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EP1442039A1 - Pyrimido (4,5-b) indole derivatives - Google Patents

Pyrimido (4,5-b) indole derivatives

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Publication number
EP1442039A1
EP1442039A1 EP02777332A EP02777332A EP1442039A1 EP 1442039 A1 EP1442039 A1 EP 1442039A1 EP 02777332 A EP02777332 A EP 02777332A EP 02777332 A EP02777332 A EP 02777332A EP 1442039 A1 EP1442039 A1 EP 1442039A1
Authority
EP
European Patent Office
Prior art keywords
pyrimido
indole
carbamoyl
amino
alkyl
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.)
Withdrawn
Application number
EP02777332A
Other languages
German (de)
French (fr)
Inventor
Timothy Lowinger
Makoto Shimazaki
Hiroki Sato
Kazuho Tanaka
Naoki Tsuno
Karsten Marx
Masaru Yamamoto
Klaus Urbahns
Florian Gantner
Hiromi Okigami
Kosuke Nakashima
Keisuke Takeshita
Kevin B. Bacon
Hiroshi Komura
Nagahiro Yoshida
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Bayer AG
Original Assignee
Bayer Healthcare AG
Priority date (The priority date 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 date listed.)
Filing date
Publication date
Priority claimed from JP2001334662A external-priority patent/JP2003137886A/en
Priority claimed from GB0214544A external-priority patent/GB2390090A/en
Application filed by Bayer Healthcare AG filed Critical Bayer Healthcare AG
Publication of EP1442039A1 publication Critical patent/EP1442039A1/en
Withdrawn legal-status Critical Current

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Classifications

    • 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
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P11/00—Drugs for disorders of the respiratory system
    • A61P11/06—Antiasthmatics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P13/00—Drugs for disorders of the urinary system
    • A61P13/12—Drugs for disorders of the urinary system of the kidneys
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/14—Drugs for disorders of the nervous system for treating abnormal movements, e.g. chorea, dyskinesia
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/28—Drugs for disorders of the nervous system for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/08—Antiallergic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A61P9/10—Drugs for disorders of the cardiovascular system for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis

Definitions

  • the present invention relates to pyrimido[4,5-b]indoles which are useful as an active ingredient of pharmaceutical preparations.
  • the pyrimido[4,5-b]indoles of the present invention have MKK7 and KK4 [MAPK (mitogen activated protein kinase) kinase
  • inhibitory activity can be used for the prophylaxis and treatment of diseases associated with MKK7 and MKK4 activity.
  • the pyrimido[4,5-b]indoles derivatives of the present invention are useful for treatment and prophylaxis of diseases as follows: inflammatory and immunoregulatory disorders, such as asthma, atopic dermatitis, rhinitis, allergic rhinitis, allergic diseases, COPD, septic shock, arthritis, joint diseases and myocardial injuries, as well as autoimmune pathologies such as rheumatoid arthritis, Grave's disease, and atherosclerosis.
  • diseases as follows: inflammatory and immunoregulatory disorders, such as asthma, atopic dermatitis, rhinitis, allergic rhinitis, allergic diseases, COPD, septic shock, arthritis, joint diseases and myocardial injuries, as well as autoimmune pathologies such as rheumatoid arthritis, Grave's disease, and atherosclerosis.
  • the compounds of the present invention are also useful for treatment of ischemia, myocardial injury, pulmonary hypertension, renal failure, Huntington's chorea and cardiac hypertrophy, as well as neurodegenerative disorders such as Parkinson's disease, Alzheimer's disease and focal ischemia, since the diseases also relate to MKK7 and/or MKK4.
  • the mitogen-activated protein kinases are a family of serine/threonine kinases involved in the transduction of signals from the cell membrane to the nucleus in response to various types of stimuli such as lipopolysaccharide (LPS), tumor necrosis factor- ⁇ (TNF- ), interleukins, CD40 and others. These kinases participate in a wide variety of signaling cascades controlling cellular events such as cell growth, differentiation, activation, apoptosis, stress responses, and transformation.
  • LPS lipopolysaccharide
  • TNF- tumor necrosis factor- ⁇
  • CD40 interleukins
  • MAPK extracellular-regulated kinases
  • ERK extracellular-regulated kinases
  • MAPK/JNK stress-activated/c-jun N-terminal kinase
  • SAPK/JNKs are activated in response to cellular "stress” such as changes in osmolarity or metabolism, ischemia, heat shock, shear stress, ceramide or inflammatory cytokines (TNF- ⁇ , IL-1).
  • stress such as changes in osmolarity or metabolism, ischemia, heat shock, shear stress, ceramide or inflammatory cytokines (TNF- ⁇ , IL-1).
  • JNKs control gene activity via phosphorylation of a variety of transcriptional factors including c-Jun, JunD, nuclear factor of activated T cells (NFAT)4, or Elk-1 , all present in immune cells and known to regulate the transcription of many genes during an inflammatory response.
  • SAPK/JNKs regulate the activation and proliferation of T and B lymphocytes, activation of mast cells
  • MKK stress kinase mitogen-activated protein kinase kinase
  • MAPKs themselves require dual phosphorylation of both threonine and tyrosine at their so-called Thr-X-Tyr motif, wliich is brought upon by the upstream regulators MAPK kinases (MKKs).
  • MKK1-MKK7 MEK1, MEK2,
  • MKK3, MKK4, MEK5, MKK6, and MKK7 are known to date with MKK7 being the most recently identified (Tournier C, Whitmarsh J., Cavanagh J., Barrett T.,
  • Mitogen-activated protein kinase kinase 7 is an activator of the c-Jun NH2- terminal kinase. Proc Natl Acad Sci USA, 94: 7337-7342, 1997), (Moriguchi T.,
  • MKK4 is also capable of phosphorylating p-38 MAP kinases.
  • p-38 kinases are also involved in the control of inflammatory gene expression, especially after stimulation of cells with lipopoly saccharide and cytokines (Han J., Lee JD., Jiang Y., Li Z., Feng L., Ulevitch RJ.
  • p38 controls the release of IL-12 and IFN ⁇ and in B cells, CD40 cross-linking leads to rapid p38 activation and thus controls proliferation, and adhesion molecule expression.
  • p38 MAPK are activated by hypoxia and, by controlling the transcription factor ATF2, play a role in neuronal development and survival (Lee JC, Kumar S., Griswold DE., Underwood DC, Notta BJ., Adams JL. Inhibition of p38 MAP kinase as a therapeutic strategy. Immunopharmacology, Al: 185-201, 2000).
  • a specific inhibitor of MKK7 and/or of MKK4, which is expected to block the synthesis of pro-inflammatory cytokines and the activation of various immune cells, should have a broad anti-inflammatory profile with potential for the treatment of inflammatory and immunoregulatory disorders and diseases, including asthma, rhinitis, allergic diseases, septic shock, joint diseases and myocardial injuries, as well as autoimmune pathologies such as rheumatoid arthritis, Grave's disease, and atherosclerosis.
  • inhibitors should also have therapeutic potential for the treatment of renal failure, Huntington's chorea, cardiac hypertrophy and neurodegenerative disorders such as Parkinson's disease, Alzheimer's disease and focal ischemia (Xia XG., Harding T., Weller M., Bieneman A., Uney JB., Schulz JB. Gene transfer of the JNK interacting protein- 1 protects dopaminergic neurons in the MPTP model of Parkinson's disease. Proc Natl Acad Sci USA, 98: 10433-10438, 2001).
  • WO 9842708 discloses anti-asthma agent represented by the general formula:
  • R a , R 2 ⁇ , R 2"la , R 2"2a , R 3'la , and R 3"2a are defined in the application.
  • WO 9626941 discloses pharmaceutical agents represented by the general formula:
  • R lc , R 2c , R 3c , R 4c , R 5c and R 6c are defined in the specification.
  • WO 9320078 discloses pharmaceutically active compound represented by the formula:
  • IN 157280 discloses the method for preparing anti hypertension agents represented by the formula:
  • R le , R 2e , R 3e , R 4e , and R 5e are defined in the specification.
  • WO 97/02266 discloses anti-hyperproliferative disease agents represented by the general formula:
  • WO 98/43973 also discloses anti-proliferative disease agents represented by the general formula:
  • Rid, R2d, R3d, R4', q and m' are defined in the application.
  • This invention is to provide a novel pyrimido[4,5-b]indole derivative of the formula (I), its tautomeric or stereoisomeric form, or a salt thereof:
  • R is hydrogen, amino, (C 1-6 )alkyl, (C 2-6 )alkenyl, (C -6 )alkynyl, halogen substituted (C 1-6 ) alkyl, cyano, cyano(C 1-6 )alkyl, (C -8 )cycloalkyl, nitro, nitro(C 1 .. 6 )alkyl or fluoro;
  • R 2 and R 3 independently represent hydrogen, (C 2 - 6 )alkenyl, (C 2-6 )alkynyl,
  • R 2 and R 3 are taken together with the attached nitrogen atom to form a 3-8 membered saturated ring optionally interrupted by one or two atoms selected from the group consisting of oxygen, sulfur and nitrogen,
  • said saturated ring is optionally fused by benzene or 3-8 membered saturated ring having 0 to 3 nitrogen atoms,
  • said saturated ring is optionally having substituents independently selected from the group consisting of halogen, hydroxy, carbamoyl, carboxy, amino, oxo, pyrrolidino, (C 1-6 )alkoxy, (C 2-6 )alkenyl, (C 2-6 )- alkynyl, (C 3-8 )cycloalkyl, (C 1-6 )alkylcarbonyl, (C 1-6 )alkylsulfonyl, arylsulfonyl, (C 1-6 )alkyl, hydroxy(C 1-6 )alkyl, (C 1-6 )alkoxy(C 1-6 )alkyl, (C 1-6 )alkoxycarbonyl, (C 1-6 )alkoxycarbonyl(C ⁇ -6 )alkyl, (C 3- )cyclo- alkyl(C 1-6 )alkyl, aryl(C 1-6 )alkyl, carbamoyl(C 1-6
  • spiro ring is optionally interrupted by 0 to 3 nitrogen or oxygen atoms
  • R 4 is halogen, hydroxy, (C 1-6 )alkyl, amino (C 1-6 )alkyl, (C 1-6 )alkoxy, cyano, carboxy, (C 1-6 )alkoxycarbony, or carbamoyl;
  • R 5 is hydrogen, or (C 1-6 )alkyl.
  • Alkyl per se and "alk” and “alkyl” in alkoxy, alkanoyl, alkylamino, alkylamino- carbonyl, alkylaminosulphonyl, alkylsulphonylamino, alkoxycarbonyl, alkoxy- carbonylamino and alkanoylamino represent a linear or branched alkyl radical having generally 1 to 6, preferably 1 to 4 and particularly preferably 1 to 3 carbon atoms, representing illustratively and preferably methyl, ethyl, n-propyl, isopropyl, tert- butyl, n-pentyl and n-hexyl.
  • Alkoxy illustratively and preferably represents methoxy, ethoxy, n-propoxy, isopropoxy, tert-butoxy, n-pentoxy and n-hexoxy.
  • Alkanoyl illustratively and preferably represents acetyl and propanoyl.
  • Alkylamino represents an alkylamino radical having one or two (independently selected) alkyl substituents, illustratively and preferably representing methylamino, ethylainino, n-propylamino, isopropylamino, tert-butylamino, n-pentylamino, n- hexyl-amino, N,N-dimethylamino, N,N-diethylamino, N-ethyl-N-methylamino, N- methyl-N-n-propylamino, N-isopropyl-N-n-propylamino, N-t-butyl-N-methylamino, N-ethyl-N-n-pentylamino and N-n-hexyl-N-methylamino.
  • Alkylaminocarbonyl or alkylcarbamoyl represents an alkylammocarbonyl radical having one or two (independently selected) alkyl substituents, illustratively and preferably representing methylaminocarbonyl, ethylaminocarbonyl, n-propylamino- carbonyl, isopropylamino-carbonyl, tert-butylaminocarbonyl, n-pentylamino- carbonyl, n-hexylaminocarbonyl, N,N-dimethylaminocarbonyl, N,N-diethylamino- carbonyl, N-ethyl-N-methylaminocarbonyl, N-methyl-N-n-propylaminocarbonyl, N- isopropyl-N-n-propylaminocarbonyl, N-t-butyl-N-methylaminocarbonyl, N-ethyl-N- n-
  • Alkylaminosulphonyl represents an alkylaminosulphonyl radical having one or two (independently selected) alkyl substituents, illustratively and preferably representing methylaminosulphonyl, ethylaminosulphonyl, n-propylaminosulphonyl, isopropyl- aminosulphonyl, tert-butylaminosulphonyl, n-pentylaminosulphonyl, n-hexyl-amino- sulphonyl, N,N-dimethylaminosulphonyl, N,N-diethylaminosulphonyl, N-ethyl-N- methylamino-sulphonyl, N-methyl-N-n-propylaminosulphonyl, N-isopropyl-N-n-propylaminosulphonyl, N-t-butyl-N-methylaminosulphonyl, N-ethy
  • Alkylsulphonylamino illustratively and preferably represents methylsulphonylamino, ethylsulphonylamino, n-propylsulphonylamino, isopropylsulphonylamino, tert-butyl- sulphonylamino, n-pentylsulphonylamino and n-hexylsulphonylamino.
  • Alkoxycarbonyl illustratively and preferably represents methoxycarbonyl, ethoxy- carbonyl, n-propoxycarbonyl, isopropoxycarbonyl, tert-butoxycarbonyl, n-pentoxy- carbonyl and n-hexoxycarbonyl.
  • Alkoxycarbonylamino illustratively and preferably represents methoxycarbonylamino, ethoxycarbonylamino, n-propoxycarbonylamino, isopropoxycarbonylamino, tert-butoxycarbonylamino, n-pentoxycarbonylamino and n-hexoxycarbonylamino .
  • Alkanoylamino illustratively and preferably represents acetylamino and ethyl- carbonylamino.
  • Cycloalkyl per se and in cycloalkylamino and in cycloalkylcarbonyl represents a cycloalkyl group having generally 3 to 8 and preferably 5 to 7 carbon atoms, illustratively and preferably representing cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and cycloheptyl.
  • Cycloalkylamino represents a cycloalkylamino radical having one or two (independently selected) cycloalkyl substituents, illustratively and preferably representing cyclopropylamino, cyclobutylamino, cyclopentylamino, cyclohexylamino and cyclo- heptylamino.
  • Cycloalkylcarbonyl illustratively and preferably represents cyclopropylcarbonyl, cyclobutylcarbonyl, cyclopentylcarbonyl, cyclohexylcarbonyl and cycloheptyl- carbonyl.
  • Aryl per se and in arylamino and in arylcarbonyl represents a mono- to tricyclic aromatic carbocyclic radical having generally 6 to 14 carbon atoms, illustratively and preferably representing phenyl, naphthyl and phenanthrenyl.
  • Arylamino represents an arylamino radical having one or two (independently selected) aryl substituents, illustratively and preferably representing phenylamino, diphenylamino and naphthylamino.
  • Arylcarbonyl illustratively and preferably represents phenylcarbonyl and naphthyl- carbonyl.
  • Heteroaryl per se and in heteroarylamino and heteroarylcarbonyl represents an aromatic mono- or bicyclic radical having generally 5 to 10 and preferably 5 or 6 ring atoms and up to 5 and preferably up to 4 hetero atoms selected from the group consisting of S, O and N, illustratively and preferably representing thienyl, furyl, pyrrolyl, thiazolyl, oxazolyl, imidazolyl, pyridyl, pyrimidyl, pyridazinyl, indolyl, indazolyl, benzofuranyl, benzothiophenyl, quinolinyl, isoquinolinyl.
  • Heteroarylamino represents an heteroarylamino radical having one or two (independently selected) heteroaryl substituents, illustratively and preferably representing thienylamino, furylamino, pyrrolylamino, thiazolylamino, oxazolylamino, imidazolyl-amino, pyridylamino, pyrimidylamino, pyridazinylamino, indolylamino, indazolylamino, benzofuranylamino, benzothiophenylamino, quinolinyl-amino, isoquinolinylamino .
  • Heteroarylcarbonyl illustratively and preferably represents thienylcarbonyl, furyl- carbonyl, pyrrolylcarbonyl, thiazolylcarbonyl, oxazolylcarbonyl, imidazolyl- carbonyl, pyridylcarbonyl, pyrimidylcarbonyl, pyridazinylcarbonyl, indolylcarbonyl, indazolylcarbonyl, benzofiiranylcarbonyl, benzothiophenylcarbonyl, quinolinyl- carbonyl, isoquinolinylcarbonyl.
  • Heterocyclyl per se and in heterocyclylcarbonyl represents a mono- or polycyclic, preferably mono- or bicyclic, nonaromatic heterocyclic radical having generally 4 to 10 and preferably 5 to 8 ring atoms and up to 3 and preferably up to 2 hetero atoms and or hetero groups selected from the group consisting of N, O, S, SO and SO 2 .
  • the heterocyclyl radicals can be saturated or partially unsaturated.
  • Heterocyclylcarbonyl illustratively and preferably represents tetrahydrofuran-2- carbonyl, pyrrolidine-2-carbonyl, pyrrolidine-3 -carbonyl, pyrrolinecarbonyl, piperidinecarbonyl, morpholinecarbonyl, perhydroazepinecarbonyl.
  • Halogen represents fluorine, chlorine, bromine and iodine.
  • This invention is also to provide a method for treating or preventing a disorder or disease associated with MKK7 and/or MKK4 activity in a human or animal subject, comprising administering to said subject a therapeutically effective amount of the pyrimido[4,5-b]indole derivative shown in the formula (I), its tautomeric or stereoisomeric form, or a physiologically acceptable salt thereof.
  • this invention is to provide a use of the pyrimido[4,5-b]indole derivative shown in the formula (I), its tautomeric or stereoisomeric form, or a physiologically acceptable salt thereof in the preparation of a medicament.
  • said medicament is suitable for treating or preventing a disorder or disease associated with MKK7 and/or MKK4 activity.
  • the compounds of the present invention surprisingly show excellent MKK7 and/or MKK4 inhibitory activity. They are, therefore, suitable for the production of medicament or medical composition, which may be useful to treat MKK7 and/or MKK4 related diseases.
  • the pyrirnido[4,5-b]indoles derivatives of the present invention inhibit MKK7 and/or MKK4, they are useful for treatment and prophylaxis of diseases as follows: inflammatory and immunoregulatory disorders, such as asthma, atopic dermatitis, rhinitis, allergic rhinitis, allergic diseases, COPD, septic shock, arthritis, joint diseases and myocardial injuries, as well as autoimmune pathologies such as rheumatoid arthritis, Grave's disease, and atherosclerosis.
  • diseases as follows: inflammatory and immunoregulatory disorders, such as asthma, atopic dermatitis, rhinitis, allergic rhinitis, allergic diseases, COPD, septic shock, arthritis, joint diseases and myocardial injuries, as well as autoimmune pathologies such as rheumatoid arthritis, Grave's disease, and atherosclerosis.
  • MKK7 and/or MKK4 is an important target and inhibition of MKK7 and/or MKK4 is likely to be effective in the treatment of such inflammatory and immunoregulatory disorders and diseases.
  • the compounds of the present invention are also useful for treatment of ischemia, myocardial injury, pulmonary hypertension, renal failure, Huntington's chorea and cardiac hypertrophy, as well as neurodegenerative disorders such as Parkinson's disease, Alzheimer's disease and focal ischemia, since the diseases also relate to MKK7 and/or MKK4.
  • the compounds of formula (I) are those wherein:
  • R 1 is amino
  • R 2 is pyrrolidyl(C 1-6 )alkyl, saturated(C 3-8 )cycloalkyl or benzyl, and
  • R 3 is hydrogen, (C 1-6 )alkyl, or benzyl
  • R 2 and R 3 are taken together with the attached nitrogen atom to form
  • a 7 membered saturated heterocyclic ring optionally interrupted by one or two atoms selected from the group consisting of oxygen, sulfur and nitrogen, a 5-7 membered saturated heterocyclic ring which is fused by benzene or saturated(C 3-8 )cycloalkyl,
  • a 8-12 membered spiro ring optionally interrupted by one or two oxygen atoms;
  • the compounds of formula (I) are those wherein:
  • R 1 is amino
  • R is carbamoyl or amino (C 1-6 )alkyl
  • R 5 is hydrogen or (C ⁇ -6 )alkyl.
