WO2014013117A1 - Chromenopyrazole diones as cannabinoid quinone derivatives having anti-tumour activity - Google Patents

Abstract

The invention relates to a compound of general formula (I) or a tautomer, pharmaceutically acceptable salt, prodrug or solvate thereof, wherein R¹ is independently selected from hydrogen or C₁-C₁₈ alkyl; R₂ is independently selected from hydrogen, C₁-C₁₈ alkyl or C₅-C₁₈ aryl; and R³ is independently selected from hydrogen or C₁-C₅ alkyl. The invention also relates to the use of said compound of formula (I) for treating cancer or a malignant tumor, such as a blastoma, carcinoma, glioma, leukemia, lymphoma and melanoma, and to a method for treating said cancer or tumour, comprising the administration of a therapeutically effective quantity of the compound of formula (I). Consequently, the invention further relates to a pharmaceutical composition comprising a therapeutically effective quantity of a compound of general formula (I) and at least one pharmaceutically acceptable excipient. In addition, the invention relates to the method for producing said compound of general formula (I) by means of oxidation with [bis(trifluoroacetoxy)iode]benzene of the corresponding precursor phenolic derivative.

Description

 CHROMENOPIRAZOLDIONAS AS CANNABINOID DERIVATIVES OF QUINONAS WITH

 ANTITUMORAL ACTIVITY

DESCRIPTION

Technical Sector

The present invention is encompassed in the field of pharmacology. Specifically, the present invention relates to chromene-derived compounds [4,3-c] pyrazol-6,9-diones, their use for the manufacture of a medicament, and the use of this medicament for an antitumor treatment.

Prior State of the Art

Despite recent advances in chemotherapy and radiation, cancer remains one of the leading causes of death at any age worldwide. Huge efforts have been made and continue to be made in order to obtain new active and safe antitumor agents to administer to patients suffering from cancer.

Quinones are compounds of natural or synthetic origin. Several of them have some physiological functions such as the vitamin K necessary for blood clotting. However, many of them are cytotoxic, which makes them candidates for antitumor drugs. The antineoplastic effects of quinones have been widely described in the literature (C. Asche, Miní-Rev. Med. Chem. 2005, 5, 449-467). For example, anthracyclines that are antibiotics from bacteria of the genus Streptomyces, are used in the treatment of many types of cancers.

Cannabinoids act on the receptors of the endocannabinoid system and can provide therapeutic benefits in various pathologies and in particular they are useful in the treatment of cancer. On the one hand, they have palliative effects: antiemetics and analgesics (RG Pertwee, Br. J. Pharmacol. 2009, 156 (3), 397- 411), and on the other hand, show antitumor properties by themselves (M. Guzman, Nat. Rev. Cancer 2003, 3, 745-765; S. Sarfaraz et al, Cancer Res. 2008, 68, 330-342). Cannabinoids have been shown to inhibit the growth of various tumor cells in laboratory animals, for example glioma (I. Galve-Roperh et al, Nat. Med. 2000, 6, 313-319), melanoma, breast and of prostate (I. Diaz-Laviada, Nat. Rev. ürol. 2011, 8 (10), 553-561). However, it has been described that a cannabinoid, (9) - tetrahydrocannabinol, enhances the growth of breast cancer and metastasis specifically in cells that express low levels of cannabinoid receptors, suggesting that exposure to cannabinoids may increase the incidence of breast cancer, as well as other cancers that do not express cannabinoid receptors (S. Sarfaraz, VM Adhami, D, N. Syed, F. Afaq, H. Muhktar, Cancer Res. 2008, 68, 339-342).

There are multiple biological processes involved in the development of tumor cells and therefore, combined therapies are often used in cancer treatment. The concept of "designed multiple ligands" to address anticancer therapy is to develop a single molecule capable of acting on several therapeutic targets. However, getting this type of molecules is very difficult because the pharmacophoric motifs are associated with different targets. The most commonly used approach consists in the union of said pharmacoforos by means of a spacer. It is much less frequent to achieve a single molecular entity capable of acting on two different targets.

Chromenopyrazole structures such as cannabinoids developed by the research group of the authors of the present invention have been the subject of a patent application (WO2010109050), of which their activity as analgesics has been disclosed, without having been attributed antitumor properties .

Quinones derived from cannabinoid structures with antitumor properties have been described for the first time by R. Mechoulam {Tetrahedron 1968, 24, 5615-5624). In fact three of these structures were the subject of a patent application (WO2005 / 067917). It is para-quinones derived from cannabidiol, of A ⁸ -tetrahydrocannabinol and cannabinol. However, these quinones of tricyclic nature, in spite of showing antiproliferative properties in human cancer cell lines, do not bind to the CB1 cannabinoid receptor as their authors specify, and therefore, exclude a cannabinoid-type antitumor mechanism. More recently, other quinones derived from cannabidiol and their use in the treatment of cancers have been claimed in another patent application (WO2008 / 107878) where anti-proliferative activity is described but in no case have they been found to have cannabinoid activity and / or cannabinoid receptor binding. That is, introducing a quinone into this type of cannabinoid structure led to the loss of cannabinoid properties.

Therefore, the present invention faces the problem of providing new compounds useful for the treatment of cancer. Due to the potential of quinones as anticancer and cannabinoids as anticancer, antiemetic and analgesic, it is of great interest to have molecules that can combine the characteristics of a quinone and a cannabinoid in the same structure. Preferably, said compounds should preserve and even improve the cytotoxic activity of known compounds while avoiding part of the associated adverse side effects.

