Substituted pyrrolo-pyridines, composition containing them, method for their producing and use thereof

[0001] The present invention particularly relates to new chemical compounds, in particular substituted pyrrolopyridines, compositions containing them, and their use as medicaments.

[0002] More specifically, according to the first aspect, the invention relates to new specific substituted pyrrolopyridines having anti-tumor activity, by modulating the activity of proteins, in particular kinases.

[0003] To date, most of the commercial compounds used in chemotherapy pose significant problems of secondary effects and patient tolerance. These effects could be reduced as long as the therapeutic agents used would act selectively on cancer cells, excluding healthy cells. One solution to reduce the undesirable effects of chemotherapy may therefore be to use a therapeutic agent acting on metabolic pathways or structural elements of these pathways, expressed mostly in cancer cells, and which would not or hardly be expressed in healthy cells.

[0004] Kinase proteins are a family of enzymes that catalyze the phosphorylation of hydroxyl groups of specific protein residues such as tyrosine, serine or threonine residues. Such phosphorylations can significantly modify the function of proteins; thus, kinase proteins play an important role in regulating a large variety of cellular processes, including in particular metabolism, cell proliferation, cell differentiation, cell migration or cell survival. Among the various cellular functions in which kinase protein activity is involved, certain processes are attractive targets for the treatment of cancer and other diseases.

[0005] Thus, one of the objects of the present invention is to propose compositions having anti-tumor activity, acting in particular on kinases. Of the kinases for which activity modulation is being studied, KDR and Tie2 are preferred.

[0006] These products correspond to the following formula (1):

AND

<img file="PL1877409T3_D0001.tif" />

Formula (I) in which:

1) A and Ar are substituted phenyl groups;

2) L is NH-CO-NH;

3) W is C (R6), one of Y and Z is selected from N and NO and the other is C (R5) and;

4) R1, R5, and R6 are H;

5) Ra is H.

[0007] Acceptable combinations of substituents include those in which

R1, R5 and R6 are H and one of Y and Z is selected from N and NO.

[0008] A is preferably substituted with a first substituent selected from the group consisting of (C1-C12) alkyl, halogenated (C1-C12) alkyl, (C3-C12) cycloalkyl, (C2-C12) alkylene, (C2-C12) alkynyl, (C8-C14) aryl, (C1-C13) heteroaryl, O- (C1-C3) alkyl, O- (C6-C14) aryl, O- (C1-C13) heteroaryl, S- (C1-C3) alkyl, S- (C6-C14) aryl, S- (C1C13) heteroaryl, each of which is optionally substituted with a substituent selected from (C1-C3) alkyl, halogen, O- (C1C3) alkyl, N (R8) (R9) ; wherein R8 and R9 are independently selected from

H, (C1-C3) alkyl, (C1-C3) alkylOH, halogenated (C1-C3) alkyl, (C1C3) alkyl-NH2, (C1-C3) alkyl-COOM, (C1-C3) alkyl-SO3M, in which when R8 and

R9 are simultaneously different from H, they can be bonded to form a 5- to 7-membered ring comprising 0 to 3 heteroatoms selected from O, N and S, in which M is H or an alkali metal cation selected from Li, Na and K.

[0009] Furthermore, A is also preferably substituted with a second substituent selected from the group consisting of F, Cl, Br, I, OH, SH, SO3M COOM, CN, NO2, CON (R8) (R9), N (R8 ) CO (R9), (C1-C3) alkyl-OH, (C1-C3) alkylN (R8) (R9), (C1-C3) alkyl- (R10), (C1-C3) alkyl-COOH, N ( R8) (R9); wherein R8 and R9 are independently selected from H, (C1-C3) alkyl, (C1-C3) alkyl3

OH, halogenated (C 1 -C 3) alkyl, (C 1 -C 3) alkyl-NH 2, (C 1 -C 3) alkyl-COOM, (C 1 -C 3) alkyl-SO 3 M; wherein when R8 and R9 are simultaneously different from H, they can be bonded to form a 5- to 7-membered ring comprising 0 to 3 heteroatoms selected from O, N and S; wherein M is H or an alkali metal cation selected from Li, Na and K; and in which

R10 is H or a non-aromatic hetero optionally substituted, comprising 2 to 7 carbon atoms, and 1 to 3 heteroatoms selected from N, O and S.

[0010] When A is disubstituted, two substituents can be bonded to each other to form a 5- to 7-membered ring containing 0 to 3 heteroatoms selected from N, O and S.

[0011] According to a preferred embodiment, A is phenyl substituted with at least one group selected from halogen, (C1C4) alkyl, halogenated (C1-C3) alkyl, O- (C1-C4) alkyl, S- (C1C4) alkyl, O- (C1-C4) halogenated alkyl, S- (C1-C4) halogenated alkyl, and in which when A is disubstituted, two substituents can be bonded to each other to form a 5- to 7-membered ring containing 0 to 3 heteroatoms selected from N , O and S.

[0012] The products of the invention may be in the form of:

1) non-chiral, or

2) racemic, or

3) enriched in stereoisomer, or

4) enriched in enantiomer;

(C1-C3) alkylCOOM, (C1-C3) alkylSO3M; wherein when R8 and R9 are simultaneously different from H, they can be bonded to form a 5- to 7-membered ring comprising 0 to 3 heteroatoms selected from O, N and S, in which M is H or an alkali metal cation selected from

Li, Na and K.

[0013] In addition, A is also preferably substituted with a second substituent selected from the group consisting of F, CI, Br, I, OH, SH,

SO3M, COOM, CN, NO2, CON (R8) (R9), N (R8) CO (R9), (C1-C3) alkyl-OH, (C1C3) alkyl-N (R8) (R9), (C1- C3) alkyl- (R10), (C1-C3) alkyl-COOH, N (R8) (R9); wherein R8 and R9 are independently selected from H, (C1-C3) alkyl, (C1C3) alkylOH, halogenated (C1-C3) alkyl, (C1-C3) alkylNH2, (C1C3) alkylCOOM, (C1-C3) alkylSO3M; wherein when R8 and R9 are simultaneously different from H, they can be bonded to form a 5- to 7-membered ring comprising 0 to 3 heteroatoms selected from O, N and S; wherein M is H or an alkali metal cation selected from Li, Na and K; and wherein R10 is H or a non-aromatic heterocycle optionally substituted, comprising 2 to 7 carbon atoms, and 1 to 3 heteroatoms selected from N, O and S.

[0014] When A is disubstituted, two substituents can be bonded to each other to form a 5- to 7-membered ring containing 0 to 3 heteroatoms selected from N, O and S.

[0015] According to a preferred embodiment, A is phenyl, pyrazolyl or isoxazolyl substituted with at least one group selected from halogen, (C1-C4) alkyl, halogenated (C1-C3) alkyl, O- (C1C4) alkyl, S- ( C1-C4) alkyl, halogenated O- (C1-C4) alkyl, halogenated S- (C1-C4) alkyl, and in which when A is disubstituted, two substituents may be bonded to each other to form a 5- to 5-ring

7-membered containing 0 to 3 heteroatoms selected from N, O and S. [0016] The products of the invention may be in the form of:

1) non-chiral, or

2) racemic, or

3) enriched in stereoisomer, or

4) enriched in enantiomer;

and possibly be converted into salt.

[0017] The product according to the invention could be used to produce a medicament useful for treating a pathological condition, in particular cancer.

[0018] The present invention also relates to a medicament comprising a product according to the invention, and therapeutic compositions comprising a product according to the invention in combination with a pharmaceutically acceptable excipient according to the chosen mode of administration. The pharmaceutical composition may be in solid, liquid or liposome form.

[0019] Among the solid compositions there may be mentioned powders, capsules, tablets. Among oral forms, solid forms protected from the acidic environment of the stomach may also be included. Carriers used for solid forms are especially mineral carriers such as phosphates, carbonates or organic carriers such as lactose, celluloses, starch or polymers. Solid forms are suspension solutions or dispersions. They contain as a dispersion carrier either water or an organic solvent (ethanol, NMP or other) or mixtures of surfactants and solvents or complexing agents and solvents.

[0020] Solid forms will preferably be injectable and therefore have a formulation acceptable for such use.

[0021] Acceptable routes of injection include intravenous, intraperitoneal, intramuscular, and subcutaneous routes, with intravenous route usually being preferred.

[0022] The dose of compounds of the invention administered will be adjusted by the physician as a function of the route of administration to the patient and the condition of the latter. [0023] The compounds of the present invention may be administered alone or in a mixture with other anti-cancer agents. Possible associations include:

• alkylating agents and especially cyclophosphamide, melphalan, ifosfamide, chlorambucil, busulfan, thiotepa, prednimustine, carmustine, lomustine, semustine, steptozotocin, decarbazine, temozolomide, procarbazine and hexamethylmaline, platinum, like platinum, like platinum especially bleomycin, mitomycin, dactinomycin • anti-microtubular agents, in particular vinblastine, vincristine, vindesine, vinorelbine, taxoids (paclitaxel and docetaxel) • anthracyclines, especially doxorubicin, daunorubicin, idarubicin, epirubicin, mitoxantrone, losoxantrone • group I and II topoisomerase inhibitors such as etoposide, teniposide, amsacrine, topotexin, tomotecanone -fluorouracil, UFT, floxuridine • cytidine analogues such as 5-azacytidine, cytarabine, gemcitabine, 6-mercaptomurine, 6-thioguanine • adenosine analogues such as pentostatin, cytarabine or fludarabine phosphate • methotrexate and folinic acid • various enzymes and compounds such as L-asparaginase, hydroxyurea, trans-retinoic acid, suramine, [0024] dexrazoxane, amifostine, herceptin and estrogen hormones, androgenic • anti-vascular agents, such as combretastatin derivatives, for example CA4P, chalcones or colchicines, for example ZD6126, and their prodrugs.

[0025] It is also possible to associate the compounds of the present invention with radiation treatment. These treatments can be administered simultaneously, separately, sequentially. The treatment will be adjusted by the doctor as a function of the disease being treated.

[0026] The products of the invention are useful as inhibitors of the kinase catalyzed reaction, in particular FAK, KDR, Tie2, Aurora A, Aurora B and CDK2. FAK, KDR and Tie2 are kinases for which the products of the invention will be particularly useful as inhibitors.

