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WO2015019037A1 - Composés pharmaceutiques - Google Patents

Composés pharmaceutiques Download PDF

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Publication number
WO2015019037A1
WO2015019037A1 PCT/GB2013/052080 GB2013052080W WO2015019037A1 WO 2015019037 A1 WO2015019037 A1 WO 2015019037A1 GB 2013052080 W GB2013052080 W GB 2013052080W WO 2015019037 A1 WO2015019037 A1 WO 2015019037A1
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WO
WIPO (PCT)
Prior art keywords
group
amino
pyrimidin
phenyl
methyl
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
PCT/GB2013/052080
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English (en)
Inventor
James Samuel Shane Rountree
Colin Roderick O'dowd
Frank Burkamp
Mark Peter Bell
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Almac Discovery Ltd
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Almac Discovery Ltd
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Publication date
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Priority to PCT/GB2013/052080 priority Critical patent/WO2015019037A1/fr
Publication of WO2015019037A1 publication Critical patent/WO2015019037A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D471/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00
    • C07D471/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00 in which the condensed system contains two hetero rings
    • C07D471/04—Ortho-condensed systems
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
    • C07D487/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains two hetero rings
    • C07D487/04—Ortho-condensed systems
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D491/00—Heterocyclic compounds containing in the condensed ring system both one or more rings having oxygen atoms as the only ring hetero atoms and one or more rings having nitrogen atoms as the only ring hetero atoms, not provided for by groups C07D451/00 - C07D459/00, C07D463/00, C07D477/00 or C07D489/00
    • C07D491/02—Heterocyclic compounds containing in the condensed ring system both one or more rings having oxygen atoms as the only ring hetero atoms and one or more rings having nitrogen atoms as the only ring hetero atoms, not provided for by groups C07D451/00 - C07D459/00, C07D463/00, C07D477/00 or C07D489/00 in which the condensed system contains two hetero rings
    • C07D491/04—Ortho-condensed systems

Definitions

  • the present invention relates to compounds that are useful as inhibitors of the activity of Wee-1 kinase.
  • the present invention also relates to pharmaceutical compositions comprising these compounds and to methods of using these compounds in the treatment of cancer and methods of treating cancer.
  • Detection of DNA damage initiates a series of events which are key in maintaining the genome.
  • Cell cycle checkpoints are designed to stop the cell cycle and allow repair of the lesion before allowing the cell to continue into mitosis .
  • Two key checkpoints have been identified, one at the end of Gl phase and the second at G2, which work in tandem to ensure all lesions, are identified and repaired.
  • the Gl checkpoint is non-functional due to mutations in the tumour suppressor gene p53.
  • the G2 check-point is seldomly mutated, and often found to be activated in cancer cells. Cancer cells exploit this to confer resistance to treatment modalities
  • Inhibitors for the kinases are currently in clinical trials .
  • Wee-1 is a nuclear tyrosine kinase, which negatively regulates entry into mitosis, at the G2/M check-point by catalysing a phosphorylation of the cdc2 / cyclin B kinase complex. The phosphorylation occurs on the
  • Wee-1 is downstream of the Chk family and is a crucial component of the checkpoint signalling cascade as it prevents cells from entering mitosis if lesions are detected .
  • anti-cancer compounds induce DNA damage; including anti-metabolites, platiniums,
  • topoisomerase inhibitors and alkylating agents are used in combination with these agents to prevent DNA repair selectively in tumour cells.
  • Compounds which work in combination with these agents to prevent DNA repair selectively in tumour cells would be extremely beneficial.
  • the tumour suppressor gene p53 is commonly mutated in tumour cell lines, therefore the
  • Irradiation is known to increase phosphorylation of the Tyrl5 and Thrl4 residues of cdc2, leading to a
  • kinase inhibitory effect for example a Wee-1 kinase inhibitory effect, are described in WO2007/126122, US2010/0063024, EP2,213,673, W02008 / 133866 and US2007/0254892.
  • the compounds of WO2005/090344 are said to show activity as protein kinase inhibitors, in particular Src family tyrosine kinase inhibitors.
  • the compounds described in Bioorg & Med Chem Lett Vol 15, pp 1931-1935 are said to be 10-100-fold more potent inhibitors of c- Src than Weel, and variation of substituents on the 6- phenyl ring does not markedly alter this preference. It is said that solubilizing substituents off the 2-anilino ring in many cases increases Weel activity, lowering this preference to about 10-fold.
  • analogues are said to be generally Weel selective, but at the expense of absolute potency.
  • compositions are provided.
  • the present invention provides a compound of Formula (I) :
  • X is an oxygen atom or a nitrogen atom
  • Y is a carbon atom or a nitrogen atom;
  • R 1 is an optionally substituted aryl group;
  • R 2 is a hydrogen atom, an optionally substituted alkyl group or an optionally substituted amino group when Y is a carbon atom and R 2 is absent when Y is a nitrogen atom;
  • R 3 is a hydrogen atom, an optionally substituted alkyl group or an optionally substituted aryl group when X is a nitrogen atom and R 3 is absent when X is an oxygen atom;
  • R 4 is an optionally substituted alkyl group or an optionally substituted aryl group.
  • the present invention provides a pharmaceutical composition
  • a pharmaceutical composition comprising the compound of formula (I), or a pharmaceutically acceptable salt or N- oxide derivative thereof, and at least one
  • the present invention provides the compound of formula (I), or a pharmaceutically acceptable salt or N-oxide derivative thereof, or a pharmaceutical composition comprising the compound of formula (I) for use in therapy.
  • the present invention provides the compound of formula (I) for use as a medicament. In a fifth aspect the present invention provides the compound of formula (I) for use in treating or preventing cancer . In a sixth aspect the present invention provides the compound of formula (I) for the manufacture of a
  • the present invention provides the use of the compound of formula (I) for the manufacture of a medicament for treating or preventing cancer.
  • the present invention provides a method of treating or preventing cancer in a human or animal patient comprising administering to a patient in need thereof an effective amount of the compound of formula (I) or a pharmaceutical composition comprising the compound of formula (I) .
  • Other preferred embodiments of the compounds according to the invention appear throughout the specification and in particular in the examples. Particularly preferred are those named compounds having greater activity as tested. Compounds having higher activity are more preferred over those having lower activity.
  • the present inventors have surprisingly found that the compounds of the present invention show an improved selectivity towards Wee-1 kinase.
  • the compounds of the present invention show an improved selectivity towards Wee-1 kinase.
  • the compounds of the invention are selective over members of the Src family of kinases, for example LCK (Lymphocyte specific protein tyrosine kinase) and c- Src.
  • LCK Lymphocyte specific protein tyrosine kinase
  • c- Src kinases
  • the present inventors have surprisingly found that the compounds of the present invention show an improved or similar kinase-inhibitory effect compared to known compounds or compositions.
  • the compounds of the present invention preferably show an improved or similar Wee-1 kinase-inhibitory effect compared to known compounds or compositions.
  • the compounds of the present invention have an improved stability in human microsomes and/or an improved tolerability compared to known compounds or compositions .
  • alkyl group refers to an aliphatic group containing at least carbon and hydrogen and containing 1 to 15 carbon atoms, such as 1 to 10 carbon atoms.
  • a " C n alkyl” group refers to an aliphatic group containing n carbon atoms.
  • a Ci-Cio alkyl group
  • An alkyl group may be straight chained or it may be branched.
  • An alkyl group may contain no ring structures or it may contain one or more rings (i.e. "a cycloalkyl group") .
  • a "cycloalkyl group” contains at least one ring. It is understood that attachment to a cycloalkyl group is via a ring atom of the cycloalkyl group.
  • Each ring may contain 3 to 10 atoms, such as 4 to 8 or 5 to 7 atoms.
  • Each ring may be independently selected to contain just carbon atoms or to contain both carbon atoms and from 1 to 4 heteroatoms selected from 0, N and S.
  • attachment to the cycloalkyl group may occur either through a carbon atom or, if one or more heteroatoms are contained in a ring, attachment may also occur through a heteroatom contained in a ring.
  • a cycloalkyl group may be mono-cyclic or bi- cyclic.
  • the bicyclic cycloalkyl group may have a spiro structure in which the two rings share one and the same ring atom, or may have a bicyclo structure in which the two rings share two or more ring atoms.
  • a " C n cycloalkyl" group contains n carbon atoms. All n carbon atoms may be contained in the ring(s) of the cycloalkyl group or one or more of the carbons may not be contained in the ring(s) and may instead form one or more chains branching from the ring.
  • the two alkyl groups contain a total number of m + n carbon atoms.
  • an alkyl group may be saturated or unsaturated.
  • the alkyl group may be an alkenyl group (i.e. contain a carbon-carbon double bond) and / or an alkynyl group (i.e. contain a carbon-carbon triple bond) .
  • the alkyl group may contain at least 2 carbon atoms. It is understood that any unsaturated portions of an alkyl group are non-aromatic (aromatic groups fall within the scope of the definition of "aryl") .
  • Any part of the alkyl group may be unsaturated, for example the straight, branched or cyclic portion of an alkyl group may contain a carbon-carbon double bond or a carbon- carbon triple bond. Attachment to an unsaturated alkyl group may occur through the unsaturated part of the alkyl group or may occur through the unsaturated part of the group .
  • an unsaturated alkyl group may contain 1 to 4 carbon-carbon double bonds or 1 to 3 carbon-carbon triple bonds or 1 to 4 of a combination of carbon-carbon double bonds and carbon-carbon triple bonds.
  • An alkyl or cycloalkyl group can be optionally
  • substituted (a "substituted alkyl” or "substituted cycloalkyl") with one or more alkyl group substituents , which can be the same or different.
  • An alkyl group substituent can be attached to the alkyl or cycloalkyl group through a carbon atom in the alkyl or cycloalkyl group, or, if one or more heteroatoms are contained in a cycloalkyl group, attachment may also occur through a heteroatom contained in the cycloalkyl group.
  • alkyl group substituent includes but is not limited to alkyl, substituted alkyl, aralkyl, substituted aralkyl, halo, cyano, amino, alkylamino, arylamino, aryl,
  • arylsulfonyl and cycloalkyl If a group, for example an alkyl group, is "optionally substituted” , it is understood that the group has one or more substituents attached (substituted) or does not have any substituents attached (unsubstituted) . Examples of unsubstituted saturated alkyl groups
  • containing no cyclic structures include methyl, ethyl, n- propyl, sec-propyl, n-butyl, sec-butyl, tert-butyl, pentyl (branched or unbranched) , hexyl (branched or unbranched) , heptyl (branched or unbranched) , octyl
  • unsubstitued saturated cycloalkyl groups include cyclopropyl, cylcobutyl, cyclopentyl and
  • unsaturated alkyl groups include ethenyl, trimethenyl, tetramethenyl , propenyl, butenyl, 2- methybutenyl , pentamethenyl , hexamethenyl and
  • cyclohexenyl examples include azetidinyl, pyrrolodinyl , piperidinyl, piperizinyl, tetrahydrofuranyl , tetrahydropyranyl , morpholine, thiomorpholine, homopiperaz ine , homo- piperidine, homomorpholine, homothiomorpholine, S,S- dioxythiomorpholine, S, S-dioxyhomothiomorpholine .
  • aryl group refers to a group containing at least one ring that is aromatic and containing 1 to 15 carbon atoms, such as 1 to 10 carbon atoms.
  • the aryl group can be optionally substituted (a
  • substituted aryl with one or more aryl group
  • aryl group substituent includes but is not limited to alkyl, substituted alkyl aryl, substituted aryl, aralkyl, hydroxyl, alkoxyl, haloalkoxyl perhaloalkoxyl , aryloxyl, aralkyloxyl, carboxyl, alkanoyl, halo, nitro,
  • alkoxycarbonyl aryloxycarbonyl , aralkoxycarbonyl , acyloxyl, cyano, acylamino, amido, carbamoyl,
  • alkylcarbamoyl dialkylcarbamoyl , arylthio and alkylthio.
  • Each ring may be independently selected to contain only carbon atoms or to contain both carbon atoms and from 1 to 4 heteroatoms selected from 0, N and S.
  • heteroaryl groups i.e. aryl groups that contain one or more heteroatoms
  • attachment to the aryl group may occur either through a carbon atom or, if one or more
  • heteroatoms are contained in a ring, attachment may also occur through a heteroatom contained in a ring.
  • heteroatoms contained in a ring of a heteroaryl group may be substituted, for example forming an iV-oxide.
  • the aromatic group may be mono-cyclic or bi- cyclic, wherein one or both of the rings of a bi-cyclic system is aromatic. If one ring is non-aromatic, the non- aromatic ring may be further substituted with one or more substituents , which can be the same or different.
  • the substituents include, but are not limited to alkyl, spirocyclic alkyl, aralkyl, substituted aralkyl, halo, cyano, amino, alkylamino, arylamino, aryl, substituted aryl, nitro, thio, acyl, hydroxyl, aryloxyl, alkoxyl, alkylthio, arylthio, aralkyloxyl, aralkylthio, carboxyl, alkoxycarbonyl , oxo, alkylsulfonyl , arylsulfonyl and cycloalkyl .
  • aryl groups include acridinyl, phenyl, carbazolyl, cinnolinyl, quinoxalinyl , pyrrazolyl, benzotriazolyl , furanyl, naphthyl, thienyl, thiazolyl, benzothienyl , benzofuranyl , quinolinyl, isoquinolinyl , oxazolyl, isoxazolyl, indolyl, pyrazinyl, pyridazinyl, pyridinyl, pyrimidinyl, pyrrolyl, tetrahydroquinoline, tetrahydroisoquinoline, benzimidazolyl and melaminyl.