Description of the invention

The authors of the present invention have found that the compounds of formula (I) act as antitumor agents. The compounds of formula (I) claimed in the present invention, consist of a series of chromenopyrazoldiones that, in addition to having quinone structures, have in their structure a pyrazole ring that proved necessary to maintain a certain affinity for cannabinoid receptors , and that differentiates them from other quinone structures known in the state of the art, such as those previously described by Mechoulam et al. {Tetrahedron 1968, 24, 5615-5624; W02005 / O 67917; WO2008 / 107878).

In a first aspect, the present invention relates to a compound of general formula (I),

Formula (I) or a tautomer, pharmaceutically acceptable salt, prodrug or solvate thereof, where:

- R ¹ is independently selected from hydrogen or alkyl

Ci ^_ Ci ₈ ;

- R ² is independently selected from hydrogen, alkyl Οχ-

Cis or aryl C ₅ -Ci ₈ ;

- R ³ is independently selected from hydrogen or alkyl

Ci-C ₅ .

The term "alkyl" refers, in the present invention, to aliphatic, linear or branched, saturated or unsaturated chains, having 1 to 18 carbon atoms, preferably 1 to 9 carbon atoms. For example, but not limited to, methyl, ethyl, n-propyl, i-propyl, n-butyl, tere-butyl, sec-butyl, n-pentyl, n-hexyl, n-heptyl, 1 ', 1' - dimethylheptyl, 1,2-dimethheptyl, Ι ', Ι'-dimethylethyl, adamantyl, cyclohexyl, 1-cyclopentylhexyl, etc. The alkyl groups may be optionally substituted by one or more substituents such as halogen, hydroxyl or carboxylic acid. Said substituted alkyl groups are for example, but not limited to, 1- (1, 3-dithiolane) hexyl, 1-hydroxyethyl, hydroxymethyl, etc.

The term "aryl" refers, in the present invention, to single or multiple aromatic rings, having between 5 and 18 links in which a proton has been removed from the ring. The aryl groups are for example, but not limited to, phenyl, naphthyl, diphenyl, indenyl, phenanthryl or anthracil. Preferably the aryl group has 5 to 7 carbon atoms and more preferably the aryl group is a phenyl. Aryl radicals may be optionally substituted by one or more substituents such as alkyl (Ci-C _6), halogen, hydroxyl or carboxylic acid. Substituted aryl radicals are, for example, but not limited to 2,4-dichlorophenyl, 1,3-dichlorophenyl, 3,5-difluorophenyl, 3- methoxyphenyl.

The term "halogen" refers, in the present invention, to bromine, chlorine, iodine or fluorine. Preferably chlorine.

According to the invention, the size of the substituents R ¹ to R ³ is important for the functionality of the compounds of formula (I), such as when they are intended to modify, among others, their pharmacokinetic, bioavailability, solubility or stability properties. Thus, possible carbon chain substitutions in R ³ are those in which said substituent does not exceed 5 carbon atoms, while in the case of the substituents of R ¹ and R ² the number of atoms can be quite greater . In fact, the positions defined by the substituents R ¹ and R ² can be functionalized with long carbon chains, such as fatty acids, where the most commonly used include chains of up to 18 carbon atoms. However, substituents such as 1,1-dimethylheptyl (of 9 carbon atoms) constitute a good example of an intermediate carbon chain between the maximum and minimum values contemplated herein. Unless otherwise indicated, the compounds of the invention they also include compounds that differ only in the presence of one or more isotopically enriched atoms. For example, the compounds having the present structures, except for the replacement of a hydrogen with a deuterium or tritium, or the replacement of a carbon with a carbon enriched in ¹³ C or ¹⁴ C or a nitrogen enriched in ¹⁵ N, are within the scope of this invention.

The term "tautomer" or "tautomeric form", as used in the present invention, refers to structural isomers of different energies that are interconvertible via a low energy barrier. For example, protonic tautomers (also known as prototropic tautomers) that include interconversions by migration of a proton, such as keto-enolic or imine-enamine isomerizations. Valencia tautomers include interconversions by reorganization of some bond electrons.

The term "pharmaceutically acceptable salts, solvates, prodrugs" refers to any pharmaceutically acceptable salt, ester, solvate, or any other compound that, when administered to a receptor, is capable of providing (directly or indirectly) a compound as described in This document. However, it will be appreciated that pharmaceutically acceptable salts are also within the scope of the invention since these may be useful in the preparation of pharmaceutically acceptable salts. The preparation of salts, prodrugs and derivatives can be carried out by methods known in the art.

For example, pharmaceutically acceptable salts of compounds provided herein are synthesized by conventional chemical methods from an original compound containing a basic or acidic moiety. Generally, such salts are prepared, for example, by reacting the free acid or base forms of the compounds with a stoichiometric amount of the appropriate base or acid in water or in an organic solvent or in a mix of the two. Generally, non-aqueous media such as ether, ethyl acetate, ethanol, isopropanol or acetonitrile are preferred. Examples of acid addition salts include mineral acid addition salts such as, for example, hydrochloride, hydrobromide, iodhydrate, sulfate, nitrate, phosphate and organic acid addition salts such as, for example, acetate, maleate, fumarate, citrate, oxalate, succinate, tartrate, malate, mandelate, methanesulfonate and p-toluenesulfonate. Examples of base addition salts include inorganic salts such as, for example, sodium, potassium, calcium, ammonium, magnesium, aluminum and lithium salts, and salts of organic bases such as, for example, ethylenediamine, ethanolamine, N, - dialkylene ethanolamine, triethanolamine, glucamine and basic amino acid salts.

Particularly preferred derivatives or prodrugs are those that increase the bioavailability of the compounds of this invention when such compounds are administered to a patient (for example, by making a compound administered orally more easily absorbed by blood), or that enhance the release of the original compound in a biological compartment (for example, the brain or lymphatic system) in relation to the original species.