[0027] The reasons why these kinases were selected are given below:

FAK [0028] FAK is a cytoplasmic tyrosine kinase that plays an important role in signal transduction in integrin-transmitted, a family of heterodimeric cell adhesion receptors. FAK and integrins are co-localized in peritoneal structures called adherent plaques. It has been shown in numerous cell types that the activation of FAK as well as its phosphorylation at tyrosine residues and in particular its autophosphorylation at tyrosine 397 were dependent on integrin binding to their extracellular ligands and thus induced during cell adhesion [Kornberg L et al. J. Biol. Chem. 267 (33): 23439-442. (1992)]. Autophosphorylation on 397 FAK tyrosine is a binding site for another tyrosine kinase, Src, through its SH2 domain [Schaller et al. Moth. Cell. Biol. 14: 1680-1688. 1994; Xing et al. Moth. Cell. Biol. 5: 413-421. 1994]. Src can thus phosphorylate FAK on tyrosine 925, thus obtaining Grb2 matching protein and triggering in some cells the activation of ras and MAP kinase associated with the control of cell proliferation [Schlaepfer et al. Nature; 372: 786791. 1994; Schlaepfer et al. Threshold. Biophy. Moth. Biol. 71: 435-478. 1999; Schlaepfer and Hunter, J. Biol. Chem. 272: 13189-13195. 1997]. Activation of FAK may also trigger the jun NH2-end kinase (JNK) signaling pathway and lead to cell progression to the G1 phase of the cell cycle [Oktay et al., J. Cell. Biol. 145: 1461-1469. 1999]. Phosphatidylositol-3-OH kinase (P13-kinase) also binds to FAK on tyrosine 397 and this interaction could be necessary for the activation of P13 kinase [Chen and Guan, Proc. Nat. Acad. Sci. USA. 91: 10148-10152. 1994; Ling et al. J. Cell. Biochem. 73: 533-544. 1999]. Complex

FAK / Src phosphorylates various substrates such as paxillin and p130CAS in fibroblasts [Vuori et al. Moth. Cell. Biol. 16: 2606-2613. 1996].

[0029] The results of numerous studies support the hypothesis that FAK inhibitors could be useful in the treatment of cancer. Studies have suggested that FAK could play an important role in cell proliferation and / or survival in vitro. For example, some authors have shown that in CHO cells overexpression of p125FAK leads to an accelerated transition of G1 to S, suggesting that p125FAK promotes cell proliferation [Zhao J.-H et al. J. Cell Biol. 143: 1997-2008. 1998]. Other authors have shown that tumor cells treated with FAK antisense oligonucleotides lose their adhesion and undergo apoptosis (Xu et al, Cell Growth Differ. 4: 413-4.18. 1996). FAK has also been shown to promote cell migration in vitro. Thus, fibroblasts insufficient for FAK expression (knockout mice for FAK) exhibit rounded morphology, cell migration deficiencies in response to chemotactic signals, and these deficiencies are removed by re-expression of FAK [DJ. Sieg et al., J. Cell Science. 112: 2677-91. 1999]. Overexpression of the FAK C-terminal domain (FRNK) blocks the stretching of adherent cells and reduces cell migration in vitro [Richardson A. and Parsons JT Nature. 380: 538-540. 1996]. Overexpression of FAK in CHO cells, COS cells or in human astrocytome cells promotes cell migration. The association of FAK with the promotion of cell proliferation and migration in numerous cell types in vitro suggests the potential role of FAK in cancer processes. Recent studies have successfully demonstrated an increase in tumor cell proliferation in vivo after expression induction

FAK in human astrocytome cells [Cary LA et al. J. Cell Sci.

109: 1787-1794. 1996; Wang D et al. J. Cell Sci. 113: 4221-4230. 2000].

In addition, immunohistochemical studies on human biopsies have shown that

FAK was overexpressed in prostate, breast, thyroid, colon, melanoma, brain and lung cancers, with the level of FAK expression being directly correlated with cancers constituting the most aggressive phenotype [Weiner TM, et al. Lancet. 342 (8878): 1024-1025. 1993; Owens et al. Cancer Research. 55: 27522755. 1995; Maung K. et al. Oncogene. 18: 6824-6828. 1999; Wang D et al. J. Cell Sci. 113: 4221-4230. 2000].

KDR [0030] KDR (Kinase insert Domain Receptor) also called VEGF-R2 (Vascular Endothelial Growth Factor Receptor 2), is expressed only in endothelial cells. This receptor binds to the angiogenic growth factor VEGF, and thus serves as a transduction signal mediator by activating its intracellular kinase domain. Direct inhibition of VEGF-R2 kinase reduces angiogenesis in the presence of exogenous VEGF (Vascular Endothelial Growth Factor) (Strawn et al., Cancer Research, 1996, vol. 56, pp. 3540-3545). This process has been especially demonstrated by VEGF-R2 mutants (Millauer et al., Cancer Research, 1996, vol. 56, pp. 1615-1620). The VEGF-R2 receptor appears to have no function in the adult other than that associated with VEGF angiogenic activity. Consequently, a selective inhibitor of VEGF-R2 kinase activity should show only slight toxicity.

[0031] In addition to this central role in the dynamic angiogenic process, recent results suggest that VEGF expression contributes to tumor cell survival after chemo- and radiotherapy, highlighting the potential synergy of KDR inhibitors with other factors (Lee et al. Cancer Research, 2000, vol. 60, pp. 5565-5570).

Tie2 [0032] Tie-2 (TEK) is a member of the tyrosine kinase receptor family that is specific for endothelial cells. Tie2 is the first receptor with tyrosine kinase activity, which is known to be an agonist (angiopoietin 1 or Ang1), which stimulates receptor autophosphorylation and cell signaling [S.

Davis et al (1996) Cell 87, 1161-1169] as well as an antagonist (angiopoietin 2 or Ang2) [PC Maisonpierre et al. (1997) Science 277, 55-60]. Angiopoietin 1. may have a synergistic effect with VEGF in the last stages of neo-angiogenesis [Asahara T. Circ. Res. (1998) 233-240].

Knock-out experiments and transgenic manipulations of Tie2 or Ang1 expression lead to animals that lack vascularization [DJ Dumont et al (1994) Genes Dev. 8, 1897-1909 et C. Suri (1996) Cell 87, 1171-1180]. Binding of Ang1 to its receptor leads to autophosphorylation of the Tie2 kinase domain, which is essential for neovascularization as well as for the acquisition and interaction of vessels with pericytes and smooth muscle cells; these phenomena contribute to the maturation and stability of newly formed vessels [PC

Maisonpierre et al (1997) Science 277, 55-60]. Lin et al (1997) J.

Clin. Invest., 100, 8: 2072-2078 et Lin P. (1998) PNAS 95, 8829-8834, showed inhibition of tumor growth and vascularization as well as reduction of lung metastasis during adenoviral infection or injection of the extracellular domain of Tie-2 (Tek) in xenograft models breast cancer and melanoma.

[0033] Tie2 inhibitors can be used in situations where neovascularization is inadequate (i.e., diabetic retinopathy, chronic inflammation, psoriasis, sarcoma)

Kaposi, chronic neovascularization due to macular degeneration, rheumatoid arthritis, childhood hemangioma and cancer).

[0034] Progression of the cell cycle is often controlled by cyclin dependent kinases (CDKs) which are activated by interaction with proteins belonging to the cyclin family; this activation terminates in phosphorylation of substrates and ultimately through cell division. In addition, endogenous CDK inhibitors that are activated (the INK4 and KIP / CIP family) negatively regulate CDK activity. The increase in normal cells is caused by a balance between CDK activators (cyclins) and endogenous CDK inhibitors. In many tumor types, the aberrant expression or activity of many of these cell cycle regulators has been described.

[0035] Cyclin E activates Cdk2 kinase, which, finally, acts on phosphorylation of pRb protein (retinoblastom protein) leading to involvement in irreversible cell division and transition to phase S (PL Toogood, Medicinal Research Reviews (2001), 21 (6) ; 487-498.

CDK2 kinase and perhaps CDK3 are necessary for progression to the G1 phase and entry into the S phase. When forming a complex with cyclin E, they maintain hyperphosphorylation of pRb to assist the progression of the G1 phase to phase S. In complexes with cyclin A, CDK2 plays a role in inactivation of E2F and is necessary for the implementation of the S phase (TD. Davies et al. (2001) Structure 9, 389-3).

[0036] The CDK1 / cyclin B complex regulates cell cycle progression between the G2 phase and the M phase. Negative regulation of the CDK / cyclin B complex prevents normal cells from entering the S phase before the G2 phase is properly and completely reached. (KK Roy and EA Sausville

Current Pharmaceutical Design, 2001, 7, 1669-1687.

[0037] There is a level of regulation of CDK activity. Cyclin-dependent kinase (CAK) activators have a positive effect on CDK regulation. CAK phosphorylates CDK at the threonine residue to make the enzyme target completely active.

[0038] The presence of deficiencies in intervening molecules in the cell cycle causes CDK activation and cycle progression; it is normal to strive to inhibit CDK enzymes to block the cell growth of cancer cells.

[0039] Numerous proteins have been identified associated with chromosome segregation and "spindle" accumulation in yeast and drosophy.

Disorganization of these proteins leads to non-segregation of chromosomes and to monopolar or disorganized "spindles". Of these proteins, certain kinases, including Aurora and IpI1, derived from Drosophila and S. cerevisiae, respectively, are necessary for chromosome segregation and centrosome separation. The human yeast Ipl1 analogue has recently been cloned and characterized by various laboratories. This kinase, called aurora2, STK15 or BTAK belongs to the serine / threonine kinase family. Bischoff et al have shown that Aurora2 is carcinogenic, and is enhanced in human colon cancers (EMBO J, 1998, 17, 3052-3065). This has also been given as an example in cancers associated with epithelial tumors such as breast cancer.

Definitions [0040] The term "halogen" refers to an element selected from F, Cl, Br and I.

[0041] The term "alkyl" refers to a saturated hydrocarbon linear or branched substituent having 1 to 12 carbon atoms. Substituents methyl, ethyl, propyl, 1-methylethyl, butyl, 1-methylpropyl, 2-methylpropyl, 1,1-dimethylethyl, pentyl, 1-methylbutyl, 2-methylbutyl, 3-methylbutyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl , 2,2-dimethylpropyl, 1-ethylpropyl, hexyl, 1-methylpentyl, 2-methylpentyl, 1-ethylbutyl, 2-ethylbutyl, 3,3-dimethylbutyl, heptyl,

1-ethylpentyl, octyl, nonyl, decyl, undecyl and dodecyl are examples of the alkyl substituent.

[0042] The term "alkylene" refers to a linear or branched hydrocarbon substituent having one or more unsaturated bonds having from 2 to 12 carbon atoms. Substituents ethylenyl, 1-methylethylene, prop-1-enyl, prop-2-enyl, Z-1-methylprop-1-enyl, E-1-methylprop-1-enyl, Z-1,2-dimethyl-prop-1-enyl , E-1,2-dimethylprop-1enyl, but-1,3-dienyl, 1-methylidenyl-prop-2-enyl, Z-2-methylbut-1,3-dienyl, E-2-methylbut-1,3-dienyl , 2-methyl-1-methylidenylprop-2-enyl, undec-1-enyl and undec-10-enyl are examples of the alkylene substituent. [0043] The term "alkynyl" refers to a linear or branched hydrocarbon substituent having at least two unsaturated bonds, the carrier of which is a pair of adjacent carbon atoms having from 2 to 12 carbon atoms. Ethynyl substituents; prop-1-ynyl;

prop-2-ynyl; and but-1-ynyl are examples of an alkynyl substituent.

[0044] The term "aryl" refers to an aromatic monolycyclic or polycyclic substituent having from 6 to 14 carbon atoms. Substituents phenyl, naphth-1-yl; naphth-2-yl; anthracen-9-yl; 1,2,3,4-tetrahydronaft-5-yl; and 1,2,3,4-tetrahydronaft-6-yl are examples of the aryl substituent.