  • heterocycle includes within its scope both cycloalkyl groups containing one or more heteroatoms within the ring system and aryl groups containing one or more heteroatoms within the ring system.
  • Heterocyclic groups may be, for example, pyrazoles, imidazoles and any of the various triazoles and may include the oxygen and/or sulphur containing analogues that is oxazoles, isoxazoles, thiazoles and isothiazoles and derivatives.
  • nitrogen-containing heterocyclic group refers to a monocyclic or bicyclic heterocyclic group containing at least one nitrogen atom, in which each ring comprises from 3 to 7 ring atoms and optionally contains, in addition to the nitrogen atom, zero or one or two or more, the same or different hetero atoms, but preferably one hetero atom selected from the group consisting of an oxygen atom, a nitrogen atom and a sulfur atom; and the heterocyclic group may be aromatic or aliphatic.
  • the bicyclic heterocyclic group may have a spiro structure of which the two rings share one and the same ring atom, or may have a bicyclo structure of which the rings share two or more ring atoms.
  • the nitrogen-containing heterocyclic group include, for example, a pyrrolyl group, an imidazolyl group, a pyrazolyl group, a
  • thiazolyl group an isothiazolyl group, an oxazolyl group, an isoxazolyl group, a triazolyl group, a
  • tetrazolyl group an oxadiazolyl group, a 1,2,3- thiadiazolyl group, a 1 , 2 , 4-thiadiazolyl group, a 1,3,4- thiadiazolyl group, a pyridyl group, a pyrazinyl group, a pyrimidinyl group, a pyridazinyl group, a 1 , 2 , 4-triazinyl group, a 1 , 3 , 5-triaz inyl group, an indolyl group, a benzimidazolyl group, a benzoxazolyl group, a
  • benzisothiazolyl group an indazolyl group, a purinyl group, a quinolyl group, an isoquinolyl group, a
  • phthalazinyl group a naphthyridinyl group, a quinoxalinyl group, a quinazolinyl group, a cinnolinyl group, a pteridinyl group, a pyrido [ 3 , 2-b] pyridyl group, an azetidinyl group, a pyrrolidinyl group, a dihydro- 1 , 2 , 4-triazolyl group, a dihydro-1 , 2 , 4-oxadiazolyl group, a dihydro-1 , 3 , 4-oxadiazolyl group, a dihydro-1 , 2 , 4- thiadiazolyl group, a dihydro-1 , 2 , 3 , 5-oxathiadiazolyl group, a piperidinyl group, a piperazinyl group, a dihydropyridyl group, a morpholinyl group, a
  • thiomorpholinyl group a 2 , 6 -diazaspiro [ 3.5 ] nonyl group, a 2 , 7-diazaspiro [ 3.5 ] nonyl group, a 2,7- diazaspiro [ 4.5 ] decyl group, or a 2,7- diazabicyclo [ 3.3.0 ] octyl group, a 3,6- diazabicyclo [ 3.3.0 ] octyl group.
  • the nitrogen-containing heterocyclic group can be
  • amino group refers to the -NH 2 group.
  • the amino group can be optionally substituted (a "substituted amino") with one or more amino group substituents, which can be the same or different, wherein “amino group substituent” includes but is not limited to alkyl and aryl .
  • alkoxy group refers to an alkyl-0 group. The alkoxy group can refer to linear, branched, or cyclic, saturated or unsaturated oxo-hydrocarbon chains,
  • alkoxy group can be optionally substituted (a "substituted alkoxy") with one or more alkoxy group substituents , which can be the same or different, wherein "alkoxy group substituent" includes but is not limited to alkyl, amino and aryl .
  • alkanoyl group refers to an organic acid group wherein the -OH of the carboxyl group has been replaced with another substituent.
  • alkanoyl groups include an acetyl group, a propionyl group, a butyryl group, an isobutyryl group, a valeryl group, an isovaleryl group, a pivaloyl group.
  • sulfonyl group refers to a sulfonic acid group wherein the wherein the -OH of the sulfonyl group has been replaced with another substituent.
  • the substitutent may be an alkyl group ("an alkylsufonyl group”) .
  • An alkylsulfonyl group can be represented by the formula S ⁇ 0) 2 ⁇ , wherein R is an alkyl group,
  • alkylsulfonyl groups include a methylsulfonyl group, an ethylsulfonyl group, a
  • propylsulfonyl group an isopropylsulfonyl group, a butylsulfonyl group, a sec-butylsulfonyl group, an isobutylsulfonyl group, a tert-butylsulfonyl group, a pentylsulfonyl group, an isopentylsulfonyl group, a hexylsulfonyl group and an isohexylsulfonyl group.
  • halo refers to a group selected from chlorine, fluorine, bromine and iodine.
  • aromaticity may indicate the presence of, for
  • a mono-, di- or tri-cyclic system that contains (4n+2) electrons where n is an integer.
  • these systems may display Hiickel aromaticity.
  • invention may possess some aspect of stereochemistry.
  • the compounds may possess chiral centres and / or planes and / or axes of symmetry.
  • the compounds may be provided as single stereoisomers, single diastereomers , mixtures of stereoisomers or as racemic mixtures.
  • Stereoisomers are known in the art to be molecules that have the same molecular formula and sequence of bonded atoms, but which differ in their spatial orientations of their atoms and / or groups.
  • the compounds of the present invention may possess tautomerism. Each tautomeric form is intended to fall within the scope of the invention.
  • the compounds of the present invention may be provided as a pro-drug.
  • Pro-drugs are transformed, generally in vivo, from one form to the active forms of the drugs described herein.
  • a prodrug may be formed by protecting the -N-H group to which R 4 is attached with a hydrolysable group that gives -NH on hydrolysis.
  • X, Y or Z and/or any moieties appended thereto taken separately or together are -N-H, one or more of these may be
  • the elements of the compounds of the present invention may be provided as isotopes.
  • hydrogen may also be deuterium.
  • the compounds of the present invention may be provided in the form of their pharmaceutically
  • the compounds may be provided having protonated amine groups.
  • pharmaceutically acceptable salt refers to ionic compounds formed by the addition of an acid to a base.
  • the term refers to such salts that are considered in the art as being suitable for use in contact with a patient, for example in vivo and pharmaceutically
  • acceptable salts are generally chosen for their non- toxic, non-irritant characteristics.
  • co-crystal refers to a multi- component molecular crystal, which may comprise non-ionic
  • compositions may be prepared by ion exchange chromatography or by reacting the free base or acidic form of a compound with
  • Salts known in the art to be generally suitable for use in contact with a patient include salts derived from inorganic and / or organic acids, including the
  • hydrobromide hydrochloride, sulphate, bisulphate, nitrate, acetate, oxalate, oleate, palmitate, stearate, laurate, benzoate, lactate, phosphate, tosylate, citrate, maleate, fumarate, succinate and tartrate.
  • alkali and alkaline earth metals such as sodium, potassium, calcium and magnesium, as well as ammonium, tetramethylammonium
  • the compounds of the present invention may sometimes exist as zwitterions, which are considered as part of the invention.
  • the present inventors have discovered that the compounds of the present invention are useful in the treatment of medical conditions associated with disordered cell growth, including, but not restricted to, cancer, in particular cancers associated with mutations in the tumour suppressor gene p53.
  • cancers include cardiac cancers, lung cancers, gastrointestinal cancers, genitourinary tract cancers, liver cancers, bone cancers, nervous system cancers, gynecological cancers, hematologic cancers, skin cancers and adrenal gland cancers.
  • cancers include adrenal tumors, bile duct, bladder, blood, bone and connective tissue, brain and central nervous system, breast, cervical, colon and rectal (colorectal), endometrial, esophageal, gallbladder, head and neck, Hodgkin's Lymphoma,
  • hypopharangeal kidney, laryngeal, leukemias, liver, lung, lymphoma, mediastinal tumors, melanoma (malignant melanoma) , mesothelioma, multiple myeloma, nasal cavity, nasopharyngeal, neuroendocrine tumors, non-Hodgkin ' s lymphoma, oral, oesophagus, oropharyngeal, ovarian, pancreas, paranasal sinus, parathyroid, penis, pituitary tumors, prostate, salivary gland, sarcoma, skin, spine, stomach, testicular, thyroid, urethra, uterine, vaginal and vulvar.
  • the compounds of the present invention are also useful in preparing a medicament that is useful in treating the diseases described above, in particular cancer.
  • the present invention is further directed to a method of inhibiting Wee-1 activity which comprises administering to a mammal in need thereof a pharmaceutically effective amount of the compound of the present invention.
  • the compounds of this invention may be administered to mammals, including humans, either alone or, in
  • the compounds can be administered orally or parenterally, including the intravenous, intramuscular,
  • the present invention also includes within its scope the use of the compounds of the present invention in
  • the present invention also includes the use of the compounds of the invention in a regime including the step of radiotherapy.
  • the radiotherapy maybe an ordinary method of treatment by x-ray, ⁇ -ray, neutron, proton or electron beam irradiation.
  • the co-administration of compounds contained in this invention may lead to the potentiation of the radiation therapy, thus classifying them as radio-sensitizers .
  • cancers often become resistant to therapy.
  • the development of resistance may be delayed or overcome by the administration of a combination of drugs that includes the compounds of the present invention for example in cancers which are known to be resistant to DNA damaging agents or radiotherapy.
  • drugs that may be used in combination with the compounds of the present invention may target the same or a similar biological pathway to that targeted by the compounds of the present invention or may act on a different or unrelated pathway.
  • the second active ingredient may include, but is not restricted to:
  • alkylating agents including cyclophosphamide
  • antimitotic agents including vinca alkaloids (vincristine, vinorelbine and vinblastine), taxanes
  • paclitaxel docetaxel
  • epothilones and inhibitors of mitotic kinases including aurora and polo kinases
  • topoisomerase inhibitors including anthracyclines , epipodophyllotoxins , camptothecin and analogues of camptothecin ; antimetabolites, including 5-fluorouracil, capecitabine , cytarabine, gemcitabine, 6-mercaptopurine, 6 -thioguanine , fludarabine, methotrexate and premetrexed; protein kinase inhibitors, including imatinib, gefitinib, sorafenib, sunitinib, erlotinib, dasatinib, and
  • proteosome inhibitors including bortezomib
  • histone deacetylase inhibitors including valproate and SAHA
  • antiangiogenic drugs including bevacizumab
  • monoclonal antibodies including trastuzumab, rituximab, alemtuzumab, tositumomab, cetuximab, panitumumab;
  • myoclonal antibodies including Gemtuzumab ozogamicin, Ibritumomab tiuxetan; hormonal therapies, including antiestrogens (tamoxifen, raloxifen,
  • the compounds of the present invention may be administered separately,
  • the present invention provides a compound of Formula ( I ) :
  • X is an oxygen atom or a nitrogen atom
  • Y is a carbon atom or a nitrogen atom
  • R 1 is an optionally substituted aryl group
  • R 2 is a hydrogen atom, an optionally substituted alkyl group or an optionally substituted amino group when Y is a carbon atom and R 2 is absent when Y is a nitrogen atom;
  • R 3 is a hydrogen atom, an optionally substituted alkyl group or an optionally substituted aryl group when X is a nitrogen atom and R 3 is absent when X is an oxygen atom;
  • R 4 is an optionally substituted alkyl group or an optionally substituted aryl group.
  • Y is a carbon atom.
  • R 2 is a hydrogen atom, an amino group or an alkyl group when Y is a carbon atom. More preferably, R 2 is a hydrogen atom, an NH 2 group or a methyl group when Y is a carbon atom. Most preferably, R 2 is a hydrogen atom when Y is a carbon atom.
  • Y is a nitrogen atom.
  • R 2 is absent when Y is a nitrogen atom.
  • R 1 is a mono-halogenated aryl group, for example a 2-chlorophenyl group .
  • X is a nitrogen atom.
  • R 3 is a hydrogen atom or an optionally
  • R 3 is a hydrogen atom or a substituted alkyl group wherein the alkyl group substituent is selected from the group consisting of an alkoxy-subst ituted aryl group, a carboxyl group, an alkoxycarbonyl group and a hydroxyl group. More
  • R 3 is a hydrogen atom, a methyl group, an ethyl group, a propenyl group, a hydroxyl-subst ituted propyl group or a cyclopropyl group. More preferably still, R 3 is a hydrogen atom or a methyl group. Most preferably, R 3 is a methyl group.
  • R 4 is a group represented by the formula a ) :
  • Z is a nitrogen atom or an optionally substituted methine group
  • R 4a is a hydrogen atom, a halo group, an optionally substituted C 1 -C6 alkyl group, an optionally substituted C 1 -C6 alkoxy group or is a nitrogen-containing
  • heterocyclic group optionally substituted with a
  • R 4 is a hydrogen atom, a halo group, an optionally substituted C 1 -C6 alkyl group or an optionally substituted C 1 -C6 alkoxy group;
  • R a and R and the ring atoms to which they are attached may form, as taken together, a C3-C7 alkyl group, in which one or two methylene groups constituting the C3- C 7 alkyl group may be each independently replaced by an oxygen atom or a group of -N(R le )-, and the C3-C7 alkyl group may be substituted with one or more substituents selected from the group consisting of a halo group and a C 1 -C6 alkyl group;
  • R 4a and R 4 and the ring atoms to which they are attached may form, as taken together, a spiro ring or a bicyclo ring to be formed of a 5-membered to 7-membered aliphatic ring and any other 3-membered to 7-membered aliphatic ring, in which one or two or more methylene groups constituting the spiro ring or the bicyclo ring may be each independently replaced by an oxygen atom, a sulfur atom, a sulfinyl group, a sulfonyl group, an oxo group or a group of -N(R lf )-, and the spiro ring or the bicyclo ring may be each independently substituted with a substituent selected from the group consisting of a halo group, a hydroxyl group or a C 1 -C6 alkyl group; and
  • R le and R lf are each independently a hydrogen atom or a Ci- C6 alkyl group optionally substituted with a substituent selected from the group consisting of a halo group, a hydroxyl group, a cyano group, an oxo group, a C 1 -C6 alkyl group, a C 1-C6 alkoxy group, an amino group, a substituted amino group and a nitrogen-containing heterocyclic group.