Any compound that is a prodrug of a compound of formula (I) is within the scope of the invention. The term "prodrug" is used in its broadest sense and encompasses those derivatives that are converted in vivo into the compounds of the invention. Such derivatives will be apparent to those skilled in the art, and include, depending on the functional groups present in the molecule and without limitation, the following derivatives of the compounds present: esters, amino acid esters, phosphate esters, salts sulfonate esters metallic, carbamates, and amides.

The compounds of formula (I) may be in crystalline form as free compounds or as solvates and it is intended that both forms are within the scope of the present invention. The methods of Solvation are generally known within the art. Suitable solvates are pharmaceutically acceptable solvates. In a particular embodiment, the solvate is a hydrate.

The compounds of formula (I) or their salts or solvates are preferably in a pharmaceutically acceptable or substantially pure form. Pharmaceutically acceptable means, among others, that they have a pharmaceutically acceptable level of purity excluding normal pharmaceutical additives such as diluents and carriers, and not including material considered toxic at normal dosage levels. The purity levels for the active ingredient are preferably greater than 50%, more preferably, greater than 70%, more preferably, greater than 90%. In a preferred embodiment, they are greater than 95% of the compound of formula (I), or of its salts, solvates or prodrugs.

The compounds of general formula (I) can be found as tautomers of formula (II) or formula (III). However, in the examples described herein, where R ³ is hydrogen, they have been obtained as compounds of formula (III) when R ² is also hydrogen, and as compounds of formula (II) when R ² is an alkyl group, as they are the methyl or ethyl groups.

According to a preferred embodiment, the present invention relates to a compound of general formula (II),

Formula (II) or a tautomer, pharmaceutically acceptable salt, prodrug or solvate thereof, wherein R ¹ , R ² and R ³ are defined as above:

- R ¹ is independently selected from hydrogen or Ci Ci ^_ _8;

- R ² is independently selected from hydrogen, Οχ-Cis alkyl or C ₅ -Ci ₈ aryl;

- R ³ is independently selected from hydrogen or C1-C5 alkyl.

In another preferred embodiment, the present invention relates to a compound of general formula (III),

Formula (III) or a tautomer, pharmaceutically acceptable salt, prodrug or solvate thereof, wherein R ¹ , R ² and R ³ are defined as above.

In a preferred example, the compound of formula (I), as well as any of its tautomeric forms of formula (II) or (III), is a compound where R ³ is a hydrogen.

In another preferred example, the compound of formula (I), as well as any of its tautomeric forms of formula (II) or (III), is a compound where R ¹ is a 1,1-dimethylheptyl group.

In another preferred example, the compound of formula (I), as well as any of its tautomeric forms of formula (II) or (III), is a compound where R ² is a group selected from hydrogen, methyl or ethyl.

In another preferred example, the compound of formula (I), as well as any of its tautomeric forms of formula (II) or (III), is a compound where R ³ is a hydrogen and R ¹ is a 1,1-dimethylheptyl group .

In another preferred example, the compound of formula (I), as well as any of its tautomeric forms of formula (II) or (III), is a compound where R ³ is a hydrogen and R ² is a group selected from hydrogen, methyl or ethyl. Even more preferred, it is a compound of formula (II) when R ² is methyl or ethyl.

In another preferred example, the compound of formula (I), as well as any of its tautomeric forms of formula (II) or (III), is a compound where R ¹ is a 1,1-dimethylheptyl group and R ² is a group selected from hydrogen, methyl or ethyl.

In another preferred example, the compound of formula (I), as well as any of its tautomeric forms of formula (II) or (III), is a compound where R ³ is a hydrogen, R ¹ is a 1,1-dimethylheptyl group and R ² is a group selected from hydrogen, methyl or ethyl.

According to a preferred embodiment, the compound of formula (I) or any of its previous preferred embodiments and examples, is selected from one of the group consisting of: a) 7- (1 ', 1' -dimethylheptyl) - 1, -dihydro-4, -dimethylchromen [4,3- c] pyrazol-6, 9-dione; b) 7- (1 ', 1' -dimethylheptyl) -2, 4-dihydro-2, 4, 4-trimethylchromen [4,3- c] pyrazol-6, 9-dione; c) 7- (1 ', 1' -dimethylheptyl) -2-ethyl-2, 4-dihydro-4, 4-dimethylchromen [4, 3-c] pyrazol-6, 9-dione; or a tautomer, pharmaceutically acceptable salt, prodrug or solvate thereof.

Said compound defined in section a) above, consists of a compound of general formula (I) that responds to the structure of formula (II) where R ¹ is 1,1-dimethylheptyl, R ² is hydrogen and R ³ is hydrogen.

Said compound defined in section b) above, consists of a compound of general formula (I) that responds to the structure of formula (III) where R ¹ is 1,1-dimethylheptyl, R ² is methyl and R ³ is hydrogen.

Said compound defined in section c) above, consists of a compound of general formula (I) that responds to the structure of formula (III) where R ¹ is 1,1-dimethylheptyl, R ² is ethyl and R ³ is hydrogen.

According to the present specification, any of the compounds defined above, ie those compounds that respond to the general formula (I) (including the compounds of general formulas (II) or (III), or any of the preferred embodiments or examples), they may also be referred to herein as "compound or compounds of the invention".