[0045] The term "heteroaryl" refers to a heteroaromatic mono- or polycyclic substituent having from 1 to 13 carbon atoms and from 1 to 4 heteroatoms. Pyrrol-1-yl substituents; pyrrol2-yl; pyrrol-3-yl; furyl; thienyl; imidazolyl; oxazolyl; thiazolyl; isoxazolyl; isothiazolyl; 1,2,4-triazolyl; oxadiazolyl;

thiadiazolyl; tetrazolyl; pyridyl; pyrimidyl; pyrazinyl; 1,3,5triazynyl; indolyl; benzo [b] furyl; benzo [b] thienyl; indazolyl;

benzimidazolyl; azaindolyl; quinoleil; isoquinoleil; carbazolyl; and acrydyl are examples of a heteroaryl substituent.

[0046] The term "heteroatom" refers herein to at least a divalent atom, different from carbon. N; ABOUT; S; et Se are examples of a heteroatom.

[0047] The term "cycloalkyl" refers to a cyclic saturated or partially unsaturated cyclic hydrocarbon substituent having from 3 to 12 carbon atoms. Cyclopropyl substituents; cyclobutyl; cyclopentyl; cyclopentenyl; cyclopentadienyl; cyclohexyl; cyclohexenyl; cycloheptyl; bicyclo [2.2.1] heptyl; cyclooctyl; bicyclo [2.2.2] octyl; adamantyl; et perhydronaphthyl are examples of a cycloalkyl substituent.

[0048] The term "heterocyclyl" refers to a cyclic saturated or partially unsaturated cyclic hydrocarbon substituent having from 1 to 13 carbon atoms and from 1 to 4 heteroatoms. Preferably, the saturated or partially unsaturated cyclic hydrocarbon substituent will be monocyclic and will contain 4 or 5 carbon atoms and 1 to 3 heteroatoms.

[0049] The term "substituted" refers to one or more substituents other than H, for example, halogen; alkyl;

aryl; heteroaryl, cycloalkyl; heterocyclyl; alkylene; alkynyl; OH; O-alkyl; O-alkylene; O-aryl; O-heteroaryl; NH2; NH-alkyl; NHarylu; NH-heteroaryl; N-alkyl-alkyl '; SH; S-alkyl; S-aryl;

S (O2) H; S (O2) -alkyl; S (O2) -aryl; SO3H; SO3-alkyl; SO3-aryl; CHO; C (O) -alkyl; C (O) -aryl; C (O) OH; C (O) O-alkyl; C (O) O-aryl; OC (O) alkyl; OC (O) -aryl; C (O) NH2; C (O) NH-alkyl; C (O) NH-aryl; NHCHO;

NHC (O) -alkyl; NHC (O) -aryl; NH-cycloalkyl; NH-heterocyclyl.

[0050] The products according to the invention can be produced starting from conventional methods of organic chemistry. Scheme 1 below is an illustration of the method used to prepare Example 1 for substituted 6-aza-indoles. In this regard, it could not constitute a limitation of the scope of the invention, including as regards the methods for preparing the claimed compounds.

Preparation of derivatives of substituted 6-aza-indole-2-carboxamide in position 3:

Diagram 1

EtOH / Na / TA

NO2

<img file="PL1877409T3_D0002.tif" />

H2 / 2b

Pd / C / EtOH

<img file="PL1877409T3_D0003.tif" />

<img file="PL1877409T3_D0004.tif" />

CO2Et

NH3 / MeOH nh₄oh

<img file="PL1877409T3_D0005.tif" />

ArNCO / THF / TA

<img file="PL1877409T3_D0006.tif" />

[0051] Scheme 2 below is an illustration of the method used to prepare examples of substituted 7-aza-indoles, in particular example 7. Therefore, it could not limit the scope of the invention, including methods of producing the claimed compounds.

Preparation of derivatives of substituted 7-aza-indole-2-carboxamide in position 3:

Diagram 2

<img file="PL1877409T3_D0007.tif" />

BuLi / CO₂/-78^aboutC fi then tBuLi / CO2 then H + 0

N&gt;

H 'f SOCl₂ / MeOH

OH

OMe

pyridine

br₃^-

Br &lt;

<img file="PL1877409T3_D0008.tif" />

N OMe

H

<img file="PL1877409T3_D0009.tif" />

NH4OH NH₃ / MeOH

<img file="PL1877409T3_D0010.tif" />

CF3

<img file="PL1877409T3_D0011.tif" />

[0052] Products of general formula (I), where Ra is different from H, can be obtained according to conventional methods known to the skilled person, for example replacing ammonia in aminolysis with the corresponding primary alkylamine.

[0053] The present invention furthermore relates to a process for the production of products of formula (I) as defined above, characterized in that the product of the following general formula (V):

<img file="PL1877409T3_D0012.tif" />

is subject to the following stages:

a) halogenation in position 3, then

b) Suzuki coupling in position 3 to obtain a product of the following general formula (III):

<img file="PL1877409T3_D0013.tif" />

and then

c) amidation of the ester in position 2 to obtain a product of the following general formula (II):

<img file="PL1877409T3_D0014.tif" />

next

d) acylation of the amino-phenyl group at the 3-position.

[0054] The present invention furthermore provides, as intermediates, compounds having the following general formula (II):

<img file="PL1877409T3_D0015.tif" />

wherein Z, Y, W are as previously defined for the preparation of products of general formula (I).

[0055] LC / MS analyzes were performed on a Micromass model LCT associated with an HP 1100 camera. The relative content of products was measured using an HP G1315A diode array detector in the 200-600 nm wave range and a Sedex 65 light dispersion detector. Mass spectra mass spectra were collected in the range of 180 to 800. Data were analyzed using Micromass MassLynx software. The division was made on a Hypersil BDS C18, 3 μm (50 x 4.6 mm) column, eluting with a linear gradient of 5 to 90% acetonitrile containing 0.05% (v / v) trifluoroacetic acid (TFA) in water containing 0.05% (v / v) TFA at 3.5 min with a flow rate of 1 mL / min. The total analysis time, including the column re-equilibration period, is 7 min.

[0056] MS spectra were taken using an electrospray (ES⁺) on a Platform II (Micromass) apparatus. The main observed ions are described.

[0057] Melting points were measured in a capillary on a Mettler FP62 apparatus, range 30 ° C to 300 ° C, increase 2 ° C per minute.

Purification by LC / MS:

[0058] The products can be purified by LC / MS using a Waters FractionsLynx system consisting of a Waters model 600 gradient pump, a Waters model 515 regenerative pump, a Waters Reagent Manager dilution pump, a Waters model 2700 autodispenser, two Rheodyne model LabPro valves, a diode array detector Waters model 996, Waters model ZMD mass spectrometer and Gilson fraction collector model 204. The system was controlled by Waters software

FractionLynx. The separation was made alternatively on two columns

Waters Symmetry (C.₁₈, 5μΜ, 19x50 mm, catalog number 186000210), where the column was being regenerated by a water / acetonitrile 95/5 (v / v) mixture containing 0.07% (v / v) trifluoroacetic acid, while the other column was in during the chapter. Column elution was performed using a 5 to 95% acetonitrile linear gradient containing 0.07% (v / v) trifluoroacetic acid in water at a flow rate of 10 mL / min. At the exit of the separation column, one thousandth of the effluent is separated by LC Packing Accurate, diluted with methyl alcohol at a flow of 0.5 mL / min and sent to detectors, using 75% to a detector with a diode system, and the remaining 25% to a mass spectrometer. The rest of the effluent (999/1000) is sent to the fraction collector, where the flow is eliminated when the expected mass of the product is not detected by the FractionLynx software. The molecular formulas of the expected products are delivered to the FractionLynx software, which causes the product to collect when the detected mass signal corresponds to the [M + H] ion⁺ and / or [M + Na]⁺. In some cases, depending on the LC / MS analytical results, when a strong ion corresponding to [M + 2H] is detected⁺⁺, a value corresponding to half the calculated molecular weight (MW / 2) is also supplied to the FractionLynx software. Under these conditions, collection also occurs when the following are detected: ion mass signal [M + 2H]⁺⁺ and / or [M + Na + H]⁺⁺. Products were collected in tared tubes. After collection, the solvents were evaporated on a Savant AES 2000 or rotary evaporator

Genevac HT8 and product weights were determined by weighing the tubes after evaporation of the solvents.

[0059] Another object of the invention relates to the products of the examples below, which illustrate in a non-limiting manner the present invention.

Example 1: 3- {4- [3- (2-fluoro-5-trifluoromethyl-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0060]

<img file="PL1877409T3_D0016.tif" />

[0061] To a solution of 90 mg 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide in 5 ml tetrahydrofuran is added, dropwise, 50 μL of 2-fluoro-5- (trifluoromethyl) phenylisocyanate. The reaction mixture is stirred for 16 hours at ambient temperature under argon, then concentrated under reduced pressure. The residue obtained is stirred for 30 minutes in 2 ml of dichloromethane. The solid in suspension is filtered and squeezed out. After drying under vacuum at 40 ° C, 115 mg of 3- {4- [3- (2-fluoro-5-trifluoromethylphenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine- 2-carboxamide with the following characteristics:

IR (KBr): 3455; 1661; 1602; 1542; 1444; 1341; 1312; 1127; 1070 et 819 cm^-1 NMR ¹H: 6.98 (s broad, 1H); 7.39 (m wide, 1H); from 7.42 to 7.56 (m, 4H); 7.60 (broad d, J = 8.0 Hz, 2H); 7.74 (s broad, 1H); 8.17 (d, J = 6.0 Hz, 1H); 8.65 (broad d, J = 7.5 Hz, 1H); 8.82 (s, 1 H); 8.94 (s broad, 1H); 9.31 (s, 1 H); 12.15 (s broad, 1H). Mass Spectrum (ES⁺): m / z = 458 [M + H]⁺

Melting point: 286 ° C (Kofler).

3- (4-Aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide:

To a solution of 600 mg of ethyl 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxylate in 62 ml of a 3N solution of ammonia in methanol is added 11 ml of a 22% aqueous solution of ammonia. The reaction mixture is stirred for 20 hours in an autoclave at 80 ° C (12 bar) and then concentrated under reduced pressure. The residue obtained is diluted in 100 ml of methanol, treated with carbon black and refluxed for 30 minutes. The mixture is hot filtered through Celite then washed with 2 x 10 mL methanol. The filtrate is concentrated under reduced pressure to obtain 490 mg of 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide in the form of a foam having the following characteristics:

Mass spectrum (EI) m / z = 252 [M]^+°, m / z = 235 [M-NH3]^+°

Ethyl 3- (4-Aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxylate:

[0063] To a solution of 1 g of ethyl 3-bromo-1H-pyrrolo [2,3-c] pyridine-2-carboxylate in 100 ml of dioxane, 773 mg of 4-aminophenyl boronic acid hydrochloride, 1.1 g of potassium fluoride in 9 are added ml of water.

The reaction mixture is stirred under argon for 15 minutes.