  • the compounds of formula (I) are those wherein:
  • R 4 is carbamoyl
  • R 5 is hydrogen
  • the compounds of formula (I) are those wherein:
  • R 1 is hydrogen, methyl or amino
  • R 4 is carbamoyl
  • R is hydrogen
  • the compounds of formula (I) are those wherein: R 2 and R 3 independently represent hydrogen, (C 1-6 )alkyl optionally substituted by amino, hydroxy, methoxy, imidazolyl, (C 1-6 )alkoxy or (C 1-6 )alkoxycarbonyl,
  • (C 3-8 )cycloalkyl optionally substituted by hydroxy or hydroxy (C 1-6 )alkyl, benzyl, or 5-7 membered saturated heterocyclic ring interrupted by one or two oxygen atoms;
  • R 2 and R 3 are taken together with the attached nitrogen atom to form a saturated heterocyclic ring selected from the group consisting of pyrrolidino, piperidino, piperazino, homopiperidino, and morpholino
  • heterocyclic ring is optionally substituted by amino, methylamino, acetamido, hydroxy, oxo, carbamoyl, benzyl, bromo, carboxy, pyrrolidino, or (C 1-6 )alkyl optionally substituted by hydroxy, amino or carboxy,
  • heterocyclic ring is optionally fused by benzene or saturated(C 3-8 )- cycloalkyl
  • R is carbamoyl or amino(C ⁇ -6 )alkyl
  • R 5 is hydrogen
  • the compounds of formula (I) are those wherein:
  • R 1 is hydrogen, methyl or amino
  • R 2 and R 3 independently represent hydrogen, (C 1-6 )alkyl optionally substituted by amino, hydroxy, methoxy, imidazolyl, (C ⁇ -6 )alkoxy or (C -8 )cycloalkyl optionally substituted by hydroxy or hydroxy (C 1-6 )alkyl, benzyl, or 5-7 membered saturated heterocyclic ring interrupted by one or two oxygen atoms;
  • R 2 and R 3 are taken together with the attached nitrogen atom to form a saturated heterocyclic ring selected from the group consisting of pyrrolidino, piperidino, piperazino, homopiperidino, and morpholino
  • heterocyclic ring is optionally substituted by amino, methylamino, acetamido, hydroxy, oxo, carbamoyl, benzyl, bromo, carboxy, pyrrolidino, or (C 1-6 )alkyl optionally substituted by hydroxy, amino or carboxy,
  • heterocyclic ring is optionally fused by benzene or saturated(C 3-8 )- cycloalkyl
  • R is carbamoyl
  • R 5 is hydrogen
  • the compounds of formula (I) are those wherein:
  • R 1 is amino
  • R 2 is saturated(C 3-8 )cycloalkyl or benzyl;
  • R 3 is hydrogen, or (C 1-6 )alkyl,
  • R 2 and R 3 are taken together with the attached nitrogen atom to form
  • heterocyclic ring is selected from the group consisting of pyrrolidino, piperidino, piperazino, homopiperidino, and morpholino,
  • R is hydrogen
  • the compounds of formula (I) are those wherein:6- Carbamoyl-4-( 1 -piperidinyl)-9H-pyrimido [4,5 -bjindole,
  • 6-Carbamoyl-4 (4-hydroxy-l-piperidinyl)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4- (cycloheptylamino)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4- ⁇ (4-methyl- 1 -piperidinyl)-9H-pyrimido [4,5 -b]indole, 6-Carbamoyl-4 (3-methyl-l-piperidinyl)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4 (3,5-dimethyl-l-pi ⁇ eridinyl)-9H-pyrimido[4,5-b]indole,
  • 6-Carbamoyl-4 [2-(hydroxymethyl)-l-pi ⁇ eridinyl]-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4- (cyclo ⁇ entylamino)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4- -(l-azepanyl)-9H-pyrimido[4,5-b]indole,
  • the present invention provides a medicament which include one of the compounds described above and optionally pharmaceutically acceptable excipient.
  • the compound of the formula (I) of the present invention can be, but not limited to be, prepared by combining various known methods.
  • one or more of the substituents, such as amino group, carboxyl group, and hydroxyl group of the compounds used as starting materials or intermediates are advantageously protected by a protecting group known to those skilled in the art. Examples of the protecting groups are described in "Protective Groups in Organic Synthesis (2nd Edition)" by Greene and Wuts.
  • the compound of the formula (I) of the present invention can be, but not limited to be, prepared by the method [A] below.
  • the compound (I) (wherein R 1 , R 2 , R 3 , R 4 and R 5 are the same as defined above) or a salt thereof, for example, can be prepared by the reaction of the compound of formula (II) (wherein R 1 , R 4 and R 5 are the same as defined above, and L represents leaving group including, for instance, halogen atom such as chlorine, bromine, or iodine atom; C 6-10 arylsulfonyloxy group such as benzenesulfonyloxy or p-toluene- sulfonyloxy; C 1-4 alkylsulfonyloxy group such as methanesulfonyloxy; and halogen substituted C 1-4 alkylsulfonyloxy group such as trifluoromethanesulfonyloxy and the like.) or a salt thereof, with a compound shown by the general formula (III) (wherein R 2 and R 3 are the same as defined above ) or
  • This reaction can be carried out without solvent or in a solvent including, for instance, alcohol such as methanol and ethanol; ethers, such as dioxane, diethyl ether, and tetrahydrofuran (THF); aromatic hydrocarbons such as benzene, toluene and xylene; nitriles such as acetonitrile; amides such as dimethylformamide (DMF) and dimethylacetamide; sulfoxides such as dimethyl sulfoxide, and others.
  • a solvent including, for instance, alcohol such as methanol and ethanol; ethers, such as dioxane, diethyl ether, and tetrahydrofuran (THF); aromatic hydrocarbons such as benzene, toluene and xylene; nitriles such as acetonitrile; amides such as dimethylformamide (DMF) and dimethylacetamide; sulfoxides such as dimethyl sul
  • the reaction temperature can be optionally set depending on the compounds to be reacted.
  • the reaction temperature is usually, but not limited to, about 10°C to 200°C and preferably about 20°C to 100°C
  • the reaction may be conducted for, usually, 30 mins to 48 hrs and preferably 1 hr to 24 hrs.
  • the reaction can be advantageously conducted in the presence of a base.
  • a base examples include an alkali metal hydride such as sodium hydride or potassium hydride; alkali metal alkoxide such as sodium methoxide or sodium ethoxide; alkali metal hydroxide such as sodium hydroxide or potassium hydroxide; carbonates such as sodium carbonate or potassium carbonate, and bicarbonates such as sodium hydrogen carbonate and potassium hydrogen carbonate; organic amines such as triethylamine.
  • the amine (III) are commercially available or can be synthesized by conventional methods. If required, R 1 , R 2 , R 3 , R 4 and R 5 can be optionally protected during the reaction and deprotected afterward.
  • the compound of the formula (I-a) can be, but not limited to be, prepared by the methods [B] or [C] below. Method [B]
  • the compound of formula (I-a) (wherein R 1 , R 2 , R 3 and R 5 are the same as defined above) can be prepared by the reaction of the compound of formula (I-b)(wherein R 1 ,
  • R 2 , R 3 and R 5 are the same as defined above, and X represents hydrogen or (C 1-6 )- alkyl) and ammonia.
  • the reaction can be carried out without solvent or in a solvent including, for instance, alcohols such as methanol and ethanol, 1 -propanol, isopropanol and tert-butanol; water; ethers such as diethyl ether, isopropyl ether, dioxane and tetrahydrofuran (THF) and 1,2-dimethoxyethane; aromatic hydrocarbons such as benzene, toluene and xylene; amides such as N, N-dimethylformamide (DMF), N, N-dimethyl- acetamide; sulfoxides such as dimethyl sulfoxide, and others.
  • a solvent including, for instance, alcohols such as methanol and ethanol, 1 -propanol, isopropanol and tert-butanol; water; ethers such as diethyl ether, isopropyl ether, dioxane and tetrahydro
  • the reaction temperature can be optionally set depending on the compounds to be reacted.
  • the reaction temperature is usually, but not limited to, about 0°C to 60°C
  • the reaction may be conducted for, usually, 30 mins to 48 hrs and preferably 1 hr to 24 hrs.
  • the reaction can be advantageously carried out using coupling agent including, for instance, carbo- diimides such as N, N-dicyclohexylcarbodiimide and l-(3-dimethylaminopropyl)-3- ethylcarbodiimide; carbonyldiazoles such as l,l'-carbonyldi(l,3-imiazole)(CDI) and l,l'-carbonyldi(l,2,4-triazole)(CDT), and others.
  • coupling agent including, for instance, carbo- diimides such as N, N-dicyclohexylcarbodiimide and l-(3-dimethylaminopropyl)-3- ethylcarbodiimide; carbonyldiazoles such as l,l'-carbonyldi(l,3-imiazole)(CDI) and l,l'-carbonyldi(l,2,4-triazole)(CD
  • the compound of formula (I-a) (wherein R , R 2 , R 3 and R 5 are the same as defined above) can be prepared by the hydrolysis of the compound of formula (I-c) (wherein R 1 , R 2 , R 3 and R 5 are the same as defined above).
  • the reaction can be carried out in a solvent including, for instance, alcohols such as methanol and ethanol, 1 -propanol, isopropanol, n-butanol and tert-butanol; water; ketone such as acetone; ethers such as diethyl ether, isopropyl ether, dioxane and tetrahydrofuran (THF) and 1,2-dimethoxyethane; aromatic hydrocarbons such as benzene, toluene and xylene; amides such as N, N-dimethylformamide (DMF), N, N- dimethylacetamide, and others.
  • a solvent including, for instance, alcohols such as methanol and ethanol, 1 -propanol, isopropanol, n-butanol and tert-butanol; water; ketone such as acetone; ethers such as diethyl ether, isopropyl ether
  • the reaction temperature can be optionally set depending on the compounds to be reacted.
  • the reaction temperature is usually, but not limited to, about 0°C to 60°C
  • the reaction may be conducted for, usually, 30 mins to 48 hrs and preferably 1 hr to 24 hrs.
  • the reaction can be advantageously conducted in the presence of a base, including, for instance, an alkali metal alkoxide such as sodium methoxide, sodium ethoxide and potassium tert-butoxide; alkali metal hydroxide such as sodium hydroxide and potassium hydroxide; alkali metal carbonates such as sodium carbonate and potassium carbonate; alkali metal hydrogen carbonates such as sodium hydrogen carbonate and potassium hydrogen carbonate; alkali metal phosphate such as sodium phosphate, and others.
  • a base including, for instance, an alkali metal alkoxide such as sodium methoxide, sodium ethoxide and potassium tert-butoxide; alkali metal hydroxide such as sodium hydroxide and potassium hydroxide; alkali metal carbonates such as sodium carbonate and potassium carbonate; alkali metal hydrogen carbonates such as sodium hydrogen carbonate and potassium hydrogen carbonate; alkali metal phosphate such as sodium phosphate, and others.
  • the reaction can be advantageously conducted in the presence of oxidating agent, for instance, hydrogen peroxide, manganese dioxide, dimethyl dioxirane, sodium percarbonate, sodium perborate, oxone, and the others.
  • oxidating agent for instance, hydrogen peroxide, manganese dioxide, dimethyl dioxirane, sodium percarbonate, sodium perborate, oxone, and the others.
  • the reaction can be advantageously conducted in the presence of an acid including, for instance, trifluoroacetic acid, hydrochloric acid and sulfonic acid, and others.
  • an acid including, for instance, trifluoroacetic acid, hydrochloric acid and sulfonic acid, and others.
  • compound 2 can be prepared by the reaction of compound 1 (wherein L' represents leaving group including, for instance, halogen atom such as chlorine, bromine, or iodine atom; C 6-10 arylsulfonyloxy group such as benzenesulfonyloxy or p-toluenesulfonyloxy; C 1-4 alkylsulfonyloxy group such as methanesulfonyloxy; and halogen substituted C 1- alkylsulfonyloxy group such as trifluoromethanesulfonyloxy and the like, and R and R 5 are the same as defined above) with ethyl cyanoacetate using a base, for instance, sodium hydride.
  • L' represents leaving group including, for instance, halogen atom such as chlorine, bromine, or iodine atom
  • C 6-10 arylsulfonyloxy group such as benzenesulfonyloxy or p
  • the reaction may be carried out in a solvent including, for instance, ethers such as diethyl ether, isopropyl ether, dioxane and tetrahydrofuran (THF) and 1,2- dimethoxyethane; aromatic hydrocarbons such as benzene, toluene and xylene; amides such as N, N-dimethylformamide (DMF), N, N-dimethylacetamide and N- methylpyrrolidone; sulfoxides such as dimethylsulfoxide (DMSO); alcohols such as methanol, ethanol, 1 -propanol, isopropanol and tert-butanol,; and others.
  • a solvent including, for instance, ethers such as diethyl ether, isopropyl ether, dioxane and tetrahydrofuran (THF) and 1,2- dimethoxyethane; aromatic hydrocarbons such as benzene, tolu
  • Compound 3 (wherein R and R 5 are the same as defined above) can be prepared by reducing nitro group of compound 2 using agent including, for instance, metals such as zinc and iron in the presence of acid including, for instance, hydrochloric acid and acetic acid.
  • agent including, for instance, metals such as zinc and iron in the presence of acid including, for instance, hydrochloric acid and acetic acid.
  • the reaction can be carried out without solvent or in a solvent including, for instance; aromatic hydrocarbons such as benzene, toluene and xylene, and others.
  • the reaction may be carried out, usually, at room temperature to 100 °C for 30 mins to 12 hrs.
  • Compound 4a (wherein R 1 represents amino, (C 1-6 )alkyl, (C 2-6 )alkenyl, (C 2-6 )alkynyl, halogen substituted (C 1-6 ) alkyl, cyano, cyano(C 1-6 )alkyl, (C 3-8 )cycloalkyl, nitro- (C ⁇ _ 6 )alkyl or fluoro and R 4 and R 5 are the same as defined above) can be prepared by the reaction of compound 3 with appropriate cyano compounds (R ⁇ CN) (wherein R 1' is the same as defined above).
  • the reaction can be carried out in a solvent including, for instance, alcohols such as methanol, ethanol, 1 -propanol, isopropanol and tert- butanol; ethers such as diethyl ether, isopropyl ether, dioxane and tetrahydrofuran (THF) and 1,2-dimethoxyethane; aromatic hydrocarbons such as benzene, toluene and xylene, and others.
  • a solvent including, for instance, alcohols such as methanol, ethanol, 1 -propanol, isopropanol and tert- butanol; ethers such as diethyl ether, isopropyl ether, dioxane and tetrahydrofuran (THF) and 1,2-dimethoxyethane; aromatic hydrocarbons such as benzene, toluene and xylene, and others.
  • the reaction may be carried out, usually,
  • Cyano compounds are commercially available or can be synthesized by conventional methods.
  • Compound 4b (wherein R 1 is hydrogen and R 4 and R 5 are the same as defined above) can be prepared by the reaction of compound 3 with ammonium formate in a solvent such as formamide. The reaction may be carried out, usually, at 40°C to 180°C for 2 hrs to two days. If desired, the resulting 4b can be further modified to introduce nitro group at the position of R 1 .
  • Ammonium formate and formamide are commercially available or can be synthesized by conventional methods.
  • the compound of formula (II) (wherein R 1 , R 4 and R 5 are the same as defined above and L represents leaving group including, for instance, halogen atom such as chlorine, bromine, or iodine atom; C 6-10 arylsulfonyloxy group such as benzenesulfonyloxy or p-toluenesulfonyloxy; C 1- alkylsulfonyloxy group such as methanesulfonyloxy; and halogen substituted C 1-4 alkylsulfonyloxy group such as trifluoro- methanesulfonyloxy and the like.) can be prepared for instance, by the reaction of compound 4 with appropriate halogenating reagent including, for instance, POCl 3 , PC1 5 , SOCl 2 , and the like; or can be prepared, for instance, by the reaction of compound 4 with appropriate sulfonyl chloride.
  • appropriate halogenating reagent including,
  • the reaction may be carried out without solvent or in a solvent including, for instance, halogenated hydrocarbons such as dichloromethane, chloroform and 1,2- dichloroethane;such as ethers such as dioxane and tetrahydrofuran (THF) and 1,2- dimethoxyethane; aromatic hydrocarbons such as benzene, toluene, and xylene, and others.
  • halogenated hydrocarbons such as dichloromethane, chloroform and 1,2- dichloroethane
  • ethers such as dioxane and tetrahydrofuran (THF) and 1,2- dimethoxyethane
  • aromatic hydrocarbons such as benzene, toluene, and xylene, and others.
  • two or more of the solvents selected from the listed above can be mixed and used.
  • the reaction can be advantageously conducted in the presence of a base, including, for instance, such as pyridine, triethylamine and N,N-diiso- propylethylamine, dimethylaniline, diethylaniline, and others.
  • a base including, for instance, such as pyridine, triethylamine and N,N-diiso- propylethylamine, dimethylaniline, diethylaniline, and others.
  • the reaction temperature is usually, but not limited to, about 40°C to 200°C and preferably about 20°C to 180°C
  • the reaction may be conducted for, usually, 30 minutes to 48 hrs and preferably 2 hr to 12 hrs.
  • halogenating reagents and sulfonyl chlorides are commercially available or can be synthesized by conventional methods.
  • Typical salts of the compound shown by the formula (I) include salts prepared by reaction of the compounds of the present invention with a mineral or organic acid, or an organic or inorganic base. Such salts are known as acid addition and base addition salts, successively.
  • Acids to form salts include inorganic acids such as, without limitation, sulfuric acid, phosphoric acid, hydrochloric acid, hydrobromic acid, hydriodic acid like, and organic acids, such as, without limitation, p-toluenesulfonic acid, methanesulfonic acid, oxalic acid, p-bromophenylsulfonic acid, succinic acid, citric acid, benzoic acid, acetic acid, and the like.
  • inorganic acids such as, without limitation, sulfuric acid, phosphoric acid, hydrochloric acid, hydrobromic acid, hydriodic acid like
  • organic acids such as, without limitation, p-toluenesulfonic acid, methanesulfonic acid, oxalic acid, p-bromophenylsulfonic acid, succinic acid, citric acid, benzoic acid, acetic acid, and the like.
  • Base addition salts include those derived from inorganic bases, such as, without limitation, ammonium hydroxide, alkaline metal hydroxide, alkaline earth metal hydroxides, carbonates, bicarbonates, and the like, and organic bases, such as, without limitation, ethanolamine, triethylamine, tris(hydroxymethyl)aminomethane, and the like.
  • inorganic bases include, sodium hydroxide, potassium hydroxide, potassium carbonate, sodium carbonate, sodium bicarbonate, potassium bicarbonate, calcium hydroxide, calcium carbonate, and the like.
  • the compound of the present invention or a salts thereof, depending on its substituents, may be modified to form lower alkylesters or known other esters; and/or hydrates or other solvates. Those esters, hydrates, and solvates are included in the scope of the present invention.
  • the compound of the present invention may be administered in oral forms, such as, without limitation normal and enteric coated tablets, capsules, pills, powders, granules, elixirs, tinctures, solution, suspensions, syrups, solid and liquid aerosols and emulsions. They may also be administered in parenteral forms, such as, without limitation, intravenous, intraperitoneal, subcutaneous, intramuscular, and the like forms, well-known to those of ordinary skill in the pharmaceutical arts.
  • the compounds of the present invention can be administered in intranasal form via topical use of suitable intranasal vehicles, or via transdermal routes, using transdermal delivery systems well-known to those of ordinary skilled in the art.
  • the dosage regimen with the use of the compounds of the present invention is selected by one of ordinary skill in the arts, in view of a variety of factors, including, without limitation, age, weight, sex, and medical condition of the recipient, the severity of the condition to be treated, the route of administration, the level of metabolic and excretory function of the recipient, the dosage form employed, the particular compound and salt thereof employed.
  • the compounds of the present invention are preferably formulated prior to administration together with one or more pharmaceutically-acceptable excipients. Excipients are inert substances such as, without limitation carriers, diluents, flavoring agents, sweeteners, lubricants, solubilizers, suspending agents, binders, tablet disintegrating agents and encapsulating material.
  • compositions of the present invention are pharmaceutical formulation comprising a compound of the invention and one or more pharmaceutically- acceptable excipients that are compatible with the other ingredients of the formulation and not deleterious to the recipient thereof.
  • Pharmaceutical formulations of the invention are prepared by combining a therapeutically effective amount of the compounds of the invention together with one or more pharmaceutically-acceptable excipients.
  • the active ingredient may be mixed with a diluent, or enclosed within a carrier, which may be in the form of a capsule, sachet, paper, or other container.
  • the carrier may serve as a diluent, which may be solid, semi-solid, or liquid material wliich acts as a vehicle, or can be in the form of tablets, pills, powders, lozenges, elixirs, suspensions, emulsions, solutions, syrups, aerosols, ointments, containing, for example, up to 10% by weight of the active compound, soft and hard gelatin capsules, suppositories, sterile injectable solutions and sterile packaged powders.
  • a diluent which may be solid, semi-solid, or liquid material wliich acts as a vehicle, or can be in the form of tablets, pills, powders, lozenges, elixirs, suspensions, emulsions, solutions, syrups, aerosols, ointments, containing, for example, up to 10% by weight of the active compound, soft and hard gelatin capsules, suppositories, sterile injectable solutions and sterile packaged powder
  • the active ingredient may be combined with an oral, and non-toxic, pharmaceutically-acceptable carrier, such as, without limitation, lactose, starch, sucrose, glucose, sodium carbonate, mannitol, sorbitol, calcium carbonate, calcium phosphate, calcium sulfate, methyl cellulose, and the like; together with, optionally, disintegrating agents, such as, without limitation, maize, starch, methyl cellulose, agar bentonite, xanthan gum, alginic acid, and the like; and optionally, binding agents, for example, without limitation, gelatin, natural sugars, beta-lactose, corn sweeteners, natural and synthetic gums, acacia, tragacanth, sodium alginate, carboxymethylcellulose, polyethylene glycol, waxes, and the like; and, optionally, lubricating agents, for example, without limitation, magnesium stearate, sodium stearate, stearic acid, sodium oleate, sodium benzoate,
  • the carrier may be a finely divided solid which is in admixture with the finely divided active ingredient.
