Classifications

• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D417/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00
  • C07D417/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings
  • C07D417/04—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings directly linked by a ring-member-to-ring-member bond
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P35/00—Antineoplastic agents
  • A61P35/04—Antineoplastic agents specific for metastasis
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D277/00—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings
  • C07D277/02—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings not condensed with other rings
  • C07D277/20—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings not condensed with other rings having two or three double bonds between ring members or between ring members and non-ring members
  • C07D277/22—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings not condensed with other rings having two or three double bonds between ring members or between ring members and non-ring members with only hydrogen atoms, hydrocarbon or substituted hydrocarbon radicals, directly attached to ring carbon atoms
  • C07D277/28—Radicals substituted by nitrogen atoms
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D277/00—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings
  • C07D277/02—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings not condensed with other rings
  • C07D277/20—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings not condensed with other rings having two or three double bonds between ring members or between ring members and non-ring members
  • C07D277/32—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings not condensed with other rings having two or three double bonds between ring members or between ring members and non-ring members with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
  • C07D277/38—Nitrogen atoms
  • C07D277/42—Amino or imino radicals substituted by hydrocarbon or substituted hydrocarbon radicals
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D417/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00
  • C07D417/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings
  • C07D417/10—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings linked by a carbon chain containing aromatic rings
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D417/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00
  • C07D417/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings
  • C07D417/12—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings linked by a chain containing hetero atoms as chain links
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D417/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00
  • C07D417/14—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing three or more hetero rings

Definitions

• the invention relates to a novel compound having Tie2 and KDR inhibiting activity, to a method of inhibiting the activity of one or more protein kinases by using the compound, to a pharmaceutical composition comprising such a compound, to the use of the compound as a medicament particular as an inhibitor of protein kinase activity most preferably as an protein kinase inhibitor of both Tie-2 and KDR, accordingly an eminent use for the inhibition of the progression of a disease state in a patient, preferably cancer, venous malformations and angiogenesis dependent disorders .
• Angiogenesis is a multistep process for the formation of new blood vessels from existing vasculature that normally occurs only during embryonic development, breast lactation, endometrail regulation and wound repair.
• endothelial cells release enzymes that degrade the basement membrane, migrate through the membrane to form a sprout, and proliferate to extend the vessel (for review see Carmeliet and Jain, 2000) .
• a number of growth factors are involved in vascular development (reviewed in Yancopoulos et al . , 2000) . They include at least five members of the vascular endothelium growth factor (VEGF) family, at least four members of the angiopoietin (Ang) family, and at least one member of the large ephrin family. To form functional vessels all of these factors have to act in a coordinated manner. VEGF can initiate vessel formation in adult animals, and Ang-1 further stabilizes and protects the adult vasculature.
• VEGF vascular endothelium growth factor
• Ang-1 angiopoietin
• RTK receptor tyrosine kinase
• VEGFR-1 The receptors that respond to VEGFs form a family of three closely related RTKs termed VEGFR-1 (Flt-1) , VEGFR-2 (KDR or Flk-1) and VEGFR-3 (Flt-4) (reviewed in Tallquist et al . , 1999) .
• Their extracellular portion all contain seven immunglobulin-like (Ig) domains and a split intracellular kinase domain. While the major growth and permeability actions of VEGF are mediated by VEGFR-2, growth factor signalling is suppressed by VEGFR-1 because it probably acts as a decoy receptor.
• VEGFR-2 die between day 8.5 and 9.5 during embryogenesis due to very few enothelial cells and failure to develop a vasculature. Mice lacking VEGFR-1 form excess endothelial cells and disorganized blood vessels also die between E8.5 and E9.5.
• VEGFR-3 knockout embryos show a cardiovascular failure between E10 and E12 from defects in remodeling the primary vessel networks into larger blood vessels. VEGFR-3 seems to play a role in lymphangiogenesis since its expression is critical for lymphatic vessels (Valtola et al . , 1999).
