WO2010103473A1

WO2010103473A1 - Method of treatment of polycystic diseases and chronic lymphocytic leukemia

Info

Publication number: WO2010103473A1
Application number: PCT/IB2010/051035
Authority: WO
Inventors: Hervé Galons; Nassima Oumata; Laurent Meijer; Christian Berthou; Karine Lester
Original assignee: Chu De Brest; Universite Paris Descartes; Institut National De La Sante Et De La Recherche Medicale (Inserm); Centre National De La Recherche Scientifique; Universite De Bretagne Occidentale
Priority date: 2009-03-10
Filing date: 2010-03-10
Publication date: 2010-09-16

Abstract

The present invention concerns a method of treatment of polycystic diseases or of reducing and/or avoiding cyst formation in a patient with cystic disease, which comprises at least one step consisting in administering to said patient an effective amount of a 2,6,9-trisubstituted purine compound of formula (VII): wherein A represents alternatively a (A.1) or (A.2) group.

Description

METHOD OF TREATMENT OF POLYCYSTIC DISEASES AND CHRONIC LYMPHOCYTIC LEUKEMIA

FIELD OF THE INVENTION

The present invention generally relates to methods and compositions for the treatment of polycystic kidney disease (PKD). The compounds and compositions according to the invention may be used to inhibit or reduce the formation of cysts in patient with polycystic kidney disease.

BACKGROUND OF THE INVENTION

Polycystic kidney disease (PKD) is a progressive, genetic disorder of the kidneys. The causes of the disease are mainly inheritable genetic DNA mutation(s). PKD is characterized by the presence of multiple cysts (hence, "polycystic") in both kidneys, and in other organs (e.g. liver, pancreas). These cysts grow and multiply over time, also causing the mass of the kidney to increase dramatically.

About 10 to 15 percent of the people with PKD may have alterations in the vasculature with intracranial aneurysms. In time, virtually all of the nephrons of both kidneys either become cystic or are compressed, distorted and rendered increasingly ineffective by the pressure of adjacent cysts. In the process, both kidneys enlarge massively, while function decreases until end-stage renal failure. PKD appears to occur all over the world among all socioeconomic and ethnic groups. Nevertheless, the severity of the renal disease in PKD is highly variable with a large interfamilial and intrafamilial variability. Men and women are affected equally. Autosomal-dominant PKD (ADPKD) (incidence 1 in 800 live births) is the most frequent form of inherited PKD which also comprises autosomal recessive PKD (ARPKD) (incidence 1 in 20,000 live births). Acquired PKD also exists but is much rarer than inherited PKD.

PKD is one of the most common, life-threatening genetic disease, affecting 600,000 Americans, about 800,000 Europeans, 85,000 French people and roughly 12.5 million people worldwide. 6,000 new cases are diagnosed each year in the US.

PKD associated symptoms include abnormally high blood pressure (hypertension), hematuria, fatigue, pain or urinary infections. Ultimately, the disease leads to the progressive loss of kidney function, causing end-stage renal disease (ESRD) for which dialysis and transplantation are the only forms of treatment.

No effective treatment is available to prevent the onset and development of the disease. However, some treatments aim at the reduction of symptoms and the prevention of complications, to ease the PKD symptoms and prolong life. Treatment is confined to complications such as infection, stones, bleeding and hypertension. Blood pressure control and prevention of kidney stones and infections have improved the prognosis for those with PKD. Dialysis and transplantation are the only alternatives. The last decade, however, has witnessed a significant effort to improve the prognosis of patients with ADPKD. Patients with chronic renal failure are now offered different therapies such as a low-protein diet, angiotensin II converting enzyme inhibitors or receptor blockers, and statins.

At a cellular level, PKD is primarily characterized by abnormal cell proliferation and apoptosis causing the growth of fluid-filled cysts in renal tubules. At the molecular level, gene mutations are responsible for abnormal cell polarity and fluid secretion, abnormal adhesion and cell-matrix interactions, abnormal signal transduction (cell differentiation).

PKD most frequently mutated proteins are Polycystin-1 and Polycystin-2. Both form complexes that are found at the cell-matrix interface, cell-cell contacts, and luminal cilium. Epithelial cilia function as sensors of the extracellular environment and interact with membrane and cytoskeleton. They transduce signals by means of intracellular phosphorylation cascades.

Mutated genes (PKDl, PKD2) have been identified in 1994 and 1995. After almost 15 years of exploratory research, PKD pathogenesis is still incompletely understood.

It follows that even if several potential therapies are now emerging that promise to effectively prevent cyst formation and progression (including signal transduction modulators, hormone modulators, anti-inflammatory agents, ACE inhibitors or anti-oxidants), altogether, there is an important need for an efficient treatment of PKD.

Protein kinases constitute a large family of structurally related enzymes that are responsible for the control of a wide variety of signal transduction processes within the cell. These enzymes function by catalyzing the transfer of a phosphate group from ATP to serine, threonine or tyrosine amino acid residues of substrate proteins.

It has been recently reported in Bukanov, N. O et al., 2006 ["Long- lasting arrest of murine polycystic kidney disease with CDK inhibitor Roscovitine" Nature, 2006, vol. 444, 949-952] that CDK inhibitors may be used to treat renal diseases such as polycystic kidney disease. In WO 98/05335, 2,6,9-trisubstituted purine compounds are described, which are useful for inhibiting cell proliferation disorders and as antifungal agent.

In WO 2003/022805, heterocycle substituted purines are disclosed as antiproliferative agents.

Moreover WO 2008/051502 describes methods to treat or ameliorate cystic diseases comprising administering purine derivatives. However, Roscovitine inhibits the formation of cysts at a much higher dose than the compounds of the invention (which are at least 20 times more potent than Roscovitine).

SUMMARY OF THE INVENTION

It has now been found that purine derivatives of formula (I) as defined in formula (I) are cyclin-dependent kinases (CDKs) inhibitors and demonstrate efficient cyst formation inhibition in a MDCK cell lines model as illustrated in the experimental data herein after and, on the basis of such activity, the compounds will be useful in the treatment of polycystic kidney disease.

As compounds of the invention strongly inhibit CDKs (CDKl, CDK2, CDK5, CDK9), they are useful in the treatment of polycystic kidney diseases. Other key targets include CKl and DYRKlA, and GSK-3 to a lesser extent.

The present invention therefore relates to a method of treatment of polycystic diseases or of reducing and/or avoiding cyst formation in a patient with cystic disease and in particular with polycystic kidney disease, which comprises at least one step consisting in administering to said patient an effective amount of a 2,6,9-trisubstituted purine compound as defined in formula (I) below or one of its pharmaceutically acceptable salts.

The present invention further relates to some particular 2,6,9-purine derivatives compounds as such, as defined below.

The present invention also provides pharmaceutical compositions comprising at least one of said particular compounds.

The present invention also provides a method of treatment of polycystic diseases or of reducing and/or avoiding cyst formation in a patient with cystic disease and in particular with polycystic kidney disease, which comprises at least one step consisting in administering to said patient an effective amount of at least one of said particular compounds. DETAILED DESCRIPTION OF THE INVENTION

The compounds of the present invention are shown to inhibit the action of certain protein kinases. More precisely the compounds of the invention are shown to be cyclin-dependent kinases (CDKs) inhibitors as illustrated in the pharmacological data below.