In the present invention it has been found that the compounds of the invention have antiproliferative activity. And in addition, the cannabinoid activity of the compounds of formula (I) is described based on in vitro tests of binding to human CB1 or CB2 receptors. The compounds of formula (I) and their pharmaceutically acceptable salts, prodrugs or solvates thereof, can therefore be used in the prevention and / or treatment of a disorder or a proliferative disease. Pharmaceutical compositions containing a therapeutically effective amount of a compound of formula (I), its pharmaceutically acceptable salts, prodrugs or solvates thereof, together with pharmaceutically acceptable carriers, constitute an additional aspect of the present invention That is, a second aspect of the invention relates to a pharmaceutical composition comprising at least one pharmaceutically acceptable carrier and a therapeutically effective amount of at least one compound of the invention. Hereinafter, said pharmaceutical composition may also be referred to as "pharmaceutical composition of the invention".

The term "vehicle" refers to a diluent, adjuvant or excipient with which the active substance is administered. Such pharmaceutical vehicles may be sterile liquids, such as water and oils, including those of petroleum, animal, vegetable or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil and the like. Preferably water or aqueous solutions of saline solution and aqueous solutions of dextrose and glycerol are used as vehicles, particularly for injectable solutions. Suitable pharmaceutical vehicles are described in "Remington's Pharmaceutical Sciences" by EW Martin, 1995. Preferably, the vehicles of the invention are approved by the regulatory agency of a state government or a federal government, or are listed in the United States Pharmacopoeia, in the European Pharmacopoeia or other pharmacopoeia generally recognized for use in animals, and more particularly in humans.

The amount of compound of the invention, its pharmaceutically acceptable salts, prodrugs or solvates thereof, therapeutically effective to be administered (also referred to herein as therapeutically effective or effective amount), as well as its dosage to treat a pathological state with said Compounds will depend on numerous factors, including age, patient status, disease severity, route and frequency of administration, modulator compound to be used, etc.

The compounds and pharmaceutical compositions of this invention can be used alone or together with other drugs to provide a combination therapy. The other drugs can form part of the same pharmaceutical composition, or be provided as a separate pharmaceutical composition, for administration at the same time or at a different time. Examples of pharmaceutical compositions include any solid composition (tablets, pills, capsules, granules, etc.) or liquid (solutions, suspensions or emulsions) for oral, topical or parenteral administration.

A third aspect of the present invention relates to the use of a compound of the invention for the manufacture of a pharmaceutical composition.

According to a preferred embodiment, the compounds of formula (I) have an antiproliferative activity.

According to the characteristics of some embodiments, antiproliferative activity refers to the proliferation of malignant tumors.

According to the characteristics of some embodiments, malignant tumors refer to blastoma, carcinoma, glioma, leukemia, lymphoma, melanoma.

A fourth aspect of the invention thus relates to a compound of the invention, or a pharmaceutical composition of the invention, for use in the treatment of a cancer or a malignant tumor, where preferably the cancer or malignant tumor is selected from blastoma. , carcinoma, glioma, leukemia, lymphoma and melanoma. Taking into account the properties of the compounds of the invention, their use in the treatment of cancers or malignant tumors may be advantageous because they combine said cancer or tumor treatment with additional effects derived from their cannabinoid activity (e.g. antiemetic and / or analgesic effects), where such additional effects may be beneficial for the patient by reducing side effects of a cancer treatment such as emesis and / or pain. In a preferred embodiment, said cancer or malignant tumor is selected from a malignant liver tumor (eg a hepatocarcinoma) or a malignant prostate tumor (eg a prostate cancer). According to the present description, the compound of the invention, or the pharmaceutical composition of the invention, for use in the treatment of a cancer or a malignant tumor, can obviously be understood as a method of treating said cancer or tumor, which comprises administration to a subject of a therapeutically effective amount of said compound or pharmaceutical composition of the invention. In other words, the present invention also relates to a method of treating a cancer or a malignant tumor (preferably selected from blastoma, carcinoma, glioma, leukemia, lymphoma and melanoma; and even more preferably, between a hepatocarcinoma or a cancer prostate), which comprises administering to a subject the compound of the invention in a therapeutically effective amount, or a pharmaceutical composition of the invention comprising the compound of the invention in a therapeutically effective amount.

Also, this aspect of the present invention also contemplates the use of the compound of the invention, or of the pharmaceutical composition of the invention, for the manufacture of a pharmaceutical composition, for the treatment of said cancers or malignant tumors. This means that the use of a compound of the invention for the manufacture of a pharmaceutical composition for the treatment of a cancer or a malignant tumor, where the cancer or malignant tumor is preferably selected from blastoma, carcinoma, is also the object of the invention. glioma, leukemia, lymphoma and melanoma; and more preferably said cancer or tumor is a hepatocarcinoma or a prostate cancer.

A fifth aspect of the invention relates to a process for obtaining a compound of general formula (I), which comprises an oxidation with [bis (trifluoroacetoxy) iodine] benzene of a compound of general formula (IV),

Formula (IV) where the substituents R ¹ , R ² and R ³ are defined as above:

R ¹ is independently selected from hydrogen or Ci Ci ^_ _8;

R ² is independently selected from hydrogen, Οχ-Cis alkyl or C ₅ -Ci ₈ aryl;

R is independently selected from hydrogen or C1-C5 alkyl.

Brief description of the figures

Figure 1. Effect of the compounds of examples 1, 2 and 3 of the present description on cell viability. The HepG2 cell lines (derived from hepatocellular carcinoma), LNCaP (derived from androgen dependent prostate cancer) and PC-3 (derived from androgen independent prostate cancer), were treated with increasing doses of the compounds of examples 1, 2 and 3 for 48 hours after which the viability with MT was determined. The data are averages of two experiments performed in triplicate + D.E. (Standard deviation)

Figure 2. Effect of the compound of example 1 of the present description on the cell cycle. The LNCaP and PC-3 cell lines were treated for 24 hours with the doses of the compound of the Example 1 to which a 50% decrease in cell viability was obtained. A, After treatment, the cell cycle was analyzed by flow cytometry. B, After the treatment, the cells were stained with Annexin V and with propidium iodide, determining the different cell populations by flow cytometry in order to detect the cells in apoptosis. The image is representative of two independent tests performed in duplicate.

