan-acetic acid ethylester and elementary sulphur in watery and alcoholic medium between 60 and According to the process - as shown in the above figure - an oxo- compound of the formula (II) containing a six-membered ring is condensed with cyan-acetic acid ethylester and elementary sulfur in watery and alcoholic medium between 60 and 100⁰C then the produced amine derivative of the formula (III) is transformed with formamide between 60 and 100⁰C in a ring closing reaction to a thieno-pyrimidine compound of the formula (IV) where R2 is as mentioned above then its hydroxy 1 group in the position 5 is changed to halogen with excessively applied phosphor- oxychloride by heating to 80 to 100⁰C then the compound is brought into reaction with a reagent of the formula R1-NH2 where Rl is as mentioned above between 50 to 100⁰C in alcohol and the compounds of formula (I) marked above with 1- 12 where Rl R2 and A are as mentioned above are produced by an optionally following salification.

The production and the application of the compounds are shown by examples of execution below.

Example 1

The production of compounds marked with 1-12.

Ia.

The compound of the formula (III) is produced as follows:

1 mole of cyclohexanone and 1 mole of cyan-acetic acid ethylester are dissolved in 00 milliliter of ethyl alcohol, 1 mole of sulfur powder 35 milliliter of distilled water and 35 milliliter of triethylamine are added. The reaction mixture is mixed at 100 centigrades during two hours then let to cool to room temperature. The precipitated crystalline material is filtered then washed with water and n-hexane. The yield is 60 to 77%. Ib.

The compound of formula (IV) is produced as follows:

I I I IV

10 millimoles of compound of formula (III) is dissolved in 15 milliliter of formamide then mixed at 100 centigrades during 3 hours. The reaction mixture is then cooled to room temperature then poured onto ice, the precipitated crystalline material is filtered and washed with water and n-hexane.

The yield is 65%.

Ic.

The compounds of the formula (I) marked with 1-12 are produced as follows:

1 millimole of starting compound of formula (IV) is dissolved at room temperature in 5 milliliters of phosphorus oxychloride (POC13), the solution is mixed at 100 centigrades during 3 hours then cooled to room temperature and poured onto ice of twentyfold quantity. The precipitated solid material is filtered, washed with n-hexane then dried. The compound gained in this way and 2 millimoles of Rl -amine are dissolved in 2.5 milliliters of isopropil alcohol then mixed at 100 centigrades during 6 hours in a closed glass recipient. The reaction mixture is poured onto ice, neutralized with IN hydrochloric acid until pH=7 then the precipitated solid material is filtered and washed with water and n-hexane.

The yield is 66 to 85%.

The compounds produced in this way are as follows:

The analytic characteristics of the produced compounds are as follows:

2), 3.16(bs,2H,CH2), 7.69(d,2H₅ArH₅J=8.46), 7.89(d,2H,ArH,J=8.40).

8.49(S₅ IH₅ArH), 8.56(S₅IH₅NH)

The applied methods of analysis are as follows:

HPLC-MS

The analysis LCMS was performed with a liquid chromatography mass-spectrometer Waters chromatograph of type ZMD of high efficiency equipped with Waters 996 DAD UV detector, Waters 2700 automatic sampler and Waters 600 control card A column of type Supelco Discovery RP- Amide was used in gradient mode with flow rate of 3 ml/min.

The starting (A) solvent was: 10% of acetonitrile, 90% of water and 0.05% of

HCOOH.

The B solvent was 100% of acetonitrile.

The gradients were 0% B, until 30 sec between 0 and 80% from 30 to 120 sec, 80% until 240 sec between 80 and 0% from 240 to 260 sec and 0% until 306 sec.

The injection was 5μL.

The solvents were purchased from the Riedel-deHaen Company (Acetonitrile G

Chromasoly (34998), Formic acid extra pure (27001)). The distilled water was produced by a MiIi-Q Academic equipment.