According to a first aspect, a subject-matter of the present invention relates to a method of treatment of polycystic diseases or of reducing and/or avoiding cyst formation in a patient with cystic disease and in particular with polycystic kidney disease, which comprises at least one step consisting in administering to said patient an effective amount of a 2,6,9-trisubstituted purine compound of formula (VII):

wherein

A represents alternatively a (A.I) or (A.2) group,

(A.1) (A.2) with, when A is (A.I):

R¹, R², R³ and R⁴ independently represent a hydrogen atom or a group chosen among a (Ci-C₃)alkyl group, a (Ci-C₃)fluoroalkyl group, a (Ci-C₃)fluroalkoxy group, a (Ci-C₃)alkoxy(Ci-C₃)alkyl group and a (Ci-C₃)alkenyl group, said group being optionally substituted by a hydroxy group, provided that at the most three hydroxy group are comprised in the -CHR¹-C(OH)R²-CR³R⁴(OR⁵) group, at the most three fluorine atoms are comprised in the -CHR'-QOH^-CRVCOR⁵) group and at the most one of R¹, R², R³ and R⁴ comprises a (Ci-C3)alkoxy group or a (Ci-C3)fluroalkoxy group,

R⁵ represents a hydrogen atom, a (Ci-C₃)alkyl group optionally substituted by an hydroxy group or a -COR⁶ group, wherein R⁶ represents a group derived from a natural or unnatural amino acid or a piperidyl group of formula (B)

R⁸

R⁷

(B) wherein R⁸ represents a hydrogen atom, a halogen atom, a (Ci-C₃)alkyl group, a hydroxy(Ci-C₃)alkyl group or a -NR^aR^b group, wherein R^a and R^b independently represent a hydrogen atom or a (Ci-C₃)alkyl group and R⁷ represents a hydrogen atom, a (Ci-C3)alkyl group, a -N(Me)₂ group, a piperidyl group or a morpholinyl group, with, when A is (A.2):

R¹¹ and R¹² independently represent a hydroxy group, a (Ci-C₃)alcoxy(Ci-C₃)alkyl group or a (Ci-C₃)alkyl substituted by one or two hydroxy group(s),

X and Y independently represent a phenyl group or a heteroaryl group, it being possible for the said phenyl and heteroaryl group to be substituted by one or two groups independently chosen among a (Ci-C₂)alkyl group, a (Ci-C₂)alkoxy group, a halogen atom, a (Ci-C2)fiuoroalkyl group, a (Ci-C2)fiuoroalkoxy group, a hydroxy group, a -COOH group, a CONHR⁹ and a -NR^aR^b group, wherein R^a and R^b are as defined above,

R⁹ represents a hydrogen atom, a (Ci-C₃)alkyl group optionally substituted by one to three hydroxy groups or a -NR^aR^b group, wherein R^a and R^b are as defined above, said heteroaryl group being chosen among a thienyl group, a pyridyl group, a pyrrolyl group, a furanyl group, a pyrimidyl group and a thiazolyl group, with the proviso that at least one of the X and Y groups is an heteroaryl group, or anyone of its pharmaceutically acceptable salt.

The present invention more particularly concerns a compound of formula (I):

wherein R₁, R₂, R3, R₄, R5, X and Y are as above-defined.

In the framework of the present invention, the term "patient" may extend to humans or mammals, such as cats or dogs.

According to one aspect, the present invention relates to a compound of formula (VII) or (I), wherein the heteroaryl group is chosen among a thienyl group and a pyridyl group.

According to another aspect, the group derived from a natural or unnatural aminoacid can be derived from the natural 20 aminoacids. Among said natural amino acids, one may more particularly cite valine, serine, threonine, leucine, asparagine, aspartic acid. Among unnatural aminoacids, which are derivatives of natural aminoacids, one may cite their acylated derivatives, i.e. comprising instead of the terminal -COOH group, a (Ci-C₃)alkylcarbonyl or a (Ci-C₄)alkoxycarbonyl group, in particular a methylcarbonyl group, giving a acetylated derivative or a tertbutoxycarbonyl group, giving a Boc derivative. D-aminoacids may also be used.

According to another aspect, R⁶ represents one of the following formulas (a) to

(e)

(C)

and their enantiomers.

The compounds of the invention may exist in the form of free bases or of addition salts with pharmaceutically acceptable acids.

Suitable physiologically acceptable acid addition salts of compounds of formula (VII) or (I) include hydrochloride, hydrobromide, tartrate, fumarate, citrate, trifluoroacetate, ascorbate, and malate.

The compounds of formula (VII) or (I) and or salts thereof may form solvates (e.g. hydrates) and the invention includes all such solvates.

In the context of the present invention, the term:

- "halogen" is understood to mean chlorine, fluorine, bromine, or iodine, and in particular denotes chlorine, fluorine or bromine,

- "(Ci-C₃)alkyl" as used herein respectively refers to C₁-C₃ normal, secondary or tertiary saturated hydrocarbon. Examples are, but are not limited to, methyl, ethyl, 1 -propyl, 2-propyl,

- "(Ci-C3)alkenyl" as used herein respectively refers to C1-C3 normal, secondary or tertiary hydrocarbon with one unsaturation. Examples are ethylene, 1 -propylene or 2-propylene,

- "phenyl group" encompasses a phenyl group (monovalent) and a phenylene group when it is divalent, and

- "heteroaryl group" encompasses a heteroaryl group (monovalent) and a arylene group when it is divalent, said heteroaryl group being a ring, aromatic or not, comprising one or two heteroatoms such as nitrogen, oxygen and sulphur,

- "fluoroalkyl group" and "fluoroalkoxy group" refers respectively to alkyl group and alkoxy group as above-defined, said groups being substituted by at least one fluorine atom. Examples are perfluoroalkyl groups, such as trifluoromethyl or perfluoropropyl. In one particular variant, the present invention is directed to a compound of formula (VII) or (I) wherein, when X and/or Y are substituted, each of X and Y may comprise one or two substitution groups, said substitution group being in particular chosen among a (Ci-C₂)alkyl group, a (Ci-C₂)fluoroalkyl group, a (Ci-C₂)alkoxy group, a (Ci-C2)fluoroalkoxy group, a halogen atom, a hydroxy group and a -COOH group.

Still in said particular variant, when Y is a heteroaryl group and is monosubstituted, the substitution takes advantageously place in position 3 or 5.

In one other particular variant, the piperidyl group of formula (A) is a

According to one embodiment, the present invention more particularly concerns the compound of formula (I) according to the present invention which is defined as follows:

R¹, R², R³ and R⁴ independently represent a hydrogen atom or a (Ci-C₃)alkyl group optionally substituted by a hydroxy group, provided that at the most three hydroxy group are comprised in the -CHR¹-C(OH)R²-CR³R⁴(OR⁵) group, and

X and Y are such as defined above and are not substituted and in particular independently represent a phenyl group, a pyridyl or a thienyl group, advantageously provided that X and Y are not simultaneously a thienyl group and a pyridyl group.