  • R 4 is a group represented by the formula ( c ) :
  • R 4a is a hydrogen atom, a halo group, a C 1-C6 alkyl group or an optionally substituted C 1-C6 alkoxy group, or is a nitrogen-containing heterocyclic group optionally substituted with a substituent selected from the group consisting of a halo group, a C 1-C6 alkyl group, an oxo group and a group of -Q 1 -N (R la ) R l ;
  • R 4 is a hydrogen atom, a halo group, an optionally substituted C 1-C6 alkyl group or an optionally substituted C1-C6 alkoxy group;
  • R la and R l each independently is a hydrogen atom, a C1-C6 alkyl group, or as taken together, they may form an optionally substituted C2-C6 alkyl group;
  • Q 1 is a single bond or a C 1-C3 alkyl group.
  • R 4a is a substituted C 1-C6 alkoxy group or a nitrogen-containing heterocyclic group
  • R 4 is a hydrogen atom.
  • R 4a is a substituted C 1-C6 alkoxy group, preferably a substituted C i_C2 alkoxy group, the
  • R a is a nitrogen-containing heterocyclic group optionally substituted with a C1-C6 alkyl group
  • R 4 is a hydrogen atom, a methyl group, a methoxy group or a methyl group substituted by a hydroxy group.
  • R 4a is a nitrogen-containing heterocyclic group optionally substituted with a methyl group
  • R 4 is a hydrogen atom, a methyl group, a methoxy group or a methyl group substituted by a hydroxy group.
  • R 4 is a group represented by the formula d) :
  • R m is selected from the group consisting of a hydrogen atom and an optionally substituted C1-C6 alkyl group. More preferably R lm is a hydrogen atom or a methyl group .
  • R 4 is a substituted alkyl group wherein the alkyl group contains no ring structures or is a cycloalkyl group, said cycloalkyl group containing carbon atoms and 0 or 1 or 2 heteroatoms elected from 0, N and S, more preferably containing carbon atoms and 1 nitrogen atom.
  • R 4 is a substituted alkyl group
  • at least one of the alkyl group substituents is an optionally substituted Ci-6 alkyl group or a substituted aryl group containing carbon atoms and 0 or 1 or 2 heteroatoms elected from 0, N and S, more preferably containing carbon atoms and 1 nitrogen atom.
  • R 1 is a group represented by the formula (e) :
  • R 2a is a hydrogen atom, a halo group, a hydroxyl group, a cyano group, a C 1-C6 alkyl group, a C 1-C6 alkoxy group or a C 1-C6 alkoxy-Ci-C6 alkyl group;
  • R 3a and R 3 are each independently selected from the group consisting of a hydrogen atom, a hydroxyl group, an amino group, an alkoxy group and a C 1-C6 alkyl group;
  • T is a nitrogen atom, or a methine group which may be substituted with a halo group, a hydroxyl group, a cyano group, a C 1-C6 alkyl group, a C 1-C6 alkoxy group or a C1-C6 alkoxy-Ci-C6 alkyl group.
  • R a and R are each
  • T is a methine group and R 3a is a hydroxyl group.
  • R 1 is a group represented by the formula (f ) :
  • R a and R are each independently selected from the group consisting of a hydrogen atom and a
  • R 3a and R 3 are a hydrogen atom and the other is a hydroxyl group. More preferably R 3a is a hydroxyl group and R 3 is a hydrogen atom.
  • R 1 is a group represented by the formula (g) :
  • R 2a is a hydrogen atom, a halo group, a hydroxyl group, a cyano group, a C 1-C6 alkyl group, a C 1-C6 alkoxy group or a C 1-C6 alkoxy-Ci-C6 alkyl group;
  • T is a nitrogen atom, or a methine group which may be substituted with a halo group, a hydroxyl group, a cyano group, a C 1-C6 alkyl group, a C 1-C6 alkoxy group or a C1-C6 alkoxy-Ci-C6 alkyl group.
  • R a is a halo group, a C 1-C6 alkyl group or a C 1-C6 alkoxy group
  • T is a methine group or is a methine group
  • R 2a is a halo group
  • T is a methine group substituted with a halo group.
  • R 1 is a 2 , 6-dichlorophenyl group, that is wherein R 1 wherein is a group represented by the formula (g) and R 2a is a chlorine atom and T is a methine group substituted with a chlorine atom.
  • Y is a nitrogen atom and R 1 is a group represented by the formula h) :
  • R 2a is a hydrogen atom, a hydroxyl group, a cyano group, a C1-C6 alkyl group, a C1-C6 alkoxy group or a C1-C6 alkoxy-Ci-C6 alkyl group;
  • R 3a and R 3 are each independently selected from the group consisting of a hydrogen atom, a hydroxyl group, an amino group, an alkoxy group and a C1-C6 alkyl group; and T' is a nitrogen atom, or a methine group which may be substituted with a halo group, a hydroxyl group, a cyano group, a Ci ⁇ C6 alkyl group, a Ci ⁇ C6 alkoxy group or a Ci ⁇ C6 alkoxy-Ci-C6 alkyl group. More preferably, R 2a , R 3a and R 3 are independently a hydrogen atom, and T' is a methine group substituted with a halo group. Most preferably, Y is a nitrogen atom and R 1 is a 2- chlorophenyl group.
  • the present invention provides a compound of Formula (II) :
  • R 1 is an optionally substituted aryl group
  • R 2 is a hydrogen atom
  • R 3 is a hydrogen atom or an optionally substituted alkyl group
  • R 4 is an optionally substituted alkyl group or an optionally substituted aryl group.
  • R 1 is a group represented by the formula (e) as defined above and/or R 4 is a group represented by the formula (a) as defined above. More preferably, R 1 is a group represented by the formula (f) or the formula (g) as defined above and/or R 4 is a group represented by the formula (c) or formula (d) as defined above. More preferably still, R 1 is a group represented by the formula (g) as defined above and/or R 4 is a group represented by the formula (d) as defined above.
  • R 1 is a 2 , 6-dichlorophenyl group or a 4-hydroxyphenyl group and/or R 2 is a hydrogen atom and/or R 3 is a hydrogen atom, a methyl group, an ethyl group, a propenyl group, a hydroxyl-substituted propyl group or a cyclopropyl group and/or R 4 is a group
  • the present invention provides a compound of Formula (III) :
  • R 1 is an optionally substituted aryl group
  • R 3 is a hydrogen atom or an optionally substituted alkyl group
  • R 4 is an optionally substituted aryl group.
  • R 1 is a group represented by the formula (h) as defined above and/or R 4 is a group represented by the formula (a) as defined above. More preferably, R 1 is a group represented by the formula (h) as defined above and/or R 4 is a group represented by the formula (c) as defined above. More preferably still, R 1 is a group represented by the formula (h) as defined above and/or R 3 is a hydrogen atom or a methyl group and/or R 4 is a group represented by the formula (c) wherein R 4a is a nitrogen-containing
  • R 4 is a hydrogen atom, a methyl group, a methoxy group or a methyl group substituted by a hydroxy group. More preferably still, R 1 is a 2-chlorophenyl group and/or R 3 is a methyl group and/or R 4 is a group re resented by the formula (i) :
  • R 5a is a hydrogen atom, a methyl group, a methoxy group or a methyl group substituted by a hydroxy group
  • R 6a is a hydrogen atom or a methyl group.
  • R 1 is a 2-chlorophenyl group and/or R 3 is a methyl group and/or R 4 is a group represented by the formula (i) as defined above, wherein R 5a is a hydrogen atom, a methyl group, a methoxy group or a methyl group substituted by a hydroxy group; and R 6a is a hydrogen atom .
  • the compound of formula (I) or formula (II) is selected from the following:
  • the compound of formula (I) or formula (III) is selected from the following:
  • Suitable pharmaceutically acceptable excipients would be known by the person skilled in the art, for example, fats, water, physiological saline, alcohol (e.g.,
  • glycerol glycerol
  • polyols aqueous glucose solution
  • extending agent disintegrating agent, binder, lubricant, wetting agent, stabilizer, emulsifier, dispersant, preservative, sweetener, colorant, seasoning agent or aromatizer, concentrating agent, diluent, buffer
  • substance, solvent or solubilizing agent, chemical for achieving storage effect salt for modifying osmotic pressure, coating agent or antioxidant, saccharides such as lactose or glucose; starch of corn,
  • fatty acids such as stearic acid
  • inorganic salts such as magnesium metasilicate aluminate or anhydrous calcium phosphate
  • synthetic polymers such as polyvinylpyrrolidone or polyalkylene glycol
  • alcohols such as stearyl alcohol or benzyl alcohol
  • synthetic cellulose derivatives such as methylcellulose
  • hydroxypropylmethylcellulose hydroxypropylmethylcellulose
  • other conventionally used additives such as gelatin, talc, plant oil and gum arabic .
  • composition comprising the compound of formula (I), or a pharmaceutically acceptable salt or N- oxide derivative thereof, and at least one
  • composition comprising the compound of formula (I) comprising one or more further
  • composition comprising the compound of formula (I) for use in therapy.
  • the compound of formula (I) for use as a medicament. In a preferred embodiment there is provided the compound of formula (I) for use in treating or preventing cancer.
  • composition comprising the compound of formula (I) for use as a medicament and/or for use in treating or preventing cancer.
  • a method of treating or preventing cancer in a human or animal patient comprising administering to a patient in need thereof an effective amount of a compound of formula ⁇ 1 ⁇ or a pharmaceutical composition comprising formula (I) .
  • the compounds of the present invention have an IC 5 o value for Wee-1 kinase of about 1 nM to about 7000 nM, more preferably from about 1 nM to about 1500 nM, or from about 1 nM to about 500 nM, or from about 1 nM to about 250 nM, or from about 1 nM to about 100 nM, or from about 1 nM to about 50 nM, or from about 1 nM to about 10 nM, most preferably less than ⁇ .
  • IC 5 o corresponds to an IC 5 o in the range of 100 nM to 10 ⁇ and '++' corresponds to an IC 5 o of ⁇ 100 nM.
  • HT29 pCDC2 activity is classified as the following: '+' corresponds to an IC 50 >500 nM and '++' corresponds to an IC 50 ⁇ 500 nM.
  • '+' corresponds to an IC 50 >4 ⁇
  • '++' corresponds to an IC 50 in the range of 1 ⁇ to 4 ⁇
  • '+++' corresponds to an IC 50 ⁇ 1 ⁇ .
  • Representative compounds (e.g. Example 2) caused dose dependent inhibition of phosphorylation of cdc2 in HT29 and HCT-116 cells, and inhibition of proliferation of
  • HT29 cells at concentrations ⁇ 10 ⁇ .
  • inhibitor MK1775 has a CLint value of 28uL/min/mg .
  • microwave experiments were carried out using a CEM DiscoverTM/Explorer24TM system controlled by Synergy 1.5 software. In other cases a Biotage InitiatorTM Eight was used. Both machines give good reproducibility and control at temperature ranges from 60-250°C and pressures of up to maximum of 20 bar.
  • Biotage KP-Sil SNAP cartridge columns (10-340 g) were used along with the stated solvent system and an appropriate solvent gradient depending on compound polarity. In the case of more polar and basic compounds, Biotage KP-NH SNAP cartridge columns (11 g) were used.
  • Method A The system consists of an Agilent Technologies 6140 single quadrupole mass spectrometer linked to an Agilent Technologies 1290 Infinity LC system with UV diode array detector and autosampler.
  • the spectrometer consists of a multimode ionization source ( electrospray and atmospheric pressure chemical ionizations) operating in positive and negative ion mode.
  • LCMS experiments were performed on each sample submitted using the following conditions: LC Column: Zorbax Eclipse Plus C18 RRHD 1.8 micron 50 x 2.1 mm maintained at 40°C. Mobile phases: A) 0.1% (v/v) formic acid in water; B) 0.1% (v/v) formic acid in acetonitrile .
  • Method B The system consisted of a ThermoFinnigan LCQ Advantage Mass Spectrometer with Surveyor LC system and 200 position autosampler. The LC system was coupled to an inline Surveyor DAD detector and ESI source operating in positive and negative ion mode. LCMS experiments were performed on each sample submitted using the following conditions: LC Column: Luna 3 micron C18 50 x 2 mm.
  • the system consisted of an Agilent Technologies 6120 single quadrupole mass spectrometer linked to an Agilent Technologies 1200 Preparative LC system with Multiple Wavelength detector and autosampler.
  • Acidic conditions LC Column: Waters XBridgeTM Prep C18 5 ⁇ OBDTM 19 x 50 mm column at RT .
  • Mobile phase A) Water 0.1% (v/v) formic acid in water; B) 0.1% (v/v) formic acid in 95:5, acetonitrile/water .