• Another group of angiogenic receptors is formed by the two closely related RTKs, Tie-1 (Partanen et al . , 1992) and Tie-2 (Ziegler et al . , 1993). These are proteins of approximately 125 kD with a single putative transmembrane region.
• the extracellular domain contains at least three epidermal growth factor (EGF) -like regions of cystein expression, at least two immunglobulin G (IgG) -like domains and at least three regions with fibronectin Ill-like repeats.
• EGF epidermal growth factor
• IgG immunglobulin G
• Tie-2 contains a tyrosine kinase domain with about 40 % sequence identity to that of FGFR-1, PDGFR and c-Kit with the typical motifs for ATP binding (GXGXXG) and tyrosine phosphorylation (HRDLAAR and DFGL) .
• the Tie receptors are specifically expressed in developing vascular endothelial cells. Embryos deficient in Tie-1 fail to establish structural integrity of vascular endothelial cells, resulting in oedema and subsequently localized haemorrhage. However, analyses of embryos deficient in Tie-2 showed that it is important in angiogenesis, particularly for vascular network formation in endothelial cells, indicating that the structurally related receptor tyrosine kinases Tie-1 and Tie-2 have important but distinct roles in the formation of blood vessels (Dumont et al . , 1994; Korhonen et al . , 1994; Puri et al., 1995; Sato et al. , 1995) .
• Angiopoietin-1 (Ang-1) binds and induces the tyrosine phosphorylation of Tie-2, it does not directly promote the growth of cultured endothelial cells but is essential for normal vascular development in the mouse (Davis et al . , 1996) .
• Mice engineered to lack Angiopoietin-1 display angiogenic deficits reminiscent of those previously seen in mice lacking Tie-2, demonstrating that Angiopoietin-1 is a primary physiologic ligand for Tie-2 and that it has critical in vivo angiogenic actions that are distinct from VEGF (Suri et al . , 1996).
• Ang-1 Transgenic overexpression of Ang-1 in the skin of mice produces larger, more numerous, and more highly branched vessels (Suri et al . , 1998) . This finding supports a more direct role of Ang-1 in angiogenesis and vascular remodelling.
• Angiopoietin-2 (Ang-2) was identified by homology screening and showed to be a naturally occurring antagonist for Ang-1 and Tie-2. Therefore, transgenic overexpression of Ang-2 disrupts blood vessel formation in the mouse embryo. In adult mice and humans, Ang-2 is expressed only at sites of vascular remodeling (Maisonpierre et al . , 1997).
• mice embryos knocked out for VEGFR-2 show lethal defects in vasculogenesis that are earlier than a corresponding disruption of Tie-2.
• angiopoietins can be used, alone or in combination with VEGF, to promote therapeutic angiogenesis.
• blocking or moderating of the Tie receptor system may block or moderate angiogenesis and further proliferation of tumor cells.
• a Tie-specific amplified cDNA band was obtained by RT- PCR from melanoma metastases but not from normal skin. These results suggest a role for the Tie receptor system in angiogenesis associated with melanoma metastases (Kaipainen et al. , 1994) .
• Ad-ExTek a soluble adenoviral expressed extracellular domain of Tie-2
• the inhibition of Tie-2 function by ExTek may be a consequence of sequestration of the angiopoietin ligand and/or heterodimerisation with the native Tie-2 receptor. This study demonstrates that disruption of Tie-2 signalling pathways, first, may be well tolerated in healthy organisms and, second, may provide therapeutic benefit.