Figure 3. In vivo antitumor effect of the compound of example 1 of the present description on prostate cancer lines. Nu / nu atomic mice were injected subcutaneously with LNCaP cells (A) or PC-3 cells (B). When the tumors reached a size of 150 mm ³ , a 15-day treatment with vehicle (Control) or 2 mg / Kg of the example 1 compound administered intraperitoneally was started. The tumor was measured daily. The figure shows the growth curves of the tumors from the beginning of the treatment. The results are expressed as both by one of the initial tumor size and correspond to the mean ± SE (Standard Error) of 6 mice per group. To the right of each graph is shown a representative image of a control tumor and one treated with the compound of example 1 of each of the two cell lines used

The following examples and drawings are provided by way of illustration, and are not intended to be limiting of the present invention.

EXAMPLES

For the synthesis of the compounds of formula (I) of the present invention, 4,4-dimethyl-7- (H or alkyl) -dihydrochromen [4, 3-c] pyrazol-9- has been used as starting products corresponding oles. The 4,4-dimethyl-7- (H or alkyl) -dihydrochromen [4, 3- c] pyrazole-9-starting olives can be obtained by a combination of synthetic reactions known in the state of the art such as those mentioned in the article by Cumella J. et al. ChemMedChem 2012, 7, 452-463.

General procedure for the synthesis of para-q inones derived from chromene [4,3-cJpirazol.

The 4,4-dimethyl dihydrochromen [4,3-c] pyrazol-6,9-diones of formula (I) of the present invention are obtained by oxidation of the 4,4-dimethyl-7- (H or alkyl) -dihydrochromen [4,3-c] pyrazol-9-oles corresponding. This oxidation occurs regiospecifically due to the use of [bis (trifluoroacetoxy) iodine] benzene (BTIB) which allows to obtain the desired para-quinone.

This procedure is summarized in the following scheme (I)

 SCHEME (I)

Example 1.- Preparation and obtaining 7- (1 ', 1' -dimethylheptyl) -1, 4- dihydro-4, 4-dimethylchromen [4, 3-c] pyrazol-6, 9-dione

To a solution of 7- (1 ', 1' -dimethylheptyl) -1, 4-dihydro-4, 4-dimethylchromen [4, 3-c] pyrazol-9-ol (0.1330 g, 0.38 mmol) in CH ₃ CN / H ₂ 0 (6: 1, 2.5 mL) a solution of [bis (trifluoroacetoxy) iodine] benzene (BTIB) (0.490 g, 1.14 mmol) in 2 mL of CH ₃ is added dropwise CN / H ₂ 0 (6: 1). The reaction is stirred for 15 minutes at room temperature observing that the solution acquires an intense red color. The reaction mixture is neutralized with a saturated solution of NaHC0 ₃ and extracted with diethyl ether. The organic phase is washed with water and dried over anhydrous MgSO ₄ . The solvent is evaporated in vacuo and the resulting mixture is purified by chromatographic column using as eluent Hex / AcOEt (1: 2). Rto. : 21% Red solid, mp: 85.7 ° C. ¹ H-NMR (CDC1 ₃ , 300 MHz) δ: 8.41 (sa, 1H, H); 7.40 (s, 1 H, 3-H); 6.69 (s, 1H, 8-H); 1.59 (sa, 6H, OC (CH ₃ ) ₂ ); 1.55-1.48 (m, 2H, ^2'- H); 1.30 (s, 6H, C (CH _{3) 2);} 1.27 to 1.23 (m, 6H, 3 ^'- H, 4'-H, 5'-H); 1.19 (sa, 2H, 6'H); 0.86-0.82 (m, 3H, 7'-H) ppm. ¹³ C-NMR (CDC1 ₃ ) 5: 184.1 (6-C); 180.9 (9-C); 160.2 (5a-C); 161.2 (7-C); 137.8 (9b-C); 132.0 (8-C); 130.4 (9a-C); 129.5 (3-C); 113.8 (3a-C); 78.6 (OC (CH ₃ ) ₂ ); 43.3 ( ^2'- C); 30.9 (l ^'-C); 29.6; 28.7 and 27.4 (3 ^{'-C 4'} -C and 5 ^'-C); 25.1 (C (CH ₃ ) ₂ ); 23.2 (OC (CH ₃ ) ₂ ); 21.8 ^(6' - C); 14.0 (7 ^'- C) ppm. HPLC / MS *: Gradient: 70% A to 100% A; MS (ES ⁺ ) m / z: 357 (98%) [M + H] ⁺ ; t _R (retention time): 3.37 min. Anal .: C ₂ iH ₂₈ N ₂ 0 ₃ (356, 46 g / mol) Theoretical: C, 70.76%; H, 7.92%. Found: C, 71.03%; H, 8.24%.