According to another embodiment, the present invention more particularly concerns the compound of formula (VII) according to the present invention which is defined as follows:

A represents a group (A.2) wherein R¹¹ and R¹² are hydroxy groups, and X and Y are such as defined above and are not substituted and in particular independently represent a phenyl group, a pyridyl or a thienyl group, advantageously provided that X and Y are not simultaneously a thienyl group and a pyridyl group. According to a particular embodiment, an additional subject-matter of the present invention is a method according to the present invention wherein the administered compound is a compound of formula (Ia)

wherein

R¹, R², R³, R⁴ and R⁵ groups are as defined in formula (I), and in particular represent a hydrogen atom or a (Ci-C₃)alkyl group optionally substituted by a hydroxy group, provided that at the most three hydroxy group are comprised in the -CHR'-CCOH^-CRVCOR⁵) group,

R⁹ and R¹⁰ independently represent a hydrogen atom, a (Ci-C₂)alkyl group, a (Ci-C2)fluoroalkyl group, a (Ci-C2)alkoxy group, a (Ci-C2)fluoroalkoxy group, a halogen atom, a hydroxy group or a -COOH group,

G represent -CH=, or -N=, and when G represents -CH=, then one of W and Z represents -N= and the other represents -CH=, and when G represents -N=, then W and Z represent -CH=, or one of its pharmaceutically acceptable salt.

According to a particular embodiment, an additional subject-matter of the present invention is a method according to the present invention wherein the administered compound is a compound of formula (Ib):

wherein

R¹, R², R³ and R⁴ groups are as defined in formula (I), and in particular represent a hydrogen atom or a (Ci-C₃)alkyl group optionally substituted by a hydroxy group, provided that at the most three hydroxy group are comprised in the -CHR'-CCOH^-CRVCOR⁵) group,

R⁵ represents a hydrogen atom or a -COR₆ group, wherein R₆ represents one of the formula (a) to (e) as described above, or one its enantiomers and more particularly (b) or (c) or one of their enantiomers.

R⁹ and R¹⁰ independently represent a hydrogen atom, a (Ci-C₂)alkyl group, a (Ci-C2)fluoroalkyl group, a (Ci-C2)alkoxy group, a (Ci-C2)fluoroalkoxy group, a halogen atom, a hydroxy group or a -COOH group, or one of its pharmaceutically acceptable salt.

According to a particular embodiment, an additional subject-matter of the present invention is a method according to the present invention wherein the administered compound is a compound of formula (Ic)

wherein

According to a particular embodiment, an additional subject-matter of the present invention is a method according to the present invention wherein the administered compound is a compound of formula (Id),

wherein

R¹, R², R³ and R⁴ groups are as defined in formula (I), and in particular represent a hydrogen atom or a (Ci-C₃)alkyl group optionally substituted by a hydroxy group, provided that at the most three hydroxy group are comprised in the -CHR¹-C(OH)R²-CR³R⁴(OR⁵) group,

R⁹ and R¹⁰ independently represent a hydrogen atom, a (Ci-C₂)alkyl group, a (Ci-C2)fluoroalkyl group, a (Ci-C2)alkoxy group, a (Ci-C2)fluoroalkoxy group, a halogen atom, a hydroxy group or a -COOH group, or one of its pharmaceutically acceptable salt. According to a particular embodiment, an additional subject-matter of the present invention is a method according to the present invention wherein the administered compound is a compound of formula (Ie)

wherein

R¹, R², R³ and R⁴ groups are as defined in formula (I), and in particular represent a hydrogen atom or a (Ci-C₃)alkyl group optionally substituted by a hydroxy group, provided that at the most three hydroxy group are comprised in the -CHR'-CCOH^.CRVCOR⁵) group,

R⁹ represents a hydrogen atom, a (Ci-C₂)alkyl group, a (Ci-C₂)fiuoroalkyl group, a (Ci-C2)alkoxy group, a (Ci-C2)fluoroalkoxy group, a halogen atom, a hydroxy group or a -COOH group, or one of its pharmaceutically acceptable salt.

The compounds of formulae (VII), (I), (Ia), (Ib), (Ic), (Id) and (Ie) can comprise one or more asymmetric carbon atoms. They can thus exist in the form of enantiomers or of diastereoisomers. These enantiomers, diastereoisomers and their mixtures, including the racemic mixtures, are encompassed within the scope of the present invention.

According to a preferred embodiment of the present invention, the compound is chosen from:

- 2[(-2S)-dihydroxypropylamino]-9-ώopropyl-6-[4-(2-pyridyl) phenylmethylamino]purine (1),

- 2 [(-25)-dihydroxypropylamino] -9-ώopropyl-6- [4-(3 -pyridyl) phenylmethylamino]purine (2), - 2[(-2S)-dihydroxypropylamino]-9-ώopropyl-6-[4-(phenyl)3-pyridyl)] methylamino]purine (3),

2[(-2S)-dihydroxypropylamino]-9-ώopropyl-6-[4-(3-thiophenyl)-(phenyl) methylamino]purine (4), and

( 1 S,2R,3R)-3 -[ [9-ώopropyl-6- [ [4-(2-pyridyl)phenyl]methylamino]purin-2- yl]amino]cyclohexane- 1 ,2-diol (5),

2[(2i?)-dihydroxypropylamino]-9-ώopropyl-6-[4-(2-pyridyl)phenylmethyl amino]purine (6),

- [(2i?)-2-hydroxy-3-[[9-ώopropyl-6-[[4-(2-pyridyl)phenyl]methylamino]purin- 2-yl]amino]propyl] (25)-2-(tert-butoxycarbonylamino)-3-methyl-butanoate (7)

- [(2i?)-2-hydroxy-3-[[9-ώopropyl-6-[[4-(2-pyridyl)phenyl]methylamino]purin- 2-yl]amino]propyl] (2S)-2-amino-3-methyl-butanoate (8)

- [(25)-2-hydroxy-3-[[9-ώopropyl-6-[[4-(2-pyridyl)phenyl]methylamino]purin- 2-yl]amino]propyl] (2i?)-2-amino-3-methyl-butanoate (9)

- their pharmaceutically acceptable salts.

Said compounds (1) to (9), which are also reported in table I below, form part of the invention, as well as their pharmaceutically acceptable salts, such as hydrochloride, tartrate and fumarate.

The two following compounds also form part of the invention.

2-(2,3-dihydroxybutylamino)-9-isopropyl-6(4-phenyl-pyridin-3- ylmethylamino)purine

OH

2-(2,3-dihydroxybutylamino)-9-isopropyl-6[4-(2-pyridyl)-phenylmethylamino] purine

OH

as well as their pharmaceutically acceptable salts, such as hydrochloride, tartrate and fumarate.

The compounds of the present invention can be prepared by conventional methods of organic synthesis practiced by those skilled in the art. The general reaction sequences outlined below represent a general method useful for preparing the compounds of the present invention and are not meant to be limiting in scope or utility.

The compounds of general formula (VII) or (I) can be prepared according to scheme 1 below.

Scheme 1

(VII) (H) The synthesis is based on a three steps sequence starting from 2,6-dichloropurine of formula (VI).

According to this process, an amine of formula (V), wherein X and Y are as defined above, can be reacted with the compound of formula (VI), for example in the presence of triethylamine (NEt₃), for example in a solvent such as butanol, for example at a temperature ranging between 80 and 100⁰C, to obtain a compound of formula (IV). Compounds of formula (III) can be obtained by reaction of compounds of formula (IV) with 2-bromopropane for example in the presence of potassium carbonate, for example in a solvent such as dimethylsulfoxyde (DMSO), for example at a temperature ranging between 15 and 20 ⁰C. Compounds of formula (VII) can be obtained by reaction of a compound of formula (III) with a compound of formula (II), wherein R¹, R², R³ and R⁴ are as defined above for example with no added solvent, for example at a temperature ranging between 110 and 160⁰C.