  • the active ingredient may be mixed with a carrier having binding properties in suitable proportions and compacted in the shape and size desired to produce tablets.
  • the powders and tablets preferably contain from about 1 to about 99 weight percent of the active ingredient which is the novel composition of the present invention.
  • Suitable solid carriers are magnesium carboxymethyl cellulose, low melting waxes, and cocoa butter.
  • Sterile liquid formulations include suspensions, emulsions, syrups and elixirs.
  • the active ingredient can be dissolved or suspended in a pharmaceutically acceptable carrier, such as sterile water, sterile organic solvent, or a mixture of both sterile water and sterile organic solvent.
  • the active ingredient can also be dissolved in a suitable organic solvent, for example, aqueous propylene glycol.
  • a suitable organic solvent for example, aqueous propylene glycol.
  • Other compositions can be made by dispersing the finely divided active ingredient in aqueous starch or sodium carboxymethyl cellulose solution or in a suitable oil.
  • the formulation may be in unit dosage form, which is a physically discrete unit containing a unit dose, suitable for administration in human or other mammals.
  • a unit dosage form can be a capsule or tablets, or a number of capsules or tablets.
  • a "unit dose" is a predetermined quantity of the active compound of the present invention, calculated to produce the desired therapeutic effect, in association with one or more excipients.
  • the quantity of active ingredient in a unit dose may be varied or adjusted from about 0.1 to about 1000 milligrams or more according to the particular treatment involved.
  • Typical oral dosages of the present invention when used for the indicated effects, will range from about 0.0 lmg ⁇ cg/day to about 100 mg/kg/day, preferably from 0.1 mg/kg/day to 30 mg/kg/day, and most preferably from about 0.5 mg/kg/day to about 10 mg/kg/day.
  • parenteral administration it has generally proven advantageous to administer quantities of about 0.001 to lOOmg /kg/day, preferably from 0.01 mg/kg/day to 1 mg/kg/day.
  • the compounds of the present invention may be administered in a single daily dose, or the total daily dose may be administered in divided doses, two, three, or more times per day. Where delivery is via transdermal forms, of course, administration is continuous.
  • MS MS data were recorded on a Micromass Platform LC with Shimadzu Phenomenex ODS column(4.6 mm ⁇ X 30 mm) flushing a mixture of acetonitrile- water (9:1 to 1:9) at 1 ml/min of the flow rate.
  • Mass spectra were obtained using electrospray (ES) ionization techniques (micromass Platform LC).
  • TLC was performed on a precoated silica gel plate (Merck silica gel 60 F-254).
  • Silica gel WAKO-gel C-200 (75-
  • a plasmid containing human MKK7 open reading frame was cloned into a pGEM-T Easy vector (Promega, Madison, WI) and further into a pGEX-6P-2 vector (Pharmacia) to construct human GST(Glutathione-S-transferase)-
  • MKK7 fusion protein This construct was coexpressed with human MEKKc (catalytic domain of MEKK (MEK (Map kinase kinase) kinase) on plasmid pBB131, in E.coli (BL21(DE3)pLysS).
  • MEKKc catalytic domain of MEKK (MEK (Map kinase kinase) kinase) on plasmid pBB131, in E.coli (BL21(DE3)pLysS).
  • the resulting GST-MKK7 was purified with the use of a glutathione column
  • GST-KN-SAPK ⁇ was purified with the use of glutathione column (Amersham Pharmacia Biotech AB, Uppsala, Sweden) according to the manufacturer's instruction. The purity of the protein was confirmed to be more than 90% by SDS-PAGE.
  • Test compounds (2.5 ⁇ l) at various concentrations (in 1% DMSO) were added to 15 ⁇ l of reaction buffer (20mM HEPES, 0.1M NaCl, 0.1 mM NasNO4, lOmM MgCk, ImM DTT, lmg/ml BSA, pH 7.5)) containing
  • a plasmid containing human MKK4 open reading frame was cloned into a pGEX-2T vector (Pharmacia) to construct human GST(Glutathione-S- transferase)-MKK4 fusion protein. This construct was coexpressed with human MEKKc (catalytic domain of MEKK on plasmid pBB131) in E.coli (BL21(DE3)pLysS).
  • the resulting GST-MKK4 was purified with the use of glutathione column (Amersham Pharmacia Biotech AB, Uppsala, Sweden) according to the manufacturer's instruction. The purity of the protein was confirmed to be more than 90% by SDS-PAGE.
  • GST-KN-SAPK ⁇ was purified with the use of glutathione column (Amersham Pharmacia Biotech AB, Uppsala, Sweden) according to the manufacturer's instruction. The purity of the protein was confirmed to be more than 90% by SDS-PAGE.
  • Biotinylation of the substrate protein was done using sulfo-NHS-LC Biotin according to the manufacturer's instructions (Pierce, Rockford, US)
  • Test compounds (5 ⁇ l) at various concentrations (in 1% DMSO) were added to 30 ⁇ l of reaction buffer (20mM HEPES, 0.1M NaCl, 0.1 mM Na3NO4, lOmM MgCk, ImM DTT, lmg/ml BSA, pH 7.5)) containing 0.5 ⁇ g/ml GST-MKK4 and 6 ⁇ M ATP.
  • reaction buffer (20mM HEPES, 0.1M NaCl, 0.1 mM Na3NO4, lOmM MgCk, ImM DTT, lmg/ml BSA, pH 7.5)
  • the kinase reaction was started by the addition of 25 ⁇ l assay buffer containing 0.48 ⁇ M SAPK ⁇ (biotinylated
  • huPBMC Human peripheral blood mononucleated cells isolated using mono-poly resolving medium (Dainippon Seiyaku, Osaka, Japan) were incubated with test compounds (various concentrations in 0.1% DMSO) for 1 hour in a 37°C CO2 incubator. Cells were then plated on 96 well plates (1x10s cell per well in 200 ⁇ l RPMI1640 cell culture medium) pre-coated for 3 hrs with 100 ⁇ l anti-CD3 antibody
  • interleukin- 2 IL-2
  • IFN- ⁇ interferon- ⁇
  • huPBMC Human peripheral blood mononucleated cells isolated using mono-poly resolving medium were either directly used for experiments (lxl 0s cells per well in 200 ⁇ l medium) or differentiated to dendritic cells (DC) in the presence of GM-SCF
  • TNF- ⁇ or IL-12 was induced by stimulation with LPS (B8, Sigma, Missouri, US) 10 ng/ml). 20 hrs later supernatant was removed and stored at -30°C in aliquots until further use.
  • TNF- ⁇ and IL-12 released from cell cultures were determined by commercially available ELISA (Genzyme Tech., Minneapolis, US) according to the manufacturer's instructions.
  • mice Male Balb/c mice (20-25 g body weight) were in injected with agonistic anti-CD3 Ab (Pharmingen, San Diego, US; 10 ⁇ g/mouse; clone 145-2C11) i.v. 5 minutes after compound application (i.v. in 10 % . 2 hrs post-challenge, mice were sacrificed and the serum cytokines IL-2, IL-4 and IFN- ⁇ were determined by ELISA (Genzyme Tech., Minneapolis, US) according to the manufacturer's instruction. Data represent mean values ⁇ SD of 5-6 animals each. * p ⁇ 0.05, ** p ⁇ 0.01 vs. vehicle control (V); Dunnett's test was used to detect differences among groups and statistics were performed using one-sided ANONA or Student's t-test if applicable.
  • MKK 7 kinase assay MKK 7 kinase assay
  • MKK4 kinase assay MKK4 kinase assay
  • the compounds of the present invention also show excellent selectivity, and strong activity in in vivo assays.
  • NBS N-bromosuccinimide
  • benzoyl peroxide 0.45 g
  • the reaction mixture was passed through a filter paper to remove resulting precipitates, that were washed with chloroform (50 ml). The filtrates were combined and washed with saturated sodium thiosulfite water solution and brine, successively.
  • Ethyl ⁇ -cyano-5-methoxycarbonyl-2-nitrophenylacetate (3.10 g) was dissolved in glacial acetic acid (30 ml) at 80°C Keeping the temperature, zinc powder (5.55 g) was added portionwise. The mixture was stirred at 90°C to 100°C for 1 h. After cooling to room temperature, the reaction mixture was passed through a filter paper to remove insoluble materials and washed with glacial acetic acid (10 ml). The filtrates were combined and evaporated in vacuo. The residue was dissolved in ethyl acetate (30 ml) and washed with saturated sodium bicarbonate water solution (30 ml X 3) and brine (30 ml), successively.
  • 6-carbamoyl-4-(l-piperidinyl)-9H-pyrimido[4,5-b]indole (0.028 g, 56%) as a colorless powder.
  • Example 1-4 to 1-32 were synthesized.
  • ethyl 2-amino-6-cyanoindole-3 -carboxylate was prepared according to the procedure described in Example 1-1. To a suspension of ethyl 2-amino-6-cyanoindole-3-carboxylate (0.96 g) and cyanamide (0.88 g) in 1,4-dioxane (50 ml) was added 36% hydrochloric acid (0.84 ml). The mixture was refluxed for 2 days and, after cooling to room temperature, concentrated in vacuo. The residue was washed with diethylether and triturated with methanol to give precipitates, that was collected by a paper filter and washed with methanol. The collected solid was dried at 85°C in vacuo to give a colorless solid (0.72 g, 66%).
  • Example 4-3 to 4-13 as shown in Table 4 were synthesized. Table 4
  • 6-benzyloxy-4-chloro-(9H)- pyrimido[4,5-b]indole hydrochloride (0.12 g, 0.355 mmol) was reacted with piperidine (1.5 mL) in 1,4-dioxane (5.0 mL) to give 6-benzyloxy ⁇ 4-(l- piperidino)-(9H)-pyrimido[4,5-b]indole as a pale orange solid (0.024 g, 18.8%).
  • Example 5 In the similar manners as described in Example5-1 above, the compounds in Example 5-2 to 5-59 as shown in Table 5 were synthesized. Table 5
  • Example 6-2 to 6-16 as shown in Table 6 were synthesized.

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Abstract

The present invention relates to pyrimido[4,5-b]indoles which are useful as an active ingredient of pharmaceutical preparations. The pyrimido[4,5-b]indoles of the present invention have MKK7 and MKK4 inhibitory activity, and can be used for the prophylaxis and treatment of diseases associated with MKK7 and MKK4 activity. Such diseases include, inflammatory and immunoregulatory disorders and diseases such as asthma, atopic dermatitis, rhinitis, allergic rhinitis, allergic diseases, COPD, septic shock, arthritis, joint diseases and myocardial injuries, as well as autoimmune pathologies such as rheumatoid arthritis, Grave's disease, and atherosclerosis. The compounds of the present invention are also useful for treatment of ischemia, myocardial injury, pulmonary hypertension, renal failure, Huntington's chorea and cardiac hypertrophy, as well as neurodegenerative disorders such as Parkinson's disease, Alzheimer's disease and focal ischemia.

Description

PYRIMIDO 5-b] INDOLE Derivatives
Detailed Description of Invention
Technical Field
The present invention relates to pyrimido[4,5-b]indoles which are useful as an active ingredient of pharmaceutical preparations. The pyrimido[4,5-b]indoles of the present invention have MKK7 and KK4 [MAPK (mitogen activated protein kinase) kinase
7 and 4] inhibitory activity, and can be used for the prophylaxis and treatment of diseases associated with MKK7 and MKK4 activity.
More specifically, the pyrimido[4,5-b]indoles derivatives of the present invention are useful for treatment and prophylaxis of diseases as follows: inflammatory and immunoregulatory disorders, such as asthma, atopic dermatitis, rhinitis, allergic rhinitis, allergic diseases, COPD, septic shock, arthritis, joint diseases and myocardial injuries, as well as autoimmune pathologies such as rheumatoid arthritis, Grave's disease, and atherosclerosis.
The compounds of the present invention are also useful for treatment of ischemia, myocardial injury, pulmonary hypertension, renal failure, Huntington's chorea and cardiac hypertrophy, as well as neurodegenerative disorders such as Parkinson's disease, Alzheimer's disease and focal ischemia, since the diseases also relate to MKK7 and/or MKK4.
Background Art
The mitogen-activated protein kinases (MAPKs) are a family of serine/threonine kinases involved in the transduction of signals from the cell membrane to the nucleus in response to various types of stimuli such as lipopolysaccharide (LPS), tumor necrosis factor-α (TNF- ), interleukins, CD40 and others. These kinases participate in a wide variety of signaling cascades controlling cellular events such as cell growth, differentiation, activation, apoptosis, stress responses, and transformation. Three subgroups of MAPK currently are known as extracellular-regulated kinases (ERK), ρ-38 MAPK, and stress-activated/c-jun N-terminal kinase (SAPK/JNK) (Pouyssegur J. An arresting start for MAPK. Science, 290: 1515-1518, 2000 ).
SAPK/JNKs are activated in response to cellular "stress" such as changes in osmolarity or metabolism, ischemia, heat shock, shear stress, ceramide or inflammatory cytokines (TNF-α, IL-1). Once activated, JNKs control gene activity via phosphorylation of a variety of transcriptional factors including c-Jun, JunD, nuclear factor of activated T cells (NFAT)4, or Elk-1 , all present in immune cells and known to regulate the transcription of many genes during an inflammatory response. Thus, among other functions such as induction of pro-inflammatory cytokines and Thl/Th2 differentiation, SAPK/JNKs regulate the activation and proliferation of T and B lymphocytes, activation of mast cells [Sasaki T., Wada T., Kishimoto, H., Irie-Sasaki J., Matsumoto G., Goto T., Yao Z. et al., The stress kinase mitogen-activated protein kinase kinase (MKK)7 is a negative regulator of antigen receptor and growth factor receptor-induced proliferation in hematopoietic cells. J. Exp Med, 194:1-14, 2001].
To become activated, MAPKs themselves require dual phosphorylation of both threonine and tyrosine at their so-called Thr-X-Tyr motif, wliich is brought upon by the upstream regulators MAPK kinases (MKKs). MKK1-MKK7 (MEK1, MEK2,
MKK3, MKK4, MEK5, MKK6, and MKK7) are known to date with MKK7 being the most recently identified (Tournier C, Whitmarsh J., Cavanagh J., Barrett T.,
Davis RJ. Mitogen-activated protein kinase kinase 7 is an activator of the c-Jun NH2- terminal kinase. Proc Natl Acad Sci USA, 94: 7337-7342, 1997), (Moriguchi T.,
Toyoshima F., Masuyama N., Hanafusa H., Gotoh Y., Nishida E. A novel
SAPK JNK kinase, MKK7, stimulated by TNFα and cellular stresses. EMBO J, 16:
7045-7053, 197) and (Lu X., Nemoto S., Lin A. Identification of c-Jun NH2-terminal protein kinase (JNK)-activating kinase 2 as an activator of JNK but not p38. J Biol Chem, 272: 24751-24754, 1997). Among this family of kinases MKK4 and MKK7 are the only ones capable of phosphorylating SAPK/_JNKs. Overexpression of dominant negative forms of these MKKs and the use of cells from mice lacking MKK4 or MKK7 have clearly shown their implication in the regulation of many inflammatory responses. Whereas MKK7 is believed to exclusively use SAPK/JNKs as substrates, MKK4 is also capable of phosphorylating p-38 MAP kinases. p-38 kinases are also involved in the control of inflammatory gene expression, especially after stimulation of cells with lipopoly saccharide and cytokines (Han J., Lee JD., Jiang Y., Li Z., Feng L., Ulevitch RJ. A MAP kinase targeted by endotoxin in mammalian cells. Science, 265: 808-811, 1994.), (Lee JC, Laydon JT., McDonnell PC, Gallagher TF., Kumar S., Green D., McNulty D., Blumenthal MJ., Heys JR., Landvatter SW., Strickler JE., McLaughlin MM., Siemens IR., Fisher SM., Livi GP.,
White JR., Adams JL., Young PR. A protein kinase involved in the regulation of inflammatory cytokine synthesis. Nature, 372: 739-746, 1994),. In T cells, p38 controls the release of IL-12 and IFNγ and in B cells, CD40 cross-linking leads to rapid p38 activation and thus controls proliferation, and adhesion molecule expression. In addition, p38 MAPK are activated by hypoxia and, by controlling the transcription factor ATF2, play a role in neuronal development and survival (Lee JC, Kumar S., Griswold DE., Underwood DC, Notta BJ., Adams JL. Inhibition of p38 MAP kinase as a therapeutic strategy. Immunopharmacology, Al: 185-201, 2000).
A specific inhibitor of MKK7 and/or of MKK4, which is expected to block the synthesis of pro-inflammatory cytokines and the activation of various immune cells, should have a broad anti-inflammatory profile with potential for the treatment of inflammatory and immunoregulatory disorders and diseases, including asthma, rhinitis, allergic diseases, septic shock, joint diseases and myocardial injuries, as well as autoimmune pathologies such as rheumatoid arthritis, Grave's disease, and atherosclerosis.
By interfering with apoptotic pathways, such inhibitors should also have therapeutic potential for the treatment of renal failure, Huntington's chorea, cardiac hypertrophy and neurodegenerative disorders such as Parkinson's disease, Alzheimer's disease and focal ischemia (Xia XG., Harding T., Weller M., Bieneman A., Uney JB., Schulz JB. Gene transfer of the JNK interacting protein- 1 protects dopaminergic neurons in the MPTP model of Parkinson's disease. Proc Natl Acad Sci USA, 98: 10433-10438, 2001).
WO 9842708 discloses anti-asthma agent represented by the general formula:
wherein
R a, R 2Λ, R2"la, R2"2a, R3'la, and R3"2a are defined in the application.
WO 9626941 discloses pharmaceutical agents represented by the general formula:
wherein
R ,2--lιbDj - Rr,2z--2^b τ R»4^--lιb0j τ R>4--2zb0j a „n„d A τ R»7,bD ^ defmed in tlιe application. WO 9519970 discloses epidermal growth-factor inhibitors of formula:
wherein
Rlc, R2c, R3c, R4c, R5c and R6c are defined in the specification.
WO 9320078 discloses pharmaceutically active compound represented by the formula:
wherein
R2-ld5 R2-2d5 R4-ld5 R4-2d5 R56-ld; R5<Wdf R56-3d> R56-4d ^ R7d ^ ^^ ^ ^ application.
IN 157280 discloses the method for preparing anti hypertension agents represented by the formula:
wherein Rle, R2e, R3e, R4e, and R5e are defined in the specification.
WO 97/02266 discloses anti-hyperproliferative disease agents represented by the general formula:
wherein
RIP, R2P, R&, q and n' are defined in the application.
WO 98/43973 also discloses anti-proliferative disease agents represented by the general formula:
wherein
Rid, R2d, R3d, R4', q and m' are defined in the application.
However, none of the references and other reference discloses pyrimido[4,5-b]- indoles derivatives having MKK7 and/or MKK4 inhibitory activity. The development of a compound, which has effective MKK7 and/or MKK4 inhibitory activity and can be used for the prophylaxis and treatment of diseases associated with MKK7 and/or MKK4 activity has been desired.