425 mg tetrakis (triphenylphosphine) palladium (0) and 630 μL triethylamine are added. The reaction mixture is stirred for 17 hours under reflux. After soot treatment, followed by filtration through Celite®, the filtrate is concentrated under reduced pressure. The residue is purified by flash column chromatography on silica (60; 35-70 μΜ), eluting with a mixture of dichloromethane, methanol and acetonitrile (90/5/5 vol.). 600 mg of ethyl 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxylate are obtained with the following characteristics:

Mass spectrum (EI) m / z = 281 [M]^+°, m / z = 235 [M-OEt]^+°

Ethyl 3-bromo-1H-pyrrolo [2,3-c] pyridine-2-carboxylate:

To a solution of 2.24 g of ethyl 1H-pyrrolo [2,3-c] pyridine-2-carboxylate in 150 ml of pyridine, a solution of 3.53g of pyridinium tribromide in 30 ml of pyridine is added dropwise at 5 ° C . The reaction mixture is then stirred for 16 hours at a temperature close to 20 ° C, then washed with 500 ml ice water. The suspension is filtered. The resulting solid is washed with water and then dried in an oven under vacuum at 40 ° C. 1.97 g of ethyl 3-bromo-1H-pyrrolo [2,3-c] pyridine-2-carboxylate are obtained with the following characteristics:

Mass spectrum (EI) m / z = 269 [M]^+°, m / z = 189 [M-Br]^+°, m / z = 144 [MOEt]^+°

Ethyl 1H-Pyrrolo [2,3-c] pyridine-2-carboxylate:

[0065] 1.8 g of palladium on carbon (10%) are charged in an autoclave, and then an inert atmosphere is created with a stream of argon. A solution of 6 g of ethyl 3- (3-nitropyridin-4-yl) -2-oxo-propionate in 72 ml of absolute ethanol is added. The reaction medium is then stirred 3 hours at 20 ° C under 2 bar hydrogen pressure. The mixture is then run through celite®. The filtrate is concentrated under reduced pressure, dried in an oven at 40 ° C to obtain 4g of ethyl 1H-pyrrolo [2,3-c] pyridine-2-carboxylate with the following characteristics:

Mass spectrum (EI) m / z = 190 [M^+°], m / z = 144 [M-OEt]^+°

Ethyl 3- (3-nitropyridin-4-yl) -2-oxo-propionate:

[0066] To a solution of 930 mg sodium in 50 ml absolute ethanol, 26 ml diethyl oxalate are quickly added. The reaction medium is stirred for 15 minutes at 20 ° C. A solution of 3.8 g of 4-methyl-3-nitropyridine in 50 ml of absolute ethanol is then added dropwise over 1 hour. The reaction mixture is stirred for 4 hours at a temperature close to 20 ° C, then concentrated under reduced pressure. The residue is taken up in 100 ml of ethyl ether and then filtered. The solid is mixed with 40 ml chlorhydrique 5N, then filtered, washed with water and dried in vacuo at 40 ° C to obtain 6.2 g of ethyl 3- (3-nitropyridin-4-yl) -2-oxo-propionate with the following characteristics:

Mass spectrum (EI) m / z = 238 [M^+°].

Example 2: 3- {4- [3- (2-methoxy-5-methyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0067]

<img file="PL1877409T3_D0017.tif" />

[0068] To a solution of 100 mg 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide in 5 ml tetrahydrofuran is added dropwise, 54.4 pL

2-methoxy-5-metylfenylizocyjanianu. The reaction mixture is stirred for 16 hours at ambient temperature under argon and then concentrated under reduced pressure. The residue obtained is stirred for 30 minutes in 2 ml of dichloromethane. The solid in suspension is filtered, washed with water and pressed. After drying under vacuum at 40 ° C, 40 mg of 3- {4- [3- (2-methoxy-5-methylphenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine- 2-carboxamide with the following characteristics:

IR (KBr), 3458; 3331; 1664; 1595; 1537; 1315;_cm^-1

NMR ¹H: 2.24 (s, 3H); 3.86 (s, 3H); 6.75 (d; from 6.85 to 6.95 (m, 2H); 7.43 (broad d,

1285; 1213;

wide, JJ = 8.5 Hz,, 5 Hz, 2H); , J = 5.5 Hz; 12.1 (ss (d, J = 5.5 Hz, 1H);

broad, 1H); 8.02 (s (s, 1H); 8.82 (s, 1H) Mass spectrum (EI): m / z Melting point:

7.58 (d broad, J = 8 broad, 1H); 8.16 (d; 9.44 (s broad, 1H) = 415 [M^+°]

227 ° C

<td> 1135</td><td> ; 1033</td>

<td> = 8,5</td><td>Hz, 1H)</td>

<td>2H);</td><td> 7,46</td>

<td> 7,73</td><td>(s</td>

<td>, 1H)</td><td> ; 8,22</td>

zeros, 1H).

Example 3: 3- {4- [3- (3-Chloro-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide trifluoroacetate.

[0069]

<img file="PL1877409T3_D0018.tif" />

[0070] To a solution of 100 mg 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide in 5 ml tetrahydrofuran is added dropwise, 45.2 pL

3-chlorophenylisocyanate. The reaction mixture is stirred for 16 hours at ambient temperature under argon and then concentrated under reduced pressure. The residue obtained is stirred for 30 minutes in 2 ml of dichloromethane. The solid in suspension is filtered, washed with water and pressed. Final purification is performed by preparative LC / MS to obtain after drying under vacuum at 40 ° C 70 mg 3- {4- [3- (3-chloro-phenyl) -ureido] -phenyl) -1H-pyrrolo [2, 3c] pyridine-2-carboxamide in the form of the trifluoroacetate salt with the following characteristics:

IR (KBr): 3390; 1672; 1592; 1537; 1483; 1203; 1138; 836; 722 cm^-1

<td>NMR</td><td>¹H</td><td>: 7.03 (m, 1H);</td><td>from 7.26</td><td>down</td><td> 7,34</td><td>(m, 2H)</td><td>; from 7.42 to 7.52 (m,</td>

<td>3H)</td><td>^;</td><td>7.63 (broad d,</td><td>J = 8.5</td><td>Hz,</td><td>2H)</td><td> ; 7,74</td><td>(s broad, 1H); 7.97</td>

<td>(D,</td><td>J</td><td>= 6.0 Hz, 1H);</td><td>8.06 (p</td><td colspan="2">wide,</td><td>1H); 8</td><td>, 31 (d, J = 6.0 Hz, 1H)</td>

<td> ; 9,</td><td> 03</td><td>(s wide, 2H)</td><td> ; 9,13</td><td>(S,</td><td>1H)</td><td> ; 13,35</td><td>(m spread out, 1H).</td>

<td colspan="2">Spectrum</td><td>mass (ES⁺): m /</td><td>z = 406</td><td>[MH⁺</td><td> ]</td><td></td><td></td>

Melting point: 221 ° C

Example 4: 3- {4- [3- (3-Chloro-4-fluoro-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide trifluoroacetate [0071]

<img file="PL1877409T3_D0019.tif" />

[0072] To a solution of 100 mg 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide in 5 ml tetrahydrofuran is added dropwise, 46.2 pL

3-chloro-4-fluorophenylisocyanate. The reaction mixture is stirred for 16 hours at ambient temperature under argon and then concentrated under reduced pressure. The residue obtained is stirred for 30 minutes in 2 ml of dichloromethane. The solid in suspension is filtered, washed with water and pressed. Final purification is performed by preparative LC / MS to obtain after drying under vacuum at 40 ° C 105 mg 3- {4- [3- (3-chloro-4-fluoro-phenyl) -ureido] phenyl} -1H- pyrrolo [2,3-c] pyridine-2-carboxamide in the form of the trifluoroacetate salt with the following characteristics:

IR (KBr): 3452; 1673; 1601; 1544; 1500; 1208; 1143; 836; 803; 722 cm^-1

NMR ¹H: from 7.32 to 7.38 (m, 2H); from 7.44 to 7.54 (m, 3H); 7.64 (broad d, J = 8.5 Hz, 2H); 7.84 (broad d, J = 7.5 Hz, 1H); 8.01 (d, J = 6.0 Hz, 1H); 8.09 (s broad, 1H); 8.32 (d, J = 6.0 Hz, 1H); 9.10 (s broad, 2H); 9.16 (s, 1 H); 13.4 (m spread out, 1H).

Mass Spectrum (ES⁺): m / z = 424 [MH⁺]

Melting point: 214 ° C

Example 5: 3- {4- [3- (2-Fluoro-5-methyl-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide trifluoroacetate [0073]

<img file="PL1877409T3_D0020.tif" />

[0074] To a solution of 100 mg 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide in 5 ml tetrahydrofuran is added dropwise, 48.3 μL of 2-fluoro-5-methylphenylisocyanate. The reaction mixture is stirred for 16 hours at ambient temperature under argon and then concentrated under reduced pressure. The residue obtained is stirred for 30 minutes in 2 ml of dichloromethane. The solid in suspension is filtered, washed with water and pressed. Final purification is performed by preparative LC / MS to obtain, after drying under vacuum, at 40 ° C, 36 mg 3- {4- [3- (2-fluoro-5-methyl-phenyl) -ureido] phenyl} -1H pyrrolo [2,3-c] pyridine-2-carboxamide in the form of the trifluoroacetate salt with the following characteristics:

IR (KBr): 3452; 1675; 1603; 1544, 1314; 1202; 1144; 836; 805; 722 cm^-1NMR ¹H: 2.29 (s, 3H); 6.82 (m, 1H); 7.12 (dd, J = 8.5 and 11.5 Hz, 1H); from 7.46 to 7.51 (m, 3H); 7.62 (broad d, J = 8.5 Hz, 2H); from 7.97 to 8.03 (m, 2H); 8.08 (s broad, 1H); 8.32 (d, J = 6.5 Hz, 1H); 8.54 (broad d, J = 2.5 Hz, 1H); 9.15 (s, 1 H); 9.25 (s, 1 H); 13.4 (m spread out, 1H)

Mass Spectrum (ES⁺): m / z = 404 [MH⁺]

Melting point: 222 ° C

Example 6: 3- [4- (3-m-tolyl-ureido) -phenyl] -1H-pyrrolo [2,3-c] pyridine-2-carboxamide trifluoroacetate [0075]

<img file="PL1877409T3_D0021.tif" />

[0076] To a solution of 100 mg 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide in 5 ml tetrahydrofuran is added dropwise, 47.8 μL of m-tolylisocyanate. The reaction mixture is stirred for 16 hours at ambient temperature under argon and then concentrated under reduced pressure. The residue obtained is stirred for 30 minutes in 2 ml of dichloromethane. The solid in suspension is filtered, washed with water and pressed. Final purification is performed by preparative LC / MS to obtain after drying under vacuum at 40 ° C, 40 mg 3- [4- (3-m-tolyl-ureido) -phenyl] -1H-pyrrolo [2,3- c] pyridine-2-carboxamide in the form of a trifluoroacetate salt with the following characteristics:

IR (KBr): 3408; 1699; 1595; 1526; 1203; 1138; 834; 797; 724 cm^-1NMR ¹H: 2.29 (s, 3H); 6.81 (broad d, J = 7.5 Hz, 1H); 7.17 (t, J =

7.5 Hz, 1H); 7.25 (broad d, J = 7.5 Hz, 1H); 7.32 (s wide, 1

H); 7.44 (s broad, 1H); 7.47 (broad d, J = 8.5 Hz, 2H); 7.62 (broad d, J = 8.5 Hz, 2H); 7.96 (m wide, 1H); 8.06 (s broad, 1H); 8.30 (d, J = 6.0 Hz, 1H); 8.67 (s, 1H); 8.86 (s, 1 H); 9.12 (s,

1H); 13.3 (m spread-out, 1H).