  • Total experiment time was ca. 10 min and an example method is given:
  • Example 1 8-Methyl-2- ( ( 4- ( 4-methylpiperazin-l- yl ) phenyl ) amino ) -6 -phenylpyrido [ 2 , 3-d] pyrimidin-5 ( 8H) -one
  • Step 1 Ethyl 4- ( ( 3-ethoxy-3-oxopropyl ) (methyl ) amino) -2- (methylthio ) pyrimidine-5-carboxylate : Ethyl 4-chloro-2- (methylthio ) pyrimidine-5-carboxylate (426 mg, 1.833 mmol) [commercially available] was added to a stirred solution of ethyl 3- (methylamino ) propanoate (240 mg, 1.83 mmol) [prepared according to the literature procedure: J. Org. Chem., 1985, 50, p3979-3982; J. Med. Chem., 2005, 48, p4100-4110] in THF (5.0 mL) at RT under nitrogen.
  • Step 2 Ethyl 5-hydroxy-8-methyl-2- (methylthio) -7, 8- dihydropyrido [ 2 , 3-d] pyrimidine-6 -carboxylate : Potassium tert-butoxide solution (1.0 M in THF) (0.767 mL, 0.767 mmol) was added to a stirred solution of ethyl 4-((3- ethoxy-3-oxopropyl ) (methyl) amino) -2-
  • Step 3 Ethyl 8-methyl-2- (methylthio ) -5-oxo-5 , 8- dihydropyrido [ 2 , 3-d] pyrimidine-6-carboxylate : Bromine
  • Step 4 8-Methyl-2- (methylthio ) -5-oxo-5 , 8- dihydropyrido [ 2 , 3-d] pyrimidine-6-carboxylic acid: 1M HC1 (aq) solution (17.4 mL, 17.4 mmol) was added to a stirred solution of ethyl 8-methyl-2- (methylthio) -5-oxo-5, 8- dihydropyrido [ 2 , 3-d] pyrimidine-6-carboxylate (1.62 g, 5.79 mmol) in 1,4-dioxane (16.0 mL) at RT under nitrogen. The temperature was increased to 95 °C.
  • Step 6 8-Methyl-2- (methylthio) -6 -phenylpyrido [2, 3- d] pyrimidin-5 ( 8H) -one : PdCl 2 (dppf ) -DCM adduct (10.4 mg, 0.013 mmol) was added to a pre-degassed solution of 6- iodo-8-methyl-2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) - one (42.3 mg, 0.127 mmol), phenylboronic acid (20.1 mg, 0.165 mmol) and sodium carbonate (40.4 mg, 0.381 mmol) in DMF (2.0 mL) in a 10 mL vial.
  • Step 7 8-Methyl-2- ( ( 4- ( 4-methylpiperazin-l- yl ) phenyl ) amino ) -6 -phenylpyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) - one: mCPBA ( ⁇ 77% pure) (17.3 mg, 0.077 mmol) in DCM (1.0 mL) was added to a stirred solution of 8-methyl-2- (methylthio ) -6 -phenylpyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one
  • Step 1 6- ( 2 , 6-Dichlorophenyl ) -8-methyl-2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one : Pd2 (dba ) 3 (4.2 mg, 4.6 pmol) was added to a pre-degassed solution of 6 -iodo-8-methyl-2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin- 5(8H)-one (153 mg, 0.460 mmol), (2,6- dichlorophenyl ) boronic acid (114 mg, 0.599 mmol), SPhos (7.6 mg, 0.018 mmol) and potassium phosphate, tribasic (293 mg, 1.38 mmol) in a, , -trifluorotoluene (2.0 mL) in a 10 mL vial. The vessel was sealed and the reaction mixture was stirred at 80 °C. After 16 h
  • Step 1 6- ( 2-Chlorophenyl ) -8-methyl-2-
  • Step 2 6- ( 2-Chlorophenyl ) -8-methyl-2- ( ( 4- ( 4- methylpiperaz in-1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5(8H)-one: mCPBA ( ⁇ 77% pure) (16.4 mg, 0.073 mmol) in DCM (0.5 mL) was added to a stirred solution of 6-(2- chlorophenyl ) -8-methyl-2- (methylthio ) pyrido [2,3- d] pyrimidin-5 ( 8H) -one (20.1 mg, 0.063 mmol) in DCM (2.0 mL) at 0 °C under nitrogen.
  • Step 1 6- ( 2-Methoxyphenyl ) -8-methyl-2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one : Pd 2 (dba)3 (3.3 mg, 3.6 pmol) was added to a pre-degassed solution of 6 -iodo-8-methyl-2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin- 5(8H)-one (30.0 mg, 0.090 mmol), ( 2-methoxyphenyl ) boronic acid (41.1 mg, 0.270 mmol), SPhos (3.0 mg, 7.2 pmol) and potassium phosphate, tribasic (57.3 mg, 0.270 mmol) in toluene (1.0 mL) in a 4 mL vial.
  • Step 2 6- ( 2-Methoxyphenyl ) -8-methyl-2- (( 4- ( 4- methylpiperaz in-1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5(8H)-one: mCPBA ( ⁇ 77% pure) (16.8 mg, 0.075 mmol) in DCM (0.5 mL) was added to a stirred solution of 6-(2- methoxyphenyl ) -8-methyl-2- (methylthio ) pyrido [2,3- d] pyrimidin-5 ( 8H) -one (20.3 mg, 0.065 mmol) in DCM (2.0 mL) at 0 °C under nitrogen.
  • Step 1 8-Methyl-2- (methylthio) -6- (o-tolyl) pyrido [2, 3- d] pyrimidin-5 ( 8H) -one : Pd 2 (dba)3 (3.3 mg, 3.6 pmol) was added to a pre-degassed solution of 6-iodo-8-methyl-2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one (30.0 mg, 0.090 mmol), o-tolylboronic acid (36.7 mg, 0.270 mmol), SPhos (3.0 mg, 7.2 pmol) and potassium phosphate, tribasic (57.3 mg, 0.270 mmol) in toluene (1.0 mL) in a 4 mL vial.
  • Step 2 8-Methyl-2- (methylsulfonyl ) -6- ( o- tolyl ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H) -one : mCPBA ( ⁇ 77% pure) (49.0 mg, 0.219 mmol) was added to a stirred solution of 8-methyl-2- (methylthio) -6- (o-tolyl) pyrido [2,3- d] pyrimidin-5 ( 8H) -one (16.9 mg, 0.057 mmol) in DCM (1.5 mL) at RT under nitrogen. After 16 h, the reaction mixture was partitioned between DCM and sodium
  • Step 3 8-Methyl-2- ( ( 4- ( 4-methylpiperazin-l- yl ) phenyl ) amino ) -6- (o-tolyl) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) - one: TFA (0.021 mL, 0.272 mmol) was added to a stirred solution of 8-methyl-2- (methylsulfonyl ) -6- ( o- tolyl ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H) -one (17.9 mg, 0.054 mmol) and 4- ( 4-methylpiperazin-l-yl ) aniline (11.4 mg, 0.060 mmol) in 2-propanol (1.0 mL) in a 10 mL vial.
  • the vessel was sealed and irradiated at 120 °C for 20 min (CEM Discover/Explorer24 ) .
  • LCMS analysis showed residual starting material.
  • the reaction was rerun under the same conditions.
  • LCMS analysis showed further conversion, but still some starting material.
  • the reaction was rerun at 150 °C for 20 min.
  • the solvents were removed in vacuo and the remaining residue was partitioned between
  • Example 6 6- ( 2 , 4-Dichlorophenyl ) -8-methyl-2- (( 4- ( 4- methylpiperaz in-1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5 ( 8H) -one
  • Step 1 6- ( 2 , 4-Dichlorophenyl ) -8-methyl-2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one : Pd 2 (dba ) 3 (3.3 mg, 3.6 pmol) was added to a pre-degassed solution of 6 -iodo-8-methyl-2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin- 5(8H)-one (30.0 mg, 0.090 mmol), (2,4- dichlorophenyl ) boronic acid (172 mg, 0.900 mmol), SPhos (3.0 mg, 7.2 pmol) and potassium phosphate, tribasic (57.3 mg, 0.270 mmol) in toluene (1.0 mL) in a 4 mL vial.
  • Step 2 6- ( 2 , 4-Dichlorophenyl ) -8-methyl-2- ( ( 4- ( 4- methylpiperaz in-1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5(8H)-one: mCPBA ( ⁇ 77% pure) (19.0 mg, 0.085 mmol) in DCM (0.5 mL) was added to a stirred solution of 6-(2,4- dichlorophenyl ) -8-methyl-2- (methylthio ) pyrido [2,3- d] pyrimidin-5 ( 8H) -one (22.1 mg, 0.063 mmol) in DCM (2.0 mL) at 0 °C under nitrogen. After 30 min, the
  • Example 7 6- ( 2 , 5-Dichlorophenyl ) -8-methyl-2- ( ( 4- ( 4- methylpiperaz in-1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5 ( 8H) -one
  • Step 1 6- ( 2 , 5-Dichlorophenyl ) -8-methyl-2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one : Pd 2 (dba ) 3 (3.3 mg, 3.6 pmol) was added to a pre-degassed solution of 6 -iodo-8-methyl-2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin- 5(8H)-one (30.0 mg, 0.090 mmol), (2,5- dichlorophenyl ) boronic acid (172 mg, 0.900 mmol), SPhos (3.0 mg, 7.2 pmol) and potassium phosphate, tribasic (57.3 mg, 0.270 mmol) in toluene (1.0 mL) in a 4 mL vial.
  • Step 2 6- ( 2 , 5-Dichlorophenyl ) -8-methyl-2- (( 4- ( 4- methylpiperaz in-1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5(8H)-one: mCPBA ( ⁇ 77% pure) (25.0 mg, 0.112 mmol) in DCM (0.5 mL) was added to a stirred solution of 6-(2,5- dichlorophenyl ) -8-methyl-2- (methylthio ) pyrido [2,3- d]pyrimidin-5 (8H) -one (29.1 mg, 0.083 mmol) in DCM (2.0 mL) at 0 °C under nitrogen.
  • Example 8 6- ( 2 , 3-Dichlorophenyl ) -8-methyl-2- ( ( 4- ( 4- methylpiperaz in-1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin-
  • Step 1 6- ( 2 , 3-Dichlorophenyl ) -8-methyl-2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one : Pd 2 (dba)3 (3.3 mg, 3.6 pmol) was added to a pre-degassed solution of 6 -iodo-8-methyl-2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin- 5(8H)-one (30.0 mg, 0.090 mmol), (2,3- dichlorophenyl ) boronic acid (172 mg, 0.900 mmol), SPhos (3.0 mg, 7.2 pmol) and potassium phosphate, tribasic
  • Step 2 6- ( 2 , 3-Dichlorophenyl ) -8-methyl-2- (( 4- ( 4- methylpiperaz in-1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5(8H)-one: mCPBA ( ⁇ 77% pure) (20.4 mg, 0.091 mmol) in DCM (0.5 mL) was added to a stirred solution of 6-(2,3- dichlorophenyl ) -8-methyl-2- (methylthio ) pyrido [2,3- d] pyrimidin-5 ( 8H) -one (20.8 mg, 0.059 mmol) in DCM (2.0 mL) at 0 °C under nitrogen.
  • Step 1 8-Methyl-2- (methylthio) -6- (thiophen-3- yl) pyrido [2, 3-d] pyrimidin-5 ( 8H) -one : Pd 2 (dba)3 (3.3 mg,
  • Step 2 8-Methyl-2- ( ( 4- ( 4-methylpiperazin-l- yl ) phenyl ) amino ) -6 - ( thiophen-3-yl ) pyrido [ 2 , 3-d] pyrimidin- 5(8H)-one: mCPBA ( ⁇ 77% pure) (21.0 mg, 0.094 mmol) was added to a stirred solution of 8-methyl-2- (methylthio) -6- (thiophen-3-yl ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H) -one (17.6 mg, 0.061 mmol) in toluene (2.0 mL) at RT under nitrogen.
  • Example 10 8-Methyl-6- ( 5-methyl-l , 3 , 4-oxadiazol-2-yl ) -2- ( ( 4- ( piperaz in- 1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5 (8H) -one
  • Step 1 Ethyl 2 , 4-dichloropyrimidine-5-carboxylate : Ethyl 4-chloro-2- (methylthio ) pyrimidine-5-carboxylate (50.0 g, 215 mmol) in sulfuryl chloride (50.0 mL, 615 mmol) was heated at 50 °C under an atmosphere of nitrogen. After 2 hours, the reaction mixture was cooled and carefully poured onto saturated sodium bicarbonate (aq) solution and was extracted using DCM (x 3) . The combined organic phase was dried (Phase Separator) and the solvents were removed in vacuo to give title compound (crude, ca. 50 g) that carried through to the next step without further purification.
  • ⁇ NMR (500 MHz, CDCI 3 ) ⁇ 9.02 (s, 1H) , 4.46 (dd, 2H) , 1.43 (t, 3H) .
  • Step 2 Ethyl 2-chloro-4- ( ( 3-ethoxy-3- oxopropyl) (methyl ) amino ) pyrimidine-5-carboxylate : Ethyl 3- (methylamino ) propanoate (17.2 g, 131 mmol) in DCM (100 mL) was added to a stirred solution of ethyl 2,4- dichloropyrimidine-5-carboxylate (crude, 25.2 g, 114 mmol) and DIPEA (43.8 mL, 251 mmol) in DCM (100 mL) in a 500 mL RB flask at 0 °C under nitrogen.