• V is H or
• R x can be independently H, alkyl, alkenyl, cycloalkyl, heteroalkyl, aryl, heteroaryl, arylalkyl, alkylaryl, N-R 6 R 7 , N-(CO)R 6 R 7 , N-R 6 (C0)R 7 or N- (CO) -0-R s R 7 ,
• R 8 can be independently H, alkyl, alkenyl, cycloalkyl, heteroalkyl, aryl, heteroaryl, arylalkyl, alkylaryl, N-R 3 R 4 , N-(CO)R 3 R 4 , N-R 3 (CO)R 4 , N- (CO) -0-R 3 R 4 , 0-R 3 , CO-R 3 , CO-OR 3 or 0-CO-R 3 ,
• R 2 , R 5 can be independently H, alkyl, alkenyl, cycloalkyl, heteroalkyl, aryl, heteroaryl, arylalkyl, alkylaryl, carboxyl, Br, Cl, F, CF 3 ,
• R 3 , R 4 , R 6/ R 7 can be independently H, alkyl, alkenyl , cycloalkyl , heteroalkyl , aryl , heteroaryl , arylalkyl, alkylaryl, COOR 5 and CO-R 5 , and may form a ring structure,
• X, Y, Z can be independently CH or N, and
• U can be independently S or NH
• W can be independently NH, O or S, and racemic-diastereomeric mixtures, optical isomers, and pharmaceutically acceptable salts thereof.
• the invention relates to a compound according to formula II,
• R x is N-R 6 R 7 ,
• R 6 is H
• R 7 is a substituted or unsubstituted alkyl, cycloalkyl, phenyl, arylalkyl, naphtyl, heteroaryl or heterocycloalkyl.
• X and Y are CH or N
• R 8 is an amine group or a mono-substituted or di-substituted alkylamine, alkylene-amine or cycloalkylamine or heterocycloalkylamine, which (cyclo) alkylamine may be substituted with an alkyl, cycloalkyl, hydroxyl, halogen, pyridinyl or alkylpyridinyl group.
• R 2 is hydrogen, heteroaryl, aralkyl or carboxylgroup
• R 5 is hydrogen, hydroxyl, halogen or alkoxy group.
• R 7 is: - phenyl C 1 -C 3 carboxylic acid, preferably para-carboxylic acid;
• R 8 is:
• alkylamine preferably di-ethyl; - hydroxypyrrolidinyl ;
• R 2 is a carboxyl group or a tetrazole group.
• the present invention provides a method of inhibiting or moderating the kinase activity of tyrosine kinases comprising the administration of a compound represented by formula (I) or (II) to said kinase in sufficient concentration to inhibit or moderate the enzyme activity of said kinase .
• the present invention further provides the use of compounds in pharmaceutical compositions with a pharmaceutically acceptable carrier or excipient.
• These pharmaceutical compositions can be administered to individuals to slow or halt the process of angiogenesis in angiogenesis-aided diseases or cancer in general.
• any heteroatom with unsatisfied valances is assumed to have the hydrogen-atom to satisfy the valances.
• alkyl or “alk” refers to a monovalent alkane (hydrocarbon) derived radical containing from 1 to 20 carbon atoms unless otherwise defined.
• An alkyl group is an optionally substituted straight, branched or cyclic saturated hydrocarbon group. When substituted, alkyl groups may substituted with R at any available point of attachment. R is defined as Ri . When the alkyl group is said to be substituted with alkyl group this is used interchangeably with “branched alkyl group” .
• Exemplary unsubstitute such groups may include but are not limited to methyl, ethyl, propyl, isopropyl, n-butyl, -butyl, isobutyl, pentyl, hexyl, isohexyl, heptyl, 4 , 4-dimethylpentyl, octyl, 2,2,4- trimethylpentyl , nonyl, decyl, undecyl, dodecyl, and the like.
• substituents may include but are not limited to one or more of the following groups: halogen (such as F, Cl, Br, I), haloalkyl (such as CC1 3 and CF 3 ) , alkoxy, alkylthio, hydroxy, carboxy (-COOH) , alkyloxycarbonyl (- C(O)R), alkylcarbonyloxy (-OCOR), amino (-NH 2 ), alkylamino, dialkylamino, carbamoyl (-NHCOOR- or - OCONHR-) , urea (- NHCONHR) or thiol (-SH) .
• R is defined as R 6 .
• Alkyl groups as defined may also comprise one or more carbon to carbon double bonds or one or more carbon to carbon triple bonds .
• alkenyl refers to a hydrocarbon radical straight, branched or cyclic containing from 2 to 20 carbon atoms and at least one carbon to carbon double bond.