Summary of the invention
As the result of extensive studies on chemical modification of pyrimido[4,5-b]- indoles derivatives, the present inventors have found that the compounds of the structure related to the present invention have unexpectedly excellent MKK7 and/or MKK4 inhibitory activity. The present invention has been accomplished based on these findings. This invention is to provide a novel pyrimido[4,5-b]indole derivative of the formula (I), its tautomeric or stereoisomeric form, or a salt thereof:
wherein
R is hydrogen, amino, (C1-6)alkyl, (C2-6)alkenyl, (C -6)alkynyl, halogen substituted (C1-6) alkyl, cyano, cyano(C1-6)alkyl, (C -8)cycloalkyl, nitro, nitro(C1..6)alkyl or fluoro;
R2 and R3 independently represent hydrogen, (C2-6)alkenyl, (C2-6)alkynyl,
(C3-8)cycloalkyl optionally substituted by hydroxy or hydroxy (C1-6)- alkyl, (C3-8)heterocycloalkyl optionally substituted by (C1-6)alkoxy- carbonyl, (C1-6)alkylcarbonyl, (C1-6)alkylsulfonyl, arylsulfonyl, heteroarylsulfonyl, or (C1-6)alkyl which is optionally substituted by hydroxy, (C1-ό)alkoxy, (C1-6)alkoxycarbonyl, (C3-8)cycloalkyl, (C3-8)- heterocycloalkyl, carbamoyl, (C1-6)alkylcarbamoyl, cyano, amino, (C1-6)alkylamino, (C1-6)alkoxycarbonylamino, N-(C1-6)alkoxy- carbonylamino(C1-6)alkyl, arylamino, heteroaryla ino, carboxyl, nitro, trifluoromethyl, aryl, piperidinyl, imidazolyl, (C1-6)alkyl- piperidinyl, (C1-6)alkylcarbonylpiperidinyl, (C1-6)alkoxycarbonyl- piperidinyl, carbamoylpiperidinyl, ((C1-6)alkylcarbamoyl)piperidinyl, arylpiperidinyl, heteroarylpiperidinyl, aryloxycarbonylpiperidinyl, heteroaryloxycarbonylpiperidinyl, (C1-6)alkylsulfonylpiperidinyl, aryl- sulfonylpiperidinyl, heteroarylsulfonylpiperidinyl, pyrrolidyl, (C1-6)- alkylpyrrolidyl, (C1-6)alkylcarbonylpyrrolidyl, (C1,6)alkoxycarbonyl- pyrrolidyl, carbamoylpyrrolidyl, ((C1-6)alkylcarbamoyl)pyrrolidyl, arylpyrrolidyl, heteroarylpyrrolidyl, aryloxycarbonylpyrrolidyl, heteroaryloxycarbonylpyrrolidyl, (Cι-6)alkylsulfonylpyrrolidyl, aryl- sulfonylpyrrolidyl, or, heteroarylsulfonylpyrrolidyl,
or
R2 and R3 are taken together with the attached nitrogen atom to form a 3-8 membered saturated ring optionally interrupted by one or two atoms selected from the group consisting of oxygen, sulfur and nitrogen,
wherein
said saturated ring is optionally fused by benzene or 3-8 membered saturated ring having 0 to 3 nitrogen atoms,
or
said saturated ring is optionally having substituents independently selected from the group consisting of halogen, hydroxy, carbamoyl, carboxy, amino, oxo, pyrrolidino, (C1-6)alkoxy, (C2-6)alkenyl, (C2-6)- alkynyl, (C3-8)cycloalkyl, (C1-6)alkylcarbonyl, (C1-6)alkylsulfonyl, arylsulfonyl, (C1-6)alkyl, hydroxy(C1-6)alkyl, (C1-6)alkoxy(C1-6)alkyl, (C1-6)alkoxycarbonyl, (C1-6)alkoxycarbonyl(Cι-6)alkyl, (C3- )cyclo- alkyl(C1-6)alkyl, aryl(C1-6)alkyl, carbamoyl(C1-6)alkyl, (C1-6)alkyl- carbamoy^C-^alkyl, cyano(C1-6)alkyl, amino (C1-6)alkyl, N-(C1-6)- alkylamino(C1-6)alkyl, carboxyl(C1-6)alkyl, nitro (C1-6)alkyl, trifluoro- methyl(C1-6)alkyl, N-(C1-6)alkylamino, N-(C1-6)alkanoylamino, N- (Cι-6)alkoxycarbonylamino, N-[amino(C1-6)alkyl]amino, N-(C1-6)- alkoxycarbonylamino N-(C1-6)alkylamino, and N-(C1-6)alkylcarboxy- ammo
or
said substituents optionally together with the carbon to which they are attached, form a 3-8 membered saturated ring that forms a spiro ring,
wherein said spiro ring is optionally interrupted by 0 to 3 nitrogen or oxygen atoms;
R4 is halogen, hydroxy, (C1-6 )alkyl, amino (C1-6 )alkyl, (C1-6)alkoxy, cyano, carboxy, (C1-6)alkoxycarbony, or carbamoyl; and
R5 is hydrogen, or (C1-6)alkyl.
The Alkyl per se and "alk" and "alkyl" in alkoxy, alkanoyl, alkylamino, alkylamino- carbonyl, alkylaminosulphonyl, alkylsulphonylamino, alkoxycarbonyl, alkoxy- carbonylamino and alkanoylamino represent a linear or branched alkyl radical having generally 1 to 6, preferably 1 to 4 and particularly preferably 1 to 3 carbon atoms, representing illustratively and preferably methyl, ethyl, n-propyl, isopropyl, tert- butyl, n-pentyl and n-hexyl.
Alkoxy illustratively and preferably represents methoxy, ethoxy, n-propoxy, isopropoxy, tert-butoxy, n-pentoxy and n-hexoxy. Alkanoyl illustratively and preferably represents acetyl and propanoyl.
Alkylamino represents an alkylamino radical having one or two (independently selected) alkyl substituents, illustratively and preferably representing methylamino, ethylainino, n-propylamino, isopropylamino, tert-butylamino, n-pentylamino, n- hexyl-amino, N,N-dimethylamino, N,N-diethylamino, N-ethyl-N-methylamino, N- methyl-N-n-propylamino, N-isopropyl-N-n-propylamino, N-t-butyl-N-methylamino, N-ethyl-N-n-pentylamino and N-n-hexyl-N-methylamino.
Alkylaminocarbonyl or alkylcarbamoyl represents an alkylammocarbonyl radical having one or two (independently selected) alkyl substituents, illustratively and preferably representing methylaminocarbonyl, ethylaminocarbonyl, n-propylamino- carbonyl, isopropylamino-carbonyl, tert-butylaminocarbonyl, n-pentylamino- carbonyl, n-hexylaminocarbonyl, N,N-dimethylaminocarbonyl, N,N-diethylamino- carbonyl, N-ethyl-N-methylaminocarbonyl, N-methyl-N-n-propylaminocarbonyl, N- isopropyl-N-n-propylaminocarbonyl, N-t-butyl-N-methylaminocarbonyl, N-ethyl-N- n-pentylamino-carbonyl and N-n-hexyl-N-methylaminocarbonyl.
Alkylaminosulphonyl represents an alkylaminosulphonyl radical having one or two (independently selected) alkyl substituents, illustratively and preferably representing methylaminosulphonyl, ethylaminosulphonyl, n-propylaminosulphonyl, isopropyl- aminosulphonyl, tert-butylaminosulphonyl, n-pentylaminosulphonyl, n-hexyl-amino- sulphonyl, N,N-dimethylaminosulphonyl, N,N-diethylaminosulphonyl, N-ethyl-N- methylamino-sulphonyl, N-methyl-N-n-propylaminosulphonyl, N-isopropyl-N-n- propylaminosulphonyl, N-t-butyl-N-methylaminosulphonyl, N-ethyl-N-n-pentyl- aminosulphonyl and N-n-hexyl-N-methylaminosulphonyl.
Alkylsulphonylamino illustratively and preferably represents methylsulphonylamino, ethylsulphonylamino, n-propylsulphonylamino, isopropylsulphonylamino, tert-butyl- sulphonylamino, n-pentylsulphonylamino and n-hexylsulphonylamino. Alkoxycarbonyl illustratively and preferably represents methoxycarbonyl, ethoxy- carbonyl, n-propoxycarbonyl, isopropoxycarbonyl, tert-butoxycarbonyl, n-pentoxy- carbonyl and n-hexoxycarbonyl. Alkoxycarbonylamino illustratively and preferably represents methoxycarbonylamino, ethoxycarbonylamino, n-propoxycarbonylamino, isopropoxycarbonylamino, tert-butoxycarbonylamino, n-pentoxycarbonylamino and n-hexoxycarbonylamino .
Alkanoylamino illustratively and preferably represents acetylamino and ethyl- carbonylamino.
Cycloalkyl per se and in cycloalkylamino and in cycloalkylcarbonyl represents a cycloalkyl group having generally 3 to 8 and preferably 5 to 7 carbon atoms, illustratively and preferably representing cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and cycloheptyl.
Cycloalkylamino represents a cycloalkylamino radical having one or two (independently selected) cycloalkyl substituents, illustratively and preferably representing cyclopropylamino, cyclobutylamino, cyclopentylamino, cyclohexylamino and cyclo- heptylamino.
Cycloalkylcarbonyl illustratively and preferably represents cyclopropylcarbonyl, cyclobutylcarbonyl, cyclopentylcarbonyl, cyclohexylcarbonyl and cycloheptyl- carbonyl.
Aryl per se and in arylamino and in arylcarbonyl represents a mono- to tricyclic aromatic carbocyclic radical having generally 6 to 14 carbon atoms, illustratively and preferably representing phenyl, naphthyl and phenanthrenyl.
Arylamino represents an arylamino radical having one or two (independently selected) aryl substituents, illustratively and preferably representing phenylamino, diphenylamino and naphthylamino. Arylcarbonyl illustratively and preferably represents phenylcarbonyl and naphthyl- carbonyl.
Heteroaryl per se and in heteroarylamino and heteroarylcarbonyl represents an aromatic mono- or bicyclic radical having generally 5 to 10 and preferably 5 or 6 ring atoms and up to 5 and preferably up to 4 hetero atoms selected from the group consisting of S, O and N, illustratively and preferably representing thienyl, furyl, pyrrolyl, thiazolyl, oxazolyl, imidazolyl, pyridyl, pyrimidyl, pyridazinyl, indolyl, indazolyl, benzofuranyl, benzothiophenyl, quinolinyl, isoquinolinyl.
Heteroarylamino represents an heteroarylamino radical having one or two (independently selected) heteroaryl substituents, illustratively and preferably representing thienylamino, furylamino, pyrrolylamino, thiazolylamino, oxazolylamino, imidazolyl-amino, pyridylamino, pyrimidylamino, pyridazinylamino, indolylamino, indazolylamino, benzofuranylamino, benzothiophenylamino, quinolinyl-amino, isoquinolinylamino .
Heteroarylcarbonyl illustratively and preferably represents thienylcarbonyl, furyl- carbonyl, pyrrolylcarbonyl, thiazolylcarbonyl, oxazolylcarbonyl, imidazolyl- carbonyl, pyridylcarbonyl, pyrimidylcarbonyl, pyridazinylcarbonyl, indolylcarbonyl, indazolylcarbonyl, benzofiiranylcarbonyl, benzothiophenylcarbonyl, quinolinyl- carbonyl, isoquinolinylcarbonyl.
Heterocyclyl per se and in heterocyclylcarbonyl represents a mono- or polycyclic, preferably mono- or bicyclic, nonaromatic heterocyclic radical having generally 4 to 10 and preferably 5 to 8 ring atoms and up to 3 and preferably up to 2 hetero atoms and or hetero groups selected from the group consisting of N, O, S, SO and SO2. The heterocyclyl radicals can be saturated or partially unsaturated. Preference is given to 5- to 8-membered monocyclic saturated heterocyclyl radicals having up to two hetero atoms selected from the group consisting of O, N and S, such as illustratively and preferably tetrahydrofuran-2-yl, ρyrrolidin-2-yl, pyrrolidin-3-yl, pyrrolinyl, piperidinyl, morpholinyl, perhydroazepinyl.
Heterocyclylcarbonyl illustratively and preferably represents tetrahydrofuran-2- carbonyl, pyrrolidine-2-carbonyl, pyrrolidine-3 -carbonyl, pyrrolinecarbonyl, piperidinecarbonyl, morpholinecarbonyl, perhydroazepinecarbonyl.
Halogen represents fluorine, chlorine, bromine and iodine.
This invention is also to provide a method for treating or preventing a disorder or disease associated with MKK7 and/or MKK4 activity in a human or animal subject, comprising administering to said subject a therapeutically effective amount of the pyrimido[4,5-b]indole derivative shown in the formula (I), its tautomeric or stereoisomeric form, or a physiologically acceptable salt thereof.
Further this invention is to provide a use of the pyrimido[4,5-b]indole derivative shown in the formula (I), its tautomeric or stereoisomeric form, or a physiologically acceptable salt thereof in the preparation of a medicament. Preferably, said medicament is suitable for treating or preventing a disorder or disease associated with MKK7 and/or MKK4 activity.
The compounds of the present invention surprisingly show excellent MKK7 and/or MKK4 inhibitory activity. They are, therefore, suitable for the production of medicament or medical composition, which may be useful to treat MKK7 and/or MKK4 related diseases.
More specifically, since the pyrirnido[4,5-b]indoles derivatives of the present invention inhibit MKK7 and/or MKK4, they are useful for treatment and prophylaxis of diseases as follows: inflammatory and immunoregulatory disorders, such as asthma, atopic dermatitis, rhinitis, allergic rhinitis, allergic diseases, COPD, septic shock, arthritis, joint diseases and myocardial injuries, as well as autoimmune pathologies such as rheumatoid arthritis, Grave's disease, and atherosclerosis.
Therefore, MKK7 and/or MKK4 is an important target and inhibition of MKK7 and/or MKK4 is likely to be effective in the treatment of such inflammatory and immunoregulatory disorders and diseases.
The compounds of the present invention are also useful for treatment of ischemia, myocardial injury, pulmonary hypertension, renal failure, Huntington's chorea and cardiac hypertrophy, as well as neurodegenerative disorders such as Parkinson's disease, Alzheimer's disease and focal ischemia, since the diseases also relate to MKK7 and/or MKK4.
In one embodiment, the compounds of formula (I) are those wherein:
R1 is amino;
R2 is pyrrolidyl(C1-6)alkyl, saturated(C3-8)cycloalkyl or benzyl, and
R3 is hydrogen, (C1-6)alkyl, or benzyl,
or
R2 and R3 are taken together with the attached nitrogen atom to form
a 7 membered saturated heterocyclic ring optionally interrupted by one or two atoms selected from the group consisting of oxygen, sulfur and nitrogen, a 5-7 membered saturated heterocyclic ring which is fused by benzene or saturated(C3-8)cycloalkyl,
or
a 8-12 membered spiro ring, optionally interrupted by one or two oxygen atoms;
In another embodiment, the compounds of formula (I) are those wherein:
R1 is amino;
R is carbamoyl or amino (C1-6 )alkyl; and
R5 is hydrogen or (Cι-6)alkyl.
In another embodiment, the compounds of formula (I) are those wherein:
R4 is carbamoyl; and
R5 is hydrogen.
In another embodiment, the compounds of formula (I) are those wherein:
R1 is hydrogen, methyl or amino;
R4 is carbamoyl; and
R is hydrogen.
In another embodiment, the compounds of formula (I) are those wherein: R2 and R3 independently represent hydrogen, (C1-6)alkyl optionally substituted by amino, hydroxy, methoxy, imidazolyl, (C1-6)alkoxy or (C1-6)alkoxycarbonyl,
(C3-8)cycloalkyl optionally substituted by hydroxy or hydroxy (C1-6)alkyl, benzyl, or 5-7 membered saturated heterocyclic ring interrupted by one or two oxygen atoms;
or
R2 and R3 are taken together with the attached nitrogen atom to form a saturated heterocyclic ring selected from the group consisting of pyrrolidino, piperidino, piperazino, homopiperidino, and morpholino
wherein
said heterocyclic ring is optionally substituted by amino, methylamino, acetamido, hydroxy, oxo, carbamoyl, benzyl, bromo, carboxy, pyrrolidino, or (C1-6 )alkyl optionally substituted by hydroxy, amino or carboxy,
or
said heterocyclic ring is optionally fused by benzene or saturated(C3-8)- cycloalkyl;
R is carbamoyl or amino(Cι-6 )alkyl; and
R5 is hydrogen.
In yet another embodiment, the compounds of formula (I) are those wherein:
R1 is hydrogen, methyl or amino; R2 and R3 independently represent hydrogen, (C1-6)alkyl optionally substituted by amino, hydroxy, methoxy, imidazolyl, (Cι-6)alkoxy or (C -8)cycloalkyl optionally substituted by hydroxy or hydroxy (C1-6)alkyl, benzyl, or 5-7 membered saturated heterocyclic ring interrupted by one or two oxygen atoms;
or
R2 and R3 are taken together with the attached nitrogen atom to form a saturated heterocyclic ring selected from the group consisting of pyrrolidino, piperidino, piperazino, homopiperidino, and morpholino
wherein
said heterocyclic ring is optionally substituted by amino, methylamino, acetamido, hydroxy, oxo, carbamoyl, benzyl, bromo, carboxy, pyrrolidino, or (C1-6 )alkyl optionally substituted by hydroxy, amino or carboxy,
or
said heterocyclic ring is optionally fused by benzene or saturated(C3-8)- cycloalkyl;
R is carbamoyl; and
R5 is hydrogen.
In yet another embodiment, the compounds of formula (I) are those wherein:
R1 is amino;
R2 is saturated(C3-8)cycloalkyl or benzyl; R3 is hydrogen, or (C1-6)alkyl,
or
R2 and R3 are taken together with the attached nitrogen atom to form
a homopiperidino,
a 5-7 membered saturated heterocyclic ring which is fused by benzene or saturated(C3-8)cycloalkyl,
wherein said heterocyclic ring is selected from the group consisting of pyrrolidino, piperidino, piperazino, homopiperidino, and morpholino,
or
1, 4 dioxa-8-azaspiro[4,5]decane; and
R is hydrogen.
In yet another embodiment, the compounds of formula (I) are those wherein:6- Carbamoyl-4-( 1 -piperidinyl)-9H-pyrimido [4,5 -bjindole,
6-Carbamoyl-4-ι (l-morpholinyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4 (4-hydroxy-l-piperidinyl)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4- (cycloheptylamino)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-ι (4-methyl- 1 -piperidinyl)-9H-pyrimido [4,5 -b]indole, 6-Carbamoyl-4 (3-methyl-l-piperidinyl)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4 (3,5-dimethyl-l-piρeridinyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4 [2-(hydroxymethyl)-l-piρeridinyl]-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4- (cycloρentylamino)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4- -(l-azepanyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4- -[[(2-aminoethyl)amino]-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4- -[4-(2-hydroxyethyl)-l-piρeridinyl]-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4- -(cyclohexylamino)-9H-pyrimido [4, 5 -bjindole,
6-Carbamoyl-4- -[[(2-hydroxy ethy l)amino] -9H-pyrimido [4, 5 -b] indole,
6-Carbamoyl-4- -[(2-hydroxyethyl)(methyl)amino]-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4--(4-benzyl-4-hydroxy- 1 -piperidinyl)-9H-pyrimido [4,5-b]indole,
6-Carbamoyl-4- -((l-pyrrolidinyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4- -[(2-hydroxy- 1 -methylethyl)amino]-9H-pyrimido[4,5-b]indole,
66--CCaarrbbaammooyvll--44--((2-ethyl- 1 -piperidinyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4- -[4-(hydroxymethyl)-l-piperidinyl]-9H-ρyrimido[4,5-b]indole, 4-(3-amino-l-piperidinyl)-6-carbamoyl-9H-pyrimido[4,5-b]indole, tert-butyl 1 -[6-carbamoyl~9H-pyrimido[4,5-b]indol-4-yl]-3 -piperidinylcarbamate, 4-[(3-aminopropyl)amino]-6-carbamoyl-9H-pyrimido[4,5-b]indole, tert-butyl 3-{[6-carbamoyl-9H-pyrimido[4,5-b]indol-4 yl]amino}propylcarbamate,
4-(3-amino-l-pyrrolidinyl) -6-carbamoyl-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-(dipropylamino)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-[3-(methylamino)-l-pyrrolidinyl]-9H-pyrimido[4,5-b]indole, l-[6-carbamoyl-9H-ρyrimido[4,5-b]indol-4-yl]-4-piperidinecarboxylic acid, 6-Carbamoyl-4-[4-(acetylamino)- 1 -piperidinyl]-9H-pyrimido[4,5-b]indole,
2-Amino-6-carbamoyl-4-(l-piperidinyl)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-2-methyl-4-(l-piperidinyl)-9H-pyrimido[4,5-b]indole, and their tautomeric and stereoisomeric form, and salts thereof.
Further, the present invention provides a medicament which include one of the compounds described above and optionally pharmaceutically acceptable excipient. EMBODIMENT OF THE INVENTION
The compound of the formula (I) of the present invention can be, but not limited to be, prepared by combining various known methods. In some embodiments, one or more of the substituents, such as amino group, carboxyl group, and hydroxyl group of the compounds used as starting materials or intermediates are advantageously protected by a protecting group known to those skilled in the art. Examples of the protecting groups are described in "Protective Groups in Organic Synthesis (2nd Edition)" by Greene and Wuts.
The compound of the formula (I) of the present invention can be, but not limited to be, prepared by the method [A] below.
Method [A]
(II) (I)
The compound (I) (wherein R1, R2, R3, R4 and R5 are the same as defined above) or a salt thereof, for example, can be prepared by the reaction of the compound of formula (II) (wherein R1, R4 and R5 are the same as defined above, and L represents leaving group including, for instance, halogen atom such as chlorine, bromine, or iodine atom; C6-10 arylsulfonyloxy group such as benzenesulfonyloxy or p-toluene- sulfonyloxy; C1-4 alkylsulfonyloxy group such as methanesulfonyloxy; and halogen substituted C1-4 alkylsulfonyloxy group such as trifluoromethanesulfonyloxy and the like.) or a salt thereof, with a compound shown by the general formula (III) (wherein R2 and R3 are the same as defined above ) or a salt thereof. This reaction can be carried out without solvent or in a solvent including, for instance, alcohol such as methanol and ethanol; ethers, such as dioxane, diethyl ether, and tetrahydrofuran (THF); aromatic hydrocarbons such as benzene, toluene and xylene; nitriles such as acetonitrile; amides such as dimethylformamide (DMF) and dimethylacetamide; sulfoxides such as dimethyl sulfoxide, and others.