The starting compounds of formula (V) are commercially available or can be prepared according to methods known to the person skilled in the art, such as described in Oumata N; Ferandin, Y; Meijer. L; Galons in Org Proc Res & Dev online February 26, 2009 (http://pubs.acs.org/doi/pdf/10.1021/op800284k).

The compound of formula (VI) is commercially available.

The chemical structures and physical data of some compounds of formula (VII) of the invention are illustrated in the following Table I.

Table I

The following examples illustrate in detail the preparation of compounds (1) to (9) according to the invention. The structures of the products obtained have been confirmed by NMR spectra.

Example 1 : 2[(-25Vdihydroxypropylamino1-9-/sopropyl-6-[4-(2-pyridyl) phenylmethylamino]purine or (S)-3-[9-Isopropyl-6-(4-pyridin-2-yl-benzylamino)-9H- purin-2-ylamino] -propane- 1 ,2-diol (1)

1. 2-Chloro-6-[4-(2-pyridyl)phenylmethylamino]-purine (TVa)

To a solution of 2,6-dichloropurine (23 g, 0.1 mol) in 200 mL n-BuOΗ was added the primary amine 4-(2-pyridyl)benzylamine (0.12 M) and NEt₃. After 3 hours heating at 110⁰C, the mixture is cooled to 20⁰C and the solid was filtrated, washed with 5 mL cold n-BuOΗ and dried in vacuo.

2. Alkylation of chloropurine (IVa), synthesis of 2-chloro-9-isopropyl-6-[4-(2- pyridyl)phenylmethylamino] purine (UIa)

To a solution of (IVa) (0.08 mol) in DMSO (100 mL) at 18-20 ⁰C was added K2CO3 (35.52 g, 0.24 mol) and 2-bromopropane (19 mL, 0.2 mol). After 5 hours stirring at 18-20⁰C, 2-bromopropane (4.7 mL, 0.05 mL) were added and the stirring pursued at the same temperature for 5 hours. After addition of 200 mL cold (5°C) H₂O, the mixture was extracted with EtOAc (3x100 mL) and the combined organic layer were washed with brine (3x50 mL), dried and dried over Na₂SO₄. Derivative (HIa) which crystallized upon concentration was triturated once with 5 mL 2-propanol and was collected by filtration.

Yield 85%, mp 180-182 ⁰C. IH NMR(DMSO) lδ 1.58 (d, 6H, J = 6.8 Hz, CH(CHs)₂), 4.90 (hept, IH, CH(CH₃)₂), 6-7.25-7.50 (m, 5H, Hphenyl), 8.16 (s, IH, H-8). ¹³C-NMR (DMSO) 5 21.81, 43.19, 46.04, 117.98, 126.44, 126.85, 127.67, 137.03, 137.97, 148.93, 153.03, 154.45.

3. Animation by nucleophilic substitution of 2-chloro-9-isopropyl-6-[4-(2- pyridyl)phenylnιethylanιino] purine (Ilia).

A mixture of chloropurine (Ilia) (0.138 mol) and (25)-2-aminopropanediol (104.1OmL, 1.11 mol) was heated, under N₂, at 160⁰C for 8 hours. After cooling, H₂O (100 mL) was added and the mixture was extracted with EtOAc (4x10OmL). The organic layer was washed with warm (50 ⁰C) H₂O (2x50 mL) dried and evaporated until dryness. Crystallization occurred after addition of 10 mL EtOAc. The crystals of trisubstituted purine (1) were collected by filtration.

Yield 92%, mp. 89-93 ⁰C. IH NMR(CDCl₃): 1.52 (d, 6H, J = 6.56 Hz, CH(CHs)₂), 3.57 (m, 4H, CH₂-OH₅CH₂-NH), 3.78(m, IH, CH-(CH₂)₂), 4.58 (hept, IH, CH(CHs)₂), 4.78 (bs, 2H, NHCH₂), 5.2 (bs, IH, CHNH), 6.22 (bs, IH, NHCH₂), 7.22 (t, IH, J = 6.31 Hz, Hpyridyl), 7.45 (d, 2H, J = 8.08 Hz, Hphenyl), 7.51 (s, IH, 8-H), 7.71 (m, 2H, Hpyridyl), 7.83 (d, 2H, J = 8.08 Hz, Hphenyl), 8.67 (d, IH, J = 5.05 Hz, Hpyridyl). 13C NMR (CDCl₃): δ 10.89, 22.43, 24.76, 46.77, 55.68, 109.95, 115.71, 136.54, 150.83, 157.15, 157.76, 158.38, 159.04

Example 2: 2 [(-25)-dihydroxypropylamino] -9-ώopropyl-6- [4-(3 -pyridyl) phenylmethylamino]purine (2)

OH

The same procedure was performed as for example 1 above except that the primary amine used was 4-(3-pyridyl)benzylamine (as in step 1.2) Yield 92%, mp. 115-120⁰C. ¹H NMR(CDCl₃): 1.53 (d, 6H, J = 6.56 Hz, CH(CHs)₂), 3.61 (m, 4H, CH₂-OH₅CH₂-NH), 3.85(m, IH, CH-(CH₂)₂), 4.60 (hept, IH, J = 6.56Hz, CH(CHs)₂), 4.8 (bs, 2H, NHCH₂), 5.40 (bs, IH, CHNH), 6.35 (bs, IH, NHCH₂), 7.35 (m, lH,Hpyndyi), 7.46 (d, 2H, J = 7.57Hz, H_phenyl), 7.53 (d, 2H, J = 7.57Hz,H_phenyl), 7.55 (s, IH, S-H), 7.84 (d, IH, J = 8.08 Hz, H_pyπdyi), 8.57 (m, IH, H_pyπdyi), 8.81 (s, IH, Hpyπdyi). ¹³C NMR (CDC13): 22.57, 44.74, 45.76, 46.64, 63.60, 123.62, 127.35, 128.36, 134.36, 148.09, 148.29.

Example 3 : 2[(-2S)-dihydroxypropylamino]-9-ώopropyl-6-[4-(phenyl)3- pyridyl)]methylamino]purine or (2S)-3- {9-Isopropyl-6-[(6-phenyl-pyridin-3-ylmethyl)- amino] -9H-purin-2-ylamino } -propane- 1 ,2-dio 1 (3)

The same procedure was performed as for example 1 above except that the primary amine used was 6-phenyl-pyridin-3-ylmethylamine (as in step 1.2)

Yield 92%, mp. 110-115 ⁰C. ¹H NMR(CDCl₃): 1.52 (d, 6H, J = 6.56 Hz, CH(CHs)₂), 3.58 (m, 4H, CH₂-OH₅CH₂-NH), 3.8(m, IH, CH-(CH₂)₂), 4.58 (hept, IH, J = 6.56Hz, CH(CHs)₂), 4.7 (bs, 2H, NHCH₂), 5.23 (bs, IH, CHNH), 6.33 (bs, IH, NHCH₂), 7.44 (m, 3H,H_phenyi), 7.52 (s, IH, 8-H), 7.66 (d, IH, J = 8.08Hz,H_pyπdyi), 7.74 (d, IH, J = 8.08Hz, Hpyπdyi), 7.95 (d, 2H, J = 7.32 Hz, H_phenyl), 8.7 (d, IH, J = 5.05 Hz, H_pyπdyl). ¹³C NMR (CDCl₃): δ , 22.54, , 41.37, 44.66, 46.40, 63.54, 72.41, 120.33, 126.79, 128.71, 128.92, 132.82, 134.62, 136.22, 138.92, 149.22, 154.72, 156.47, 160.05. Example 4: 2[(-2S)-dihydroxypropylamino]-9-ώopropyl-6-(4-thiophen-3- yl)phenyl)methylamino]purine (4)