• alkynyl refers to a hydrocarbon radical straight, branched or cyclic containing from 2 to 20 carbon atoms and at least one carbon to carbon triple bond.
• Cycloalkyl is a specie of alkyl containing from 3 to 15 carbon atoms, without alterning or resonating double bonds between carbon atoms. It may contain from 1 to 4 rings. Exemplary unsubstituted such groups include cyclopropyl, cyclobutyl, cyclopentyl, cydohexyl, adamantyl, etc. Exemplary substituents may include but are not limited to one or more of the following groups: halogen, alkyl, alkoxy, alkyl hydroxy, amino, alkylamino, dialkylamino, nitro, cyano, thiol and/or alkylthio.
• Cycloalkenyl is a specie of alkenyl containing from 3 to 15 carbon atoms, without alterning or resonating double bonds between carbon atoms and at least one carbon to carbon double bond. It may contain from 1 to 4 rings. Exemplary unsubstituted such groups include cyclopropyl, cyclobutyl, cyclopentyl, cydohexyl, adamantyl, etc. Exemplary substituents may include but are not limited to one or more of the following groups: halogen, alkyl, alkoxy, alkyl hydroxy, amino, alkylamino, dialkylamino, nitro, cyano, thiol and/or alkylthio.
• Cycloalkynyl is a specie of alkyl containing from 3 to 15 carbon atoms, without alterning or resonating double bonds between carbon atoms and at least one carbon to carbon triple bond. It may contain from 1 to 4 rings. Exemplary unsubstituted such groups include cyclopropyl, cyclobutyl, cyclopentyl, cy ohexyl, adamantyl, etc. Exemplary substituents may include but are not limited to one or more of the following groups: halogen, alkyl, alkoxy, alkyl hydroxy, amino, alkylamino, dialkylamino, nitro, cyano, thiol and/or alkylthio.
• heterocycloalkyl refers to a cycloalkyl group (nonaromatic) in which one to three of the carbon atoms in the ring are replaced by a heteroatom selected from 0, S or N.
• heterocycloalkenyl refers to a cycloalkenyl group (nonaromatic) in which one to three of the carbon atoms in the ring are replaced by a heteroatom selected from 0, S or N.
• heterocycloalkynyl refers to a cycloalkynyl group (nonaromatic) in which one to three of the carbon atoms in the ring are replaced by a heteroatom selected from 0, S or N.
• aryl refers to monocyclic, bicyclic, tricyclic or tetracyclic aromatic rings, e.g. phenyl, substituted phenyl and the like, as well as groups which are fused, e.g. naphtyl, substituted naphtyl, phenanthrenyl or substituted phenanthrenyl and the like.
• An aryl group thus contains at least one ring having at least 6 atoms, with up to five such rings being present, containing up to 22 atoms therein, with alternating (resonating) double bonds between adjacent carbon atoms or suitable heteroatoms.
• heteroaryl refers to a monocyclic aromatic hydrocarbon group having 5 or 6 ring atoms, a bicyclic aromatic group having 8 to 10 atoms, or a tricyclic aromatic group having 11 to 14 atoms containing at least one heteroatom, 0, S, or N, in which a carbon or nitrogen atom is the point of attachment, and in which one to three additional carbon atoms is optionally replaced by a heteroatom selected from O, N, or S, said heteroaryl group being optionally substituted as described herein.
• heteroaryl groups may include but are not limited to the following: thienyl, furyl, pyrrolyl, pyridinyl, imidazolyl, oxazolyl, pyrrolidinyl , piperidinyl , thiazolyl, pyrazinyl, pyridazinyl, pyrimidinal, triazinylazepinyl , indolyl, isoindolyl, quinolinyl, isoquinolinyl , benzothiazolyl , benzoxazolyl, benzimidazolyl , benzoxadiazolyl, benzofuranzanyl tetrahydropyranyl and the like.
• arylalkyl denotes an aromatic ring bonded to an alkyl group as described above.