The reaction temperature can be optionally set depending on the compounds to be reacted. The reaction temperature is usually, but not limited to, about 10°C to 200°C and preferably about 20°C to 100°C The reaction may be conducted for, usually, 30 mins to 48 hrs and preferably 1 hr to 24 hrs.
The reaction can be advantageously conducted in the presence of a base. Examples of the base include an alkali metal hydride such as sodium hydride or potassium hydride; alkali metal alkoxide such as sodium methoxide or sodium ethoxide; alkali metal hydroxide such as sodium hydroxide or potassium hydroxide; carbonates such as sodium carbonate or potassium carbonate, and bicarbonates such as sodium hydrogen carbonate and potassium hydrogen carbonate; organic amines such as triethylamine.
The amine (III) are commercially available or can be synthesized by conventional methods. If required, R1, R2, R3, R4 and R5 can be optionally protected during the reaction and deprotected afterward.
The compound of the formula (I-a) can be, but not limited to be, prepared by the methods [B] or [C] below. Method [B]
The compound of formula (I-a) (wherein R1, R2, R3 and R5 are the same as defined above) can be prepared by the reaction of the compound of formula (I-b)(wherein R1 ,
R2, R3 and R5 are the same as defined above, and X represents hydrogen or (C1-6)- alkyl) and ammonia.
The reaction can be carried out without solvent or in a solvent including, for instance, alcohols such as methanol and ethanol, 1 -propanol, isopropanol and tert-butanol; water; ethers such as diethyl ether, isopropyl ether, dioxane and tetrahydrofuran (THF) and 1,2-dimethoxyethane; aromatic hydrocarbons such as benzene, toluene and xylene; amides such as N, N-dimethylformamide (DMF), N, N-dimethyl- acetamide; sulfoxides such as dimethyl sulfoxide, and others. Optionally, two or more of the solvents selected from the listed above can be mixed and used.
The reaction temperature can be optionally set depending on the compounds to be reacted. The reaction temperature is usually, but not limited to, about 0°C to 60°C The reaction may be conducted for, usually, 30 mins to 48 hrs and preferably 1 hr to 24 hrs.
When X is hydrogen in compound of formula (I-b), the reaction can be advantageously carried out using coupling agent including, for instance, carbo- diimides such as N, N-dicyclohexylcarbodiimide and l-(3-dimethylaminopropyl)-3- ethylcarbodiimide; carbonyldiazoles such as l,l'-carbonyldi(l,3-imiazole)(CDI) and l,l'-carbonyldi(l,2,4-triazole)(CDT), and others. Method [C]
(l-c) (I-a)
Alternatively, the compound of formula (I-a) (wherein R , R2, R3 and R5 are the same as defined above) can be prepared by the hydrolysis of the compound of formula (I-c) (wherein R1, R2, R3 and R5 are the same as defined above).
The reaction can be carried out in a solvent including, for instance, alcohols such as methanol and ethanol, 1 -propanol, isopropanol, n-butanol and tert-butanol; water; ketone such as acetone; ethers such as diethyl ether, isopropyl ether, dioxane and tetrahydrofuran (THF) and 1,2-dimethoxyethane; aromatic hydrocarbons such as benzene, toluene and xylene; amides such as N, N-dimethylformamide (DMF), N, N- dimethylacetamide, and others. Optionally, two or more of the solvents selected from the listed above can be mixed and used.
The reaction temperature can be optionally set depending on the compounds to be reacted. The reaction temperature is usually, but not limited to, about 0°C to 60°C The reaction may be conducted for, usually, 30 mins to 48 hrs and preferably 1 hr to 24 hrs.
The reaction can be advantageously conducted in the presence of a base, including, for instance, an alkali metal alkoxide such as sodium methoxide, sodium ethoxide and potassium tert-butoxide; alkali metal hydroxide such as sodium hydroxide and potassium hydroxide; alkali metal carbonates such as sodium carbonate and potassium carbonate; alkali metal hydrogen carbonates such as sodium hydrogen carbonate and potassium hydrogen carbonate; alkali metal phosphate such as sodium phosphate, and others.
When base is used, the reaction can be advantageously conducted in the presence of oxidating agent, for instance, hydrogen peroxide, manganese dioxide, dimethyl dioxirane, sodium percarbonate, sodium perborate, oxone, and the others.
The reaction can be advantageously conducted in the presence of an acid including, for instance, trifluoroacetic acid, hydrochloric acid and sulfonic acid, and others.
Preparation of intermediates
The compound of formula (II) (wherein R1, R4 and R5 are the same as defined above) can be prepared by the following procedures.
lkyl, etc.,) dl) 4b (R1' = H)
Thus, compound 2 can be prepared by the reaction of compound 1 (wherein L' represents leaving group including, for instance, halogen atom such as chlorine, bromine, or iodine atom; C6-10 arylsulfonyloxy group such as benzenesulfonyloxy or p-toluenesulfonyloxy; C1-4 alkylsulfonyloxy group such as methanesulfonyloxy; and halogen substituted C1- alkylsulfonyloxy group such as trifluoromethanesulfonyloxy and the like, and R and R5 are the same as defined above) with ethyl cyanoacetate using a base, for instance, sodium hydride.
The reaction may be carried out in a solvent including, for instance, ethers such as diethyl ether, isopropyl ether, dioxane and tetrahydrofuran (THF) and 1,2- dimethoxyethane; aromatic hydrocarbons such as benzene, toluene and xylene; amides such as N, N-dimethylformamide (DMF), N, N-dimethylacetamide and N- methylpyrrolidone; sulfoxides such as dimethylsulfoxide (DMSO); alcohols such as methanol, ethanol, 1 -propanol, isopropanol and tert-butanol,; and others. Optionally, two or more of the solvents selected from the listed above can be mixed and used. The reaction may be carried out, usually, at room temperature to 100 °C for 4 hrs to 12 hrs.
Compound 1 and ethyl cyanoacetate are commercially available or can be synthesized by conventional methods.
Compound 3 (wherein R and R5 are the same as defined above) can be prepared by reducing nitro group of compound 2 using agent including, for instance, metals such as zinc and iron in the presence of acid including, for instance, hydrochloric acid and acetic acid. The reaction can be carried out without solvent or in a solvent including, for instance; aromatic hydrocarbons such as benzene, toluene and xylene, and others. The reaction may be carried out, usually, at room temperature to 100 °C for 30 mins to 12 hrs.
Compound 4a (wherein R1 represents amino, (C1-6)alkyl, (C2-6)alkenyl, (C2-6)alkynyl, halogen substituted (C1-6) alkyl, cyano, cyano(C1-6)alkyl, (C3-8)cycloalkyl, nitro- (Cι_6)alkyl or fluoro and R4 and R5 are the same as defined above) can be prepared by the reaction of compound 3 with appropriate cyano compounds (R^CN) (wherein R1' is the same as defined above). The reaction can be carried out in a solvent including, for instance, alcohols such as methanol, ethanol, 1 -propanol, isopropanol and tert- butanol; ethers such as diethyl ether, isopropyl ether, dioxane and tetrahydrofuran (THF) and 1,2-dimethoxyethane; aromatic hydrocarbons such as benzene, toluene and xylene, and others. The reaction may be carried out, usually, at 40°C to 180°C for 2 hrs to two days.
Cyano compounds are commercially available or can be synthesized by conventional methods.
Compound 4b (wherein R1 is hydrogen and R4 and R5 are the same as defined above) can be prepared by the reaction of compound 3 with ammonium formate in a solvent such as formamide. The reaction may be carried out, usually, at 40°C to 180°C for 2 hrs to two days. If desired, the resulting 4b can be further modified to introduce nitro group at the position of R1.
Ammonium formate and formamide are commercially available or can be synthesized by conventional methods.
The compound of formula (II) (wherein R1, R4 and R5 are the same as defined above and L represents leaving group including, for instance, halogen atom such as chlorine, bromine, or iodine atom; C6-10 arylsulfonyloxy group such as benzenesulfonyloxy or p-toluenesulfonyloxy; C1- alkylsulfonyloxy group such as methanesulfonyloxy; and halogen substituted C1-4 alkylsulfonyloxy group such as trifluoro- methanesulfonyloxy and the like.) can be prepared for instance, by the reaction of compound 4 with appropriate halogenating reagent including, for instance, POCl3, PC15, SOCl2, and the like; or can be prepared, for instance, by the reaction of compound 4 with appropriate sulfonyl chloride.
The reaction may be carried out without solvent or in a solvent including, for instance, halogenated hydrocarbons such as dichloromethane, chloroform and 1,2- dichloroethane;such as ethers such as dioxane and tetrahydrofuran (THF) and 1,2- dimethoxyethane; aromatic hydrocarbons such as benzene, toluene, and xylene, and others. Optionally, two or more of the solvents selected from the listed above can be mixed and used. The reaction can be advantageously conducted in the presence of a base, including, for instance, such as pyridine, triethylamine and N,N-diiso- propylethylamine, dimethylaniline, diethylaniline, and others.
The reaction temperature is usually, but not limited to, about 40°C to 200°C and preferably about 20°C to 180°C The reaction may be conducted for, usually, 30 minutes to 48 hrs and preferably 2 hr to 12 hrs.
The halogenating reagents and sulfonyl chlorides are commercially available or can be synthesized by conventional methods.
When the compound shown by the formula (I) or a salt thereof has tautomeric isomers and/or stereoisomers (e.g., geometrical isomers and conformational isomers), each of their separated isomers and mixtures are also included in the scope of the present invention.
Typical salts of the compound shown by the formula (I) include salts prepared by reaction of the compounds of the present invention with a mineral or organic acid, or an organic or inorganic base. Such salts are known as acid addition and base addition salts, successively.
Acids to form salts include inorganic acids such as, without limitation, sulfuric acid, phosphoric acid, hydrochloric acid, hydrobromic acid, hydriodic acid like, and organic acids, such as, without limitation, p-toluenesulfonic acid, methanesulfonic acid, oxalic acid, p-bromophenylsulfonic acid, succinic acid, citric acid, benzoic acid, acetic acid, and the like. Base addition salts include those derived from inorganic bases, such as, without limitation, ammonium hydroxide, alkaline metal hydroxide, alkaline earth metal hydroxides, carbonates, bicarbonates, and the like, and organic bases, such as, without limitation, ethanolamine, triethylamine, tris(hydroxymethyl)aminomethane, and the like. Examples of inorganic bases include, sodium hydroxide, potassium hydroxide, potassium carbonate, sodium carbonate, sodium bicarbonate, potassium bicarbonate, calcium hydroxide, calcium carbonate, and the like.
The compound of the present invention or a salts thereof, depending on its substituents, may be modified to form lower alkylesters or known other esters; and/or hydrates or other solvates. Those esters, hydrates, and solvates are included in the scope of the present invention.
The compound of the present invention may be administered in oral forms, such as, without limitation normal and enteric coated tablets, capsules, pills, powders, granules, elixirs, tinctures, solution, suspensions, syrups, solid and liquid aerosols and emulsions. They may also be administered in parenteral forms, such as, without limitation, intravenous, intraperitoneal, subcutaneous, intramuscular, and the like forms, well-known to those of ordinary skill in the pharmaceutical arts. The compounds of the present invention can be administered in intranasal form via topical use of suitable intranasal vehicles, or via transdermal routes, using transdermal delivery systems well-known to those of ordinary skilled in the art.
The dosage regimen with the use of the compounds of the present invention is selected by one of ordinary skill in the arts, in view of a variety of factors, including, without limitation, age, weight, sex, and medical condition of the recipient, the severity of the condition to be treated, the route of administration, the level of metabolic and excretory function of the recipient, the dosage form employed, the particular compound and salt thereof employed. The compounds of the present invention are preferably formulated prior to administration together with one or more pharmaceutically-acceptable excipients. Excipients are inert substances such as, without limitation carriers, diluents, flavoring agents, sweeteners, lubricants, solubilizers, suspending agents, binders, tablet disintegrating agents and encapsulating material.
Yet another embodiment of the present invention is pharmaceutical formulation comprising a compound of the invention and one or more pharmaceutically- acceptable excipients that are compatible with the other ingredients of the formulation and not deleterious to the recipient thereof. Pharmaceutical formulations of the invention are prepared by combining a therapeutically effective amount of the compounds of the invention together with one or more pharmaceutically-acceptable excipients. In making the compositions of the present invention, the active ingredient may be mixed with a diluent, or enclosed within a carrier, which may be in the form of a capsule, sachet, paper, or other container. The carrier may serve as a diluent, which may be solid, semi-solid, or liquid material wliich acts as a vehicle, or can be in the form of tablets, pills, powders, lozenges, elixirs, suspensions, emulsions, solutions, syrups, aerosols, ointments, containing, for example, up to 10% by weight of the active compound, soft and hard gelatin capsules, suppositories, sterile injectable solutions and sterile packaged powders.
For oral administration, the active ingredient may be combined with an oral, and non-toxic, pharmaceutically-acceptable carrier, such as, without limitation, lactose, starch, sucrose, glucose, sodium carbonate, mannitol, sorbitol, calcium carbonate, calcium phosphate, calcium sulfate, methyl cellulose, and the like; together with, optionally, disintegrating agents, such as, without limitation, maize, starch, methyl cellulose, agar bentonite, xanthan gum, alginic acid, and the like; and optionally, binding agents, for example, without limitation, gelatin, natural sugars, beta-lactose, corn sweeteners, natural and synthetic gums, acacia, tragacanth, sodium alginate, carboxymethylcellulose, polyethylene glycol, waxes, and the like; and, optionally, lubricating agents, for example, without limitation, magnesium stearate, sodium stearate, stearic acid, sodium oleate, sodium benzoate, sodium acetate, sodium chloride, talc, and the like.
In powder forms, the carrier may be a finely divided solid which is in admixture with the finely divided active ingredient. The active ingredient may be mixed with a carrier having binding properties in suitable proportions and compacted in the shape and size desired to produce tablets. The powders and tablets preferably contain from about 1 to about 99 weight percent of the active ingredient which is the novel composition of the present invention. Suitable solid carriers are magnesium carboxymethyl cellulose, low melting waxes, and cocoa butter.
Sterile liquid formulations include suspensions, emulsions, syrups and elixirs. The active ingredient can be dissolved or suspended in a pharmaceutically acceptable carrier, such as sterile water, sterile organic solvent, or a mixture of both sterile water and sterile organic solvent.
The active ingredient can also be dissolved in a suitable organic solvent, for example, aqueous propylene glycol. Other compositions can be made by dispersing the finely divided active ingredient in aqueous starch or sodium carboxymethyl cellulose solution or in a suitable oil.
The formulation may be in unit dosage form, which is a physically discrete unit containing a unit dose, suitable for administration in human or other mammals. A unit dosage form can be a capsule or tablets, or a number of capsules or tablets. A "unit dose" is a predetermined quantity of the active compound of the present invention, calculated to produce the desired therapeutic effect, in association with one or more excipients. The quantity of active ingredient in a unit dose may be varied or adjusted from about 0.1 to about 1000 milligrams or more according to the particular treatment involved. Typical oral dosages of the present invention, when used for the indicated effects, will range from about 0.0 lmg Λcg/day to about 100 mg/kg/day, preferably from 0.1 mg/kg/day to 30 mg/kg/day, and most preferably from about 0.5 mg/kg/day to about 10 mg/kg/day. In the case of parenteral administration, it has generally proven advantageous to administer quantities of about 0.001 to lOOmg /kg/day, preferably from 0.01 mg/kg/day to 1 mg/kg/day. The compounds of the present invention may be administered in a single daily dose, or the total daily dose may be administered in divided doses, two, three, or more times per day. Where delivery is via transdermal forms, of course, administration is continuous.
Examples
The present invention will be described in detail below in the form of examples, but they should by no means be construed as defining the metes and bounds of the present invention.
In the examples below, all quantitative data, if not stated otherwise, relate to percentages by weight.
Melting points are uncorrected. Liquid Chromatography - Mass spectroscopy (LC-
MS) data were recorded on a Micromass Platform LC with Shimadzu Phenomenex ODS column(4.6 mmφ X 30 mm) flushing a mixture of acetonitrile- water (9:1 to 1:9) at 1 ml/min of the flow rate. Mass spectra were obtained using electrospray (ES) ionization techniques (micromass Platform LC). TLC was performed on a precoated silica gel plate (Merck silica gel 60 F-254). Silica gel (WAKO-gel C-200 (75-
150 μm)) was used for all column chromatography separations. All chemicals were reagent grade and were purchased from Sigma-Aldrich, Wako Pure Chemical Industries, Ltd., Tokyo Kasei Kogyo Co., Ltd., Nacalai tesque, Inc., Watanabe Chemical hid. Ltd., Maybridge pic, Lancaster Synthesis Ltd., Merck KgaA, Kanto Chemical Co., Ltd. The effect of the present compounds were examined by the following assays and pharmacological tests.
[The measurement of MKK7 kinase activity]
(1) Preparation of MKK7 protein
A plasmid containing human MKK7 open reading frame was cloned into a pGEM-T Easy vector (Promega, Madison, WI) and further into a pGEX-6P-2 vector (Pharmacia) to construct human GST(Glutathione-S-transferase)-
MKK7 fusion protein. This construct was coexpressed with human MEKKc (catalytic domain of MEKK (MEK (Map kinase kinase) kinase) on plasmid pBB131, in E.coli (BL21(DE3)pLysS).
The resulting GST-MKK7 was purified with the use of a glutathione column
(Amersham Pharmacia Biotech AB, Uppsala, Sweden) according to the manufacturer's instructions.
(2) A construct containing the MKK7 substrate rat GST-KN-SAPKα (GST + kinase negative rat SAPKα2) was inserted into a pGEX-SAP plasmid
(Amersham Pharmacia Biotech AB, Uppsala, Sweden) and transformed into E.coli BL21(DE3)pLysS. Using this expression strain, GST-KN-SAPKα was purified with the use of glutathione column (Amersham Pharmacia Biotech AB, Uppsala, Sweden) according to the manufacturer's instruction. The purity of the protein was confirmed to be more than 90% by SDS-PAGE.
Biotinylation of the substrate protein was done using sulfo-NHS-LC Biotin according to the manufacturer's instructions (Pierce, Rockford, US) (3) The measurement of MKK7 kinase activity
All Test compounds (2.5 μl) at various concentrations (in 1% DMSO) were added to 15 μl of reaction buffer (20mM HEPES, 0.1M NaCl, 0.1 mM NasNO4, lOmM MgCk, ImM DTT, lmg/ml BSA, pH 7.5)) containing
0.5 μg/ml GST-MKK7 and 0.8 μM SAPK α (biotinylated GST-KΝ-SAPKα fusion protein). The kinase reaction was started by the addition of 12.5 μl of 12 μM ATP. After one hour incubation period at room temperature, the reaction was stopped by the addition of 40 μl stop solution (0.1M EDTA, pH 8.0).
60 μl of this reaction mixture were transferred to a well of the streptavidine- coated detection plate (SA-plate, Steffens: 08114E14.FWD) and 40 μl Tris- buffered saline (TBS, 50 mM Tris-HCl (pH8.0), 20 mM EDTA, 1 % BSA, 1 M ΝaCl, 0.05% tween 20) were added. This mixture was incubated for 30 minutes and washed 3 times with 0.05% tween20 in (TBS), before 100 μl of Eu-labeled anti-phosphothreonine-proline antibody (LAΝCE) was added. After incubation for 30 minutes, plates were again washed 3 times with TBS, and 100 μl of the enhancement solution (Amersham Pharmacia Biotech) was added. One hour later, time-resolved fluorescence was measured by a multi- label counter (ARNO, Wallac Oy, Finland) using 340 nm for excitation and 615 nm for emission with 400 ms of delay and 400 ms of window.
[The measurement of MKK4 kinase activity]
(1) Preparation of MKK4 protein
A plasmid containing human MKK4 open reading frame was cloned into a pGEX-2T vector (Pharmacia) to construct human GST(Glutathione-S- transferase)-MKK4 fusion protein. This construct was coexpressed with human MEKKc (catalytic domain of MEKK on plasmid pBB131) in E.coli (BL21(DE3)pLysS).
The resulting GST-MKK4 was purified with the use of glutathione column (Amersham Pharmacia Biotech AB, Uppsala, Sweden) according to the manufacturer's instruction. The purity of the protein was confirmed to be more than 90% by SDS-PAGE.
(2) A construct containing the MKK4 substrate rat GST-KN-SAPKα (GST + kinase negative rat SAPKα2) was inserted into a pGEX-SAP plasmid
(Amersham Pharmacia Biotech AB, Uppsala, Sweden) and transformed into E.coli BL21(DE3)pLysS. Using this expression strain, GST-KN-SAPKα was purified with the use of glutathione column (Amersham Pharmacia Biotech AB, Uppsala, Sweden) according to the manufacturer's instruction. The purity of the protein was confirmed to be more than 90% by SDS-PAGE.