The same procedure was performed as for example 1 above except that the primary amine used was 4-(thiophen-3-yl)benzylamine (as in step 1.2)

Yield 92%, mp. 152 ⁰C. ¹H NMR(CDCl₃): 1.54 (d, 6H, J = 6.56 Hz, CH(CHs)₂), 3.61 (m, 4H, CH₂-OH, CH₂-NH), 3.83(m, IH, CH-(CH₂)₂), 4.60 (hept, IH, J = 6.82Hz, CH(CHs)₂), 4.74 (bs, 2H, NHCH₂), 5.27 (bs, IH, CHNH), 6.25 (bs, IH, NHCH₂), 7.35 (m, 2H), 7.39(m, 2H), 7.43 (m, IH, H), 7.54 (d, 2H, S-H), 7.56 (s, IH, 8-H). 13C NMR (CDCl₃): δ ¹³C NMR (CDCl₃): δ 22.66, 44.77, 46.56, 63.60, 72.50, 126.24, 126.65, 128.13.

Example 5: (15',2i?,3i?)-3-[[9-ώopropyl-6-[[4-(2-pyridyl)phenyl]methylamino] purin-2-yl]amino]cyclohexane-l,2-diol (5)

The same procedure was performed as for example 1 except that (1S,2R,3R)- aminopropane-l,2-diol was used in the last amination step.

RMN ¹H (DMSO) δ ppm: 1.5 (s, 6H, 2CH₃), 1,8 (m, 2H, CH₂-CH₂-CH₂), 2 (m, 2H, CH₂-CH₂-CH₂), 2.9 (m, IH, CHNH), 3.45 (m, IH, CHOH), 3,7 (m, IH, CH₂-NH-Ar), 3.9-4 (m, IH, CH-OH), 4.6(m,lH, N-H-N), 4.75 (m,lH, CH-iPr), 6 (s, IH, NH), 7.1 (m, IH, H-pyr),7.4-7.6 (m, 2H, H-Aro), 7.7 (m, 2H, H-pyr), 8 (m, 2H, H-Aro), 8.5 (m, IH, H- pyr)

Example 6: 2[(2i?)-dihydroxypropylamino]-9-ώopropyl-6-[4-(2-pyridyl) phenylmethylamino]purine (6)

This compound was prepared as product 1 except that in the last amination step, (7?)-3-amino propane- 1,2-diol was used.

Yield 76%. ¹H NMR(CDCl₃): 1.5 (d, 6H, J = 6.5 Hz, CH(CH₃)₂), 3.6 (m, 4H, CH₂OH₅CH₂-NH), 3.78(m, IH, CH-(CH₂)₂), 4.6 (hept, IH, CH(CH₃)₂), 4.78 (bs, 2H, NHCH₂), 5.2 (bs, IH, CHNH), 6.20 (bs, IH, NHCH₂), 7.20 (t, IH, J = 6.2 Hz, H_pyπdyi), 7.45 (d, 2H, J = 8.08 Hz, H_phen_yi), 7.5 (s, IH, S-H), 7.7 (m, 2H, H_pyridyi), 7.8 (d, 2H, J = 8.0 Hz, Hphenyi), 8.7 (d, IH, J = 5.05 Hz, Hpyridyi). ¹³C NMR (CDCl₃): δ 22.4, 24.8, 46.8, 55.7, 110, 115.5, 136.5, 150.8, 157, 158, 158.5, 159.

Example 7: [(2i?)-2-hydroxy-3-[[9-ώopropyl-6-[[4-(2-pyridyl)phenyl]methyl amino]purin-2-yl]amino]propyl] (25)-2-(tert-butoxycarbonylamino)-3-methyl-butanoate

(7).

Dicyclohexycarbodiimide (0.763) was added to a cool (0⁰C) solution of L-Boc- valine (0.8 g) and N-hydroxybenzotriazole (0.49 g). The mixture was stirred at 20⁰C for 2 h and filtrated. The solid was rinsed with 3 mL of EtOAc. The combined filtrates were added to a solution of compound 6 in 20 mL THF and 1 mL NEt₃. After stirring for 12 h, the mixture was concentrated under vacuo and the residue was extracted with EtOAc (30 mL). The organic solution was washed first with 10 mL of a 2 M aqueous solution of citric acid and then with 5 mL of a saturated sodium carbonate solution and finally with water after drying, the orhganic layer was evaporated and the ester purified by column chromatography (Eluent : EtOAc-EtOH-THF-NEt₃ 95: 1 : 3: 1).

Yield 76 %. ¹H-NMR (CDCl₃) ppm: 0.95 (2d, 6H, 2CH₃ Valine), 1.45 (s, 9H, t- but ),1.55 (d, 6H, 2CH₃ iPr), 2.4 (m, IH, NH-CH-iPr), 3.47 (m, IH, CH₂-NH); 3.66 (m, IH, CH₂-NH); 4.05 (m, IH, CHOH); 4.21 (m, 2H, CH₂OCO-), 4.6 (hept, 1H,CH(CH₃)₂),

4.8 (s large, 2H, CF^-phe), 5.05 (M, IH, CH-iPr), 7.21 (t, IH, pyr), 7.5 (d, 2H, Haro), 7.52 (s, IH, 8-H), 7.75 (m, 2H, pyr), 7.95 (d, 2H, phe), 8.7 (m, IH, H-pyr).

¹³C -NMR(CDCl₃): 17.62, 19.04, 22.53, 28.32, 46.59, 120.44, 122.08, 125.27, 127.13, 128.03, 128.22, 129.01, 136.74, 137.85, 139.32, 149.65 Example 8: [(2i?)-2-hydroxy-3-[[9-ώopropyl-6-[[4-(2-pyridyl)phenyl]methyl amino]purin-2-yl]amino]propyl] (25)-2-amino-3-methyl-butanoate (8)

An ethereal 2M HCl solution (5 rnL) was added to a solution of 7 (0.5g) in 10 rnL AcOEt. After 2 hours standing at 20⁰C the solid was collected by filtration and rinced with anhydrous ether.

Yield 86 %. ¹H-NMR (DMSO d₆) δ ppm: 0.9-1 (m, 6H, 2CH₃), 1.5 (d, 6H, 2CH₃), 2.1 (m, IH, NH₂-CH-iPr), 3.9-4.1 (m, 4H, CH₂-CH-CH₂), 4.2 (m, IH, CHOH),

4.75 (m, IH, NCHN-), 4.8 (m, 2H, CH₂-VaI), 4.9 (m, IH, CH-iPr), 7.65 (m, IH, pyr), 7.9 (s, IH, NH), 8.1 (d, 2H, H-Ar), 8.25-8.5 (m, 2H, H-Ar), 8.65 (m, 2H, H-Ar) , 8.8 (d, IH, pyr )

¹³C-NMR (DMSO): 9.53, 17.16, 17.87, 20.11, 28.95, 33.758, 38.32, 57.056, 127.44, 150.64, 168.22.