• alkylaryl denotes an alkyl group bonded to an aromatic ring as described above.
• alkoxy or “alkylthio”, as used herein, denote an alkyl group as described above bonded through an oxygen linkage (-0-) or a sulfur linkage (-S-) , respectively.
• alkenyloxy or "alkenylthio” , as used herein, denote an alkenyl group as described above bonded through an oxygen linkage (-0-) or a sulfur linkage (-S-), respectively.
• alkynyloxy or alkynylthio denote an alkynyl group as described above bonded through an oxygen linkage (-0-) or a sulfur linkage (-S-), respectively.
• alkynyloxy or alkynylthio denote an alkynyl group as described above bonded through an oxygen linkage (-0-) or a sulfur linkage (-S-), respectively.
• cycloalkoxy or "cycloalkylthio” , as used herein, denote an cycloalkyl group as described above bonded through an oxygen linkage (-0-) or a sulfur linkage (-S-), respectively.
• cycloalkenyloxy or "cycloalkenylthio” , as used herein, denote a cycloalkenyl group as described above bonded through an oxygen linkage (-0-) or a sulfur linkage (-S-) , respectively.
• cycloalkynyloxy or "cycloalkynylthio” , as used herein, denote a cycloalkynyl group as described above bonded through an oxygen linkage (-0-) or a sulfur linkage (-S-), respectively.
• heterocycloalkoxy or “heterocycloalkylthio” , as used herein, denote a heterocycloalkyl group as described above bonded through an oxygen linkage (-0-) or a sulfur linkage (-S-) , respectively.
• heterocycloalkenyloxy or “heterocycloalkenylthio” , as used herein, denote a heterocycloalkenyl group as described above bonded through an oxygen linkage (-0-) or a sulfur linkage (-S-), respectively.
• heterocycloalkynyloxy or “heterocycloalkynylthio” , as used herein, denote an heterocycloalkynyl group as described above bonded through an oxygen linkage (-0-) or a sulfur linkage (-S-), respectively.
• aryloxy or "aryllthio” , as used herein, denote an aryl group as described above bonded through an oxygen linkage (-0-) or a sulfur linkage (-S-) , respectively.
• heteroalkyloxy or “heteroalkyllthio” , as used herein, denote an heteroalkyl group as described above bonded through an oxygen linkage (-0-) or a sulfur linkage (-S-), respectively.
• heteroalkenyloxy or “heteroalkenyllthio” , as used herein, denote an heteroalkenyl group as described above bonded through an oxygen linkage (-0-) or a sulfur linkage (-S-), respectively.
• heteroalkynyloxy or “heteroalkynyl1thio” , as used herein, denote an heteroalkynyl group as described above bonded through an oxygen linkage (-0-) or a sulfur linkage (-S-) , respectively.
• arylalkoxy or "arylalkylthio” , as used herein, denote an arylalkyl group as described above bonded through an oxygen linkage (-0-) or a sulfur linkage (-S-) , respectively.
• alkylaryloxy or "alkylaryllthio” , as used herein, denote an alkylaryl group as described above bonded through an oxygen linkage (-0-) or a sulfur linkage (-S-) , respectively.
• alkylamin or "alkyloxycarbonyl”, as used herein, denote an alkyl group as described above bonded through an nitrogen linkage (-N-) or denotes an alkoxy group bonded through a carbonyl group, respectively.
• alkenylamin or "alkenyloxycarbonyl” , as used herein, denote an alkenyl group as described above bonded through an nitrogen linkage (-N-) or denotes an alkoxy group bonded through a carbonyl group, respectively.
• alkynylamin or “alkynyloxycarbonyl” , as used herein, denote an alkynyl group as described above bonded through an nitrogen linkage (-N-) or denotes an alkoxy group bonded through a carbonyl group, respectively.
• cycloalkamin or "cycloalkyloxycarbonyl” , as used herein, denote an cycloalkyl group as described above bonded through an nitrogen linkage (-N-) or denotes an alkoxy group bonded through a carbonyl group, respectively.