Biotinylation of the substrate protein was done using sulfo-NHS-LC Biotin according to the manufacturer's instructions (Pierce, Rockford, US)
(3) The measurement of MKK4 kinase activity
All Test compounds (5 μl) at various concentrations (in 1% DMSO) were added to 30 μl of reaction buffer (20mM HEPES, 0.1M NaCl, 0.1 mM Na3NO4, lOmM MgCk, ImM DTT, lmg/ml BSA, pH 7.5)) containing 0.5 μg/ml GST-MKK4 and 6μM ATP. The kinase reaction was started by the addition of 25 μl assay buffer containing 0.48 μM SAPKα (biotinylated
GST-KΝ-SAPKα fusion protein). After a two hours incubation period at room temperature, the reaction was stopped by the addition of 80 μl stop solution (0.1M EDTA, pH 8.0).
120 μl of this reaction mixture were transferred to a well of the streptavidine- coated detection plate (SA-plate, Steffens: 08114E14.FWD) and 40 μl Tris- buffered saline (TBS, 50 mM Tris-HCl (ρH8.0), 20 mM EDTA, 1 % BSA, 1 M NaCl, 0.05% tween 20) were added. This mixture was incubated for 30 min and washed 3 times with 0.05% tween20 in (TBS), before 100 μl of Eu-labeled anti-phosphotyrosine antibody (5ng/well;4G10, Upstate Bio- technology, Lake Placid, NY, US)) was added. After incubation for 30 min., plates were again washed 3 times with TBS, and 100 μl of the enhancement solution (Amersham Pharmacia Biotech) was added. One hour later, time- resolved fluorescence was measured by a multi-label counter (ARNO, Wallac Oy, Finland) using 340 nm for excitation and 615 nm for emission with 400 ms of delay and 400 ms of window.
[Cell-based assays]
IL-2 and IFN-γ release in human PBMC
Human peripheral blood mononucleated cells (huPBMC) isolated using mono-poly resolving medium (Dainippon Seiyaku, Osaka, Japan) were incubated with test compounds (various concentrations in 0.1% DMSO) for 1 hour in a 37°C CO2 incubator. Cells were then plated on 96 well plates (1x10s cell per well in 200 μl RPMI1640 cell culture medium) pre-coated for 3 hrs with 100 μl anti-CD3 antibody
(ΝU-T3: Nichirei) (4 μg/ml)) or without any coating (unstimulated controls). Solution was removed and plates were washed three times with 200 μl/well phosphate buffered saline (PBS). Anti-CD28 antibody (KOLT-2: Nichirei, Tokyo, Japan) and goat anti mouse kappa antibody (Bethyl Laboratories, Inc., Montgomery, Texas, US) was added to the wells at final concentrations of 1.5 μg/ml and 2 μg/ml, respectively. Plates were incubated for 20 hrs in the incubator. Supernatant was removed and stored at -30°C in aliquots until further use. The amount of interleukin- 2 (IL-2) and interferon-γ (IFN-γ) released from huPBMC was determined by commercially available ELISA (Genzyme Tech., Minneapolis, US) according to the manufacturer's instructions. TNF-a and IL-12 release in human PBMC and human dendritic cells
Human peripheral blood mononucleated cells (huPBMC) isolated using mono-poly resolving medium were either directly used for experiments (lxl 0s cells per well in 200 μl medium) or differentiated to dendritic cells (DC) in the presence of GM-SCF
(Pepro Tech., New Jersey, US, 25 ng/ml) + IL-4 (Pepro Tech., New Jersey, US, 10 ng/ml) over 7 days, then collected, counted and plated at a density of 2x104 cells per well per 200 μl). Cells were incubated with test compounds (various concentrations in 0.1 % DMSO) for 1 hr in a 37°C CO2 incubator and then plated on 96 well plates (1x10s cell per well in 200 μl RPMI1640 cell culture medium). Induction of
TNF-α or IL-12 was induced by stimulation with LPS (B8, Sigma, Missouri, US) 10 ng/ml). 20 hrs later supernatant was removed and stored at -30°C in aliquots until further use.
The amounts of TNF-α and IL-12 released from cell cultures were determined by commercially available ELISA (Genzyme Tech., Minneapolis, US) according to the manufacturer's instructions.
[Systemic inflammatory response syndrome in mice]
Male Balb/c mice (20-25 g body weight) were in injected with agonistic anti-CD3 Ab (Pharmingen, San Diego, US; 10 μg/mouse; clone 145-2C11) i.v. 5 minutes after compound application (i.v. in 10 % . 2 hrs post-challenge, mice were sacrificed and the serum cytokines IL-2, IL-4 and IFN-γ were determined by ELISA (Genzyme Tech., Minneapolis, US) according to the manufacturer's instruction. Data represent mean values ± SD of 5-6 animals each. * p<0.05, ** p< 0.01 vs. vehicle control (V); Dunnett's test was used to detect differences among groups and statistics were performed using one-sided ANONA or Student's t-test if applicable.
Results of MKK 7 kinase assay (MKK7) and MKK4 kinase assay (MKK4) are shown in Examples and tables of the Examples below. The data corresponds to the compounds as yielded by solid phase synthesis and thus to levels of purity of about 40 to 90%. For practical reasons, the compounds are grouped in three classes of activity as follows:
ICso = A lμM < B 10 μM < C
The compounds of the present invention also show excellent selectivity, and strong activity in in vivo assays.
Exan nple 1-1
(1) 3-Fluoro-4-nitrobenzyl bromide
To a solution of 3-fluoro-4-nitrotoluene (4.83 g) in carbon tetrachloride (50 ml) were added N-bromosuccinimide (NBS, 12.6 g) and benzoyl peroxide (0.45 g). The mixture was refluxed overnight and additional NBS (6.3 g) and benzoyl peroxide (0.15 g) were added to reflux for another 10 h. After cooling, the reaction mixture was passed through a filter paper to remove resulting precipitates, that were washed with chloroform (50 ml). The filtrates were combined and washed with saturated sodium thiosulfite water solution and brine, successively. The organic layer was dried over sodium sulfate and evaporated in vacuo. The residue was purified by silica gel column chromatography (hexane/ethyl acetate = 15/1 to 5/1) to obtain 3-fluoro-4- nitrobenzyl bromide as a yellow oil (3.43 g, 47%). (2) 3-Fluoro-4-nitrobenzyl alcohol
A mixture of 3-fluoro-4-nitrobenzyl bromide (3.43 g) and calcium carbonate (7.63 g) in a mixture of water (40 ml) and 1,4-dioxane (40 ml) was refluxed overnight. After cooling to room temperature, the reaction mixture was passed through a paper filter to remove insoluble materials, that were washed with 1,4-dioxane (20 ml). The filtrates were combined and evaporated in vacuo. The residue was dissolved in ethyl acetate (40 ml) and washed with IN hydrochloric acid, saturated sodium bicarbonate water solution and brine, successively, to be dried over sodium sulfate. When the solvent was removed in vacuo and the residue was triturated with hexane, 3-fluoro-4-nitrobenzyl alcohol was obtained as colorless powders (2.07 g, 83%).
(3) 3-Fluoro-4-nitrobenzoic acid
To a solution of 3-fluoro-4-nitrobenzyl alcohol (2.97 g) in acetone (60 ml) was added Jones reagent (13 ml), prepared from chromic acid (26.7 g) and sulfuric acid (23 ml) in water (100 ml), dropwise at 0°C. The mixture was stirred on a ice-bath for 0.5 h and quenched with isopropanol (20 ml) to be concentrated in vacuo. The residue was dissolved in ethyl acetate (30 ml) and washed with water (30 ml X 3), and brine (30 ml X 1), successively. The organic layer was dried over sodium sulfate and concentrated in vacuo to obtain a yellow solid, that was triturated with hexane to give 3-fluoro-4- nitrobenzoic acid as a pale yellow solid (2.94 g, 92%).
(4) Methyl 3-fluoro-4-nitrobenzoate
To a solution of 3-fluoro-4-nitrobenzoic acid (5.00 g) in methanol (50 ml) was added 98% sulnxric acid (1 ml). The mixture was refluxed overnight and concentrated in vacuo. The residue was dissolved in ethyl acetate (30 ml) and washed with saturated sodium bicarbonate water solution (30 ml) and brine (30 ml), successively. The organic layer was dried over sodium sulfate and evaporated in vacuo. The residue was purified by silica gel column chromatography (hexane/ethyl acetate = 5/1) to give methyl 3-fluoro-4- nitrobenzoate (5.0 g, 93%) as a colorless solid.
(5) Ethyl α-cyano-5-methoxycarbonyl-2-nitrophenylacetate
To a suspension of 60% sodium hydride (2.01 g) in N,N-dimethylformamide (DMF, 15 ml) was added a solution of ethyl cyanoacetate (5.68 g) in DMF (5 ml) at 0°C The mixture was stirred at room temperature for 0.5 hrs and a solution of methyl 3-fluoro-4-nitrobenzoate (5.00 g) in DMF (5 ml) was added. The mixture was stirred at room temperature for another 3 hrs and poured into a mixture of ethyl acetate (100 ml) and IN hydrochloric acid (200 ml). The organic layer was separated and washed with water (100 ml X
2) and brine (100 ml), successively, to be dried over sodium sulfate. The solvent was removed in vacuo to obtain a brown solid, that was purified by silica gel column chromatography (hexane/ethyl acetate = 5/1 to 2/1) to give ethyl α-cyano-5-methoxycarbonyl-2-nitrophenylacetate as a pale yellow solid (5.74 g, 78%).
(6) Methyl 2-Amino-3-ethoxycarbonyl-lH-indole-5-carboxylate
Ethylα-cyano-5-methoxycarbonyl-2-nitrophenylacetate (3.10 g) was dissolved in glacial acetic acid (30 ml) at 80°C Keeping the temperature, zinc powder (5.55 g) was added portionwise. The mixture was stirred at 90°C to 100°C for 1 h. After cooling to room temperature, the reaction mixture was passed through a filter paper to remove insoluble materials and washed with glacial acetic acid (10 ml). The filtrates were combined and evaporated in vacuo. The residue was dissolved in ethyl acetate (30 ml) and washed with saturated sodium bicarbonate water solution (30 ml X 3) and brine (30 ml), successively. The organic layer was dried over sodium sulfate and evaporated in vacuo. The residue was triturated with isopropyl ether to give methyl 2- amino-3-ethoxycarbonyl-lH-indole-5-carboxylate as a colorless solid (2.37 g, 85%). (7) Methyl 4-Hydroxy-9H-pyrimido[4,5-b]indole-6-carboxylate
A mixture of methyl 2-amino-3-ethoxycarbonyl-lH-indole-5 -carboxylate (0.80 g) and ammonium formate (0.20 g) in formamide (4 ml) was stirred at 175°C under the atmosphere of argon gas stream overnight. After cooling to room temperature, the reaction mixture was diluted with water (40 ml) to give precipitates, that were washed with methanol (20 ml) to obtain pure methyl 4- hydroxy-9H-pyrimido[4,5-b]indole-6-carboxylate (0.413 g, 56%).
(8) Methyl 4-Chloro-9H-pyrimido[4,5-b]indole-6-carboxylate
To a solution of methyl 4-hydroxy-9H-pyrimido[4,5-b]indole-6-carboxylate (0.35 g) and N,N-dimethylaniline (0.52 g) in 1,4-dioxane (2 ml) was added phosphoryl chloride (1.4 ml). The mixture was stirred at 100°C for 6 hrs and, after cooling to room temperature, poured into crushed ice. When this quenching was completed, precipitates resulted in to be collected by a paper filter and dried at 80°C in vacuo for 5 hrs. Resulting solid was suspended in methanol and passed tlirough a filter paper to obtain methyl 4-chloro-9H- pyrimido[4,5-b]indole-6-carboxylate as a pale yellow solid (0.298 g, 79%).
(9) Methyl 4-(l-piperidinyl)-9H-pyrimido[4,5-b]indole-6-carboxylate
A mixture of methyl 4-chloro-9H-pyrimido[4,5-b]indole-6-carboxylate (0.29 g) and piperidine (0.5 ml) in 1,4-dioxane (1 ml) was stirred at 100°C for 0.5 hrs and poured into water (10 ml). Resulting mixture was extracted with chloroform (10 ml X 3). The organic layer was washed with water (10 ml X 3) and dried over sodium sulfate. The solvent was removed in vacuo and the residue was purified by silica gel column chromatography (chlorofo m/- methanol = 20/1) and triturated with isopropylether to give methyl 4-(l- piperidinyl)-9H-pyrimido[4,5-b]indole-6-carboxylate as a colorless solid (0.224 g, 64%).
Mp 224-227°C; LC-MS (ESI): Retention time: 2.4 min (YMC-Pack Pro C18, 2.0 mmφ 35 mm, acetonitrile-water (9:1 to 1:9), 1 ml/min), Calcd [M+l]: 311, Found: m/z 311. Molecular weight: 310.36 (10) 4-(l-Piperidinyl)-9H-pyrimido[4,5-b]indole-6-carboxylic acid
To a solution of methyl 4-(l-piperidinyl)-9H-pyrimido[4,5-b]indole-6-carb- oxylate (0.20 g) in methanol (2 ml) was added 4N NaOH aqueous solution
(1 ml). The mixture was refluxed for 2.5 hrs and acidified with acetic acid
(0.3 ml). When the solution was diluted with water (30 ml), precipitates were given to be collected to give 4-(l-piperidinyl)-9H-pyrimido[4,5-b]indole-6- carboxylic acid (0.172 g, 90%).
Mp >300°C; LC-MS (ESI): Retention time: 1.0 min (YMC-Pack Pro C18, 2.0 mmφ x 35 mm, acetonitrile-water (9:1 to 1:9), 1 ml/min), Calcd [M+1]: 297,
Found: m z 297.
Molecular weight: 296.33
(11) 6-Carbamoyl-4-(l -ρiperidinyl)-9H-pyrimido[4,5-b]indole
4-(l-Piperidinyl)-9H-pyrimido[4,5-b]indole-6-carboxylic acid (0.05 g) was dissolved in DMF (2 ml) and carbonyldiimidazole (0.03 g) was added at 0°C. The mixture was stirred at room temperature for 0.5 hrs and 28% ammonium aqueous solution (0.5 ml) was added to be stirred at room temperature for 0.5 hrs. The reaction mixture was extracted with ethyl acetate (5 ml X 3) and the organic layer was washed with brine (10 ml) to be dried over sodium sulfate and evaporated. The residue was triturated with isopropyl ether to give
6-carbamoyl-4-(l-piperidinyl)-9H-pyrimido[4,5-b]indole (0.028 g, 56%) as a colorless powder.
Mp 293-294 °C; LC-MS (ESI): Retention time: 3.1 min, Calcd [M+1]: 296, Found: m/z 296. Molecular weight: 295.35
Activity grade MKK7: A
Example 1-2
6-Carbamoyl-4-(l-piperidinyl)-9H-pyrimido[4,5-b]indole Hydrochloride Salt
To a suspension of 6-carbamoyl-4-(l-piperidinyl)-9H-pyrimido[4,5-b]indole (0.082 g) in 1,4-dioxane (1 ml) and methanol (1 ml) was added 4N hydrochloride 1,4-dioxane solution (0.15 ml). The mixture was allowed to stand for 5 min and water (0.5 ml) was added. When the mixture was warmed by heat gun, the clouded reaction mixture turned to a clear solution, that was concentrated in vacuo. The residue was recrystallized from a mixture of ethanol and water to obtain 6- carbamoyl-4-(l-piperidinyl)-9H-pyrimido[4,5-b]indole hydrochloride Salt as colorless crystals (94 mg, >99%). Mp 278°C (dec); LC-MS (ESI): Retention time: 2.4 min (YMC-Pack Pro C18,
2.0 mmφ x 35 mm, acetonitrile-water (9:1 to 1:9), 1 ml/min) min, Calcd [M+1]: 296, Found: m/z 296. Molecular weight: 331.81 Activity grade MKK7: A Example 1-3
(1) 6-Cyano-4-(l-mo horyl)-9H-pyrimido[4,5-b]indole
According to the procedure for the synthesis of methyl 4-(l-piρeridinyl)-9H- pyrimido[4,5-b]indole-6-carboxylate from methyl 3-fluoro-4-nitrobenzoate, that was described in Example 1-1, 6-cyano-4-(l-piperidinyl)-9H-ρyrimido-
[4,5-b]indole was prepared from 3-bromo-4-nitrobenzonitrile.
Mp >300°C (dec); LC-MS (ESI): Retention time: 3.70 min, Calcd [M+1]:
280, Found: m/z 280.
Molecular weight: 279.30
(2) 6-Carbamoyl-4-(l-morphoryl)-9H-pyrimido[4,5-b]indole
To a solution of 6-cyano-4-(l-morpholino)pyrimido[4,5-b]indole (Illb) in the mixture of 30% hydrogen peroxide (2 ml) and methanol (2 ml) was added IN sodium hydroxide (1 ml) at 0°C. The mixture was stirred at room temperature for 1 hr and extracted with dichloromethane (5 ml X 3). The organic layer was washed with brine and dried over sodium sulfate to be concentrated in vacuo. The residue was applied onto preparative silica gel TLC (dichloromethane / methanol = 95/5) to obtain 6-carbamoyl-4-(l-morphoryl)- 9H-pyrimido[4,5-b]indole (0.015 g, 28%) as a pale yellow solid Mp >300°C (dec); LC-MS (ESI): Retention time: 3.06 min, Calcd [M+1]: 298, Found: m/z 298.
Molecular weight: 279.32 Activity grade MKK7: A
In a similar method according to the Examplel -1, 1-2 and 1-3 above, the compounds in Example 1-4 to 1-32 were synthesized.
Table 1
Example 2-1
(1) 2-Amino-6-cyano-4-hydroxy-9H-pyrimido[4,5-b]indole Hydrochloride Salt
The starting material, ethyl 2-amino-6-cyanoindole-3 -carboxylate was prepared according to the procedure described in Example 1-1. To a suspension of ethyl 2-amino-6-cyanoindole-3-carboxylate (0.96 g) and cyanamide (0.88 g) in 1,4-dioxane (50 ml) was added 36% hydrochloric acid (0.84 ml). The mixture was refluxed for 2 days and, after cooling to room temperature, concentrated in vacuo. The residue was washed with diethylether and triturated with methanol to give precipitates, that was collected by a paper filter and washed with methanol. The collected solid was dried at 85°C in vacuo to give a colorless solid (0.72 g, 66%).
(2) 2-Amino-4-chloro-6-cyano-9H-pyrimido[4,5-b]indole Hydrochloride Salt
To a solution of 2-amino-6-cyano-4-hydroxy-9H-pyrimido[4,5-b]indole (0.49 g) and N,N'-dimethylaniline (0.8 ml) in 1,4-dioxane (0.5 ml) was added phosphoryl chloride (3 ml) dropwise. The reaction mixture was stirred at 100°C overnight and, after cooling to room temperature, poured into crushed ice to give yellow precipitates, that was collected by a filter paper and dried at 80°C in vacuo to give a yellow solid (0.42 g, 69%).
(3) 2-Amino-6-cyano-4-(l -piperidinyl)-9H-pyrimido[4,5-b]indole
A mixture of 2-amino-4-chloro-6-cyanopyrimido[4,5-b]pyrimido[4,5-b]- indole (0.42 g) and piperidine (4 ml) was stirred at 100°C for 4 hrs. After cooling to room temperature, the reaction mixture was concentrated in vacuo and the residue was dissolved in dichloromethane (10 ml) and washed with brine. The organic layer was dried over sodium sulfate and concentrated in vacuo. The residue was purified by silica gel column chromatography (dichloromethane/methanol = 10/0 to 98/2) to obtain a yellow solid (0.13 g, 29%).
(4) 2-Amino-6-carbamoyl-4-(l-piperidinyl)-9H-pyrimido[4,5-b]indole
To a solution of 2-amino-6-cyano-4-(l-piperidinyl)-9H-pyrimido[4,5-b]- indole (0.08 g) in ethanol (3 ml) were added 4N sodium hydroxide water solution (0.6 ml) and 30% hydroperoxide (3 ml), successively. The mixture was stirred at room temperature for 2 h and poured into water (30 ml) to give yellow precipitates, that were collected by a filter paper and washed with diethylether and methanol, successively, to obtain 2-amino-6-carbamoyl-4-(l- piperidinyl)-9H-pyrimido[4,5-b]indole as a yellow solid (0.07 mg, 80%). Mp >184°C; LC-MS (ESI): Retention time: 3.45 min, Calcd [M+1]: 311, Found: m/z 311. Molecular weight: 310.36 Activity grade MKK7: A
Example 3-1
(1) 3-Ethyl 5-methyl 2-(ethanimidoylamino)-lH-indole-3,5-dicarboxylate hydrochloride
A mixture of methyl 2-amino-3-ethoxycarbonyl-lH- indole-5-carboxylate (0.48 g), obtained in step (6) of Example 1-1, and 4 N hydrochloric acid in 1,4-dioxane (5 ml) in acetonitrile (5 ml) was stirred at room temperature overnight. The resulting precipitate was collected by filtration, washed with 1,4-dioxane and acetonitrile to give 3-ethyl 5-methyl 2-(ethanimidoylamino)- lH-indole-3,5-dicarboxylate hydrochloride as a white solid (0.537 g, 87%).