Example 9: [(25)-2-hydroxy-3-[[9-ώopropyl-6-[[4-(2-pyridyl)phenyl]methyl amino]purin-2-yl]amino]propyl] (2i?)-2-amino-3-methyl-butanoate (9)

This product was obtained, starting from compound 1 in the same two step procedure used for the synthesis of 8. Boc-D-valine was used in the acylation step.

Example 10: Pharmacological data

The compounds of the invention have been the subject of pharmacological tests which have demonstrated their relevance as active substances in therapy and in particular in the treatment of polycystic kidney disease.

Example 10.1

The inhibitory activity of the compounds according to the invention on CdKs was firstly measured. The following materials and methods have been used.

MATERIAL AND METHODS

Buffers

Buffer A: 10 mM MgCl₂, 1 mM Ethylene Glycol Tetraacetic Acid (EGTA), 1 mM DTT, 25 mM Tris-HCl pH 7.5, 50 μg heparin/ml.

Buffer C: 60 mM β-glycerophosphate, 15 mM p-nitrophenylphosphate, 25 mM 3-(N-morpholino)propanesulfonic acid (MOPS) (pH 7.2), 5 mM EGTA, 15 mM MgCl₂, 1 mM Dithiothreitol (DTT), 1 mM sodium vanadate, 1 mM phenylphosphate.

Kinases preparations and assays

Kinase activities were assayed in Buffer A or C, at 30 ⁰C, at a final ATP concentration of 15 μM. Blank values were subtracted and activities expressed in % of the maximal activity, i.e. in the absence of inhibitors. Controls were performed with appropriate dilutions of dimethylsulfoxide.

CDKl/cvclin B (M phase starfish oocytes, native) and CDK5/p25 (human, recombinant) were prepared as previously described (Leclerc S. et al, J Biol Chem 2001; 276:251-60). Kinase activity was assayed in buffer C, with 1 mg histone Hi/ml, in the presence of 15 μM [γ-³³P] ATP (3,000 Ci/mmol; 10 mCi/ml) in a final volume of 30 μl. After 30 min. incubation at 30⁰C, 25 μl aliquots of supernatant were spotted onto 2.5 x 3 cm pieces of Whatman P81 phosphocellulose paper, and, 20 sec. later, the filters were washed five times (for at least 5 min. each time) in a solution of 10 ml phosphoric acid/liter of water. The wet filters were counted in the presence of 1 ml ACS (Amersham) scintillation fluid. CDK2/cvclin A (human, recombinant, expressed in insect cells) was assayed as described for CDKl/cyclin B.

CDK9/cvclin T (human, recombinant, expressed in insect cells) was assayed as described for CDKl/cyclin B, but using a pRB fragment (amino acids.773-928) (3.5 μg/assay) as a substrate.

GSK-3a/β (porcine brain, native, affinity purified) was assayed, as described for CDKl but in Buffer A and using a GSK-3 specific substrate (GS-I : YRRAAVPPSPSLSRHSSPHQSpEDEEE) (Sp stands for phosphorylated serine) (Bach S. et al. J Biol Chem 2005; 280:31208-19).

CKl δ/ε (porcine brain, native, affinity purified) was assayed as described for CDKl but using the CKl -specific peptide substrate RRKHAAIGSpAYSITA (Reinhardt J. et al. Protein Expr & Purif 2001; 54:101-9).

RESULTS

The results are gathered in the following table II. They show that the compounds inhibit CDKs but also CKl and DYRKlA, and to a lesser extent GSK-3.

Example 10.2

The compounds were also tested on MDCK cells. Cysts are formed starting from MDCK cells. They were also tested for their antiproliferative / cell death inducing properties using the human neuroblastoma SH-SY5Y cell line. Finally, they were tested for their ability to trigger apoptotic cell death of B-lymphocytes derived from patients afflicted with chronic lymphocytic leukemia (CLL).

MATERIAL AND METHODS

A. In vitro cysts formation

Cells, reagents and culture media

The MDCK (Madin-Darby Canine Epithelial Cells) cell line (ref. CCL34) was obtained from ATCC (American Type Culture Collection).

Rat tail collagen I (A1048301) was from Invitrogen and diluted at 5 mg/ml.

The cell culture medium EMEM 1OX (M0275) was from Sigma. It was supplemented with 0.2 g sodium bicarbonate / 10 ml, followed by filtration. The cell culture medium EMEM IX (M2279) was from Sigma. It was supplemented with 10 % fetal bovine serum and 1% glutamine.

Methods

The method used is derived from Montesano et al. (Montesano R, Matsumoto K, Nakamura T, Orci L., 1991. Identification of a fϊbroblast-derived epithelial morphogen as hepatocyte growth factor. Cell. 1991 Nov 29;67(5):901-8).

MDCK cells are dissociated with trypsin/EDTA (3 min at 37°C), counted, adjusted at the desired concentration and centrifuged in 15 ml Falcon tubes. Optimal concentrations are 5 x 104 to 10 x 104 cells in 2600 μl gel/well (60 mm).

The cell pellet, maintained on ice, is resuspended in a cold 3 mg/ml collagen solution prepared as follows:

8 volumes of the stock collagen solution are mixed with 1 volume EMEM 1OX and 1 volume of filtered sodium bicarbonate solution (11.76 mg/ml). The solution is kept in ice to prevent premature gel formation.

1300 μl are gently added (no bubbles) in each well of Falcon 6 well plates. Incubation for 20 minutes at 37°C allows the gel to form. 700 μl of the same preparation without cells are then added to cover the cell containing layer. After overnight incubation at 37°C, 2 ml of IX culture medium are added. Cell aggregates, small cysts develop first and numerous cysts appear after 15 days.

B. SH-SY5Y Antiproliferative/cell survival assays

SH-SY5Y human neuroblastoma cells were grown in DMEM medium (Invitrogen, Cergy Pontoise, France). The media were supplemented with antibiotics (penicillin- streptomycin) from Lonza and 10% volume of fetal calf serum from Invitrogen. Cells were cultured at 37°C with 5% CO₂. Drug treatments were performed on exponentially growing cultures at the indicated time and concentrations. Control experiments were carried out using appropriate dilutions of DMSO.

Cell death and cell viability assessments

Cell viability was determined by measuring the reduction of 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H- tetrazolium (MTS). Cell death was determined by measuring the level of lactate dehydrogenase activity (LDH) released upon cell lysis. Both procedures have been previously described in detail (Ribas J. et al. Oncogene 2006, 25, 6304-18).

C. CLL lymphocyte cell death assays

Patient and cell purification

CLL cells were isolated from the heparinized blood of 21 untreated patients following informed consent. This study was approved by the Local Ethics Committee ("Comite de Protection des Personnes de Brest").

Mononuclear cells were isolated after density gradient centrifugation by Lymphosep (Biowest). All B-CLL samples had a Matutes's score of 4 or 5. Percentage of B-CLL cells was evaluated by flow cytometry after CD 19-PE (clone J4.119) and CD5-PC5 (clone BLIa) labeling and analyzed using a FACS Flow cytometer in (EPICS XL, Beckman Coulter, France). When the percentage of CD19+/CD5+cells was lower than 90%, B-CLL cells were enriched using the B cell Isolation Kit II by immunomagnetic depletion of monocytes, NK, granulocytes and T lymphocytes (Miltenyi Biotech).