Example 2.- Preparation and obtaining 7- (1 ', 1' -dimethylheptyl) -2, 4- dihydro-2, 4, 4-trimethylchromen [4, 3-c] pyrazol-6, 9-dione

The synthesis of this compound is based on the procedure described in Example 1 following the reaction conditions detailed below: 7- (1 ', 1' -dimethylheptyl) -2, 4-dihydro-2, 4, 4-trimethylchromen [ 4, 3-c] pyrazol-9-ol (0.035 g, 0.1 mmol),

[bis (trifluoroacetoxy) iodine] benzene (BTIB) (0.129 g, 0.3 mmol). Eluent Hex / AcOEt (1: 4). Rto. : 33% Red solid, mp: 138.2 ° C. ¹ H-NMR (CDCI ₃ , 300 MHz) δ: 7.36 (s, 1H, 3-H); 6.61 (s, 1H, 8-H); 3.95

(s, 3H, NCH ₃ ,); 1.84 (s, 6H, OC (CH ₃ ) ₂ ); 1.70 (sa, 2H, 2'-H); 1.33 (s, 6H, C (CH ₃ ) ₂ ); 1.26 (sa, 6H, 3'-H, 4'-H, 5'-H); 1.16-1.13 (m, 2H, 6'H); 0.85 (t, J = 6.9 Hz, 3H, 7'-H) ppm. ¹³ C-NMR (CDC1 ₃ ) 5: 186.2 (6- C); 181.4 (9-C); 160.2 (5a-C); 155.0 (7-C); 139.6 (9b-C); 135.4 (8-C); 129.3 (9a-C); 126.9 (3-C); 115.7 (3a-C); 81.5 (OC (CH ₃ ) ₂ ); 43.0

(NCH ₃ ); 40.9 (2'-C); 31.7 (l'-C);30.8; 29.3 and 28.1 (3'-C, 4'-C and 5'-C); 27.5 (C (CH ₃ ) ₂ ); 23.4 (OC (CH ₃ ) ₂ ); 22.9 (6'-C); 14.6 (7'-C) ppm. HPLC / MS *: Gradient: 80% A to 100% A; MS (ES ⁺ ) m / z: 371 (100%) [M + H] ⁺ ; t _R : 2.43 min. Anal .: C ₂₂ H ₃ oN ₂ 0 ₃ (370, 49 g / mol) Theoretical: C, 71.32%; H, 8.16%. Found: C, 70.98%; H, 8.31%.

Example 3.- Preparation and obtaining 7- (1 ', 1' -dimethylheptyl) -2-ethyl-2, 4-dihydro-4, 4-dimethylchromen [4, 3-c] pyrazol-6, 9-dione

The synthesis of this compound is based on the procedure described in Example 1 following the reaction conditions detailed in continued: 7- (1 ', 1' -dimethylheptyl) -2-ethyl-2, -dihydro-4, 4- dimethylchromen [4, 3-c] pyrazol-9-ol (0.061 g, 0.16 mmol),

[bis (trifluoroacetoxy) iodine] benzene (BTIB) (0.212 g, 0.49 mmol). Eluent Hex / AcOEt (2: 3). Rto. : 36% Red solid, mp: 83.9 ° C. ¹ H-NMR (CDC1 ₃ , 300 MHz) δ: 7.15 (s, 1H, 3-H); 6.50 (s, 1H, 8-H); 4.28

(c, J = 7.4 Hz, 2H, NCfí ₂ CH ₃ ,); 1.72 (s, 6H, OC (CH ₃ ) ₂ ); 1.61-1.58 (m, 2H, 2'-H); 1.49 (t, J = 7.4 Hz, 3H, NCH ₂ Cfí ₃ ); 1.30 (s, 6H, C (CH ₃ ) ₂ ); 1.23-1.10 (m, 8H, 3'-H, 4'-H, 5'-H and 6'-H); 0.89 (t, J ^" = 6.9 Hz, 3H, 7'-H) ppm. ¹³ C-NMR (CDC1 ₃ ) 5: 183.4 (6-C); 180.5 (9-C) ; 152.4 (5a-C); 151.4 (7-C); 137.2 (9b-C); 133.1 (8-C); 122.1 (3-C); 120.9 ( 3a-C); 110.6 (9a-C); 79.9 (OC (CH ₃ ) ₂ ); 46.6 (NCH ₂ CH ₃ ); 39.7 (2'-C); 37.6 ( 1'- C); 31.6; 29.7 and 28.7 (3'-C, 4'-C and 5'-C); 26.6 (OC (CH ₃ ) ₂ ); 24.1

(C (CH ₃ ) ₂ ); 21.5 (6'-C); 14.6 (NCH ₂ CH ₃ ); 13.0 (7'-C) ppm. HPLC / MS *: Gradient: 70% A to 100% A; MS (ES ⁺ ) m / z: 385 (96%) [M + H] ⁺ ; t _R : 3.86 min. Anal .: C ₂₃ H ₃₂ N ₂ 0 ₃ (384, 51 g / mol) Theoretical: C, 71.84%; H, 8.39%. Found: C, 72.01%; H, 8.62%.

* For all HPLC / MS data the mobile phases used are: A (MeCN + 0.08% formic acid) and B (H ₂ 0 + 1% formic acid). The gradient used is indicated in each case, in 5 minutes at a flow of 0.25 mL / min and λ = 254 nm.

C. Biological tests

C. 1. Affinity for cannabinoid receptors.

In the present invention, the cannabinoid activity of the compounds of formula (I) was assessed by carrying out in vitro tests of displacement of the cannabinoid radioligand [ ³ H] -CP55940 (10μΜ) and [ ³ H] -WIN 55,212-2 (50μΜ ) in cells transfected by human CB1 or CB2 receptors. The affinity constants of the compound of example 1 of the present invention are given below by way of illustration: 7- (1 ', 1' -dimethylheptyl) -1, 4-dihydro-4, 4-dimethylchromen [4, 3- c] pyrazol-6, 9-dione (Example 1) 324 nM (CB1), 134 nM (CB2).

C.2. Antiproliferative effect in vitro and in vivo. The antiproliferative effect of 7- (1 ', 1' -dimethylheptyl) -1, 4- dihydro-4, -dimethylchromen [4, 3-c] pyrazol-6, 9-dione (example 1), of 7- (1 ', 1' -dimethylheptyl) -2, 4-dihydro-2, 4, 4-trimethylchromen [4,3- c] pyrazol-6, 9-dione (example 2) and 7- (1 ', 1'-dimethylheptyl) -2- ethyl-2, 4-dihydro-4, 4-dimethylchromen [4, 3-c] pyrazol-6, 9-dione (eg emplo 3) of the present invention was studied in vitro in lines Cells derived from hepatocellular carcinoma and prostate cancer using the following methods.