(2) Methyl 4-hydroxy-2-methyl-9H-pyrimido[4,5-b]indole-6-carboxylate
To a suspension of 3-ethyl 5-methyl 2-(ethanimidoylamino)-lH-indole-3,5- dicarboxylate hydrochloride (0.540 g) in methanol (5 ml) was added a solution of sodium bicarbonate (0.500 g) in water (5 ml), and the mixture was stirred at 60°C for 1.5 hrs. The mixture was cooled to room temperature and the resulting precipitate was collected by filtration, washed with methanol, dried at 60°C in vacuo to give methyl 4-hydroxy-2-methyl-9H-pyrimido[4,5- b]indole-6-carboxylate as a solid (514 mg), which was used next step without purification.
(3) Methyl 4-chloro-2-methyl-9H-pyrimido[4,5-b]indole-6-carboxylate
To a solution of methyl 4-hydroxy-2-methyl-9H-pyrimido[4,5-b]indole-6- carboxylate (0.41 g) and N,N-dimethylaniline (0.57 g) in 1,4-dioxane (2 ml) was added phosphoryl chloride (2.1 ml). The mixture was stirred at 100°C overnight, and poured into crushed ice. The resulting precipitates was collected by filtration dried at 80°C in vacuo to give methyl 4-chloro-2- methyl-9H-pyrimido[4,5-b]indole-6-carboxylate as a solid (0.409 g, 94%).
(4) Methyl 2-methyl-4-(l -piperidinyl)-9H-pyrimido[4,5-b]indole-6-carboxylate
A mixture of methyl 4-chloro-2-methyl-9H-pyrimido[4,5-b]indole-6-carb- oxylate (0.41 g) and piperidine (2 ml) in 1,4-dioxane (1 ml) was stirred at 100°C for 0.5 hrs and poured into water (10 ml). Resulting precipitate was collected by filtration, and recrystallized from AcOEt and ethanol to give methyl 2-methyl-4-(l-piperidinyl)-9H-pyrimido[4,5-b]indole-6-carboxylate as a colorless crystalline solid (0.296 g, 62%).
Mp 259-260°C; LC-MS (ESI): Retention time: 4.85 min, Calcd [M+1]: 325,
Found: m/z 325.
Molecular weight: 324.39
Activity grade MKK7: A
(5) 2-Methyl-4-(l -piperidinyl)-9H-pyrimido[4,5-b]indole-6-carboxylic acid
To a solution of methyl 2-methyl-4-(l-piperidinyl)-9H-pyrimido[4,5-b]- indole-6-carboxylate (0.25 g) in methanol (2 ml) was added 4N NaOH aqueous solution (0.5 ml). The mixture was refluxed for 6 hrs, diluted with water (30 ml) and acidified with acetic acid (0.3 ml). The resulting precipitate was collected by filtration, and recrystallized from AcOH and ethanol to give
2-methyl-4-(l-piperidinyl)-9H-pyrimido[4,5-b]indole-6-carboxylic acid as a white solid (0.107 g, 45%).
Mp >300°C; LC-MS (ESI): Retention time: 3.26, Calcd [M+1]: 311, Found: m/z 311.
Molecular weight: 310.36
Activity grade MKK7: A
Activity grade MKK4: A (6) 2-Methyl-4-(l-ρiperidinyl)-9H-pyrimido[4,5-b]indole-6-carboxamide
2-methyl-4-(l-piperidinyl)-9H-pyrimido[4,5-b]indole-6-carboxylic acid
(0.05 g) was dissolved in DMF (2 ml) and carbonyldiimidazole (0.03 g) was added at 0°C The mixture was stireed at room temperature. After 0.5 hrs, 28%o ammonium aqueous solution (0.5 ml) was added. The resulting mixture was st red at room temperature overnight, and diluted with water (30 ml). The resulting precipitate was collected by filtration, dried at 60 °C in vacuo to give 2-methyl-4-( 1 -piperidinyl)-9H-pyrimido[4,5-b]indole-6-carboxamide as a colorless solid (0.028 g, 56%).
Mp >300 °C; LC-MS (ESI): Retention time: 3.80 min, Calcd [M+1]: 310, Found: m/z 310. Molecular weight: 309.37. Activity grade MKK7: A Activity grade MKK4: A
Example 4-1
(1) 4-Benzyloxy-2-fluoronifrobenzene
A mixture of 3-fluoro-4-nitrophenol (50.00 g, 318.26 mmol), benzyl bromide (57.16 g, 334.18 mmol) and potassium carbonate (87.97 g, 636.53 mmol) was refluxed in acetone (750 mL) for 18 hrs. After cooled to room temperature, the mixture was passed through a filter and the filfrate was concentrated under reduced pressure. The residue was dissolved in ethyl acetate and washed with 5% sodium bicarbonate. The separated organic layer was washed with brine, dried over magnesium sulfate and concentrated under reduced pressure. The residue was triturated with hexane, filtered and air-dried to give 4-benzyloxy- 2-fluoronitrobenzene as a colorless solid (74.65 g, 94.9 %).
(2) Ethyl 5-benzyloxy-2-nitrophenylcyanoacetate
To the suspension of 60% sodium hydride (9.71 g, 242.69 mmol) in 300 ml of N,N-dimethylfornιamide (DMF) were added a solution of ethyl cyanoacetate (15.10 g, 133.48 mmol) in DMF (40 mL) and 4-benzyloxy-2- fluoronitrobenzene (20.0 g, 80.9 mmol), successively, at 0°C. The mixture was stirred at 70°C for 4 hrs and cooled to room temperature. The resulting suspension was poured into 10% potassium hydroxide and washed with ether. The aqueous layer was acidified with 10% hydrochloride and extracted with ether. The extract was washed with brine and dried over magnesium sulfate. The solvent was removed and the residue was purified by silica gel column chromatography (Hexane: AcOEt = 5:1) to give ethyl 5-benzyloxy-2- nifrophenylcyanoacetate (23.6 g, 84.9%) as a pale brown oil.
(3) 2-Amino-5-benzyloxy-3-ethoxycarbonyl-(lH)-indole
To a mixture of acetic acid (190 mL) and toluene (380 mL) was added ethyl 5-benzyloxy-2-nitrophenylcyanoacetate (63.99 g, 188.02 mmol). The resulting mixture was stirred at 80°C to give a clear solution. The external heating was removed and zinc powder (98.33 g, 1504.14 mmol) was added slowly, portionwise. The reaction mixture was stirred at 80°C for 3 hrs and passed through a filter, while the solution is hot, and the collected solid was washed with toluene. The filtrate was evaporated under reduced pressure and the residue was triturated with ether to give 2-amino-5-benzyloxy-3-ethoxy- carbonyl-(lH)-indole as a purplish solid (28.89 g, yield 49.5%).
(4) 5-Benzyloxy-3-ethoxycarbonyl-2-guanidyl-(lH)-indole hydrochloride
A suspension of ethyl 2-amino-5-benzyloxy-3-ethoxycarbonyl-(lH)-indole (11.36 g, 36.60 mmol), cyanamide (2.46 g, 58.55 mmol), and 36% HCl (3 ml) in 1,4-dioxane (300 ml) was refluxed for 2 days. The reaction mixture was cooled to room temperature and the precipitated solid was collected on a filter and washed with dry ether to give 5-benzyloxy-3-ethoxycarbonyl-2-guanidyl- (lH)-indole hydrochloride (2.99 g, 21%), that was used for the following reaction without any further purification.
(5) 2-Amino-6-benzyloxy-4-hydroxy-(9H)-pyrimido[4,5-b]indole
A mixture of 5-benzyloxy-3-ethoxycarbonyl-2-guanidyl-(lH)-indole hydrochloride (4.54 g, 11.67 mmol) and sodium hydroxide (4.67 g, 116.72 mmol) was refluxed in water (50 mL) for 6 hrs. After cooled to room temperature, the precipitate was collected on a filter and air-dried to give 2-amino-6- benzyloxy-4-hydroxy-(9H)-pyrimido[4,5-b]indole as a crude product, that was used for the following reaction without any further purification.
(6) 2-Amino-6-benzyloxy-4-chloiO-(9H)-pyrimido[4,5-b]indole hydrochloride
A mixture of 2-amino-6-benzyloxy-4-hydroxy-(9H)-pyrimido[4,5-b]indole (1.00 g, 3.26 mmol) obtained in the step (5) and N,N-dimethylaniline (1.19 g, 9.79 mmol) was refluxed in phosphoryl chloride (3.0 g) for 2 hrs. The reaction mixture was concentrated under reduced pressure and the residual syrup was treated with ice water. The resulting solid was collected on a filter and washed with ethanol and ether to give 2-amino-6-benzyloxy-4-chloro- (9H)-pyrimido[4,5-b]indole hydrochloride as a pale green solid. This crude product was used for the following reaction without further purification.
(7) 2-Amino-6-benzyloxy-4-(l-homopiperidino)-(9H)-pyrimido[4,5-b]indole
The mixture of 2-amino-6-benzyloxy-4-chloro-(9H)-pyrimido[4,5-b]indole hydrochloride (0.18 g, 0.50mmol) obtained in the step (6) and homo- piperidine (0.99 g, 10.0 mmol) was refluxed in 1,4-dioxane (5 mL) for 18 hrs and cooled to room temperature. The solvent was removed in vacuo and the residue was dissolved in chloroform and washed with water. The chloroform solution was dried over magnesium sulfate and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (CHCl3:MeOH = 50:1) to give 2-amino-6-benzyloxy-4-(l-homopiperidino)- (9H)-pyrimido[4,5-b]indole (0.012 g, 6.2 %). (8) 2-Amino-6-hydroxy-4-(l-homopiperidino)-(9H)-pyrimido[4,5-b]indole
A mixture of 2-amino-6-benzyloxy-4-(l-homopiperidino)-(9H)-pyrimido[4,5- b]indole (0.010 g, 0.023 mmol) in methanol (2 mL) were added acetic acid
(1.0 mL) and trifluoroboran diethyl etherate (0.01 g, 0.069 mmol). 10% Pd-C
(100 mg) was added to the mixture and stirred at room temperature for 2 hrs under a hydrogen atmosphere (3 atm). The mixture was passed through a
Celite filter and the filtrate was concentrated under reduced pressure. The residue was triturated with ether and purified by prep TLC (silica gel)
(CHCl3:MeOH = 5:1) to give 2-amino-6-hydroxy-4-(l-homoρiperidino)-
(9H)-pyrimido[4,5-b]indole (0.006 g, 86.9%) as a colorless solid.
Mp >240°C; LC-MS (ESI): Retention time: 3.48 min, Calcd [M + H]+ 298,
Found: 298.
Molecular weight: 297.36
Activity grade MKK7: A
Activity grade MKK4: A Example 4-2
(1) 2-amino-6-benzyloxy-4-(l-isoindolino)-(9H)-pyrimido[4,5-b]indole
The mixture of 2-amino-6-benzyloxy-4-chloro-(9H)-pyrimido[4,5-b]indole hydrochloride (0.10 g, 0.28 mmol) obtained in the example 4-1 step (6) and isoindoline (0.66 g, 5.54 mmol) was refluxed in 1,4-dioxane (5 mL) for 18 hrs and cooled to room temperature. The solvent was removed in vacuo and the residue was dissolved in chloroform and washed with water. The chloroform solution was dried over magnesium sulfate and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (CHCl3:MeOH = 50:1) to give 2-amino-6-benzyloxy-4-(l-isoindolino)-(9H)- pyrimido[4,5-b]indole (0.040 g, 35.5 %).
(2) 2-amino-6-hydroxy-4-(l -isoindolino)-(9H)-pyrimido[4,5-b]indole
A mixture of 2-amino-6-benzyloxy-4-(l-isoindolino)-(9H)-pyrimido[4,5- b]indole (0.038 g, 0.09 mmol) in methanol (3 mL) were added acetic acid (1.0 mL) and trifluoroboran diethyl etherate (0.04 g, 0.28 mmol). 10% Pd-C (100 mg) was added to the mixture and stirred at room temperature for 2 hrs under a hydrogen atmosphere (3 atm). The mixture was passed through a Celite filter and the filtrate was concentrated under reduced pressure. The residue was triturated with ether and purified by prep TLC (silica gel) (CHCl3:MeOH = 5:1) to give 2-amino-6-hydroxy-4-(l-isoindolino)-(9H)- pyrimido[4,5-b]indole (0.006 g, 86.9%) as a colorless solid. Mp 230°C; LC-MS (ESI): Retention time: 3.46 min, Calcd [M + H]+ 318, Found: 318. Molecular weight: 317.35
Examples 4-3 to 4-39
In the similar manners as described in Example 4-1 and 4-2 above, the compounds in
Example 4-3 to 4-13 as shown in Table 4 were synthesized. Table 4
Example 5
(1) 6-Benzyloxy-4-hydroxy-(9H)-pyrimido[4,5-b]indole
To a dry NaOMe prepared from Na (60 mg) and absolute methanol (2 mL) was added a solution of 2-amino-5-benzyloxy-3-ethoxycarbonyl-(lH)-indole (0.50 g, 1.61 mmol) obtained in the step (3) of Example 4-1 in formamide (15 mL). The mixture was refluxed for 18 hrs and cooled to room temperature. The reaction mixture was poured into water (100 mL) and the precipitated materials were collected on a filter, that was purified by silica gel column chromatography (CHC13 : MeOH = 50 :1) to give 6-benzyloxy-4- hydroxy-(9H)-pyrimido[4,5-b]indole (0.137g, 29.2%)
(2) 6-Benzyloxy-4-chloro-(9H)-pyrimido[4,5-b]indole hydrochloride
A mixture of 6-benzyloxy-4-hydroxy-(9H)-pyrimido[4,5-b]indole (0.06 mg, 0.213 mmol) and N,N-dimethylaniline (0.08 g, 0.638 mmol) was refluxed in phosphoryl chloride (1 g) for 5 hrs. The reaction mixture was evaporated and the residual syrup was treated with ice water. The precipitated solid was collected on a filter and washed with ether to give 6-benzyloxy-4-chloro- (9H)-pyrimido[4,5-b]indole hydrochloride as a brown solid (0.039g, 52.9%).
(3) 6-Benzyloxy-4-(l-piperidino)-(9H)-pyrimido[4,5-b]indole
According to the step (7) of Example 4-1, 6-benzyloxy-4-chloro-(9H)- pyrimido[4,5-b]indole hydrochloride (0.12 g, 0.355 mmol) was reacted with piperidine (1.5 mL) in 1,4-dioxane (5.0 mL) to give 6-benzyloxy~4-(l- piperidino)-(9H)-pyrimido[4,5-b]indole as a pale orange solid (0.024 g, 18.8%).
(4) 6-Hydroxy-4-(l-piperidino)-(9H)-pyrimido[4,5-b]indole
According to the step (8) of Example 4-1, 6-benzyloxy-4-(l-piperidino)-(9H)- pyrimido[4,5-b]indole (0.02g, 0.05 mmol) was hydrogenated to deprotect 6- benzyloxy group to give 6-hydroxy-4-(l-piperidino)-(9H)-pyrimido[4,5- b]indole (0.01 g, 74.2 %) as a colorless solid.
Mp 270°C; LC-MS (ESI): Retention time: 3.24 min, Calcd [M + H]+ 269,
Found: 269.
Molecular weight: 268.32
Activity grade MKK7: A
Examples 5-2 to 5-59
In the similar manners as described in Example5-1 above, the compounds in Example 5-2 to 5-59 as shown in Table 5 were synthesized. Table 5
Example 6
( 1 ) 6-B enzyloxy-4-hydroxy-2-methyl-9H-pyrimido [4, 5 -b]indole
A mixture of 2-amino-5-benzyloxy-3-ethoxycarbonyl-(lH)-indole (5.00 g, 16.11) obtained in the example 4-1 step (3) and 4 N hydrochloric acid in 1,4- dioxane (100 ml) in acetonitrile (100 ml) was stirred at room temperature overnight. The resulting precipitate was collected by filtration, washed with 1,4-dioxane and acetonitrile and dissolved in a mixture of ethanol (85 mL) and H2O (5 mL). To the mixture was added a solution of sodium hydroxide (1.58 g, 39.44 mmol) in water (10 ml), and the mixture was stirred at 50°C for 2 hrs. The mixture was cooled to room temperature and the resulting precipitate was collected by filfration, washed with ether and dried at 60°C in vacuo to give 6-benzyloxy-4-hydroxy-2-methyl-9H-pyrimido[4,5-b]indole as a white solid (4.26 g, 87.4 %).
(2) 6-Benzyloxy-4-chloro-2-methyl-9H-pyrimido[4,5-b]indole
To a suspension of 6-benzyloxy-4-hydroxy-2-methyl-9H-pyrimido[4,5-b]- indole (1.15 g, 3.77 mmol) in phosphoryl chloride (20 ml) was added N,N- dimethylaniline (1.37 g, 11.30 mmol) at room temperature. The mixture was stirred at 80°C for 4 hrs. The reaction mixture was concentrated in vacuo, poured into ice and stined at 0°C to give the brown precipitate. The precipitate was dried in vacuo at 65°C overnight to give 6-benzyloxy-4- chloro-2-methyl-9H-pyrimido[4,5-b]indole (1.22 g, 100%), wliich was used for next step without further purification.
(3) 6-benzyloxy-4-[(2-hydroxy- 1 -methylethyl)amino]-2-methyl-9H-pyrimido[4,5- b]indole
A mixture of 6-benzyloxy-4-chloro-2-methyl-9H-pyrimido[4,5-b]indole (0.18 g, 0.361 mmol) and DL-alaninol (0.81 g, 10.84 mmol) in 1,4-dioxane (2 ml) was stined at 100°C overnight. After toluene was added there, the solvent was evaporated. The residue was dissolved in CH2C12 and the organic layer was washed with aq. NaHCO3, dried over MgSO4 and concentrated in vacuo. The residue was purified by silica gel column chromatography (CH2Cl2/MeOH= 0/l-19/l) to give 6-benzyloxy-4-[(2-hydroxy-l-methyl- ethyl)amino]-2-methyl-9H-pyrimido[4,5-b]indole as a white solid (0.063 g, 48.1%).
Mp 210°C; LC-MS (ESI): Retention time: 4.41 min, Calcd [M+1]: 363, Found: m/z 363. Molecular weight: 362.44 Activity grade MKK7: A
(4) 6-Hydroxy-4-[(2-hydroxy-l-methylethyl)amino]-2-methyl-(9H)-pyrimido- [4,5-b]indole
To a solution of 6-benzyloxy-4-[(2-hydroxy-l-methylethyl)amino]-2-methyl- 9H-pyrimido[4,5-b]indole (0.056 g, 0.155 mmol) in THF (1.5 mL) and acetic 0.5 mL) was added 10% Pd-C (100 mg). Hydrogen gas was charged in the flask. The mixture was stirred at room temperature overnight under a hydrogen atmosphere (3 atm). The mixture was passed through a filter and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (CH2Cl2/MeOH = 19/1-9/1) to give 6- hydroxy-4- [(2-hydroxy- 1 -methylethyl)amino]-2-methyl-(9H)-pyrimido[4,5- b]indole (0.031 g, 72.5%) as a white solid.
Mp 297°C; LC-MS (ESI): Retention time: 3.25 min, Calcd [M + H]+ 273, Found: 273.
Molecular weight: 272.31 Activity grade MKK7 : A
Activity grade MKK4: A
Examples 6-2 to 6-16
In the similar manners as described in Example 6-1 above, the compounds in
Example 6-2 to 6-16 as shown in Table 6 were synthesized.