Apoptosis

3.105 B-CLL cells/wells were cultured at 37°C in RPMI 1640 (Lonza) containing 10% FCS (InVitrogen) with various concentrations of (R)-roscovitine and analogues. After incubation, B-CLL were collected, washed in PBS and resuspended in lOOμL binding buffer Ix containing FITC-conjugated annexin V and Propidium Iodide (Beckman Coulter Apoptosis Detection Kit). After 10 min incubation on ice, the cells were analyzed by flow cytometry.

RESULTS

The results are gathered in the following table II.

The compounds according to the invention present an IC50 value generally of less than 1 μM in terms of prevention of cysts formation. They also show antiproliferative effects as illustrated with the human neuroblastoma SH-SY5Y cell survival assays. They are able to induce apoptotic cell death in B-lymphocytes obtained from CLL patients. Table II

IC50 values were calculated from dose-response curves and are shown in μM.

"NT": Not tested

Therefore, the result of the tests carried out on the compounds disclosed in the present invention show that, in vitro, they exhibit the property to regulate an excess of cell proliferation & cell death.

For this purpose an effective amount of a said compound may be administered to a patient with cystic disease and in particular with polycystic kidney disease.

The present invention is also related to a method for treating, preventing or avoiding cyst formation in a patient with a cystic disease, comprising at least one step consisting in administering to said patient an effective amount of a compound of formula (I) according to the present invention, said cystic disease including but not limited to renal cystic diseases such as: acquired renal cystic disease (ARCD), dialysis-associated cystic disease, autosomal dominant polycystic kidney disease (ADPKD), autosomal recessive polycystic kidney disease (ARPKD), congenital multicystic kidney (CMK), multicystic dysplastic kidney, end-stage renal disease (ESRD), medullary sponge kidney (MSK), nephronophthisis-medullary cystic kidney disease complex (NMCD), nephronophthisis- uremic medullary cystic disease complex, juvenile nephronophthisis, medullary cystic disease, renal cell carcinoma (RCC), tuberous sclerosis (TS), von Hippel-Lindau syndrome (VHLS) and polycystic ovary syndrome.

The present invention is also related to the use of a compound of anyone of formula (VII), (I), (Ia), (Ib), (Ic), (Id), (Ie) and (1) to (9) or one of its pharmaceutically acceptable salts according to the present invention for the manufacture of a pharmaceutical composition intended for the treatment of cystic disease, and in particular of polycystic kidney disease or for the inhibition of the formation of cysts in a patient with polycystic kidney disease.

The present invention is also related to a compound of anyone of formula (I), (VII), (Ia), (Ib), (Ic), (Id), (Ie) and (1) to (9) or one of its pharmaceutically acceptable salts according to the present invention as a medicament for the treatment and/or the prevention of cystic disease, and in particular of polycystic kidney disease or for the inhibition of the formation of cysts in a patient with polycystic kidney disease.

The present invention also encompasses pharmaceutical compositions comprising at least a compound chosen among compound (1) to (9) as defined above or any pharmaceutically acceptable salt thereof.

Thus, these pharmaceutical compositions contain an effective amount of said compound, and one or more pharmaceutical excipients.

The aforementioned excipients are selected according to the dosage form and the desired mode of administration.

In this context they can be present in any pharmaceutical form which is suitable for enteral or parenteral administration, in association with appropriate excipients, for example in the form of plain or coated tablets, hard gelatine, soft shell capsules and other capsules, suppositories, or drinkable, such as suspensions, syrups, or injectable solutions or suspensions, in doses which enable the daily administration of from 0.1 to 1000 mg of active substance.

The compounds of the present invention are also useful in the treatment of Chronic Lymphocytic Leukemia (CLL), in particular of type B, i.e. affecting the B cell.

Therefore, the present invention further relates to a method of treatment of Chronic Lymphocytic Leukemia, in particular of type B, which comprises at least a step of administration to a patient suffering thereof of an effective amount of a compound of anyone of formula (VII), (I), (Ia), (Ib), (Ic), (Id), (Ie) and (1) to (9) or one of its pharmaceutically acceptable salts.

The present invention is also related to a compound of anyone of formula (VII), (I), (Ia), (Ib), (Ic), (Id), (Ie) and (1) to (9) or one of its pharmaceutically acceptable salts according to the present invention as a medicament for the treatment of Chronic Lymphocytic Leukemia, in particular of type B.

Claims

1. A method of treatment of polycystic diseases or of reducing and/or avoiding cyst formation in a patient with cystic disease, which comprises at least one step consisting in administering to said patient an effective amount of a 2,6,9-trisubstituted purine compound of formula (VII):

wherein A represents alternatively a (A.I) or (A.2) group,

(A.1) (A.2) with, when A is (A.1):

R¹, R², R³ and R⁴ independently represent a hydrogen atom or a group chosen among a (Ci-C₃)alkyl group, a (Ci-C₃)fluoroalkyl group, a (Ci-C₃)fluroalkoxy group, a (Ci-C₃)alkoxy(Ci-C₃)alkyl group and a (Ci-C₃)alkenyl group, said group being optionally substituted by a hydroxy group, provided that at the most three hydroxy group are comprised in the -CHR¹-C(OH)R²-CR³R⁴(OR⁵) group, at the most three fluorine atoms are comprised in the -CHR¹-C(OH)R²-CR³R⁴(OR⁵) group and at the most one of R¹, R², R³ and R⁴ comprises a (Ci-C3)alkoxy group or a (Ci-C3)fluroalkoxy group, R⁵ represents a hydrogen atom, a (Ci-C₃)alkyl group optionally substituted by an hydroxy group or a -COR⁶ group, wherein R⁶ represents a group derived from a natural or unnatural amino acid or a piperidyl group of formula (B)

wherein R represents a hydrogen atom, a halogen atom, a (Ci-C₃)alkyl group, a hydroxy(Ci-C₃)alkyl group or a -NR^aR^b group, wherein R^a and R^b independently represent a hydrogen atom or a (Ci-C₃)alkyl group and R⁷ represents a hydrogen atom, a (Ci-C3)alkyl group, a -N(Me)₂ group, a piperidyl group or a morpholinyl group, or with, when A is (A.2):

R¹¹ and R¹² independently represent a hydroxy group or a (Ci-C₃)alcoxy(Ci-C₃)alkyl group or a (Ci-C₃)alkyl substituted by one or two hydroxy group(s),

X and Y independently represent a phenyl group or a heteroaryl group, it being possible for the said phenyl and heteroaryl group to be substituted by one or two groups independently chosen among a (Ci-C₂)alkyl group, a (Ci-C₂)alkoxy group, a halogen atom, a (Ci-C2)fluoroalkyl group, a (Ci-C2)fiuoroalkoxy group, a hydroxy group, a -COOH group, a CONHR⁹ and a -NR^aR^b group, wherein R^a and R^b are as defined above,

2. A method according to claim 1 wherein the 2,6,9-trisubstituted purine compound is of formula (I):

wherein R¹, R², R³, R⁴ ' R⁵, X and Y are as defined in claim 1.

3. The method according to claim 1, wherein the cystic disease is polycystic kidney disease.

4. The method according to claim 1, wherein the heteroaryl group is chosen among a thienyl group and a pyridyl group.

5. The method according to claim 1, wherein R⁶ represents one of the following formulas (a) to (e)

and their enantiomers.

6. A method according to claim 1, wherein, X and/or Y are substituted, each of X and Y comprising one or two substitution groups, said substitution group being chosen among a (Ci-C₂)alkyl group, a (Ci-C₂)fluoroalkyl group, a (Ci-C₂)alkoxy group, a (Ci-C2)fluoroalkoxy group, a halogen atom, a hydroxy group and a -COOH group.