  • Step 3 Ethyl 2-chloro-5-hydroxy-8-methyl-7 , 8- dihydropyrido [ 2 , 3-d] pyrimidine-6-carboxylate : Potassium tert-butoxide (1 M in THF) solution (114 mL, 114 mmol) was added to a stirred solution of ethyl 2-chloro-4- ( ( 3- ethoxy-3-oxopropyl ) (methyl ) amino ) pyrimidine-5-carboxylate (36.0 g, 114 mmol) in toluene (360 mL) at RT under nitrogen.
  • Step 4 Ethyl 2-chloro-8-methyl-5-oxo-5 , 8- dihydropyrido [ 2 , 3-d] pyrimidine-6 -carboxylate : Bromine (5.87 mL, 114 mmol) in chloroform (200 mL) was added to a stirred solution of ethyl 2-chloro-5-hydroxy-8-methyl- 7 , 8-dihydropyrido [ 2 , 3-d] pyrimidine-6 -carboxylate (30.7 g, 114 mmol) and triethylamine (47.7 mL, 342 mmol) in chloroform (400 mL) at 0 °C under nitrogen.
  • Step 5 Ethyl 2- ( ( 4- ( 4- ( tert-butoxycarbonyl ) piperaz in-1- yl ) phenyl ) amino ) -8-methyl-5-oxo-5 , 8-dihydropyrido [2,3- d] pyrimidine-6 -carboxylate : tert-Butyl 4- (4- aminophenyl ) piperazine-l-carboxylate (1.12 g, 4.03 mmol) was added to a stirred solution of ethyl 2-chloro-8- methyl-5-oxo-5 , 8-dihydropyrido [ 2 , 3-d] pyrimidine-6 - carboxylate (1.08 g, 4.03 mmol) in DMF (10.0 mL) in a 50 mL RB flask.
  • Step 6 Sodium 2- (( 4- ( 4- (tert-butoxycarbonyl) piperazin-1- yl ) phenyl ) amino ) -8-methyl-5-oxo-5 , 8-dihydropyrido [2,3- d] pyrimidine-6 -carboxylate : 2 M sodium hydroxide (aq) solution (4.92 mL, 9.83 mmol) was added to a stirred solution of ethyl 2- (( 4- ( 4- ( tert- butoxycarbonyl ) piperaz in-1-yl ) phenyl ) amino ) -8-methy1-5- oxo-5 , 8-dihydropyrido [ 2 , 3-d] pyrimidine-6 -carboxylate (1.0 g, 1.966 mmol) in 1,4-dioxane (5.0 mL) in a 50 mL RB flask. The reaction mixture was heated at 50 °C. After 16
  • Step 7 8-Methyl-6- ( 5-methyl-l , 3 , 4-oxadiazol-2-yl ) -2- ( ( 4- (piperaz in-1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5(8H)-one: Acetohydrazide (8.1 mg, 0.109 mmol) was added to sodium 2- (( 4- ( 4- (tert-butoxycarbonyl) piperazin-1- yl ) phenyl ) amino ) -8-methyl-5-oxo-5 , 8-dihydropyrido [2,3- d] pyrimidine-6 -carboxylate (50.0 mg, 0.100 mmol) in POCI 3 (1.0 mL, 10.7 mmol) at RT in a 10 mL vial. The vessel was sealed and the temperature was increased to 100 °C with stirring. After 2 h, the reaction mixture was poured onto ice water, neutralised using 2 M Na
  • Step 1 6- ( 3-Chloropyridin-2-yl ) -8-methyl-2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one :
  • Example 12 6- ( 2 , 6 -Dichlorophenyl ) -8-methyl-2- ( ( 4- (piperaz in-1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5 ( 8H) -one :
  • Step 1 tert-Butyl 4- ( 4- ( ( 6-iodo-8-methyl-5-oxo-5, 8- dihydropyrido [ 2 , 3-d] pyrimidin-2- yl ) amino ) phenyl ) piperaz ine-l-carboxylate : To a stirred solution of 6 -iodo-8-methyl-2- (methylthio ) pyrido [ 2 , 3- d] pyrimidin-5 ( 8H) -one (100 mg, 0.30 mmol) in toluene (2 mL) at 0 °C under nitrogen was added mCPBA ( ⁇ 77% pure) (78 mg, 0.35 mmol) in DCM (2 mL).
  • Step 2 tert-Butyl 4- ( 4- (( 6- ( 2 , 6-dichlorophenyl ) -8- methyl-5-oxo-5 , 8-dihydropyrido [ 2 , 3-d] pyrimidin-2- yl ) amino ) phenyl ) piperaz ine-l-carboxylate : Pd 2 (dba)3 (3.9 mg, 3.91 pmol) was added to a pre-degassed solution of 6- iodo-8-methyl-2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) - one (44 mg, 0.078 mmol), ( 2 , 6-dichlorophenyl ) boronic acid (149 mg, 0.782 mmol), SPhos (2.6 mg, 6.26 pmol) and potassium phosphate, tribasic (50 mg, 0.235 mmol) in a, , -trifluorotoluene (
  • Step 3 6- ( 2 , 6-Dichlorophenyl ) -8-methyl-2- ( ( 4- (piperazin 1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one : To a solution of tert-butyl 4- ( 4- (( 6- ( 2 , 6-dichlorophenyl ) -8- methyl-5-oxo-5 , 8-dihydropyrido [ 2 , 3-d] pyrimidin-2- yl ) amino ) phenyl ) piperaz ine-l-carboxylate (20 mg, 0.034 mmol) in DCM (2 mL) at 0 °C was added TFA (1 mL, 12.98 mmol) .
  • Example 13 8-Allyl-6- ( 2 , 6 -dichlorophenyl ) -2- ( ( 4- ( 4- methylpiperaz in-1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5 ( 8H) -one
  • Step 1 Ethyl 3- ( allylamino ) propanoate : Prop-2-en-l-amine (15.1 mL, 202 mmol) was added dropwise to a solution of ethyl acrylate (20 mL, 184 mmol) in EtOH (100 mL) and the resulting solution was stirred for 18 h at RT . The reaction mixture was concentrated in vacuo to afford the title compound (28 g, 97%) which was used without further purification.
  • Step 2 Ethyl 4- ( allyl ( 3-ethoxy-3-oxopropyl ) amino ) -2- (methylthio ) pyrimidine-5-carboxylate : Following the procedure for ethyl 4- ( ( 3-ethoxy-3- oxopropyl ) (methyl ) amino ) -2- (methylthio ) pyrimidine-5- carboxylate, ethyl 3- ( allylamino ) propanoate (28 g, 178 mmol) was reacted to afford the title compound as a yellow oil which was used without further purification (61 g, 97%).
  • Step 3 Ethyl 8-allyl-5-hydroxy-2- (methylthio ) -7 , 8- dihydropyrido [ 2 , 3-d] pyrimidine-6 -carboxylate : Following the procedure for ethyl 5-hydroxy-8-methyl-2-
  • Step 4 Ethyl 8-allyl-2- (methylthio ) -5-oxo-5 , 8- dihydropyrido [ 2 , 3-d] pyrimidine-6 -carboxylate : Following the procedure for ethyl 8-methyl-2- (methylthio ) -5-oxo- 5 , 8-dihydropyrido [ 2 , 3-d] pyrimidine-6 -carboxylate , ethyl 8-allyl-5-hydroxy-2- (methylthio ) -7 , 8-dihydropyrido [2,3- d] pyrimidine-6 -carboxylate (20.0 g, 65 mmol) was reacted to afford the title compound (19 g, 96%) as a yellow solid which was used without further purification.
  • Step 5 8-Allyl-2- (methylthio ) -5-oxo-5 , 8- dihydropyrido [ 2 , 3-d] pyrimidine-6 -carboxylie acid :
  • Step 6 8-Allyl-6- ( 2 , 6 -dichlorophenyl ) -2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one : To a solution of 8-allyl-2- (methylthio ) -5-OXO-5 , 8- dihydropyrido [ 2 , 3-d] pyrimidine-6-carboxylic acid (50 mg, 0.18 mmol) in toluene (2 mL) and DMA (0.22 mL) was added 1 , 3-dichloro-2-iodobenzene (98 mg, 0.36 mmol) and silver carbonate (49.7 mg, 0.18 mmol) .
  • the suspension was degassed for 5 min and (oxybis(2,l- phenylene) ) bis (diphenylphosphine) (9.7 mg, 0.018 mmol) and palladium (II) chloride (1.6 mg, 9.0 pmol) added.
  • the reaction mixture was then heated in a microwave
  • Step 7 8-Allyl-6- ( 2 , 6 -dichlorophenyl ) -2- ( ( 4- ( 4- methylpiperaz in-1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5(8H)-one: Following the procedure for 8-methyl-2- ( ( 4- ( 4- methylpiperaz in-1-yl ) phenyl ) amino ) -6 -phenylpyrido [2,3- d] pyrimidin-5 ( 8H) -one, 8-allyl-6- (2, 6 -dichlorophenyl ) -2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one (50.0 mg, 0.13 mmol) was reacted to give crude material which was purified by flash chromatography using a KP-NH column (0- 100% EtOAc in cyclohexane) to afford the title compound (4.5
  • Step 1 Ethyl 3- (( 4-methoxybenzyl ) amino ) propanoate :
  • Step 2 Ethyl 4- ( ( 3-ethoxy-3-oxopropyl ) (4- methoxybenzyl ) amino ) -2- (methylthio ) pyrimidine-5- carboxylate :
  • Step 3 Ethyl 5-hydroxy-8- ( 4-methoxybenzyl ) -2- (methylthio ) -7 , 8-dihydropyrido [ 2 , 3-d] pyrimidine-6 - carboxylate: Following the procedure for ethyl 5-hydroxy- 8-methyl-2- (methylthio) -7, 8-dihydropyrido [2,3- d] pyrimidine-6 -carboxylate , ethyl 4- ( ( 3-ethoxy-3- oxopropyl ) ( 4-methoxybenzyl ) amino ) -2-
  • Step 4 Ethyl 8- ( 4-methoxybenzyl ) -2- (methylthio ) -5-oxo- 5, 8-dihydropyrido [ 2 , 3-d] pyrimidine-6 -carboxylate :
  • Step 5 8- ( 4-Methoxybenzyl ) -2- (methylthio ) -5-oxo-5 , 8- dihydropyrido [ 2 , 3-d] pyrimidine-6 -carboxylie acid :
  • Step 7 6- ( 2 , 6-Dichlorophenyl ) -8- ( 4-methoxybenzyl ) -2- (( 4- ( 4-methylpiperaz in-1-yl ) phenyl ) amino ) pyrido [2,3- d] pyrimidin-5 ( 8H ) -one :
  • Example 16 6- ( 2 , 6 -Dichlorophenyl ) -8-ethyl-2- ( ( 4- ( 4- methylpiperaz in- 1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5 (8H) -one
  • Example 17 Ethyl 2- ( 6- ( 2 , 6 -dichlorophenyl ) -2- ( ( 4- ( 4- methylpiperaz in-l-yl ) phenyl ) amino ) -5-oxopyrido [2,3- d] pyrimidin-8 (5H -yl) acetate
  • Example 19 6- ( 2 , 6 -Dichlorophenyl ) -8- ( 3-hydroxypropyl ) -2- ( ( 4- ( 4-methylpiperaz in-l-yl ) phenyl ) amino ) pyrido [2,3- d] pyrimidin-5 ( 8H -one
  • Example 20 6- ( 2 , 5-Dimethyl-lH-pyrrol-l-yl ) -8- ( 4- methoxybenzyl ) -2- ( ( 4- ( 4-methylpiperazin-l-
  • Step 1 tert-Butyl ( 8- ( 4-methoxybenzyl ) -2- (methylthio) -5- oxo-5 , 8-dihydropyrido [ 2 , 3-d] pyrimidin-6 -yl ) carbamate : To a yellow solution of 8- ( 4-methoxybenzyl ) -2- (methylthio ) - 5-oxo-5, 8-dihydropyrido [ 2 , 3-d] pyrimidine-6 -carboxy1 ic acid (0.5 g, 1.40 mmol) in tert-butanol (10 mL) was added DIPEA (0.244 mL, 1.40 mmol) followed by diphenyl
  • Step 2 tert-Butyl ( 8- ( 4-methoxybenzyl ) -2- (( 4- ( 4- methylpiperaz in-l-yl ) phenyl ) amino ) -5-oxo-5, 8- dihydropyrido [ 2 , 3-d] pyrimidin-6-yl ) carbamate : Following the procedure for 8-methyl-2- ( ( 4- ( 4-methylpiperazin-l- yl ) phenyl ) amino ) -6 -phenylpyrido [ 2 , 3-d] pyrimidin-5 ( 8H) - one, tert-butyl ( 8- ( 4-methoxybenzyl ) -2- (methylthio ) -5- oxo-5 , 8-dihydropyrido [ 2 , 3-d] pyrimidin-6 -yl ) carbamate (0.084 mg, 0.19 mmol) was reacted to give the title compound (74 mg, 67%) as
  • Step 3 6-Amino-8- ( 4-methoxybenzyl ) -2- (( 4- ( 4- methylpiperaz in-l-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5(8H)-one: To a solution of tert-butyl (8-(4- methoxybenzyl ) -2- ( ( 4- ( 4-methylpiperaz in-1- yl ) phenyl ) amino ) -5-oxo-5, 8-dihydropyrido [ 2 , 3-d] pyrimidin- 6 -yl ) carbamate (74 mg, 0.131 mmol) in DCM (1 mL) was added TFA (1 mL) .