Mass Spectrum (ES⁺): m / z = 386 [MH⁺]

Example 7: 3- {4- [3- (2-fluoro-5 trifluoromethyl-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide [0077]

<img file="PL1877409T3_D0022.tif" />

[0078] To a solution of 130 mg 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide in 5 ml tetrahydrofuran is added, dropwise, 85 μL of 2-fluoro-5- (trifluoromethyl) phenylisocyanate. The reaction mixture is then stirred for 16 hours at ambient temperature under an argon atmosphere and then concentrated under reduced pressure. The residue obtained is chromatographed on a silica column (eluant dichloromethane-methanol 9-1 vol.). Fractions containing the expected product are concentrated under reduced pressure.

237 mg of 3- {4- [3- (2-fluoro-5-trifluoromethyl-phenyl) ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide are obtained in the form of a white solid the following characteristics:

IR (KBr): 1659; 1623; 1542; 1443; 1339; 1316; 1119 cm^-1

NMR ¹H: 7.08 (m spread out, 1H); 7.14 (dd, J = 5.0 and 8.0 Hz, 1H); 7.40 (m, 1H); 7.46 (broad d, J = 8.5 Hz, 2H); 7.51 (m, 1H); 7.57

<td colspan="2">(broad d, J = 8.5 Hz, 2H)</td><td>; from</td><td> 7,</td><td> 55</td><td>down</td><td> 7,</td><td colspan="2">60 (m masked, 1H)</td>

<td>7.92 (d</td><td>wide, J = 8.0 Hz,</td><td>1H)</td><td> ;</td><td> 8,</td><td> 38</td><td>(d</td><td>wide,</td><td>J = 5.0 Hz, 1</td>

<td>8.64 (d</td><td>wide, J = 7.5 Hz,</td><td>1H)</td><td> ;</td><td> 9,</td><td> 01</td><td>(s</td><td>wide,</td><td>1H); 9.36 (p</td>

<td>wide,</td><td colspan="2">1H); 12.1 (s wide,</td><td>1H)</td><td></td><td></td><td></td><td></td><td></td>

Mass Spectrum (ES⁺): m / z = 458 [M + H⁺]

Melting point: 232 ° C (Kofler).

3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide

[0079] To a solution of 260 mg of methyl 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxylate in 30 ml of a 7N ammonia in methanol solution is added 5 ml of aqueous ammonia (22%). The reaction mixture is then stirred for 20 hours in an autoclave at 80 ° C (12.6 bar) and then concentrated under reduced pressure. The residue obtained is chromatographed on a silica column (eluant dichloromethane-methanol 9-1 vol.). Fractions containing the expected product are concentrated under reduced pressure. 140 mg are obtained

3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide in the form of a pale yellow solid with the following characteristics:

Melting point: 139 ° C

Methyl 3- (4-Aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxylate:

[0080] To a solution of 0.64 g of methyl 3-bromo-1H-pyrrolo [2,3-b] pyridine-2-carboxylate in 50 ml of toluene and 50 ml of methanol, 1.08 g of 4-aminophenyl hydrochloride boronate and 0.9 ml triethylamine. The reaction mixture is then stirred under argon for 15 minutes. 144 mg tetrakis (triphenylphosphine) palladium (0), 0.3 g lithium chloride, 0.66 g sodium carbonate, and 7.5 ml distilled water are added successively. The reaction mixture is stirred for 8 hours under reflux. After filtration through Celit®, the filtrate is concentrated under reduced pressure. The residue is purified by silica column chromatography eluting with a mixture of ethyl acetate and cyclohexane (7-3 vol.). 400 mg of methyl 3- (4-aminophenyl) -1H-pyrrolo [2,3b] pyridine-2-carboxylate are obtained in the form of a yellow solid with the following characteristics:

Melting point: 236 ° C

Methyl 3-Bromo-1H-pyrrolo [2,3-b] pyridine-2-carboxylate:

To a solution of 3.2 g of methyl 1H-pyrrolo [2,3-b] pyridine-2-carboxylate hydrochloride in 165 ml of pyridine, a solution of 5.04 g of pyridinium tribromide in 35 ml of pyridine is added dropwise at 0 ° C under an argon atmosphere. . The reaction mixture is then stirred at 0 ° C, then poured onto a mixture of 250 g crushed ice and 750 ml distilled water. The suspension is filtered, the solid is washed twice with 25 ml of distilled water, then dried in the open air. 0.87 g of methyl 3-bromo-1H-pyrrolo [2,3-b] pyridine-2-carboxylate is obtained in the form of a beige solid with the following characteristics:

Mass Spectrum (ES⁺): m / z = 256 [M + H⁺]

Methyl 1H-pyrrolo [2,3-b] pyridine-2-carboxylate hydrochloride:

[0082] To a solution of 4 g of 1H-pyrrolo [2,3-b] pyridine-2-carboxylic acid hydrochloride in 100 ml of methanol is added dropwise 6 ml of thionyl chloride at ambient temperature. The reaction mixture is then stirred for 5 hours at ambient temperature and then concentrated under reduced pressure. The residue obtained is triturated in 50 ml of ethyl ether, then dried under vacuum at 40 ° C. 3.22 g of methyl 1H-pyrrolo [2,3-b] pyridine-2-carboxylate hydrochloride are obtained in the form of a pale yellow solid used as such in the next step.

1H-Pyrrolo [2,3-b] pyridine-2-carboxylic acid hydrochloride [0083] To a solution cooled to -70 ° C, 6.03 g of 1H-pyrrolo [2,3b] pyridine in 75 ml of anhydrous THF is added dropwise. 33 ml solution

1.6M n-butyllithium in hexane. After 15 minutes of stirring at -70 ° C, 20g of dry ice in pieces are added to the solution. It is then allowed to return to ambient temperature and then concentrated under reduced pressure. 8.4 g of a white solid are obtained, which is dissolved in 175 ml of tetrahydrofuran. This solution is cooled to -70 ° C and then 35 ml of solution are added dropwise

1.5M t-butyllithium in hexane. After 30 min of stirring at - 70 ° C, 20 g of dry ice in pieces in solution are added. It is then allowed to return to ambient temperature, then this reaction mixture is poured onto 50 ml of distilled water cooled to 0 ° C.

The tetrahydrofuran is evaporated under reduced pressure. The residual aqueous solution is diluted with 150 ml distilled water, washed twice with 100 ml dichloromethane, acidified to pH 1 by the addition of 30 ml aqueous 5N hydrochloric acid solution and then concentrated under reduced pressure. 10.01 g of a pasty solid are obtained, which is recrystallized in 50 ml of methanol. The solid obtained is treated with a mixture of 50 ml isopropanol in hydrochloric acid (7N) and 50 ml isopropyl ether. After air drying, 5.71 g of 1H-pyrrolo [2,3-b] pyridine-2-carboxylic acid hydrochloride is obtained in the form of a cream solid.

Mass Spectrum (EI): m / z = 162 [M^+°]

Example 8: 3- {4- [3- (2-fluoro-5-trifluoromethyl-phenyl) -ureido] phenyl} -7-oxy-1H-pyrrolo [2,3-b] pyridine-2-carboxamide [0084]

F

<img file="PL1877409T3_D0023.tif" />

[0085] To a solution of 50 mg 3- {4- [3- (2-fluoro-5-trifluoromethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide in a 0 ° C solution 2 ml of chloroform is added dropwise 0.31 ml of a solution of 0.7M metachloroperbenzoic acid in chloroform. The solution is stirred at 0 ° C for 4 hours and then at ambient temperature for 16 hours. The reaction mixture is diluted with 3 ml of dichloromethane, filtered through a N4 glass frit, the solid obtained is washed twice with 3 ml of dichloromethane and then air-dried. 40 mg of 3- {4- [3- (2-fluoro-5-trifluoromethyl-phenyl) ureido] -phenyl} -7-oxy-1H-pyrrolo [2,3-b] pyridine-2-carboxamide in forms of a light yellow solid with the following characteristics:

IR (KBr): 3352; 1671; 1609; 1545; 1442; 1340; 1315; 1239; 1119; 1069 and 885 cm^-1

NMR ¹H: 7.16 (m, 1H); from 7.35 to 7.58 (m, 7H); 7.63 (m spread-out, 1H); 7.77 (m spread out, 1H); 8.31 (broad d, J = 6.0 Hz, 1H); 8.65 (broad d, J = 8.5 Hz, 1H); 8.94 (s broad, 1H); 9.29 (s, 1 H); from 12.5 to 13.2 (m very stretched, 1H).

Mass Spectrum (ES⁺): m / z = 474 [M + H⁺]

Melting point: 220 ° C (Kofler).

Example 9: 3- {4- [3- (2-fluoro-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3b] pyridine-2-carboxamide [0086]

<img file="PL1877409T3_D0024.tif" />

66.6 mg of a beige solid 3- {4- [3- (2-fluoro-phenyl) ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide is prepared as described in example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 2-fluoro-phenylisocyanate. Melting point = 268.7 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺ = 390

Retention time (min): 3.71

Example 10: 3- {4- [3- (2-methoxy-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3b] pyridine-2-carboxamide [0088]

<img file="PL1877409T3_D0025.tif" />

[0089] 83.6 mg of a beige solid 3- {4- [3- (2-methoxy-phenyl) ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide is prepared as described in example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 2-methoxy-phenylisocyanate. Melting point: 227.1 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺ = 402

Retention time (min): 3.77

Example 11: 3- {4- [3- (4-trifluoromethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide [0090]

F

<img file="PL1877409T3_D0026.tif" />

77.6 mg of white solid 3- {4- [3- (4-trifluromethylphenyl) -ureido] -phenyl} 1H-pyrrolo [2,3-b] pyridine-2-carboxamide is prepared as described in Example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 4-trifluoromethylphenylisocyanate.

Melting point: 296.2 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺ = 440

Retention time (min): 4.24

Example 12: 3- {4- [3- (2-Chloro-5-trifluoromethyl-phenyl) -ureido] phenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide.

[0092]

<img file="PL1877409T3_D0027.tif" />

40.56 mg of a white solid 3- {4- [3- (2-chloro-5-trifluoromethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide is prepared as described in Example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 2-chloro-5-trifluoromethyl-phenylisocyanate.

Melting point: 188.3 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺ = 474 Retention time (min): 4.51

Example 13: 3- {4- [3- (2-fluoro-3-trifluoromethyl-phenyl) -ureido] phenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide.

[0094]

F

<img file="PL1877409T3_D0028.tif" />

-F

F [0095] 79 mg of a white solid 3- {4- [3- (2-fluoro-310 trifluoromethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide is prepared as described in Example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 2-fluoro-3-trifluoromethyl-phenylisocyanate.

Melting point: 265.4 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺ = 458

Retention time (min): 4.24

Example 14: 3- {4- [3- (4-fluoro-3-trifluoromethyl-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-b) pyridine-2-carboxamide.

[0096]

<img file="PL1877409T3_D0029.tif" />

, Oh₂ [0097] 76.5 mg of brown solid 3- {4- [3- (4-fluoro-3-trifluoromethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide prepared as described in Example 7 starting from 325 (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 4-fluoro-3-trifluoromethyl-phenylisocyanate.