7. A method according to claim 1, wherein

R¹, R², R³ and R⁴ independently represent a hydrogen atom or a (Ci-C₃)alkyl group optionally substituted by a hydroxy group, provided that at the most three hydroxy group are comprised in the -CHR¹-C(OH)R²-CR³R⁴(OR⁵) group, X and Y independently represent a phenyl group, a pyridyl or a thienyl group, provided that X and Y are not simultaneously a thienyl group and a pyridyl group.

8. A method according to claim 1, wherein the administered compound is a compound of formula (Ia)

wherein:

R¹, R², R³, R⁴ and R⁵ groups are as defined in claim 1, and in particular represent a hydrogen atom or a (Ci-C₃)alkyl group optionally substituted by a hydroxy group, provided that at the most three hydroxy group are comprised in the -CHR¹-C(OH)R².CR³R⁴(OR⁵) group,

G represent -CH=, or -N=, and when G represents -CH=, then one of W and Z represents -N= and the other represents-CH=, and when G represents -N=, then W and Z represent -CH=, or one of its pharmaceutically acceptable salt.

9. A method according to claim 1, wherein the administered compound is a compound of formula (Ib):

wherein

10. A method according to claim 1, wherein the administered compound is a compound of formula (Ic)

wherein R¹, R², R³ and R⁴ groups are as defined in claim 1, and in particular represent a hydrogen atom or a (Ci-C₃)alkyl group optionally substituted by a hydroxy group, provided that at the most three hydroxy group are comprised in the -CHR^C(OH)RICR³R⁴COR⁵) group,

11. A method according to claim 1 , wherein the administered compound is a compound of formula (Id),

wherein:

R¹, R², R³ and R⁴ groups are as defined in claim 1, and in particular represent a hydrogen atom or a (Ci-C₃)alkyl group optionally substituted by a hydroxy group, provided that at the most three hydroxy group are comprised in the -CHR¹-C(OH)R².CR³R⁴(OR⁵) group,

12. A method according to claim 1, wherein the administered compound is a compound of formula (Ie),

wherein

R¹, R², R³ and R⁴ groups are as defined in claim 1, and in particular represent a hydrogen atom or a (Ci-C₃)alkyl group optionally substituted by a hydroxy group, provided that at the most three hydroxy group are comprised in the -CHR'-CCOH^-CRVCOR⁵) group,

R⁹ represents a hydrogen atom, a (Ci-C₂)alkyl group, a (Ci-C₂)fluoroalkyl group, a (Ci-C2)alkoxy group, a (Ci-C2)fluoroalkoxy group, a halogen atom, a hydroxy group or a -COOH group, or one of its pharmaceutically acceptable salt.

13. A compound chosen among :

- 2 [(-25)-dihydroxypropylamino] -9-ώopropyl-6- [4-(3 -pyridyl) phenylmethylamino]purine (2),

- 2 [(-25)-dihydroxypropylamino]-9-ώopropyl-6-[4-(phenyl)3 -pyridyl)] methylamino]purine (3),

2[(-25)-dihydroxypropylamino]-9-ώopropyl-6-[4-(3-thiophenyl)-(phenyl) methylamino]purine (4), and

(15',2i?,3i?)-3-[[9-ώopropyl-6-[[4-(2-pyridyl)phenyl]methylamino]purin-2- yl]amino]cyclohexane- 1 ,2-diol (5),

2[(2i?)-dihydroxypropylamino]-9-ώopropyl-6-[4-(2- pyridyl)phenylmethylamino]purine (6),

- [(2i?)-2-hydroxy-3-[[9-ώopropyl-6-[[4-(2-pyridyl)phenyl]methylamino]purin- 2-yl]amino]propyl] (25)-2-(tert-butoxycarbonylamino)-3-methyl-butanoate (7) - [(2i?)-2-hydroxy-3-[[9-ώopropyl-6-[[4-(2-pyridyl)phenyl]methylamino]purin- 2-yl]amino]propyl] (2S)-2-amino-3-methyl-butanoate (8)

- their pharmaceutically acceptable salts.

14. A pharmaceutical composition comprising at least one compound as defined in the preceding claim.

15. A method of treatment of Chronic Lymphocytic Leukemia, which comprises at least a step of administration to a patient suffering thereof of an effective amount of a compound of formula (I),

wherein

R¹, R², R³ and R⁴ independently represent a hydrogen atom or a group chosen among a (Ci-C₃)alkyl group, a (Ci-C₃)fluoroalkyl group, a (Ci-C₃)fluroalkoxy group, a (Ci-C₃)alkoxy(Ci-C₃)alkyl group and a (Ci-C₃)alkenyl group, said group being optionally substituted by an hydroxy group, provided that at the most three hydroxy group are comprised in the -CHR¹-C(OH)R²-CR³R⁴(OR⁵) group, at the most three fluorine atoms are comprised in the -CHR¹-C(OH)R²-CR³R⁴(OR⁵) group and at the most one of R¹, R², R³ and R⁴ comprises a (Ci-C3)alkoxy group or a (Ci-C3)fluroalkoxy group,

R⁵ represents a hydrogen atom, a (Ci-C₃)alkyl group optionally substituted by an hydroxy group or a -COR⁶ group, wherein R⁶ represents a group derived from a natural or unnatural amino acid or a piperidyl group of formula (A)

(A) wherein R⁸ represents a hydrogen atom, a halogen atom, a (Ci-C₃)alkyl group, a hydroxy(Ci-C₃)alkyl group or a -NR^aR^b group, wherein R^a and R^b independently represent a hydrogen atom or a (Ci-C₃)alkyl group and R⁷ represents a hydrogen atom, a (Ci-C3)alkyl group, a -N(Me)₂ group, a piperidyl group or a morpholinyl group,

X and Y independently represent a phenyl group or a heteroaryl group, it being possible for the said phenyl and heteroaryl group to be substituted by one or two groups independently chosen among a (Ci-C₂)alkyl group, a (Ci-C₂)alkoxy group, a halogen atom, a (Ci-C2)fluoroalkyl group, a (Ci-C2)fluoroalkoxy group, a hydroxy group, a -COOH group, a CONHR⁹ and a -NR^aR^b group, wherein R^a and R^b are as defined above,

R⁹ represents a hydrogen atom, a (Ci-C₃)alkyl group optionally substituted by one to three hydroxy groups or a -NR^aR^b group, wherein R^a and R^b are as defined above, said heteroaryl group being chosen among a thienyl group, a pyridyl group, a pyrrolyl group, a furanyl group, a pyrimidyl group and a thiazole group, with the proviso that at least one of the X and Y groups is an heteroaryl group, or anyone of its pharmaceutically acceptable salt.

16. A compound of formula (VII) as defined in claim 1, a compound of formula (I) as defined in claim 2, a compound of formula (Ia) as defined in claim 8, a compound of formula (Ib) as defined in claim 9, a compound of formula (Ic) as defined in claim 10, a compound of formula (Ib) as defined in claim 11, a compound of formula (Ie) as defined in claim 12, a compound chosen among compounds (1) to (9) as defined in claim 13 or anyone of their pharmaceutically acceptable salts as a medicament for the treatment and/or the prevention of cystic disease, and in particular of polycystic kidney disease or for the inhibition of the formation of cysts in a patient with polycystic kidney disease.