Maintenance and cell culture. LNCaP and PC-3 cells were routinely grown in RPMI 1640 medium supplemented with 10% fetal bovine serum (FBS), 100 IU / ml sodium penicillin G, 100 g / ml streptomycin sulfate and 0.25 g / μΐ amphotericin B (Invitrogen, Paisley, UK). The cells were kept in a humid environment, at 37 ° C and with 5% C0 ₂ .

Cell proliferation test. After treatment of the cells with the different compounds, they were incubated with 0.3 mg / ml of 3- (4,5-dimethylthiazol-2-yl) 2, 5-diphenyl tetrazolium bromide (MTT) at 37 ° C In darkness for lh. Subsequently the crystals were solubilized with 500 μΐ of acid isopropanol and gentle agitation. Once the coloration was homogeneous, the different absorbances at 570 nm were evaluated with a plate spectrophotometer (ELX 800 Bio-Tek Instruments, INC). The absorbance measurement obtained in the control well was considered as 100% of living cells, and therefore the values obtained in the different treatments were referred to this data.

Analysis of the cell cycle by flow cytometry. To perform this technique the cells were seeded, treated for the estimated time and subsequently lifted with trypsin. They were then centrifuged at 400 g for 5 minutes and resuspended in 70% ethanol. The cell suspension was maintained for 30 minutes at -20 ° C so that the cells were fixed and permeabilized. After that time the cells were centrifuged again to remove the ethanol and be able to be washed with PBS. Finally, they were resuspended in PBS with 20 μg / μl of propidium iodide and 0.2 mg / ml of RNAse (in order to degrade the RNA so that it does not interfere with the extent of flowering). Cells were analyzed with a FACSCalibur cytometer (Becton Dickinson Labware, New Yersey, USA). For the quantification of the percentages of cells in the different phases of the cell cycle, the WinMDI 2.8 program was used, following its instructions.

Union of propidium iodide and Annex V-FITC. The Annexin V-FITC apoptosis detection kit (BD, New Jersey) commercial kit was used. The cells were seeded in six-well plates with a density of 5,000 cells / cm ² . Once the treatments were carried out, the cells were trypsinized and centrifuged at 400 g for 5 minutes. Subsequently, the cells were incubated 15 minutes in the dark with 5 μΐ of Annexin V, conjugated with fluorescein isothiocyanate (FITC) and 5 μΐ of propidium iodide dissolved in 100 μΐ of binding buffer (0.1 M Hepes pH 7.4 , 1.4 M NaCl, 25 mM CaCl ₂ ). After the incubation time, 400 μΐ of binding buffer was added and analyzed with a FACSCalibur cytometer.

The in vivo antiproliferative effect of 7- (1 ', 1'-dimethylheptyl) -1,4-dihydro-4,4-dimethylchromen [4,3-c] pyrazol-6,9-dione (example 1) was performed according to the method described below, by way of illustration.

In vivo studies Four-week-old male nu / nu (BALB / c01aHsd-Foxnl ^nu ) male mice (Harían Ibérica, Barcelona, Spain) were used. The animals were kept in a pathogen-free zone (SPF) in the Animal Experimentation Center of the University of Alcalá, with constant light / dark cycles of 12 hours.

The studies were carried out in accordance with the Spanish regulation for the use and care of experimental animals and according to the guidelines of the European Union related to animal research (Royal Decree 1201/2005 of October 10). The Mice were injected subcutaneously in the right flank, with 0.1 ml of PBS buffer with 10 million PC-3 cells or LNCaP. Approximately one week later, when the tumors reached a size of 150 mm ³ , the animals were divided into two groups (n = 8) that were treated daily with vehicle (control) or with 1.5 mg / Kg of compound object of the This patent administered intraperitoneally. The tumor size (which is calculated with the formula 0.523xW ² xL; W being the width of the tumor and L the length thereof) was controlled and calculated daily using a calibrator. After fifteen days of treatment the animals were sacrificed and the tumors were removed calculating the weight and size of the tumor at the end of the treatment.

Biological Results

In vitro antiproliferative activity. The results of the in vitro antiproliferative assays of 7- (1 ', 1' -dimethylheptyl) -1, 4-dihydro-4, 4-dimethylchromen [4, 3-c] pyrazole- are presented below and by way of illustration 6, 9-dione (example 1), of 7- (l ', l'-dimethylheptyl) -2, 4-dihydro-2, 4, 4-trimethylchromen [4, 3-c] pyrazole-6, 9- dione (example 2) and 7- (1 ', 1' -dimethylheptyl) -2-ethyl-2, 4- dihydro-4, 4-dimethylchromen [4, 3-c] pyrazol-6, 9-dione ( Example 3) of the present invention on cell proliferation.