Melting point: 234.7 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺ = 458

Retention time (min): 4.22

Example 15: 3- {4- [3- (3-fluoro-5-trifluoromethyl-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide [0098]

F

<img file="PL1877409T3_D0030.tif" />

78.1 mg of a beige solid 3- {4- [3- (3-fluoro-5-trifluoromethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide is prepared as described in Example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 3-fluoro-5-trifluoromethyl-phenylisocyanate.

Melting point: 257.5 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺ = 458

Retention time (min): 4.42

Example 16: 3- {4- [3- (4-trifluoromethoxy-phenyl) -ureidol-phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide.

[0100]

<img file="PL1877409T3_D0031.tif" />

[0101] 92.3 mg of chestnut powder 3- {4- [3- (4-trifluoromethoxyphenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide is prepared as described in Example 7 starting from 3- (4aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 4-trifluoromethoxy-phenylisocyanate.

Melting point: 258.9 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺ = 456

Retention time (min): 4.29

Example 17: 3- {4- [3- (3,4-dimethoxy-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide [0102]

<img file="PL1877409T3_D0032.tif" />

79 mg of a beige solid 3- {4- [3- (3,4-dimethoxy-phenyl) ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide is prepared as described in Example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 3,4-dimethoxyphenylisocyanate.

Melting point: 223.7 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺ = 432

Retention time (min): 3.27

Example 18: 3- {4- [3- (2,5-dimethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide [0104]

<img file="PL1877409T3_D0033.tif" />

75.9 mg of a white solid 3- {4- [3- (2,5-dimethyl-phenyl) ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide produces as described in Example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 2,5-dimethyl-phenylisocyanate. Melting point: 308.8 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺ = 400

Retention time (min): 3.90

Example 19: 3- {4- [3- (3-methoxy-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide [0106]

<img file="PL1877409T3_D0034.tif" />

[0107] 55.5 mg of a beige solid 3- {4- [3- (3-methoxy-phenyl) ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide is prepared as described in example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 3-methoxy-phenylisocyanate. Melting point: 306.2 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺ = 402

Retention time (min): 3.39

Example 20: 3- {4- [3- (3-trifluoromethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide [0108]

<img file="PL1877409T3_D0035.tif" />

Η

ΝΗ₂

F [0109] 56.5 mg of a white solid 3- {4- [3- (3-trifluoromethylphenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide is prepared as described in example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 3-trifluoromethyl-phenylisocyanate.

Melting point: 263.6 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺ = 440

Retention time (min): 3.95

Example 21: 3- {4- [3- (3,4-dimethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide [0110]

<img file="PL1877409T3_D0036.tif" />

45.2 mg of a white solid 3- {4- [3- (3,4-dimethyl-phenyl) ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide is prepared as described in Example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 3,4-dimethyl-phenylisocyanate. Melting point: 274.7 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺ = 400

Retention time (min): 3.75

Example 22: 3- {4- [3- (2-methoxy-5-methyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide.

[0112] ο

<img file="PL1877409T3_D0037.tif" />

44.9 mg of a beige solid 3- {4- [3- (2-methoxy-5-methylphenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide produces as described in Example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 2-methoxy-5-methyl-phenylisocyanate.

Melting point: 327.7 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺ = 416

Retention time (min): 3.76

Example 23: 3- [4- (3-m-tolyl-ureido) -phenyl] -1H-pyrrolo [2,3-b] pyridine-2-carboxamide [0114]

<img file="PL1877409T3_D0038.tif" />

N '[0115] 62.5 mg of a beige solid 3- [4- (3-m-tolyl-ureido) phenyl] -1H-pyrrolo [2,3-b] pyridine-2-carboxamide is prepared as described in Example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3b] pyridine-2-carboxamide and m-tolylisocyanate.

Melting point: 266 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺ = 386

Retention time (min): 3.60

Example 24: 3- {4- [3- (4-fluoro-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide [0116]

<img file="PL1877409T3_D0039.tif" />

49.7 mg of a beige solid 3- {4- [3- (4-fluoro-phenyl) ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide is prepared as described in Example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 4-fluoro-phenylisocyanate. Melting point: 299.9 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺ = 390

Retention time (min): 3.45

Example 25: 3- [4- (3-p-tolyl-ureido) -phenyl] -1H-pyrrolo [2,3-b] pyridine-2-carboxamide

<img file="PL1877409T3_D0040.tif" />

[0119] 68.4 mg of a beige solid 3- [4- (3-p-tolyl-ureido) phenyl] -1H-pyrrolo [2,3-b] pyridine-2-carboxamide is prepared as described in Example 7 starting from from 3- (4-aminophenyl) -1H-pyrrolo [2,3b] pyridine-2-carboxamide and p-tolylisocyanate.

Melting point: 293 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺ = 386

Retention time (min): 3.58

Example 26: 3- {4- [3- (4-methyl-3-trifluoromethyl-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide [0120]

<img file="PL1877409T3_D0041.tif" />

47.1 mg of a white solid 3- {4- [3- (4-methyl-3-trifluoromethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide is prepared as described in Example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 4-methyl-3-trifluoromethyl-phenylisocyanate.

Melting point: 285 ° C

Mass Spectrum (ES⁺): [M + H]⁺ = 454

Retention time (min): 4.10

Example 27: 3- {4- [3- (4-difluoromethoxy-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide [0122]

<img file="PL1877409T3_D0042.tif" />

47.5 mg of a white solid 3- {4- [3- (4-difluoromethoxyphenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide is prepared as described in Example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 4-difluoromethoxy-phenylisocyanate.

Melting point: 283.5 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺ = 438 Retention time (min): 3.64

Example 28: 3- {4- [3- (3,5-dimethoxy-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide [0124]

<img file="PL1877409T3_D0043.tif" />

[0125] 59.2 mg of a beige solid 3- {4- [3- (3,5-dimethoxy-phenyl) ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide is prepared as described in example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 3,5-dimethoxyphenylisocyanate.

Melting point: 266.5 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺ = 432

Retention time (min): 3.45

Example 29: 3- {4- [3- (4-chloro-3-trifluoromethyl-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide [0126]

cl

NH₂ 29.8 mg of a solid 3- {4- [3- (4-chloro-3-trifluoromethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide is prepared as described in example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 4-chloro-3-trifluoromethyl-phenylisocyanate.

Melting point: 311.1 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺ = 474

Retention time (min): 4.22

Example 30: 3- {4- [3- (2,5-dimethoxy-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide [0128] /

<img file="PL1877409T3_D0044.tif" />

[0129] 33.1 mg of yellow 3- {4- [3- (2,5-dimethoxy-phenyl) ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide yellow lyophilisate is prepared as described in Example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 2,5-dimethoxyphenylisocyanate.

Mass Spectrum: LC-MS-DAD-ELSD: 432 (+) = (M + H) (+); 430 (-) = (MH) (-)

Retention time (min): 3.53

Example 31: 3- {4- [3- (3-fluoro-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide.

[0130]

F

<img file="PL1877409T3_D0045.tif" />

Ν '

NH₂ 31.5 mg of 3- {4- [3- (3-fluoro-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide white lyophilisate is prepared as described in Example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3b] pyridine-2-carboxamide and 3-fluoro-phenylisocyanate.

LC-MS-DAD-ELSD mass spectrum: 390 (+) = (M + H) (+); 388 (-) = (MH) (-)

Retention time (min): 3.55

Example 32: 3- {4- [3- (2-methoxy-5-trifluoromethyl-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide [0132]

<img file="PL1877409T3_D0046.tif" />

Ν '[0133] 50 mg of a beige solid 3- {4- [3- (2-methoxy-5-trifluoromethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide is prepared as described in Example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-b] pyridine-2-carboxamide and 2-methoxy-5-trifluoromethyl-phenylisocyanate.

Melting point: 221 ° C (Kofler-sublimation)

LC-MS-DAD-ELSD mass spectrum: 470 (+) = (M + H) (+) 468 (-) = (MH) (-)

Example 33: 3- {4- [3- (2-Acetylamino-5-trifluoromethylphenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide trifluoroacetate.

[0134]

F

<img file="PL1877409T3_D0047.tif" />

-F

F

<img file="PL1877409T3_D0048.tif" />

[0135] 12 mg of yellow solid 3- {4- [3- (2-acetylamino-5-trifluoromethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide trifluoroacetate is prepared as described in example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 2-acetylamino-5-trifluoromethyl-phenylisocyanate.

Mass Spectrum (ES⁺): [M + H]⁺ = 497

Retention time (min): 2.63

Example 34: 3- {4- [3- (2-methoxy-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0136]

<img file="PL1877409T3_D0049.tif" />

[0137] 25 mg of a yellow solid 3- {4- [3- (2-methoxy-phenyl) ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in Example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 2-methoxy-phenylisocyanate. Melting point: 216 ° C (Kofler)

Mass Spectrum (ES⁺): [M + H]⁺ = 402

Retention time (min): 3.06

Example 35: 3 {4- [3- (2-trifluoromethyl-phenyl) -ureido] -phenyl} -1Hpyrolo [2,3-c] pyridine-2-carboxamide.

[0138]

<img file="PL1877409T3_D0050.tif" />

[0139] 80 mg of a yellow solid 3- {4- [3- (2-trifluoromethylphenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in Example 1 starting from 3- (4aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 2-trifluoromethyl-phenylisocyanate.

Melting point: 228 ° C (Kofler)

Mass Spectrum (ES⁺): [M + H]⁺ = 440

Retention time (min): 3.17

Example 36: 3- {4- [3- (3-trifluoromethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0140]

<img file="PL1877409T3_D0051.tif" />

[0141] 77 mg of a yellow solid 3- {4- [3- (3-trifluoromethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in Example 1 starting from 3- (420 aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 3-trifluoromethyl-phenylisocyanate.

Melting point: 256 ° C (Buchi B-545)

Mass Spectrum (ES⁺): [M + H]⁺ = 440 Retention time (min): 3.48

Example 37: 3- {4- [3- (4-fluoro-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0142]

<img file="PL1877409T3_D0052.tif" />

73 mg of a yellow solid 3- {4- [3- (4-fluoro-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in Example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3c] pyridine-2-carboxamide and 4-fluoro-phenylisocyanate.

Melting point: 271 ° C (Buchi B-545)

Mass Spectrum (ES⁺): [M + H]⁺ = 390

Retention time (min): 2.93

Example 38: 3- {4- [3- (4-trifluoromethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0144]

<img file="PL1877409T3_D0053.tif" />

[0145] 91 mg of a yellow solid 3- {4- [3- (4-trifluoromethylphenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in Example 1 starting from 3- (4aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 4-trifluoromethyl-phenylisocyanate.

Melting point: 289 ° C

Mass Spectrum (ES⁺): [M + H]⁺ = 440

Retention time (min): 3.48

Example 39: 3- [4- (3-p-tolyl-ureido) -phenyl] -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0146]

<img file="PL1877409T3_D0054.tif" />

ρ-Ο "

76 mg of a yellow solid 3- [4- (3-p-tolyl-ureido) -phenyl] 1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in Example 1 starting from 3 - (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and p-tolylisocyanate.