Table 6

Claims

Claims
(1) A pyrimido[4,5-b]indole derivative of the formula (I), its tautomeric or stereoisomeric form, or a salt thereof:
wherein
R is hydrogen, amino, (Cι-6)alkyl, (C2-6)alkenyl, (C2-6)alkynyl, halogen substituted (C1-6) alkyl, cyano, cyano(C1-6)alkyl, (C3-8)cycloalkyl, nitro, nitro(C1-6)alkyl or fluoro;
R2 and R3 independently represent hydrogen, (C2-6)alkenyl, (C2-6)alkynyl, (C3-8)cycloalkyl optionally substituted by hydroxy or hydroxy (C1-6)alkyl, (C3-8)heterocycloalkyl optionally substituted by (C1-6)- alkoxycarbonyl, (C1-6)alkylcarbonyl, (C1-6)alkylsulfonyl, arylsulfonyl, heteroarylsulfonyl, or (C1-6)alkyl which is optionally substituted by hydroxy, (C1-6)alkoxy, (C1-6)alkoxycarbonyl, (C3-8)cycloalkyl, (C3-8)- heterocycloalkyl, carbamoyl, (C1-6)alkylcarbamoyl, cyano, amino, (C1-6)alkylamino, (C1-6)alkoxycarbonylamino, N-(C1-6)alkoxy- carbonylamino(C1-6)alkyl, arylamino, heteroarylamino, carboxyl, nitro, trifluoromethyl, aryl, piperidinyl, imidazolyl, (C1-6)alkyl- piperidinyl, (C1-6)alkylcarbonylpiperidinyl, ( -^alkoxycarbonyl- piperidinyl, carbamoylpiperidinyl, ((C1-6)alkylcarbamoyl)piperidinyl, arylpiperidinyl, heteroarylpiperidinyl, aryloxycarbonylpiperidinyl, heteroaryloxycarbonylpiperidinyl, (C1.6)alkylsulfonylpiperidinyl, aryl- sulfonylpiperidinyl, heteroarylsulfonylpiperidinyl, pynolidyl, (C1-6)- alkylpynolidyl, (C1-6)alkylcarbonylpynolidyl, (C1-6)alkoxycarbonyl- pynolidyl, carbamoylpynohdyl, ((C1-6)alkylcarbamoyl)pynolidyl, arylpynolidyl, heteroarylpynolidyl, aryloxycarbonylpynolidyl, heteroaryloxycarbonylpynolidyl, (C1-6)alkylsulfonylpynolidyl, aryl- sulfonylpynolidyl, or, heteroarylsulfonylpynolidyl,
or
R2 and R3 are taken together with the attached nitrogen atom to form a 3-8 membered saturated ring optionally interrupted by one or two atoms selected from the group consisting of oxygen, sulfur and nitrogen,
wherein
said saturated ring is optionally fused by benzene or 3-8 membered saturated ring having 0 to 3 nifrogen atoms,
or
said saturated ring is optionally having substituents independently selected from the group consisting of halogen, hydroxy, carbamoyl, carboxy, amino, oxo, pynolidino, (C1-6)alkoxy, (C2-6)alkenyl, (C -6)- alkynyl, (C3-8)cycloalkyl, (C1-6)alkylcarbonyl, (C1-6)alkylsulfonyl, arylsulfonyl, (C1-6)alkyl, hydroxy(C1-6)alkyl, (C1-6)alkoxy(C1-6)alkyl, (C1-6)alkoxycarbonyl, (C1-6)alkoxycarbonyl(C1-6)alkyl, (C3- )cyclo- alkyl(C1-6)alkyl, aryl(C1-6)alkyl, carbamoyl(C1-6)alkyl, (C1-6)alkyl- carbamoyl(C1-6)alkyl, cyano(C1-6)alkyl, amino (C1-6)alkyl, N- (C1-6)alkylamino(C1-6)alkyl, carboxyl(C1-6)alkyl, nitro (C1-6)alkyl, trifluoromethyl(C1-6)alkyl, N-(C1-6)alkylamino, N-(C1-6)alkanoyl- amino, N-(C1-6)alkoxycarbonylamino, N-[amino(C1-6)alkyl]amino, N- (C1-6)alkoxycarbonylamino N-(C1-6)alkylamino, and N-(C1-6)alkyl- carboxyamino
or
said substituents optionally together with the carbon to which they are attached, form a 3-8 membered saturated ring that forms a spiro ring,
wherein said spiro ring is optionally interrupted by 0 to 3 nitrogen or oxygen atoms;
R4 is halogen, hydroxy, (C1-6 )alkyl, amino (C1-6 )alkyl, (C1-6)alkoxy, cyano, carboxy, (C1-6)alkoxycarbony, or carbamoyl; and
R5 is hydrogen, or ( ^alkyl.
(2) The pyrimido[4,5-b]indole derivative, its tautomeric or stereoisomeric form, or a salt thereof as claimed in claim 1,
wherein
R1 is amino,
R2 is pynolidyl(C1-6)alkyl, saturated(C3-8)cycloalkyl or benzyl, and
R3 is hydrogen, (C1-6)alkyl, or benzyl,
or
R and R are taken together with the attached nifrogen atom to form a 7 membered saturated heterocyclic ring optionally interrupted by one or two atoms selected from the group consisting of oxygen, sulfur and nifrogen,
a 5-7 membered saturated heterocyclic ring which is fused by benzene or saturated(C3-8)cycloalkyl,
or
a 8-12 membered spiro ring, optionally interrupted by one or two oxygen atoms.
(3) The pyrimido[4,5-b]indole derivative, its tautomeric or stereoisomeric form, or a salt thereof as claimed in claim 1,
wherein
R1 is amino,
R4 is carbamoyl or amino (C1-6 )alkyl; and
R is hydrogen or (C1-6)alkyl.
(4) The pyrimido[4,5-b]indole derivative, its tautomeric or stereoisomeric fonn, or a salt thereof as claimed in claim 1 ,
wherein
R is carbamoyl; and
R5 is hydrogen. (5) The pyrimido[4,
5-b]indole derivative, its tautomeric or stereoisomeric form, or a salt thereof as claimed in claim 1,
wherein
R1 is hydrogen, methyl or amino;
R is carbamoyl; and
R5 is hydrogen.
(6) The pyrimido[4,5-b]indole derivative, its tautomeric or stereoisomeric form, or a salt thereof as claimed in claim 1,
wherein
R2 and R3 independently represent hydrogen, (C1-6)alkyl optionally substituted by amino, hydroxy, methoxy, imidazolyl, (C -6)alkoxy or (C1-6)alkoxycarbonyl, (C3-8)cycloalkyl optionally substituted by hydroxy or hydroxy (C1-6)alkyl, benzyl, or 5-7 membered saturated heterocyclic ring interrupted by one or two oxygen atoms;
or
R2 and R3 are taken together with the attached nifrogen atom to form a saturated heterocyclic ring selected from the group consisting of pynolidino, piperidino, piperazino, homopiperidino, and morpholino
wherein said heterocyclic ring is optionally substituted by amino, methyl- amino, acetamido, hydroxy, oxo, carbamoyl, benzyl, bromo, carboxy, pynolidino, or (C1-6 )alkyl optionally substituted by hydroxy, amino or carboxy,
or
said heterocyclic ring is optionally fused by benzene or saturated- (C3-8)cycloalkyl;
R is carbamoyl or amino(C1-6 )alkyl; and
R5 is hydrogen.
(7) The pyrimido[4,5-b]indole derivative, its tautomeric or stereoisomeric form, or a salt thereof as claimed in claim 1,
wherein
R1 is hydrogen, methyl or amino;
R2 and R3 independently represent hydrogen, (C1-6)alkyl optionally substituted by amino, hydroxy, methoxy, imidazolyl, (C1-6)alkoxy or (C1-6)alkoxycarbonyl, (C3-8)cycloalkyl optionally substituted by hydroxy or hydroxy (C1-6)alkyl, benzyl, or 5-7 membered saturated heterocyclic ring interrupted by one or two oxygen atoms;
or R2 and R3 are taken together with the attached nifrogen atom to form a saturated heterocyclic ring selected from the group consisting of pynolidino, piperidino, piperazino, homopiperidino, and morpholino
wherein
said heterocyclic ring is optionally substituted by amino, methyl- amino, acetamido, hydroxy, oxo, carbamoyl, benzyl, bromo, carboxy, pynolidino, or (C1-6 )alkyl optionally substituted by hydroxy, amino or carboxy,
or
said heterocyclic ring is optionally fused by benzene or saturated- (C3-8)cycloalkyl;
R is carbamoyl; and
R is hydrogen.
(8) The pyrimido[4,5-b]indole derivative, its tautomeric or stereoisomeric form, or a salt thereof as claimed in claim 1,
wherein
R1 is amino;
R2 is saturated(C3-8)cycloalkyl or benzyl;
R3 is hydrogen, or (C1-6)alkyl,
or R2 and R3 are taken together with the attached nifrogen atom to form
a homopiperidino,
a 5-7 membered saturated heterocyclic ring which is fused by benzene or saturated(C3-8)cycloalkyl,
wherein said heterocyclic ring is selected from the group consisting of pynolidino, piperidino, piperazino, homopiperidino, and morpholino,
or
1, 4 dioxa-8-azaspiro[4,5]decane; and
R is hydrogen.
(9) The pyrimido[4,5-b]indole derivative, its tautomeric or stereoisomeric form, or a salt thereof as claimed in claim 1, wherein said derivative is selected from the group consisting of the following compounds:
6-Carbamoyl-4 (l-piperidinyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4-ι (l-mo holinyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4-ι (4-hydroxy-l-piperidinyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4 (cycloheptylamino)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4 (4-methyl- 1 -piperidinyl)-9H-pyrimido [4,5-b]indole,
6-Carbamoyl-4-ι (3-methyl-l-piperidinyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4-ι (3,5-dimethyl- 1 -piperidinyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4- [2-(hydroxymethyl)-l-piperidinyl]-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4- (cyclopentylamino)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4 (l-azepanyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4 [(2-aminoethyl)amino]-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-[4-(2-hydroxyethyl)-l-piperidmyl]-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-(cyclohexylamino)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-[(2-hydroxyethyl)amino]-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-[(2-hydroxyethyl)(methyl)amino]-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-(4-benzyl-4-hydroxy- 1 -piperidmyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4-(l-pynolidinyl)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-[(2-hydroxy-l-memylemyl)arnino]-9H-pyrimido[4,5-b]m^ 6-Carbamoyl-4-(2-ethyl-l-piperidinyl)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-[4-(hydroxymethyl)-l-piperidinyl]-9H-pyrimido[4,5-b]indole, 4-(3-amino-l-piperidinyl)-6-carbamoyl-9H-pyrimido[4,5-b]indole, tert-butyl l-[6-carbamoyl--9H-pyrinήdo[4,5-b]__ndol-4-yl]-3-piperidinylcarbamate, 4-[(3-aminoproρyl)amino]-6-carbamoyl-9H-pyrimido[4,5-b]indole, tert-butyl 3-{[6-carbamoyl-9H-pyrimido[4,5-b]indol-4 yl]am__no}propylcarbamate, 4-(3 -amino- 1 -pynolidinyl) -6-carbamoyl-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-(dipropylamino)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4-[3-(methylamino)-l-pynolidinyl]-9H-pyrimido[4,5-b]indole, l-[6-carbamoyl-9H-pyrimido[4,5-b]indol-4-yl]-4-piperidinecarboxylic acid, 6-Carbamoyl-4-[4-(acetylamino)-l-piperidinyl]-9H-pyrimido[4,5-b]indole, 2-Amino-6-carbamoyl-4-(l-piperidinyl)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-2-methyl-4-(l -piperidinyl)-9H-pyrimido[4,5-b]indole, and their tautomeric and stereoisomeric form, and salts thereof.
(10) A medicament comprising the pyrimido[4,5-b]indole derivative, its tautomeric or stereoisomeric form, or a physiologically acceptable salt thereof as claimed in claim 1 as an active ingredient.
(11) The medicament as claimed in claim 10, further comprising one or more pharmaceutically acceptable excipients.
(12) The medicament as claimed in claim 10, wherein the pyrimido[4,5-b]indole derivative, its tautomeric or stereoisomeric form, or a physiologically acceptable salt thereof is a MKK7 inhibitor.
(13) The medicament as claimed in claim 10, wherein the pyrimido[4,5-b]indole derivative, its tautomeric or stereoisomeric form, or a physiologically acceptable salt thereof is a MKK4 inhibitor.
(14) An agent to treat or prevent a inflammatory or immunoregulatory disorder; comprising the pyrimido[4,5-b]indole derivative, its tautomeric or stereoisomeric form, or a physiologically acceptable salt thereof as claimed in claim 1 as an active ingredient.
(15) An agent to freat or prevent asthma, rhinitis, and allergic diseases, and auto- immune pathologies such as rheumatoid arthritis, Grave's disease, and atherosclerosis; comprising the pyrimido[4,5-b]indole derivative, its tautomeric or stereoisomeric form, or a physiologically acceptable salt thereof as claimed in claim 1 as an active ingredient.
(16) An agent to treat or prevent a neurodegenerative disorders, Alzheimer's disease, or focal ischemia; comprising the pyrimido[4,5-b]indole derivative, its tautomeric or stereoisomeric form, or a physiologically acceptable salt thereof as claimed in claim 1 as an active ingredient.
(17) An agent to freat a disease selected from the group consisting of ischemia, myocardial injury, pulmonary hypertension, renal failure, Huntington's chorea and cardiac hypertrophy; comprising the pyrimido[4,5-b]indole derivative, its tautomeric or stereoisomeric form, or a physiologically acceptable salt thereof as claimed in claim 1 as an active ingredient.
(18) A method for freating or preventing disorder or disease associated with MKK7 activity in a human or animal subject, comprising administering to said subject a therapeutically effective amount of the pyrimido [4,5-b]indole derivative, its tautomeric or stereoisomeric form, or a physiologically acceptable salt thereof as claimed in claim 1.
(19) A method for treating or preventing disorder or disease associated with MKK4 activity in a human or animal subject, comprising administering to said subject a therapeutically effective amount of the pyrimido [4,5-b]indole derivative, its tautomeric or stereoisomeric form, or a physiologically acceptable salt thereof as claimed in claim 1.
(20) The method of claim 18 or 19, wherein said disorder or disease is a inflammatory or immunoregulatory disorder or disease.
(21) The method of claim 18 or 19, wherein said disorder or disease is selected from the group consisting of asthma, rhinitis, and allergic diseases, and autoimmune pathologies such as rheumatoid arthritis, Grave's disease, and atherosclerosis.
(22) The method of claim 18 or 19, wherein said disorder or disease is selected from the group consisting of neurodegenerative disorders, Alzheimer's disease, focal ischemia, ischemia, myocardial injury, pulmonary hypertension, renal failure, Huntington's chorea and cardiac hypertrophy.
(23) The method of claim 18 or 19, wherein said pyrimido [4,5-b]indole derivative is selected from the group consisting of:
6-Carbamoyl-4-(l-piperidinyl)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-(l -moφholinyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4-(4-hydroxy-l-piperidinyl)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4- -(cycloheptylamino)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4- -(4-methyl- 1 -piperidinyl)-9H-pyrimido [4,5-b]indole,
6-Carbamoyl-4- -(3-methyl-l-piperidinyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4- -(3,5-dimethyl-l-piperidinyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4- -[[2-(hydroxymethyl)-l-piperidinyl]-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4- -(cyclopentylamino)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4- -(l-azepanyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4- -[(2-aminoethyl)amino]-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4- -[4-(2-hydroxyethyl)-l-ρiperid_inyl]-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4- -((cyclohexylamino)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4- -[(2-hydroxyethyl)amino]-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4- -[(2-hydroxyethyl)(methyl)amino]-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4--((4-benzyl-4-hydroxy- 1 -piperidinyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4- -(l-pynolidinyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4-[[((2-hya^oxy-l-me ylemyl)ammo]-9H-pyriimdo[4,5-b]indole,
6-Carbamoyl-4- -(2-ethyl-l-piperidinyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4- -[4-(hydroxymethyl)-l-piperidinyl]-9H-pyrimido[4,5-b]indole, 4-(3-amino-l-piperidinyl)-6-carbamoyl-9H-pyrimido[4,5-b]indole, tert-butyl l-[6-carbamoyl--9H-pyrimido[4,5-b]indol-4-yl]-3-piperidinylcarba^ 4-[(3-aminopropyl)amino]-6-carbamoyl-9H-pyrimido[4,5-b]indole, tert-butyl 3-{[6-carbamoyl-9H-pyrimido[4,5-b]mdol-4-yl]amino}propylcarbamate, 4-(3-amino-l-pynolidinyl) -6-carbamoyl-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-(dipropylamino)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-[3-(methylamino)-l-pynolidinyl]-9H-pyrimido[4,5-b]indole, l-[6-carbamoyl-9H-pyrimido[4,5-b]indol-4-yl]-4-piperidinecarboxylic acid, 6-Carbamoyl-4-[4-(acetylamino)-l-piperidinyl]-9H-pyrimido[4,5-b]indole, 2-Amino-6-carbamoyl-4-(l -piperidinyl)-9H-pyrimido[4,5-b]indole, and 6-Carbamoyl-2-methyl-4-(l-piperidinyl)-9H-pyrimido[4,5-b]indole.
(24) The method of claim 18 or 19, wherein said pyrimido [4,5-b]indole derivative, its tautomeric or stereoisomeric form, or a physiologically acceptable salt thereof is administered with one or more pharmaceutically acceptable excipients.
(25) Use of the pyrimido [4,5-b]indole derivative, its tautomeric or stereoisomeric form, or a physiologically acceptable salt thereof as claimed in claim 1 in the preparation of a medicament.
(26) Use of pyrimido [4,5-b]indole derivative, its tautomeric or stereoisomeric form, or a physiologically acceptable salt thereof as claimed in claim 1 in the preparation of a medicament for treating or preventing disorder or disease associated with MKK7 activity.
(27) Use of the pyrimido [4,5-b]indole derivative, its tautomeric or stereoisomeric form, or a physiologically acceptable salt thereof as claimed in claim 1 in the preparation of a medicament for treating or preventing disorder or disease associated with MKK4 activity.
(28) The use of claim 26 or 27, wherein said disorder or disease is a inflammatory or immunoregulatory disorder or disease.
(29) The use of claim 26 or 27, wherein said disorder or disease is selected from the group consisting of asthma, rhinitis, and allergic diseases, and autoimmune pathologies such as rheumatoid arthritis, Grave's disease, and atherosclerosis.
(30) The use of claim 26 or 27, wherein said disorder or disease is selected from the group consisting of neurodegenerative disorders, Alzheimer's disease, focal ischemia, ischemia, myocardial injury, pulmonary hypertension, renal failure, Huntington's chorea and cardiac hypertrophy.
(31) The use of claim 26 or 27, wherein said pyrimido [4,5-b]indole derivative is selected from the group consisting of:
6-Carbamoyl-4-(l-piperidinyl)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-(l -morpholinyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4-(4-hydroxy-l-piperidinyl)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-(cycloheptylamino)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-(4-methyl-l-piperidinyl)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-(3-methyl-l-piperidinyl)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-(3,5-dimethyl-l-piperidinyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4-[2-(hydroxymethyl)-l-piperidinyl]-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-(cyclopentylamino)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-(l-azepanyl)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-[(2-aminoethyl)amino]-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-[4-(2-hydroxyethyl)-l -piperidmyl]-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4-(cyclohexylamino)-9H-ρyrimido[4,5-b]indole, 6-Carbamoyl-4-[(2-hydroxyethyl)amino]-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-[(2-hydroxyethyl)(methyl)amino]-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-(4-beri2yl-4-hydroxy-l-piperidmyl)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-(l -pynolidinyl)-9H-pyrimido[4,5-b]indole,
6-Carbamoyl-4-[(2-hydroxy-l-me ylemyl)amino]-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-(2-ethyl- 1 -piperidinyl)-9H-pyrimido [4, 5 -b] indole, 6-Carbamoyl-4-[4-(hydroxymethyl)-l-piperidinyl]-9H-pyrimido[4,5-b]indole, 4-(3-amino-l-piperidinyl)-6-carbamoyl-9H-pyrimido[4,5-b]indole, tert-butyl 1 -[6-carbamoyl--9H-pyriιmdo[4,5-b]indol-4-yl]-3-piρeridinylcarbamate,
4-[(3-aminopropyl)amino]-6-carbamoyl-9H-pyrimido[4,5-b]indole, tert-butyl 3-{[6-carbamoyl-9H-pyrimido[4,5-b]indol-4-yl]amino}propylcarbamate, 4-(3-amino-l-pynolidinyl) -6-carbamoyl-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-(dipropylamino)-9H-pyrimido[4,5-b]indole, 6-Carbamoyl-4-[3-(methylamino)-l-pynolidinyl]-9H-pyrimido[4,5-b]indole, l-[6-carbamoyl-9H-pyrimido[4,5-b]indol-4-yl]-4-piperidinecarboxylic acid, 6-Carbamoyl-4-[4-(acetylamino)-l-piperidinyl]-9H-pyrimido[4,5-b]indole, 2-Amino-6-carbamoyl-4-(l-piperidinyl)-9H-pyrimido[4,5-b]indole, and 6-Carbamoyl-2-methyl-4-(l-piperidinyl)-9H-pyrimido[4,5-b]indole.
(32) The use of claim 26 or 27, wherein said pyrimido [4,5-b]indole derivative, its tautomeric or stereoisomeric form, or a physiologically acceptable salt thereof is formulated with one or more pharmaceutically acceptable excipients.
EP02777332A 2001-10-31 2002-10-29 Pyrimido (4,5-b) indole derivatives Withdrawn EP1442039A1 (en)

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JP2001334662 2001-10-31
JP2001334662A JP2003137886A (en) 2001-10-31 2001-10-31 PYRIMIDO[4,5-b]INDOLE
GB0214544A GB2390090A (en) 2002-06-24 2002-06-24 Pharmaceutically active 6-carbamoylpyrimido[4,5-b]indoles
GB0214544 2002-06-24
PCT/EP2002/012057 WO2003037898A1 (en) 2001-10-31 2002-10-29 Pyrimido [4,5-b] indole derivatives

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