The effect of the compounds examples 1, 2 and 3 on cell viability was analyzed using three cell lines derived from human tumors. The HepG2 cell line comes from a differentiated hepatocellular carcinoma, the LNCaP cell line comes from an androgen-dependent prostate tumor and the PC-3 cell line comes from an androgen-independent and therefore more aggressive prostate cancer. As can be seen in Figure 1, the three compounds were more effective in prostate cell lines than in hepatocellular carcinoma, highlighting the example 1 compound of the present patent as the most potent in all cell lines with an IC50 of 30 μΜ in HepG2 and 15 μΜ cells in LNCaP and PC-3 cells. In order to characterize the effect of the quinone example 1 of the present patent on the cell cycle, The cells were treated with the IC50 of said compound (15 μΜ) for 24 hours, followed by a cycle analysis by flow cytometry. The results shown in Figure 2A indicate that compound PM49 produces an increase in the subGO cell population corresponding to cells that have undergone apoptosis. This increase is greater in LNCaP cells than in PC-3s. Likewise, cytometry analysis of cells stained with Annexin V and propidium iodide revealed that the example 1 compound induced apoptosis both in the LNCaP cell line and in the cells. PC-3, as doubly stained cells increased (upper right quadrant of each figure). Again, the effect was superior in LNCaP cells, in which the population of cells in early apoptosis (lower right quadrant) quadrupled and cells in late apoptosis (upper right quadrant) tripled (Figure 2B).

Antitumor activity in vivo. Illustratively, the antitumor activity of 7- (1 ', 1' -dimethylheptyl) -1, 4-dihydro-4, 4-dimethylchromen [4, 3-c] pyrazol-6, 9-dione (example 1) is presented ) in vivo. It was analyzed by subcutaneous induction of xenograph tumors in immunocompromised mice administering the example compound of the present patent (2 mg / Kg) dissolved in olive oil, intraperitoneally for 15 days consecutively. Control animals were given the same volume of olive oil

(vehicle) than to treated animals. As seen in Figure 3, administration of the compound example 1 of the present patent reduced tumor growth by almost 40% in PC-3 cells while in LNCaP cells almost completely blocked tumor growth, the tumors remaining at the same size as at the beginning of the treatment. The images of the dissected tumors at the end of the treatment (to the right of each graph) show the reduction in the size of the tumors of the animals treated with 7-

(1 ', 1' -dimethylheptyl) -1, 4-dihydro-4, 4-dimethylchromen [4, 3-c] pyrazol-6, 9-dione (example 1) compared to control animals.

These results indicate that cannabinoid-derived quinones have an antitumor effect on various cell lines. This The effect is especially evident as we illustrate with the compound example 1 of the present patent and the cell line derived from prostate cancer LNCaP, one of the lines generally resistant to various compounds. Therefore the results shown here indicate that the compounds claimed in the present patent could have therapeutic applications for the treatment of different tumors being particularly effective for androgen-dependent prostate tumors.

Claims

one . A compound of general formula (I),

Formula (I) or a tautomer, pharmaceutically acceptable salt, prodrug or solvate thereof, where:

- R ¹ is independently selected from hydrogen or alkyl

Ci ^_ Ci ₈ ;

- R ² is independently selected from hydrogen, alkyl Οχ-

Cis or aryl C ₅ -Ci ₈ ;

- R ³ is independently selected from hydrogen or Ci-C ₅ alkyl.

2. A compound according to claim 1 of general formula (II),

Formula (II) or a tautomer, pharmaceutically acceptable salt, solvate prodrug thereof, wherein R ¹ , R ² and R ³ are defined as in claim 1.

3 . A compound according to claim 1 of general formula (III)

Formula (III) or a tautomer, pharmaceutically acceptable salt, solvate prodrug thereof, wherein R ¹ , R ² and R ³ are defined as in claim 1.

A compound according to one of claims 1 to 3, wherein a hydrogen

5 . A compound according to one of claims 1 to 4, wherein R ¹ is a 1,1-dimethylheptyl group.

6. A compound according to one of claims 1 to 5, wherein R ² is a group selected from hydrogen, methyl or ethyl.

A compound according to one of claims 1 to 6, which is selected from the group consisting of: a) 7- (1 ', 1' -dimethylheptyl) -1, 4-dihydro-4, 4-dimethylchromen [4,3- c] pyrazole-6,9-dione; b) 7- (1 ', 1' -dimethylheptyl) -2, -dihydro-2,4, -trimethylchromen [4,3- c] pyrazol-6, 9-dione; c) - (1 ', 1' -dimethylheptyl) -2-ethyl-2, 4-dihydro-4, 4-dimethylchromen [4, 3-c] pyrazol-6, 9-dione.

8. A pharmaceutical composition characterized in that it comprises at least one pharmaceutically acceptable excipient and a therapeutically effective amount of at least one compound of formula generates (I) defined in any one of claims 1 to 7.

9. Use of a compound defined in any one of claims 1 to 7 for the manufacture of a pharmaceutical composition.

10. Use of a compound defined in any one of claims 1 to 7 for the manufacture of a pharmaceutical composition for treating a cancer or a malignant tumor.

eleven . Use of a compound according to claim 10, wherein the cancer or malignant tumor is selected from blastoma, carcinoma, glioma, leukemia, lymphoma and melanoma.

12. Use of a compound according to claim 10, wherein the cancer or malignant tumor is selected from a hepatocarcinoma or a prostate cancer.

13. A compound defined in any one of claims 1 to 7, or a pharmaceutical composition defined in claim 8, for use in the treatment of a cancer or a malignant tumor.

14. A compound or pharmaceutical composition according to claim 13, wherein the cancer or malignant tumor is selected from blastoma, carcinoma, glioma, leukemia, lymphoma and melanoma.

fifteen . A compound or a pharmaceutical composition according to claim 13, wherein the cancer or malignant tumor is selected between a hepatocarcinoma or a prostate cancer.

16. A method of treating a cancer or a malignant tumor comprising administering to a patient affected by said cancer or malignant tumor a therapeutically effective amount of a compound of one of claims 1 to 7.

17. A process for obtaining a compound of general formula (I), characterized in that it comprises an oxidation with [bis (trifluoroacetoxy) iodine] benzene of a compound of general formula (IV),

Formula (IV) wherein the substituents R ¹ , R ² and R ³ are defined as in claim 1.