• cycloalkenylamin or “cycloalkenyloxycarbonyl” , as used herein, denote a cycloalkenyl group as described above bonded through an nitrogen linkage (-N-) or denotes an alkoxy group bonded through a carbonyl group, respectively.
• cycloalkynylamin or "cycloalkynyloxycarbonyl”, as used herein, denote a cycloalkynyl group as described above bonded through an nitrogen linkage (-N-) or denotes an alkoxy group bonded through a carbonyl group, respectively.
• heterocycloalkamin or “heterocycloalkyloxycarbonyl” , as used herein, denote a heterocycloalkyl group as described above bonded through an nitrogen linkage (-N-) or denotes an alkoxy group bonded through a carbonyl group, respectively, such as piperazinyl, morpholinyl, thiomorpholinyl
• heterocycloalkenylamin or “heterocycloalkenyloxycarbonyl” , as used herein, denote a heterocycloalkenyl group as described above bonded through an nitrogen linkage (-N-) or denotes an alkoxy group bonded through a carbonyl group, respectively.
• heterocycloalkynylamin or “heterocycloalkynyloxycarbonyl” , as used herein, denote an heterocycloalkynyl group as described above bonded through an nitrogen linkage (-N-) or denotes an alkoxy group bonded through a carbonyl group, respectively.
• arylamin or "arylloxycarbonyl” , as used herein, denote an aryl group as described above bonded through an nitrogen linkage (-N-) or denotes an alkoxy group bonded through a carbonyl group, respectively.
• heteroalkylamin or “heteroalkylloxycarbonyl” , as used herein, denote an heteroalkyl group as described above bonded through an nitrogen linkage (-N-) or denotes an alkoxy group bonded through a carbonyl group, respectively.
• heteroalkenylloxycarbonyl denote an heteroalkenyl group as described above bonded through an nitrogen linkage (-N-) or denotes an alkoxy group bonded through a carbonyl group, respectively.
• heteroalkynylamin or
• heteroalkynylloxycarbonyl denote an heteroalkynyl group as described above bonded through an nitrogen linkage (-N-) or denotes an alkoxy group bonded through a carbonyl group, respectively.
• arylalkamin or "arylalkyloxycarbonyl” , as used herein, denote an arylalkyl group as described above bonded through an nitrogen linkage (-N-) or denotes an alkoxy group bonded through a carbonyl group, respectively.
• alkylarylamin or “alkylarylloxycarbonyl” , as used herein, denote an alkylaryl group as described above bonded through an nitrogen linkage (-N-) or denotes an alkoxy group bonded through a carbonyl group, respectively.
• heteroatom means 0, S or N, selected on an independent basis.
• halogen refers to chlorine, bromine, fluorine or iodine .
• a functional group is termed “protected”, this means that the group is in modified form to preclude undesired side reactions at the protected site.
• Suitable protecting groups of the compounds of the present invention will be recognized from the present application taking into account the level of skill in the art, and with reference to standard textbooks, such as Greene, T.W. et al . , Protective Groups in Organic Synthesis, Wiley, N.Y. (1991) .
• salts of the compounds according to the invention with inorganic or organic acids are hydrochloride, hydrobro ide, hydrosulfate, sulfate, hydrophosphate, phosphate and the like. Salts which are unsuitable for pharmaceutical uses but which can be employed, for example, for the isolation or purification of free compounds or their acceptable salts, are also included.
• Suitable salts of carboxylic groups like sodium, potassium, lithium or magnesium or other pharmaceutically acceptable salts are also included.
• All stereoisomers of the compounds of the instant invention are contemplated, either in a mixture or in pure or substantially pure form.
• the definition of the compounds according to the invention embraces all possible stereoisomers and their mixtures. It very particularly embraces the racemic forms and the isolated optical isomers having the specified activity.
• the racemic forms can be dissolved by physical methods, such as, for example fractional crystallisation, separation or crystallisation of diastereomeric derivatives or separation by chiral column chromatography.
• the individual optical isomers can be obtained from the racemates by conventional methods, such as, salt formation with an optically active acid followed by crystallisation.