  • Step 4 6- ( 2 , 5-Dimethyl-lH-pyrrol-l-yl ) -8- ( 4- methoxybenzyl ) -2- ( ( 4- ( 4-methylpiperaz in-l- yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one : To a solution of 6-amino-8- ( 4-methoxybenzyl ) -2- (( 4- ( 4- methylpiperaz in-1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5(8H)-one (0.05 g, 0.106 mmol) in DMF (1 mL) was added 4- methylbenzenesulfonic acid (0.027 g, 0.159 mmol) and hexane-2 , 5-dione (0.012 g, 0.106 mmol) . The resulting mixture was heated for 30 min at 120° C under microwave conditions,
  • Example 21 6- ( 2 , 6 -Dichlorophenyl ) -2- ( ( 6- (piperazin-1- yl ) pyridin-3-yl ) amino pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one
  • Step 1 6- ( 2 , 6-Dichlorophenyl ) -8- ( 4-methoxybenzyl ) -2- ( ( 6- (piperaz in-1-yl ) pyridin-3-yl ) amino ) pyrido [2,3- d] pyrimidin-5 ( 8H) -one :
  • Step 2 6- ( 2 , 6-Dichlorophenyl ) -2- ( ( 6- (piperazin-1- yl ) pyridin-3-yl ) amino ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one : Following the procedure for 6- ( 2 , 6 -dichlorophenyl ) -2- ( ( 4- ( 4-methylpiperaz in-l-yl ) phenyl ) amino ) pyrido [2,3- d] pyrimidin-5 ( 8H) -one, 6- ( 2 , 6 -dichlorophenyl ) -8- ( 4- methoxybenzyl ) -2- ( (6- (piperaz in-l-yl ) pyridin-3- yl ) amino ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one was reacted to give crude material which was purified by preparative HPLC to afford the
  • Example 22 6- ( 2 , 6-Dichlorophenyl ) -2- (( 4- ( 2- (diethylamino ) ethoxy ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5 (8H) -one
  • Step 1 6- (2, 6-Dichlorophenyl) -2- ( (4- (2- ( diethylamino ) ethoxy ) phenyl ) amino ) -8- ( 4- methoxybenzyl ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H) -one : Following the procedure for 8-methyl-2- ( ( 4- ( 4-methylpiperazin-l- yl ) phenyl ) amino ) -6 -phenylpyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) - one, 6- ( 2 , 6 -dichlorophenyl ) -8- ( 4-methoxybenzyl ) -2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one (55 mg, 0.116 mmol) was reacted with 4- (2- (2-
  • Step 2 6- ( 2 , 6-Dichlorophenyl ) -2- (( 4- ( 2-
  • Step 1 6- ( 2 , 6-Dichlorophenyl ) -8- ( 4-methoxybenzyl ) -2- ( (pyridin-4-ylmethyl ) amino ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H )
  • Step 2 6- ( 2 , 6-Dichlorophenyl ) -2- ( (pyridin-4- ylmethyl ) amino ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one :
  • Example 24 6- ( 2 , 6-Dichlorophenyl ) -2- (( 1-methylpiperidin- 4-yl ) amino ) pyrido [ 2 3-d] pyrimidin-5 ( 8H ) -one
  • Step 1 6- ( 2 , 6-Dichlorophenyl ) -8- ( 4-methoxybenzyl ) -2- (( 1 methylpiperidin-4-yl ) amino ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H) - one: Following the procedure for 8-methyl-2- ( ( 4- ( 4- methylpiperaz in-l-yl ) phenyl ) amino ) -6 -phenylpyrido [2,3- d] pyrimidin-5 ( 8H) -one, 6- ( 2 , 6 -dichlorophenyl ) -8- ( 4- methoxybenzyl ) -2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin- 5(8H)-one (55 mg, 0.116 mmol) was reacted with 1- methylpiperidin-4-amine (13.2 mg, 0.116 mmol) instead of 4- ( 4-methylpiperazin-l-yl )
  • Step 2 6- ( 2 , 6-dichlorophenyl ) -2- (( l-methylpiperidin-4- yl ) amino ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one : Following the procedure for 6- ( 2 , 6-dichlorophenyl ) -2- (( 4- ( 4- methylpiperaz in- 1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5 ( 8H) -one, 6- ( 2 , 6-dichlorophenyl ) -8- ( 4-methoxybenzy1 ) -2- ( ( l-methylpiperidin-4-yl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5(8H)-one was reacted to give crude material which was purified by HPLC to afford the title compound (5.1 mg, 17%) .
  • Example 25 7-Amino-6- ( 2-chlorophenyl ) -8-methyl-2- ( ( 4- ( 4- methylpiperaz in- 1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5 (8H) -one
  • Step 1 Ethyl 4- (methylamino ) -2- (methylthio ) pyrimidine-5 carboxylate: To a solution of ethyl 4-chloro-2- (methylthio ) pyrimidine-5-carboxylate (10 g, 43 mmol) in anhydrous THF (100 mL) was added triethylamine (6.0 mL, 43 mmol) and monomethylamine (2M in THF, 22 mL, 43 mmol) The reaction mixture was heated to 50 °C overnight, then diluted with water (100 mL) and extracted into ethyl acetate (3 x 100 mL) .
  • Step 2 4- (Methylamino ) -2- (methylthio ) pyrimidine-5- carboxylic acid: Lithium hydroxide (211 mg, 8.8 mmol) was added to ethyl 4- (methylamino ) -2- (methylthio ) pyrimidine- 5-carboxylate (1.0 g, 4.4 mmol) in a mixture of methanol (8 mL) and water (4 mL) to give a white suspension. This was heated to 60 °C for 3 h and then concentrated to dryness under reduced pressure. The residue was re- dissolved in water (25 mL) and acidified with acetic acid to pH 5. The precipitated solid was isolated by
  • Step 3 4- (Methylamino ) -2- (methylthio ) pyrimidine-5- carbonyl fluoride: To a solution of 4- (methylamino ) -2- (methylthio ) pyrimidine-5-carboxylic acid (610 mg, 3.1 mmol) and triethylamine (470 pL, 3.4 mmol) in anhydrous dichloromethane (6 mL) was added dropwise a solution of cyanuric fluoride (525 pL, 6.1 mmol) in anhydrous
  • Step 4 7-Amino-6- ( 2-chlorophenyl ) -8-methyl-2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one : A solution of 2- ( 2-chlorophenyl ) acetonitrile (34 mg, 0.22 mmol) in anhydrous DMF (1 mL) was cooled to 0 °C followed by the addition of sodium hydride (60% in mineral oil, 18 mg, 0.45 mmol) . The resulting mixture was stirred at RT for 10 min before re-cooling to 0 °C.
  • the reaction mixture was heated to 100 °C for 60 min then allowed to cool to RT, diluted with water (5 mL) and extracted into ethyl acetate (3 x 5 mL) .
  • the combined organic phases were washed with 1:1 water/brine (3 x 5 mL) , dried over Na 2 SC>4, filtered, and concentrated to dryness under reduced pressure.
  • the residue was purified by preparative HPLC to give the title compound (1.0 mg, 3%) as a pale yellow solid.
  • Example 26 6- ( 2-Chlorophenyl ) -7 , 8-dimethyl-2- ( ( 4- ( 4- methylpiperaz in-1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5 ( 8H) -one
  • Step 1 2- ( 2-Chlorophenyl ) -1- ( 4- (methylamino ) -2- (methylthio ) pyrimidin-5-yl ) buta-1 , 3-diene-l , 3-diol :
  • a solution of 1- ( 2-chlorophenyl ) propan-2-one (170 mg, 0.99 mmol) in anhydrous DMF (2 mL) was cooled to 0 °C followed by the addition of sodium hydride (60% in mineral oil, 80 mg, 1.99 mmol) . The resulting mixture was stirred at RT for 10 min before re-cooling to 0 °C.
  • Step 2 6- ( 2-Chlorophenyl ) -7 , 8-dimethyl-2-
  • Step 3 6- ( 2-Chlorophenyl ) -7 , 8-dimethyl-2- ( ( 4- ( 4- methylpiperaz in-1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5(8H)-one: A suspension of 6 - ( 2-chlorophenyl ) -7 , 8- dimethyl-2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one (80 mg, 0.24 mmol) in anhydrous DCM (1.5 mL) was cooled to 0 °C.
  • Step 1 2-Chloro-6- ( 2-chlorophenyl ) -7-methyl-5H- pyrano [ 2 , 3-d] pyrimidin-5-one :
  • a solution of l-(2- chlorophenyl ) propan-2-one (80 mg, 0.47 mmol) in anhydrous THF (1 mL) was cooled to 0 °C followed by the dropwise addition of tert-butylmagnesium chloride (2 M in diethyl ether, 0.24 mL, 0.47 mmol) and stirred at RT for 10 min.
  • Step 2 tert-Butyl 4- ( 4- (( 6- ( 2-chlorophenyl ) -7-methyl-5- oxo-5H-pyrano [ 2 , 3-d] pyrimidin-2- yl ) amino ) phenyl ) piperaz ine-l-carboxylate : A solution of 2-chloro-6- ( 2-chlorophenyl ) -7-methyl-5H-pyrano [2,3- d] pyrimidin-5-one (45 mg, 0.15 mmol), tert-butyl 4- (4- aminophenyl ) piperazine-l-carboxylate (41 mg, 0.15 mmol) and DIPEA (51 pL, 0.29 mmol) in anhydrous DMF (1 mL ) was heated to 100 °C under a nitrogen atmosphere for 60 min.
  • reaction mixture was allowed to cool to RT, diluted with water (5 mL) and extracted into ethyl acetate (3 x 5 mL) .
  • the combined organic phases were washed with 1:1 water/brine (3 x 5 mL) , dried over Na 2 SC>4, filtered, and concentrated to dryness under reduced pressure to give the title compound (42 mg, 52%) as a yellow solid.
  • Step 3 6- ( 2-Chlorophenyl ) -7-methyl-2- ( ( 4- (piperazin-1- yl ) phenyl ) amino ) -5H-pyrano [ 2 , 3-d] pyrimidin-5-one , 2HC1 : To a solution of tert-butyl 4- ( 4- (( 6- ( 2-chlorophenyl ) -7- methyl-5-oxo-5H-pyrano [ 2 , 3-d] pyrimidin-2- yl ) amino ) phenyl ) piperaz ine-l-carboxylate (42 mg, 0.077 mmol) in dichloromethane (1 mL) was added dropwise 4 M HC1 in 1,4-dioxane (1 mL) and the mixture stirred at RT for 90 min. The reaction mixture was then diluted with diethyl ether (5 mL) and slurried at RT for 15 min, before removal of the supernatant solvent by pipet
  • Example 28 6- ( 2-Chlorophenyl ) -2- (( 4- (piperazin-1- yl ) phenyl ) amino ) -5H-pyrano [ 2 , 3-d] pyrimidin-5-one .
  • Step 1 2- ( 2-Chlorophenyl ) acetaldehyde : A solution of methyl 2- ( 2-chlorophenyl ) acetate (3.0 g, 16 mmol) in anhydrous dichloromethane (30 mL) was cooled to -78 °C followed by the dropwise addition of DIBAL-H (1 M in toluene, 16.3 mL, 16 mmol) over a period of 30 min. The reaction mixture was stirred at -78 °C under a nitrogen atmosphere for a further 30 min, then quenched by the addition of methanol (15 mL) and poured into saturated sodium potassium tartrate (aq) solution (100 mL) .
  • Step 2 2-Chloro-6- ( 2-chlorophenyl ) -5H-pyrano [ 2 , 3- d] pyrimidin-5-one :
  • a solution of 2- (2- chlorophenyl ) acetaldehyde (730 mg, 4.7 mmol) in anhydrous THF (6 mL) was cooled to 0 °C followed by the dropwise addition of tert-butylmagnesium chloride (2 M in diethyl ether, 4.7 mL, 9.5 mmol) and stirring at RT for 10 min.
  • Step 3 tert-Butyl 4- ( 4- (( 6- ( 2-chlorophenyl ) -5-oxo-5H- pyrano [ 2 , 3-d] pyrimidin-2-yl ) amino ) phenyl ) piperaz ine-1- carboxylate: A solution of 2-chloro-6- ( 2-chlorophenyl) - 5H-pyrano [ 2 , 3-d] pyrimidin-5-one (42 mg, 0.14 mmol), tert- butyl 4- ( 4-aminophenyl ) piperazine-l-carboxylate (40 mg, 0.14 mmol) and DIPEA (50 pL, 0.29 mmol) in anhydrous DMF (1 mL) was heated to 100 °C under a nitrogen atmosphere for 60 min.
  • reaction mixture was allowed to cool to RT, diluted with water (5 mL) and extracted into ethyl acetate (3 x 5 mL) .
  • the combined organic phases were washed with 1:1 water/brine (3 x 5 mL) , dried over Na 2 S0 4 , filtered, and concentrated to dryness under reduced pressure. The residue was purified by flash
  • Step 4 6- ( 2-Chlorophenyl ) -2- ( ( 4- (piperazin-1- yl ) phenyl ) amino ) -5H-pyrano [ 2 , 3-d] pyrimidin-5-one , 2HC1 : To a solution of tert-butyl 4- ( 4- ( ( 6- ( 2-chlorophenyl) -5- oxo-5H-pyrano [ 2 , 3-d] pyrimidin-2- yl ) amino ) phenyl ) piperaz ine-l-carboxylate (50 mg, 0.094 mmol) in dichloromethane (1 mL) was added dropwise 4 M HC1 in 1,4-dioxane (1 mL) and the mixture stirred at RT for 2 h. The reaction mixture was then diluted with diethyl ether (5 mL) and slurried at RT for 15 min, before removal of the supernatant solvent by pipette.