The technical data of the mass-spectrometer are: Ionization: ES+/ES-, pre-heating temperature: 120 °C, temperature of desolvation: 350 °C, gas of desolvation: 400 L/min, pulverizing gas: 100 L/min, capillary: 3000 V, pulverizing voltage: 25V. Extractor: 3 V Rf, Lenses: 0.2 V, Scan: from 120 to 1000 m/z 1 sec, inter-scan delay : 0.1 s.

NMR

The analysis Hl-NMR of 300 MHz was performed with an apparatus of type Bruker

AC-300 at 25 ⁰C. DNSO-d6 was used as solvent.

The melting points were found with an apparatus Bϋchi Melting Point B-540.

Example 2.

Examinations of the effectiveness:

The EGFR-PTK inhibitor action and selectivity of the produced compound were found on a selectivity panel containing 20 kinase enzymes, as follows:

Control examinations:

Negative control (C-): 100 mM of EDTA, without inhibitor Positive control (C+): Buffer without inhibitor

Reaction buffer:

The materials for assay were prepared as stock-solution dissolved in dimethyl- sulfoxid (DMSO) of 100%, from which samples were taken and the solutions of desired concentration were prepared by dilution of samples with buffer solution. The kinase inhibitor action of components is given in per cents (0% - no action, 100% - complete action) in the table below. The measurements were performed with three samples (in triplicate) at each concentration the results were calculated as arithmetic average of the three samples.

The concentrations causing inhibition of 50% of the existing kinase activity were determined for the compounds having the highest selectivity and the highest effectiveness, (inhibiting concentration of 50%, IC50) The concentrations are given in units of nanomole/liter.

Claims

1. Therapeutic application of tricyclic aromatic and saturated benzo[4,5]thieno[2,3- d]pyrimidine derivatives of the formula (I) and their therapeutically acceptable salts

( D

- wherein Rl is hydrogen, phenyl, mono-, di-, tri-, tetra- or penta- substituted phenyl, five- or six-membered substituted or unsubstituted heterocyclic group containing one or more heteroatoms advantageously pyrazole, imidazole, isoxazole, furane, pyrrole, thiophene, thiazole, isothiazole, triazole, pyrane, pyridine, pyrimidine, dioxane, morpholine, thiomorpholine, pyridazine, pyrazine, or piperazine group, R2 is hydrogen, linear or branched alkyl, cycloalkyl, unsubstituted and substituted phenyl group, 'A' is methylene (CH2), methylidene (CH).

2. Use of tricyclic aromatic and saturated benzo[4,5]thieno[2,3-d]pyrimidine compounds of the formula (I) wherein Rl, R2 and 'A' are as mentioned above as active agent of therapeutic preparations of tyrosine-kinase inhibiting action.

3. The compound applied according to the Claim 1 is phenyl-(5,6,7,8-tetrahydro- benzo[4,5]thieno[2,3-d]pyrimidine-4-yl)-amine.

4. The compound applied according to the Claim 1 is (3-chlor-phenyl)-(5,6,7,8- tetrahydro-benzo[4,5]thieno[2,3-d]pyrimidine.

5. The compound applied according to the Claim 1 is 3-(5,6,7,8-tetrahydro- benzo[4,5]thieno[2,3-d]pyrimidine-4-yl-amino)-phenol.

6. The compound applied according to the Claim 1 is (5,6,7, 8-tetrahydro- benzo[4,5]thieno[2,3-d]pyrimidine-4-yl)-(3-trifluormethyl-phenyl)-amine.

7. The compound applied according to the Claim 1 is (3-nitro-phenyl)-(5,6,7,8- tetrahydro-benzo[4,5]thieno[2,3-d]pyrirnidine-4-il)-amine.

8. The compound applied according to the Claim 1 is (3-methoxy-phenyl)-(5, 6.7.8- tetrahydro-benzo[4,5]thieno[2,3-d]pyrimidine-4-yl)-amine.

9. The compound applied according to the Claim 1 is (4-bromine-phenyl)-(5,6,7,8- tetrahydro-benzo[4,5]thieno[2,3-d]pyrimidine-4-yl)-amine.