Melting point: 277 ° C (Buchi B-545)

Mass Spectrum (ES⁺): [M + H]⁺ = 386

Retention time (min): 3.13

Example 40: 3- {4- [3- (4-chloro-3-trifluoromethyl-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0148]

cl

<img file="PL1877409T3_D0055.tif" />

[0149] 103 mg of yellow solid 3- {4- [3- (4-chloro-3-trifluoromethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine -2-carboxamide is prepared as described in Example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 4-chloro-3-trifluoromethyl-phenylisocyanate.

Melting point: 228 ° C (Buchi B-545)

Mass Spectrum (ES⁺): [M + H]⁺ = 474

Retention time (min): 3.64

Example 41: 3- {4- [3- (2-Chloro-5-trifluoromethyl-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0150]

<img file="PL1877409T3_D0056.tif" />

76 mg yellow solid 3- {4- [3- (2-Chloro-5-trifluoromethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in Example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 2-chloro-5-trifluoromethyl-phenylisocyanate.

Melting point: 243 ° C (Buchi B-545)

Mass Spectrum (ES⁺): [M + H]⁺ = 474

Retention time (min): 3.56

Example 42: 3- {4- [3- (4-trifluoromethoxy-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide

<img file="PL1877409T3_D0057.tif" />

[0153] 94 mg of a yellow solid 3- {4- [3- (4-trifluoromethoxyphenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in Example 1 starting from 3- (4aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 4-trifluoromethoxy-phenylisocyanate.

Melting point: 276 ° C (Buchi B-545)

Mass Spectrum (ES⁺): [M + H]⁺ = 456

Retention time (min): 3.63

Example 43: 3- {4- [3- (4-difluoromethoxy-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0154]

<img file="PL1877409T3_D0058.tif" />

87 mg of a yellow solid 3- {4- [3- (4-difluoromethoxyphenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in Example 1 starting from 3- (4aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 4-difluoromethoxy-phenylisocyanate.

Melting point: 257 ° C

Mass Spectrum (ES⁺): [M + H]⁺ =4,38

Retention time (min): 3.23

Example 44: 3- {4- [3- (3,4-dimethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0156]

<img file="PL1877409T3_D0059.tif" />

[0157] mg of a yellow solid 3- {4- [3- (3,4-dimethyl-phenyl) ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in Example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 3,4-dimethyl-phenylisocyanate. Melting point: 230 ° C (Buchi B-545)

Mass Spectrum (ES⁺): [M + H]⁺ = 400 5 Retention time (min): 3.32

Example 45: 3- {4- [3- (3,5-dimethoxy-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0158]

<img file="PL1877409T3_D0060.tif" />

87 mg of a yellow solid 3- {4- [3- (3,5-dimethoxy-phenyl) ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 3,5-dimethoxyphenylisocyanate.

Melting point: 225 ° C (Buchi B-545)

Mass Spectrum (ES⁺): [M + H]⁺ = 432

Retention time (min): 3.07

Example 46: 3- {4- [3- (2,5-dimethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0160]

<img file="PL1877409T3_D0061.tif" />

87 mg of a yellow solid 3- {4- [3- (2,5-dimethyl-phenyl) ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 2,5-dimethylphenylisocyanate.

Melting point: 261 ° C (Buchi B-545)

Mass Spectrum (ES⁺): [M + H]⁺ = 400

Retention time (min): 3.25

Example 47: 3- {4- [3- (2-fluoro-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0162]

<img file="PL1877409T3_D0062.tif" />

[0163] 59 mg of a pale yellow solid 3- {4- [3- (2-fluoro-phenyl) ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in Example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 2-fluoro-phenylisocyanate. Melting point: 242 ° C (Buchi B-545)

Mass Spectrum (ES⁺): [M + H]⁺ = 390

Retention time (min): 2.41

Example 48: 3- {4- [3- (3-fluoro-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0164]

<img file="PL1877409T3_D0063.tif" />

[0165] mg of a light yellow solid 3- {4- [3- (3-fluoro-phenyl) ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in Example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 3-fluoro-phenylisocyanate. Melting point: 252 ° C (Buchi B-545)

Mass spectrum (ES +): [M + H] + = 390 Retention time (min): 2.55

Example 49: 3- {4- [3- (2-fluoro-3-trifluoromethyl-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0166]

<img file="PL1877409T3_D0064.tif" />

[0167] 69 mg of a light yellow solid 3- {4- [3- (2-fluoro-3-trifluoromethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in Example 1 starting from 315 (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 2-fluoro-3-trifluoromethyl-phenylisocyanate.

Melting point: 240 ° C (Buchi B-545)

Mass Spectrum (ES⁺): [M + H]⁺ = 458 Retention time (min): 2.75

Example 50: 3- {4- [3- (3-fluoro-5-trifluoromethyl-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0168]

<img file="PL1877409T3_D0065.tif" />

[0169] 69 mg of a light yellow solid 3- {4- [3- (3-fluoro-5-trifluoromethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in Example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 3-fluoro-5-trifluoromethyl-phenylisocyanate.

Melting point: 261 ° C (Buchi B-545)

Mass Spectrum (ES⁺): [M + H]⁺ = 458

Retention time (min): 2.88

Example 51: 3- {4- [3- (4-fluoro-3-trifluoromethyl-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0170]

F

<img file="PL1877409T3_D0066.tif" />

56 mg of an yellow yellow solid 3- {4- [3- (4-fluoro-3-trifluoromethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 4-fluoro-3-trifluoromethyl-phenylisocyanate.

Melting point: 201 ° C (Buchi B-545)

Mass Spectrum (ES⁺): [M + H]⁺ = 458

Retention time (min): 2.85

Example 52: 3- {4- [3- (4-methyl-3-trifluoromethyl-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0172]

<img file="PL1877409T3_D0067.tif" />

61 mg of a pale yellow 3- 3- 4- 4- [3- (4-methyl-3-trifluoromethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide solid is prepared as described in Example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 4-methyl-3-trifluoromethyl-phenylisocyanate.

Melting point: 199 ° C

Mass Spectrum (ES⁺): [M + H]⁺ = 454

Retention time (min): 2.84

Example 53: 3- {4- [3- (3-methoxy-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide trifluoroacetate

<img file="PL1877409T3_D0068.tif" />

[0175] 33.3 mg of 3- {4- [3- (3-methoxy-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide yellow lyophilisate of trifluoroacetate trifluoroacetate is prepared as described in Example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 3-methoxyphenylisocyanate.

Mass Spectrum (ES⁺): [M + H]⁺ = 402

Retention time (min): 2.60

Example 54: 3- {4- [3- (3,4-Dimethoxy-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide trifluoroacetate [0176]

<img file="PL1877409T3_D0069.tif" />

[0177] 80.5 mg of yellow 3- {4- [3- (3,4-dimethoxy-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide trifluoroacetate lyophilisate is prepared as described in Example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 3,4-dimethoxy-phenylisocyanate.

Mass Spectrum (ES⁺): [M + H]⁺ = 432

Retention time (min): 2.27

Example 55: 3- {4- [3- (2,5-Dimethoxy-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide trifluoroacetate

<img file="PL1877409T3_D0070.tif" />

[0179] 90.7 mg of yellow 3- {4- [3- (2,5-dimethoxy-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide trifluoroacetate lyophilisate is prepared as described in Example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 2,5-dimethoxy-phenylisocyanate.

Mass Spectrum (ES⁺): [M + H]⁺= 432

Retention time (min): 2.62

Example 56: 3- [4- (3-o-Tolyl-ureido) -phenyl] -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0180]

<img file="PL1877409T3_D0071.tif" />

75.3 mg of a yellow solid 3- [4- (3-o-Tolyl-ureido) phenyl] -1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in Example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and o-Tolylisocyanate.

Melting point: 270 ° C (Buchi B-545)

Mass Spectrum (ES⁺): [M + H]⁺ = 386

Retention time (min): 2.54

Example 57: 3- {4- [3- (4-methoxy-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0182]

<img file="PL1877409T3_D0072.tif" />

51.1 mg of a light yellow solid 3- {4- [3- (4-methoxy-phenyl) ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 4-methoxy-phenylisocyanate. Melting point: 275 ° C (Buchi B-545)

Mass spectrum (ES +): [M + H] + = 402

Retention time (min): 2.28

Example 58: 3- {4- [3- (3-chloro-4-difluoromethoxy-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0184]

cl

<img file="PL1877409T3_D0073.tif" />

[0185] 93 mg of a light yellow solid 3- {4- [3- (3-chloro-4-difluoromethoxy-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in Example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 3-chloro-4-difluoromethoxy-phenylisocyanate.

Melting point: 267 ° C (Buchi B-545)

Mass Spectrum (ES⁺): [M + H]⁺= 472

Retention time (min): 2.90

Example 59: 3- {4- [3- (3,5-dimethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0186]

<img file="PL1877409T3_D0074.tif" />

[0187] 61 mg of pale yellow solid 3- {4- [3- (3,5-dimethylphenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide is prepared as described in example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3-c] pyridine-2-carboxamide and 3,5-dimethylphenylisocyanate.

Melting point: 188 ° C (Buchi B-545)

Mass Spectrum (ES⁺): [M + H]⁺ = 400

Retention time (min): 2.68

Example 60: 3- {4- [3- (3-ethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide [0188]

<img file="PL1877409T3_D0075.tif" />

[0189] 61 mg of a light yellow solid 3- {4- [3- (3-ethyl-phenyl) ureido] -phenyl} -1H-pyrrolo [2,3-c] pyridine-2-carboxamide was prepared as described in Example 1 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3c] pyridine-2-carboxamide and 3-ethyl-phenylisocyanate.

Melting point: 257 ° C (Buchi B-545)

Mass Spectrum (ES⁺): [M + H]⁺ = 400

Retention time (min): 2.97

Example 61: 3- {4- [3- (3-ethyl-phenyl) -ureido] -phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide [0190]

<img file="PL1877409T3_D0076.tif" />

[0191] 0.8 mg of a white solid 3- {4- [3- (3-ethyl-phenyl) -ureido] phenyl} -1H-pyrrolo [2,3-b] pyridine-2-carboxamide is prepared as described in example 7 starting from 3- (4-aminophenyl) -1H-pyrrolo [2,3b] pyridine-2-carboxamide and 3-methyl-phenylisocyanate.

Melting point: 254 ° C (Buchi)

Mass Spectrum (ES⁺): [M + H]⁺= 400

Retention time (min): 7.18

Determination of compound activity - Experimental protocols

1. FAK [0192] Inhibitory activity of compounds against FAK is determined by measuring the inhibition of enzyme autophosphorylation using a time-separated fluorescence assay (HTRF).

[0193] Complete human FAK DNA, whose N-terminal end was tagged with histidine, was cloned into the baculovirus expression vector pFastBac HTc. The protein was expressed and purified to approximately 70% homogeneity.