  • Example 29 6- ( 2-Chlorophenyl ) -8-cyclopropyl-2- ( ( 4- ( 4- methylpiperaz in-1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5 ( 8H) -one
  • Step 1 1- ( 4-Chloro-2- (methylthio ) pyrimidin-5-yl ) -2 -( 2- chlorophenyl ) ethanone : To a solution of 2- (2- chlorophenyl ) acetic acid (2 g, 11.7 mmol) in THF (30 mL) was added tert-butylmagnesium chloride (11.7 mL, 23.5 mmol) . The brown suspension formed was stirred at RT for 30 min and ethyl 4-chloro-2- (methylthio ) pyrimidine-5- carboxylate (2.46 g, 10.6 mmol) in 10 mL THF added. The resulting yellow suspension was stirred for 16 h.
  • Step 2 2- ( 2-Chlorophenyl ) -1- ( 4- ( cyclopropylamino ) -2- (methylthio ) pyrimidin-5-yl ) ethanone :
  • Step 3 6- ( 2-Chlorophenyl ) -8-cyclopropyl-2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one :
  • Example 30 6- ( 4-Hydroxyphenyl ) -8-methyl-2- (( 4- ( 4- methylpiperaz in-1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin-
  • Step 1 6- ( 4-Hydroxyphenyl ) -8-methyl-2-
  • Step 2 4- ( 8-Methyl-2- (methylthio ) -5-oxo-5 , 8- dihydropyrido [ 2 , 3-d] pyrimidin-6-yl ) phenyl acetate: Acetic anhydride (0.014 mL, 0.147 mmol) was added to a stirred solution of 6- ( 4-hydroxyphenyl ) -8-methyl-2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one (8.8 mg, 0.029 mmol) and triethylamine (0.5 mL, 3.59 mmol) in DCM (1.0 mL) at RT under nitrogen.
  • Step 3 6- ( 4-Hydroxyphenyl ) -8-methyl-2- ( ( 4- ( 4- methylpiperaz in-1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5(8H)-one: mCPBA ( ⁇ 77% pure) (7.4 mg, 0.033 mmol) in DCM (0.5 mL) was added to a stirred solution of 4- ( 8-methyl- 2- (methylthio ) -5-oxo-5, 8-dihydropyrido [ 2 , 3-d] pyrimidin-6 - yl)phenyl acetate (9.8 mg, 0.029 mmol) in toluene (1.0 mL) at RT under nitrogen. After 15 min, DIPEA (0.015 mL, 0.086 mmol) and 4- ( 4-methylpiperazin-l-yl ) aniline (6.0 mg, 0.032 mmol) were added, sequentially, and the
  • Example 31 6- ( 3-Hydroxyphenyl ) -8-methyl-2- ( ( 4- (piperaz in-1-yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin- 5 ( 8H) -one
  • Step 1 6- ( 3-Hydroxyphenyl ) -8-methyl-2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one :
  • Step 2 3- ( 8-Methyl-2- (methylthio ) -5-oxo-5 , 8- dihydropyrido [ 2 , 3-d] pyrimidin-6-yl ) phenyl acetate: Acetic anhydride (0.023 mL, 0.247 mmol) was added to a stirred solution of 6- ( 3-hydroxyphenyl ) -8-methyl-2- (methylthio ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one (14.8 mg,
  • Step 3 tert-Butyl 4- ( 4- (( 6- ( 3-hydroxyphenyl ) -8-methyl-5- oxo-5 , 8-dihydropyrido [ 2 , 3-d] pyrimidin-2- yl ) amino ) phenyl ) piperaz ine-l-carboxylate : mCPBA ( ⁇ 77% pure) (8.8 mg, 0.039 mmol) in DCM (0.5 ml) was added to a stirred solution of 3- ( 8-methyl-2- (methylthio ) -5-oxo-5 , 8- dihydropyrido [ 2 , 3-d] pyrimidin-6-yl ) phenyl acetate (11.6 mg, 0.034 mmol) in toluene (1.0 mL) at RT under nitrogen.
  • Step 4 6- ( 3-Hydroxyphenyl ) -8-methyl-2- ( ( 4- (piperaz in-1- yl ) phenyl ) amino ) pyrido [ 2 , 3-d] pyrimidin-5 ( 8H ) -one : TFA (1.0 mL, 13.0 mmol) was added to a stirred solution of tert-butyl 4- ( 4- ( ( 6 - ( 3-hydroxyphenyl ) -8-methyl-5-oxo-5 , 8- dihydropyrido [ 2 , 3-d] pyrimidin-2- yl ) amino ) phenyl ) piperaz ine-l-carboxylate (9.7 mg, 0.018 mmol) in DCM (1.0 mL) at RT under nitrogen.
  • Step 1 1- ( 4-chloro-2- (methylthio ) pyrimidin-5-yl ) -2 -( 2- chlorophenyl ) -2- ( 2- ( 4-methoxyphenyl ) hydrazono ) ethanone : To a solution of 4-methoxyaniline (0.170 g, 1.382 mmol) in cone. HC1 (0.433 ml, 5.20 mmol) with 1.5 ml of water at 0 °C was added sodium nitrite (0.095 g, 1.382 mmol) in 0.5 ml of water.
  • Step 2 3- ( 2-chlorophenyl ) -1- ( 4-methoxyphenyl ) -7- (methylthio) pyrimido [ 4, 5-c] pyridazin-4 ( IH) -one : To a solution of 1- ( 4-chloro-2- (methylthio ) pyrimidin-5-y1 ) -2- ( 2-chlorophenyl ) -2- ( 2- ( 4-methoxyphenyl ) hydrazono ) ethanone (0.4 g, 0.894 mmol) in acetonitrile (8 ml) was added potassium carbonate (0.148 g, 1.073 mmol) .
  • Step 3 3- ( 2-chlorophenyl ) -1- ( 4-methoxyphenyl ) -7- (( 4- ( 4- methylpiperaz in-l-yl ) phenyl ) amino ) pyrimido [4,5- c] pyridazin-4 ( IH) -one : Following the procedure for 8- methyl-2- ( ( 4- ( 4-methylpiperaz in-l-yl ) phenyl ) amino ) -6 - phenylpyrido [ 2 , 3-d] pyrimidin-5 ( 8H) -one, 3- (2- chlorophenyl ) -1- ( 4-methoxyphenyl ) -7-
  • Example 33 3- ( 2-chlorophenyl ) -l-methyl-7- ( ( 4- (piperazin- 1-y1 ) phenyl ) amino ) pyrimido [ 4 , 5-c ] pyridaz in-4 ( 1H ) -one
  • Step 1 1- ( 4-chloro-2- (methylthio) pyrimidin-5-yl ) -2- (2- chlorophenyl ) ethanone : To a solution of 2- (2- chlorophenyl ) acetic acid (10 g, 58.6 mmol) in THF (120 mL) under nitrogen, was added tert-butylmagnesium
  • Step 2 di-tert-butyl 3- ( 2-chlorophenyl ) -7- (methylthio ) - 4-oxo-3 , 4-dihydropyrimido [ 4 , 5-c ] pyridaz ine-1 , 2- dicarboxylate :
  • a solution of 1- ( 4-chloro-2- (methylthio ) pyrimidin-5-yl ) -2- ( 2-chlorophenyl ) ethanone (1.0 g, 3.19mmol) in anhydrous THF (20 mL) was cooled to -78°C.
  • Step 3 3- ( 2-chlorophenyl ) -7- (methylthio ) pyrimido [ 4 , 5- c] pyridazin-4-ol : To a solution of di-tert-butyl 3-(2- chlorophenyl ) -7- (methylthio) -4-oxo-3, 4- dihydropyrimido [ 4 , 5-c ] pyridaz ine-1 , 2-dicarboxylate
  • the reaction mixture was concentrated in vacuo.
  • the residue was dissolved in acetonitrile (20 mL) and 4, 5-dichloro-3 , 6-dioxocyclohexa-l , 4-diene-l, 2- dicarbonitrile (1.79 g, 7.89 mmol) was added in a microwave sealed tube.
  • the reaction mixture was heated by microwave irradiation at 100°C for 10 minutes.
  • the reaction mixture was concentrated in vacuo.
  • the residue was triturated with water and the precipitate was filtrated and washed with water and ethyl acetate to afford the title compound as a brown powder (1 g, 42%) .
  • Step 5 tert-butyl 4- ( 4- (( 3- ( 2-chlorophenyl ) -l-methyl-4- oxo-1 , 4-dihydropyrimido [ 4 , 5-c ] pyridaz in-7- yl ) amino ) phenyl ) piperaz ine-l-carboxylate : 3- (2- chlorophenyl ) -1-methyl-7- (methylthio ) pyrimido [4,5- c] pyridazin-4 ( 1H) -one (100 mg, 0.314 mmol) was dissolved in anhydrous dichloromethane . 3-Chloroperbenzoic acid 70% (93mg, 0.378 mmol) was added portionwise at 0°C. After stirring at room temperature for 30 minutes, all the starting material was converted into the
  • Step 1 tert-butyl 5- ( ( 3- ( 2-chlorophenyl ) -l-methyl-4-oxo- 1 , 4-dihydropyrimido [ 4 , 5-c ] pyridaz in- 7-yl ) amino ) -2- morpholinobenzyl (methyl ) carbamate : Following the
  • Step 2 3- ( 2-chlorophenyl ) -l-methyl-7- (( 3- ( (methylamino ) methyl ) -4- morpholinophenyl ) amino ) pyrimido [ 4 , 5-c ] pyridaz in-4 ( 1H) - one: tert-butyl 5- (( 3- ( 2-chlorophenyl ) -l-methyl-4-oxo- 1 , 4-dihydropyrimido [ 4 , 5-c ] pyridaz in- 7-yl ) amino ) -2- morpholinobenzyl (methyl ) carbamate (25 mg, 0.042 mmol) was dissolved in dichloromethane (1 mL) .
  • Step 1 tert-butyl 4- ( 4- (( 3- ( 2-chlorophenyl ) -4-oxo-l , 4- dihydropyrimido [ 4 , 5-c ] pyridaz in-7-yl ) amino ) -2- ( hydroxymethyl ) phenyl ) piperazine-l-carboxylate : Following the procedure of tert-butyl 4- ( 4- (( 3- ( 2-chlorophenyl ) -1- methyl- 4-oxo-l , 4-dihydropyrimido [ 4 , 5-c ] pyridaz in-7- yl ) amino ) phenyl ) piperaz ine-l-carboxylate , 3- ( 2- chlorophenyl ) -7- (methylthio ) pyrimido [ 4 , 5-c ] pyridaz in- 4(lH)-one (50 mg, 0.16 mmol) was reacted with tert-
  • Step 2 3- ( 2-chlorophenyl ) -7- (( 3- (hydroxymethyl ) -4- (piperaz in-l-yl ) phenyl ) amino ) -pyrimido [ 4 , 5-c ] pyridazin- 4(lH)-one: tert-butyl 4- ( 4- (( 3- ( 2-chlorophenyl ) -4-oxo- 1 , 4-dihydropyrimido [ 4 , 5-c ] pyridaz in-7-yl ) amino ) -2- (hydroxymethyl ) phenyl ) piperazine-l-carboxylate (30 mg, 0.053 mmol) was dissolved in dichloromethane ( 1 mL) . Then TFA (1 mL, 13 mmol) was added at 0°C. The resulting solution was stirred for 16 hours at room temperature until the starting material had been consumed (LC
  • Step 1 tert-butyl 4- ( 4- (( 3- ( 2-chlorophenyl ) -l-methyl-4- oxo-1 , 4-dihydropyrimido [ 4 , 5-c ] pyridaz in-7-yl ) amino ) -2- (hydroxymethyl ) phenyl ) piperazine-l-carboxylate : Following the procedure of tert-butyl 4- ( 4- (( 3- ( 2-chlorophenyl ) -1- methyl-4-oxo-l , 4-dihydropyrimido [ 4 , 5-c ] pyridaz in-7- yl ) amino ) phenyl ) piperaz ine-l-carboxylate , 3- ( 2- chlorophenyl ) -1-methyl-7- (methylthio ) pyrimido [4,5- c] pyridazin-4 ( 1H) -one (50 mg, 0.15 mmol) was reacted with
  • Step 2 3- ( 2-chlorophenyl ) -7- (( 3- (hydroxymethyl ) -4- (piperaz in-l-yl ) phenyl ) amino ) -1-methylpyrimido [4,5- c] pyridazin-4 ( 1H) -one : tert-butyl 4-(4-((3-(2- chlorophenyl ) -l-methyl-4-oxo-l , 4-dihydropyrimido [4, 5- c ] pyridaz in-7-yl ) amino ) -2- ( hydroxymethyl ) phenyl )
  • Step 1 tert-butyl 4- ( 4- (( 3- ( 2-chlorophenyl ) -l-methyl-4- oxo-1, 4-dihydropyrimido [ 4 , 5-c ] pyridaz in-7-yl ) amino ) -2- methylphenyl ) piperazine-l-carboxylate : Following the procedure of tert-butyl 4- ( 4- (( 3- ( 2-chlorophenyl ) -1- methyl-4-oxo-l , 4-dihydropyrimido [ 4 , 5-c ] pyridaz in-7- yl ) amino ) phenyl ) piperaz ine-l-carboxylate , 3- ( 2- chlorophenyl ) -l-methyl-7- (methylthio ) pyrimido [4,5- c] pyridazin-4 ( 1H) -one (100 mg, 0.314 mmol) was reacted with tert
  • Example 38 3- ( 2-chlorophenyl ) -7- ( ( 3-methoxy-4- (piperaz in-l-yl ) phenyl ) amino ) -1-methylpyrimido [4,5- c] pyridaz in-4 ( IH) -one
  • Step 1 tert-butyl 4- ( 4- (( 3- ( 2-chlorophenyl ) -l-methyl-4- oxo-1, 4-dihydropyrimido [ 4 , 5-c ] pyridaz in-7-yl ) amino ) -2- methoxyphenyl ) piperazine-l-carboxylate : Following the procedure of tert-butyl 4- ( 4- (( 3- ( 2-chlorophenyl ) -1- methyl-4-oxo-l , 4-dihydropyrimido [ 4 , 5-c ] pyridaz in-7- yl ) amino ) phenyl ) piperaz ine-l-carboxylate , 3- ( 2- chlorophenyl ) -1-methyl-7- (methylthio ) pyrimido [4,5- c] pyridazin-4 ( IH) -one (100 mg, 0.314 mmol) was reacted with tert
  • Step 2 3- ( 2-chlorophenyl ) -7- (( 3-methoxy-4- (piperaz in-1 yl ) phenyl ) amino ) -1-methylpyrimido [ 4 , 5-c ] pyridaz in-4 ( IH ) one: Following the procedure of 3- ( 2-chlorophenyl ) -1- methyl-7- ( ( 3- ( (methylamino ) methyl ) -4- morpholinophenyl ) amino ) pyrimido [ 4 , 5-c ] pyridaz in-4 ( IH) - one, tert-butyl 4- ( 4- ( ( 3- ( 2-chlorophenyl ) -l-methyl-4-oxo- 1 , 4-dihydropyrimido [ 4 , 5-c ] pyridaz in-7-yl ) amino ) -2- methoxyphenyl ) piperazine-l-carboxylate (49 mg, 0.085
  • Step 1 (R) -tert-butyl 4- ( 4- (( 3- ( 2-chlorophenyl ) -1- methyl-4-oxo-l , 4-dihydropyrimido [ 4 , 5-c ] pyridaz in-7- yl ) amino ) phenyl ) -2-methylpiperaz ine-l-carboxylate :
  • Step 2 (R) -3- ( 2-chlorophenyl ) -l-methyl-7- (( 4- ( 3- methylpiperaz in-l-yl ) phenyl ) amino ) pyrimido [4,5- c] pyridazin-4 ( 1H) -one : (R) -tert-butyl 4-(4-((3-(2- chlorophenyl ) -l-methyl-4-oxo-l , 4-dihydropyrimido [4, 5- c] pyridaz in-7-yl ) amino ) phenyl ) -2-methylpiperaz ine-1 - carboxylate (82 mg, 0.146 mmol) was reacted with HC1 (3 mL, 4 M in dioxane) in DCM (3mL) for 4h at RT .