• solvates e.g. hydrates
• Methods of solvation are generally known in the art . Accordingly, the compounds of the instant invention may be in the free or hydrate form, and may be obtained by methods exemplified.
• the present invention relates to a new class of compounds that block or moderate kinase activity of tyrosine kinases
• An embodiment of the invention relates to a new class of compounds that block or moderate kinase activity of the tyrosine kinase Tie-2 and KDR.
• the simultaneous impairment of the VEGF/VEGF-R and of the Ang/Tie receptor systems is advantageous for cancer treatment.
• the improved effect can be attributed probably to the fact that the impairment of the Ang/Tie receptor system destabilizes the interaction between endothelial and periendothelial cells in the existing tumor blood vessels and thus sensitizes the endothelium for compounds which are directed against the VEGF/VEGF-R system.
• Follwing IC50 values were determined in the RTK ELISA using recombinant kinase domains of receptor tyrosine kinases which were expressed in baculovirus infected insect cells.
• Compounds of the invention are comprised in the following listings including their kinase activity inhibiting activity.
• Synthesis of the compound 1-37 may be as follows: Synthesis of methyl-thiazol-2-yl-amine hydrochloride (2) :
• HPLC (Column: Xterra, MS C18, 5 ⁇ m, 4.6*100 mm; 3 ml/min; 10% to 90% B gradient in 3.5 min.
• Solvent A water + 0.1% trifluoroacetic acid
• Solvent B 100% acetonitrile + 0.1% trifluoroacetic acid
• UV: 214 nm, 254 nm, 301 nm) : r 0.55 min, purity >95%.
• HPLC- S 115.1 (M+H) .
• Solvent A water + 0.1% trifluoroacetic acid
• Solvent B- 100% acetonitrile + 0.1% trifluoroacetic acid
• UV: 214 nm, 254 nm, 301 nm) : r t 1.83 min, purity >95% HPLC-MS: 246 (M+H) .
• Solvent A water + 0.1% trifluoroacetic acid
• Solvent B 100% acetonitrile + 0.1% trifluoroacetic acid
• UV: 214 nm, 254 nm, 301 nm) : r t 2.71 min, purity >85%.
• HPLC (Column: Xterra, MS C18, 5 ⁇ m, 4.6*100 mm; 3 ml/min; 10% to 90% B gradient in 3.5 min.
• Solvent A water + 0.1% trifluoroacetic acid
• Solvent B 100% acetonitrile + 0.1% trifluoroacetic acid
• UN 214 nm, 254 nm, 301 nm
• r 2.63 min, purity >90%.
• HPLC-MS 322 (M+H) .
• HPLC (Column: Xterra, MS C18, 5 ⁇ m, 4.6*100 mm; 3 ml/min; 10% to 90% B gradient in 3.5 min.
• Solvent A water + 0.1% trifluoroacetic acid
• Solvent B 100% acetonitrile + 0.1% trifluoroacetic acid
• UV 214 nm, 254 nm, 301 nm
• r t 2.79 min, purity >99%.
• HPLC-MS 367 (M+H) .
• HPLC (Column: Xterra, MS C18, 5 ⁇ m, 4.6*100 mm; 3 ml/min; 10% to 90% B gradient in 3.5 min.
• Solvent A water + 0.1% trifluoroacetic acid
• Solvent B 100% acetonitrile + 0.1% trifluoroacetic acid
• UV 214 nm, 254 nm, 301 nm
• r t 2.80 min, purity >95%.
• HPLC-MS 565 (M+H) .
• Solvent A water + 0.1% trifluoroacetic acid
• Solvent B 100% acetonitrile + 0.1% trifluoroacetic acid
• UV 214 nm, 254 nm, 301 nm
• r t 1.30 min, purity >98%.
• GC-MS 324 (M + ) .
• HPLC (Column: Xterra, MS C18, 5 ⁇ m, 4.6*100 mm; 3 ml/min; 10% to 90% B gradient in 3.5 min.