[0194] Kinase activity is determined by incubating the enzyme (6.6 pg / ml) at various concentrations of test compound in 50 mM Hepes buffer pH = 7.2, 10 mM MgCl₂, 100 μΜ Na₃VO₄ , 15 μΜ ATP for 1 hour at 37 ° C. The enzyme reaction was stopped by adding Hepes buffer pH = 7.0 containing 0.4 mM KF, 133 mM EDTA, 0.1% BSA and labeling was done for 1 to 2 hours at ambient temperature by adding anti-mystidine antibody in this buffer labeled XL665 and phosphospecific monoclonal antibody for europium cryptate conjugated tyrosine (Eu-K). The characteristics of the two fluorophores are available in: G. Mathis et al., Anticancer Research, 1997, 17, pp. 3011-3014. The energy transfer between the excited europium cryptane to the XL665 acceptor is proportional to the degree of FAK autophosphorylation. The long-lasting signal specific to XL-665 is measured on a Packard Discovery plate counter. All tests are carried out in duplicate and the average of two tests is calculated. Inhibition of FAK autophosphorylation activity with the compounds of the invention is expressed as a percentage of inhibition relative to the control whose activity is measured in the absence of test compound. For the calculation of% inhibition, the ratio [signal at 665 nm / signal at 620 nm] is considered.

2. KDR [0195] The inhibitory effect of the compounds is determined in a substrate phosphorylation assay by the KDR enzyme in vitro by scintillation (96 well plate, NEN).

[0196] The cytoplasmic domain of the human KDR enzyme was cloned in the form of a GST fusion in the pFastBac baculovirus expression vector. The protein was expressed in SF21 cells and purified to approximately 60% homogeneity.

[0197] KDR kinase activity was measured in 20 mM MOPS, 10 mM MgCl2, 10 mM MnCl2, 1 mM DTT, 2.5 mM EGTA, 10 mM b-glycerophosphate, pH = 7.2, in the presence of 10 mM MgCl₂, 100 μΜ Na₃VO₄, 1 mM NaF. 10 µl of compound is added to 70 pL kinase buffer containing 100 ng KDR enzyme at 4 ° C. The reaction was started by adding 20 pL of solution containing 2 pg of substrate (the SH2-SH3 PLOT fragment was expressed as a GST fusion protein), 2 pCi □³³P [ATP] and 2 pM cold ATP. After 1 hour incubation at 37 ° C, the reaction was stopped by adding 1 volume (100 µL) of 200 mM EDTA. The incubation buffer is withdrawn and the wells washed three times with 300 pL PBS. Radioactivity was measured in each well using a Top Count NXT radioactivity counter (Packard).

[0198] Radiation background is determined by measuring radioactivity in four different wells containing radioactive ATP and the substrate alone.

[0199] The total activity control test is measured in four different wells containing all reagents (y³³P- [ATP], KDR and substrate PLCs) but in the absence of compound.

[0200] Inhibition of KDR activity with a compound of the invention is expressed as a percentage of inhibition of control activity determined in the absence of the compound.

[0201] Compound SU5614 (Calbiochem) (1 pM) is included in each plate as a control inhibition.

3. Tie2 [0202] The sequence coding for human Tie2 corresponding to the amino acids of the intracellular domain 776-1124 was generated by PCR using cDNA extracted from human placenta as a model. This sequence was introduced into the pFastBacGT baculovirus expression vector as a GST fusion protein.

[0203] The inhibitory effect of the molecules is determined in a Tie2 phosphorylation PLC test in the presence of GST-Tie2 purified to about 80% homogeneity. The substrate is composed of SH2-SH3 PLC fragments expressed as a GST fusion protein.

[0204] Tie2 kinase activity was measured in MOPS buffer 20mM pH 7.2 containing 10 mM MgCl2, 10 mM MnCl2, 1 mM DTT, 10 mM glycerophosphate. On a 96-well FlashPlate plate kept on ice, a reaction mixture consisting of 70 pL kinase buffer containing 100 ng of GST-Tie2 enzyme per well is deposited. Finally 10 μL of the test molecule dissolved in DMSO with a maximum concentration of 10% is added. At a given concentration, each measurement was performed four times. The reaction is initiated by adding 20 μL of solution containing 2 μg GSTPLC, 2 μΜ cold ATP and 1 pCi ³³P [ATP]. After 1 hour incubation at 37 ° C, the reaction was stopped by adding 1 volume (100 µl) EDTA (200 mM). After removing the incubation buffer, the wells are washed three times with 300 pL PBS. Radioactivity was measured on MicroBeta1450 Wallac.

[0205] Inhibition of Tie2 activity is calculated and expressed as a percentage of inhibition relative to control activity determined in the absence of the compound.

4. Aurora1 and Aurora2 [0206] The inhibitory effect of compounds on the Aurora1 and Aurora2 kinases is determined by enzymatic assay using radioactivity detection.

[0207] Aurora 1 and Aurora 2 kinase activity is assessed by phosphorylation of a Numa-histidine substrate in the presence of radiolabeled ATP ([³³P] ATP) using 96-well Flashplate plates where chelate-nickel is bonded to the surface of the microplate. The amount of phosphate³³The P incorporated into the NuMA substrate is proportional to the activity of the Aurora1 or Aurora2 enzyme.

protein:

[0208] Proteins are produced in a Sanofi-Aventis group protein production laboratory.

[0209] Aurora 1: Aurora-B / INCENP-C3 recombinant complex, purified to about 50%, whose Aurora-B N-terminal end is marked with histidine.

[0210] Aurora 2: the whole recombinant protein including the histidine terminus at the N-terminus, expressed in E.coli and purified to over 82%.

[0211] NuMA (ring protein that associates with the mitotic apparatus): a fragment of 424 amino acids, expressed in E.coli, whose N-terminal end is marked with histidine and used as a substrate for two Aurora enzymes.

Protocol:

[0212] The microwells used are 96-well Flash-Plate, nickel chelate (Perkin Elmer, model SMP107).

[0213] Test products are incubated in a reaction volume of 100 pL per well in the presence of 10 nM Aurora 1 or Aurora 2, 500 nM substrate

NuMA in a buffer composed of 50 mM Tris / HCl (pH 7.5), 50 mM NaCl, 5 mM

MgCl2 (Aurora-B) or 10 mM MgCl2 (Aurora-A) and 1 mM DTT, at 37 ° C.

[0214] To each well, 80 μL enzyme / substrate incubation buffer is distributed, followed by 10 μL of test product at varying concentrations. The reaction is initiated by the addition of 1 μΜ final ATP containing 0.2 μθί [³³P] ATP (10 μΙ). After 30 minutes incubation, the reaction is stopped by simply removing the reaction buffer and each well is washed twice with 300 μl Tris / HCl buffer. Radioactivity is therefore measured in each well using a scintillation apparatus, model Packard, Top count.

[0215] The enzymatic activity of the Aurora control is expressed by the number of counts per minute obtained within 30 minutes after subtracting the background radiation (reaction mixture not containing the enzyme). The evaluation of the various products tested is expressed as a percentage of inhibition of Aurora activity relative to the control.

5. CDK2 / cyclin E:

Purification of the CDK2 / CyclinE- (His) complex₆ by IMAC (Immobilized Metal Affinity Chromatography):

[0216] Two recombinant baculoviruses having human sequences encoding CDK2 and cyclin E, respectively (the latter including the C-terminal hexa-histidine tag) are used to co-infect Sf21 insect cells. Two to three days after the beginning of co-infection, the cells are harvested by centrifugation and then preserved at -40 ° C until they are used.

After thawing and mechanical lysis of the cells, the complex present in the lysis supernatant is purified by nickel affinity chromatography (IMAC), and preserved at -80 ° C.

Flashplate CDK2 / Cyklina trial in 96-well format.

[0217] The format of 96 well plates coated with streptavidin is used to test compound activity for CDK2 / Cyclin E kinase activity.

[0218] To perform this assay, the biotinylated peptide substrate, pRb protein fragment, (biotinyl-SACPLNLPLQNNHTAADMYLSPVRSPKKKGSTTROH) is dissolved at a concentration of 1 mM in kinase buffer (HEPES / NaOH 50 mM, NaCl 1 mM, pH 7.5 m) to form a mother liquor preserved at -20 ° C as a 110 μΙ part of the sample. On the day of the test, a sample of this solution is thawed and diluted in kinase buffer containing 1 mM dithiothreitol, added to the buffer immediately before use to obtain a 14.3 μ stężenie concentration. 70 pL of this solution is added to each Flashplate well to obtain a final substrate concentration of 10 μΜ during the enzymatic reaction carried out in a final volume of 100 pL reaction medium (see below).

[0219] Intermediate dilutions of inhibitors (products of the invention) to various concentrations are prepared in DMSO starting from 10 mM mother liquors in separate tubes. Therefore, dilutions of 1000 μΜ, 333.3 μΜ, 111.1 μΜ, 37.03 μΜ, 12.35 μΜ, 4.11 μΜ and 1.37 μΜ are carried out. pL of each of these solutions (or 1 pL DMSO for the control) are transferred to the wells of the test plate.

[0220] To each well, 19 μΐ of a solution of the adenosine triphosphate (ATP) and ATPy mixture is finally added.³³P in kinase buffer at 5.26 μ 5, total ATP and 52.6 pCi / ml ³³P. The enzyme reaction was started by adding 10 pL to a well of a CDK2 / cyclin E solution (200 ΜΜ) in kinase buffer containing 1 mM dithiothreitol (or 10 pL kinase buffer containing 1 mM dithiothreitol for blind reactions).

[0221] After addition of each reagent, the final volume of each well is 100pL, the final concentration of the substrate is 10 μΜ, the final concentrations of inhibitors are 10 μΜ, 3.33 μΜ, 1.11 μΜ, 0.37 μΜ, 0.123 μΜ, 0.041 μΜ and 0.014 μΜ (according to intermediate dilution concentration), the final ATP concentration is 1 μΜ, the final amount ³³P is 1 pCi / well, final concentration of the CDK2 / cyclin E complex is 20 πΜ.

[0222] After all reagents have been added, the test plate is incubated at 30 ° C with orbital mixing at 650 rpm.

[0223] When the incubation is complete, the plate is washed three times with 300 pL per PBS well (Buffered Saline Phosphate, pH = 7.4 without calcium or magnesium, reference number 10010-015, Gibco BRL). Inclusion³³P to peptide is quantified by scintillation counting with a Packard Topcount.NXT instrument. The inhibitory activity of the products of the invention is assessed by measuring the concentration of the inhibitor allowing a reduction of the enzymatic activity by 50% (CI50).

Results:

[0224]

Table 1:

<td rowspan="2">Example</td><td colspan="6">IC 50 (nM)</td>

<td>FAK</td><td>KDR</td><td>TIE2</td><td>Aurora A.</td><td>Aurora B.</td><td>CDK2</td>

<td> 1</td><td> 164</td><td> 29</td><td> 4</td><td> 172</td><td> 138</td><td></td>

<td rowspan="2">Example</td><td></td><td></td><td></td><td>IC 50 (nM)</td><td></td><td></td>

<td>FAK</td><td>KDR</td><td>TIE2</td><td>Aurora A.</td><td>Aurora B.</td><td>CDK2</td>

<td> 2</td><td> 299</td><td> 150</td><td> 21</td><td> 222</td><td> 196</td><td></td>

<td> 5</td><td> 249</td><td> 258</td><td> 47</td><td> 131</td><td> 67</td><td></td>

<td> 7</td><td> 184</td><td> 34</td><td> 9</td><td> 553</td><td> 133</td><td></td>

V3048PL00 / KK

EP 1 877 409