  • Wee-1 activity In the measurement of Wee-1 activity, a commercial peptide Poly(Lys Tyr(4:l)) hydrobromide was purchased from Sigma Aldrich and used as the substrate. Activated Wee-1 kinase was purchased from Invitrogen (PV3817) and an ADP-Glo luminescent kit was purchased from Promega.
  • the plate was incubated at room temperature for 30 minutes before the addition of 2 pg/mL of substrate and 30 pM ATP in a 5 pL aliquot. The plate was centrifuged for one minute and incubated for 1 h at RT . 15 ⁇ 1 ⁇ L of ADP-Glo stop reagent was added to each well to quench the reaction and deplete unconverted ATP. The plate was incubated for a further 40 min in the dark at RT .
  • the plate was shaken for 1 min, and incubated in the dark for an additional hour.
  • Luminescence from each well was detected using the Biotek Synergy4 HD plate reader and the percentage inhibition of kinase activity calculated for each inhibitor tested. Positive (kinase only) and negative (no kinase) controls were added to each plate to ensure specific interaction of kinase and inhibitor. The IC 50 concentration for each inhibitor was calculated by plotting the percentage kinase inhibition against concentration of inhibitor and the curve generated by non-linear regression fitting.
  • the colorectal cancer cell lines HT-29 and HCT-116 were purchased from the ATCC and routinely maintained in
  • the cells were trypsinised from their growing vessel and counted, 100 ⁇ 1 ⁇ L of cell suspension containing 6000 cells was pipetted into black 96 well Co-star plates and incubated overnight to allow adherence to the surface at a temperature of 37°C and an atmosphere of 5% CO 2 .
  • Test compounds were formulated in DMSO and diluted in foetal calf serum supplemented medium. Incubating medium was removed by aspiration and diluted drug supplemented medium added to each well.
  • the plate was returned to the incubator for an additional eight hours at 37°C and an atmosphere of 5% CO 2 .
  • Post incubation, the drug supplemented medium was aspirated from each well and the cells were washed once in ice-cold phosphate buffered saline (PBS) .
  • PBS phosphate buffered saline
  • cell lysis buffer Cell Signalling Technologies #9803
  • 20 mM Tris 150 mM NaCl, 1 mM EDTA, 1 mM EGTA, 1% Triton- X100, 2.5 mM sodium pyrophosphate, 1 mM glycerophosphate, 1 mM Na 3 VC>4 and 1 pg/mL leupeptin was added to each well of the 96 well plate and incubated at 4 °C for 30 min. The samples on the plate were snap frozen at -80 °C until required. Immediately before the continuation of the assay, the sample plate was thawed and centrifuged at 4 °C for 10 min and the supernatant transferred to
  • Cell supernatant was mixed in a ratio of 1:1 with sample dilutent buffer and vortexed for one minute. 100 pL of diluted sample was pipetted into pre-coated plates containing a rabbit polyclonal antibody for phospho-cdc2 (Tyrl5) (Cell Signalling Technologies PathScan kit
  • the plate was sealed and incubated overnight at 4 °C.
  • kit detection antibody was added to each well and the plate re-sealed and incubated at 37°C for 1 h. Post incubation the plate was washed and processed in a similar manner to that previously
  • TMB reagent tetramethylbenz idine
  • the percentage of phospho-cdc2 was calculated compared to DMSO control and plotted versus the concentration of inhibitor using GraphPad Prism. Data was fitted using non-linear regression analysis and IC 5 o values generated.
  • Method 3 Determining the anti-proliferative properties of Wee-1 inhibitors in the HT-29 cell line Cell Titre Glo (Promega) is a highly sensitive
  • the reagent uses a stable form of luciferase to measure ATP as an output of viability.
  • the luminescent values generated in the assay are directly proportional to the number of viable cells in your assay.
  • HT-29 cells were washed, detached and re-suspended in their respective fresh media. The cells were pelleted by centrifugation (Eppendorf 5414) and the spent supernatant was discarded. The cells were re-suspended by vortex mixing, counted and seeded into clear bottom white 96 well plates at a density of 5000 cells per well. The cells were incubated overnight at 37 °C (95% O 2 / 5% CO 2 ) , and next day treated with increasing concentrations of test compound formulated in fresh medium. The plates were returned to the incubator for 72 h.
  • Cell Titre Glo (Promega) was prepared by mixing the supplied reagents as per manufacturer's instructions and left to stand at RT .
  • the cell plates were removed from the incubator and 80 pL of the Cell Titre Glo solution added to each well. The plate was shaken for five minutes to ensure homogenous mixing of reagents and cells, then left to stand for 10 min at RT .
  • the cell viability post compound treatment was determined by the luminescent intensity emitted from the drug treated wells in the plate.
  • the assay plate was placed in the Biotex Synergy 4 Hybrid plate reader and the luminescence read in each well.
  • the compound treated wells were compared to vehicle treated wells and the % inhibition of cell viability calculated.
  • the data was analysed using GraphPad Prism, with IC 5 o values generated using non-linear regression of the data set .
  • Method 4 Determination of CLint estimates using human liver microsomes.
  • AKT1 (PKB alpha) - - - -
  • GSK3B (GSK3 beta) - - -
  • PDGFRB (PDGFR beta) - - +
  • PRKCB1 (PKC beta I) - - - -
  • RPS6KA3 (RSK2) - - - RPS6KB1 (p70S6K) — — —

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Abstract

La présente invention concerne des composés qui sont utiles en tant qu'inhibiteurs de l'activité de la kinase Wee-1. La présente invention concerne en outre des compositions pharmaceutiques comprenant ces composés et des procédés d'utilisation de ces composés dans le traitement du cancer et des procédés de traitement du cancer.
PCT/GB2013/052080 2013-08-05 2013-08-05 Composés pharmaceutiques Ceased WO2015019037A1 (fr)

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Cited By (10)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2018133829A1 (fr) 2017-01-23 2018-07-26 南京明德新药研发股份有限公司 Dérivé de 1,2-dihydro-3h-pyrazolo[3,4-d]pyrimidin-3-one utilisé en tant qu'inhibiteur de wee1
WO2018183891A1 (fr) 2017-03-31 2018-10-04 Cascadian Therapeutics Combinaisons d'inhibiteurs de chk1 et wee1
WO2019085933A1 (fr) 2017-11-01 2019-05-09 南京明德新药研发股份有限公司 Composé macrocyclique servant d'inhibiteur de wee1 et ses applications
WO2020083404A1 (fr) 2018-10-26 2020-04-30 南京明德新药研发有限公司 Dérivé de pyrimidopyrazolone en tant qu'inhibiteur de wee1 et son utilisation
US10807994B2 (en) 2017-10-09 2020-10-20 Nuvation Bio Inc. Heterocyclic compounds and uses thereof
WO2020221358A1 (fr) 2019-04-30 2020-11-05 石家庄智康弘仁新药开发有限公司 Forme cristalline d'un composé inhibiteur de wee1 et son utilisation
WO2021254389A1 (fr) 2020-06-17 2021-12-23 微境生物医药科技(上海)有限公司 Dérivé de pyrazolo[3,4-d]pyrimidine-3-cétone utilisé en tant qu'inhibiteur de wee-1
US11299493B2 (en) 2017-10-09 2022-04-12 Nuvation Bio Inc. Heterocyclic compounds and uses thereof
US11332473B2 (en) 2019-04-09 2022-05-17 Nuvation Bio Inc. Substituted pyrazolo[3,4-d]pyrimidines as Wee1 inhibitors
WO2022228511A1 (fr) 2021-04-30 2022-11-03 微境生物医药科技(上海)有限公司 Composé à cycle fusionné utilisé comme inhibiteur de wee-1, son procédé de préparation et son utilisation

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WO2008055013A2 (fr) * 2006-10-31 2008-05-08 Janssen Pharmaceutica N.V. 5-oxo-5,8-dihydro-pyrido-pyrimidines comme inhibiteurs de la kinase c-fms
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WO2010067888A1 (fr) * 2008-12-12 2010-06-17 Banyu Pharmaceutical Co.,Ltd. Dérivés de dihydropyrimidopirymidine
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Cited By (12)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2018133829A1 (fr) 2017-01-23 2018-07-26 南京明德新药研发股份有限公司 Dérivé de 1,2-dihydro-3h-pyrazolo[3,4-d]pyrimidin-3-one utilisé en tant qu'inhibiteur de wee1
WO2018183891A1 (fr) 2017-03-31 2018-10-04 Cascadian Therapeutics Combinaisons d'inhibiteurs de chk1 et wee1
US10807994B2 (en) 2017-10-09 2020-10-20 Nuvation Bio Inc. Heterocyclic compounds and uses thereof
US11299493B2 (en) 2017-10-09 2022-04-12 Nuvation Bio Inc. Heterocyclic compounds and uses thereof
WO2019085933A1 (fr) 2017-11-01 2019-05-09 南京明德新药研发股份有限公司 Composé macrocyclique servant d'inhibiteur de wee1 et ses applications
US11613545B2 (en) 2017-11-01 2023-03-28 Wuxi Biocity Biopharmaceutics Co., Ltd. Macrocyclic compound serving as Wee1 inhibitor and applications thereof
WO2020083404A1 (fr) 2018-10-26 2020-04-30 南京明德新药研发有限公司 Dérivé de pyrimidopyrazolone en tant qu'inhibiteur de wee1 et son utilisation
US12180184B2 (en) 2018-10-26 2024-12-31 Wuxi Biocity Biopharmaceutics Co., Ltd. Pyrimidopyrazolone derivative as Wee1 inhibitor and use thereof
US11332473B2 (en) 2019-04-09 2022-05-17 Nuvation Bio Inc. Substituted pyrazolo[3,4-d]pyrimidines as Wee1 inhibitors
WO2020221358A1 (fr) 2019-04-30 2020-11-05 石家庄智康弘仁新药开发有限公司 Forme cristalline d'un composé inhibiteur de wee1 et son utilisation
WO2021254389A1 (fr) 2020-06-17 2021-12-23 微境生物医药科技(上海)有限公司 Dérivé de pyrazolo[3,4-d]pyrimidine-3-cétone utilisé en tant qu'inhibiteur de wee-1
WO2022228511A1 (fr) 2021-04-30 2022-11-03 微境生物医药科技(上海)有限公司 Composé à cycle fusionné utilisé comme inhibiteur de wee-1, son procédé de préparation et son utilisation

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