• Solvent A water + 0.1% trifluoroacetic acid
• Solvent B 100% acetonitrile + 0.1% trifluoroacetic acid
• UN 214 nm, 254 nm, 301 ran
• r t 1.59 min, purity >98%.
• HPLC-MS 337 (M+H) .
• the organic solvent was removed under reduced pressure and after cooling filtered if possible or extracted with methylene chloride, dried over gS0 4 and concentrated. If acidic groups are present the organic solvent was removed, the aqueous layer was brought to neutral pH followed by filtration or extractive workup as described above. The crude thiourea was used without further purification.
• X leaving group (i.e. chloride, bromide, iodide)
• SP5659 941 mg (5 mmol) 2, 4-difluorophenylthiourea and 1.22 g (5 mmol) alpha- bromo-4 -nitroacetophenone in 30 ml ethanol were reacted according to general procedure 2.
• the nitro group of the product was reduced according to general procedure 3 using 3.4 g (15 mmol) SnCl 2 x 2H 2 0 in 50 ml ethanol.
• the following in vitro assays are used to determine the ability of different compounds to inhibit the transfer of phosphate groups onto tyrosine residues of downstream substrates.
• the level of phosphorylation is measured by a monoclonal antibody which is specific for phosphorylated tyrosine residues in an enzyme-linked immunosorbent assay (ELISA) .
• ELISA enzyme-linked immunosorbent assay
• GST-FLT4 (aa725-1298) The following recombinant kinase was expressed in baculovirus infected insect cells and purchased from MoBiTec, G ⁇ ttingen:
• BSA Bovine Serum Albumine
• Enhancement factor 125-150
• DMEM Dulbeccos modified eagle medium
• 293T cells are plated at 1 x 106 cells per well in a six well plate in DMEM medium supplemented with 10 % fetal calf serum (FCS) , incubated overnight at 37 °C and transfected with 1 ⁇ g plasmid DNA of pCB ANGl by the Escort transfection reagent according to the manufacturer's protocol. After one day medium is changed to DMEM without FCS, and cell culture supernatants are harvested by centrifugation after additional 3 days.
• FCS fetal calf serum
• HAVEC Human umbilical vein endothelial cells
• ECG Endothelial cell growth (Promocell C-22010) supplemented with:
• bFGF bascic Fibroblast Growth Factor
• BSA Bovine Serum Albumine
• Endothelial cells e. g. HUNECs between passage 2 and 12 are plated at between 2 x 105 and 1 x 106 cells per well in a six well plate in supplemented ECG medium. After 24 to 48 hours the medium is changed to M199 medium containing increasing concentrations of the inhibitory compound in the individual wells. The cells are incubated at 37 °C and then treated with cell culture supernatants containing the ligand for 5 to 30 min.
• the cells are placed on ice, washed once with 1 ml PBS, lysed by the addition of 300 ⁇ l RIPA buffer and removed from the plate by a cell scraper.
• the suspension is transferred into a microcentrifuge tube, sonicated for 5 sec and boiled after addition of 100 ⁇ l 4 x SDS sample buffer for 5 min at 95 °C.
• the blots are blocked with PBS / 0.05 % Tween 20 / 1 % BSA for 1 h at room temperature, incubated with either antiphosphotyrosin or anti-Tie-2 antibody diluted 1:2000 in PBS/Tween for 1 hour and washed 3 times with PBS/Tween.
• the blot is incubated with a goat anti rabbit IgG secondary antibody/ HRP conjugate diluted 1:4000 in PBS/Tween. After washing 3 times in PBS/Tween the blot is developed by the ECL method according to the manufacturer's instruction.
• Angiopoietin-2 a natural antagonist for Tie2 that disrupts in vivo angiogenesis. Science 277, 55-60.
• the receptor tyrosine kinase TIE is required for integrity and survival of vascular endothelial cells. Embo J 14 , 5884-5891.
• VEGFR-3 and its ligand VEGF-C are associated with angiogenesis in breast cancer. Am J Pathol 154 , 1381-1390.

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