Classifications

• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D307/00—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom
  • C07D307/77—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom ortho- or peri-condensed with carbocyclic rings or ring systems
  • C07D307/78—Benzo [b] furans; Hydrogenated benzo [b] furans
  • C07D307/79—Benzo [b] furans; Hydrogenated benzo [b] furans with only hydrogen atoms, hydrocarbon or substituted hydrocarbon radicals, directly attached to carbon atoms of the hetero ring
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
  • A61P1/08—Drugs for disorders of the alimentary tract or the digestive system for nausea, cinetosis or vertigo; Antiemetics
• • 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
  • A61P25/24—Antidepressants
• • 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
  • A61P35/00—Antineoplastic 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

Definitions

• the invention relates to heterocyclic sulfonamide compounds and pharmaceutical compositions thereof, in particular heterocyclic sulfonamide compounds that are specific inhibitors of kinase activity of MEK.
• the invention also relates to the use of the compounds and compositions thereof in the management of hyperproliferative diseases like cancer and inflammation.
• Hyperproliferative diseases like cancer and inflammation are receiving a lot of attention from the scientific community and there is a strong desire to discover compounds that provide therapeutic benefits with regard to treating hyperproliferative diseases. In this regard efforts have been made to identify and target specific mechanisms which play a role in proliferating the diseases.
• MAP mitogen-activated protein
• MEK is an attractive therapeutic target because the only known substrates for MEK phosphorylation are the MAP kinases, ERKl and ERK2. Constitutive activation of MEK/ERK was been found in pancreatic, colon, lung, kidney and ovarian primary tumor samples.
• Stroke see, e.g., Evidence of efficacy in stroke models significant neuroprotection against ischemic brain injury by inhibition of the
• the invention provides the compounds (S)-N-(4,5-difluoro-6-((2- fluoro-4-iodophenyl)amino)benzofuran-7-yl)- 1 -(2,3-dihydroxypropyl)cyclopropane- 1 - sulfonamide and (R)-N-(4,5-difluoro-6-((2-fluoro-4-iodophenyl)amino)benzofuran-7-yl)- l-(2,3-dihydroxypropyl)cyclopropane-l-sulfonamide:
• a pharmaceutical composition which comprises the compounds described above, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.
• compound of the present invention refers to (S)-N-(4,5-difluoro-6-((2-fluoro-4-iodophenyl)amino)benzofuran-7- yl)- 1 -(2,3-dihydroxypropyl)cyclopropane- 1 -sulfonamide or (R)-N-(4,5-difluoro-6-((2- fluoro-4-iodophenyl)amino)benzofuran-7-yl)- 1 -(2,3-dihydroxypropyl)cyclopropane- 1 - sulfonamide, and salts thereof, as well as all isotopically labeled compounds (including deuterium substitutions), and inherently formed moieties (e.g., polymorphs, solvates and/or hydrates).
• the present invention provides compounds and pharmaceutical compositions thereof that are useful in the treatment of diseases, conditions and/or disorders modulated by the inhibition of kinase activity of MEK.
• Compounds of the present invention may be synthesized by synthetic routes that include processes analogous to those well-known in the chemical arts, particularly in light of the description contained herein.
• the starting materials are generally available from commercial sources such as Aldrich Chemicals (Milwaukee, Wis.) or are readily prepared using methods well known to those skilled in the art (e.g., prepared by methods generally described in Louis F. Fieser and Mary Fieser, Reagents for Organic Synthesis, v. 1-19, Wiley, New York (1967-1999 ed.), or Beilsteins Handbuch der organischen Chemie, 4, Aufl. ed. Springer- Verlag, Berlin, including supplements (also available via the Beilstein online database)).
• Beilsteins Handbuch der organischen Chemie 4, Aufl. ed. Springer- Verlag, Berlin, including supplements (also available via the Beilstein online database)
• compositions of the invention are capable of forming salts with bases, in particular pharmaceutically acceptable bases such as those well known in the art; suitable such salts include metal salts, particularly alkali metal or alkaline earth metal salts such as sodium, potassium, magnesium or calcium salts, or salts with ammonia or
• organic amines or heterocyclic bases such as ethanolamines, benzylamines or pyridine.
• These salts may be prepared by known salt- forming procedures.
• the present invention includes isotopically-labeled or -enriched compounds of the present invention.
• isotopes suitable for inclusion in the compounds of the invention include isotopes of hydrogen, such as 2 H and 3 H, carbon, such as n C, 13 C and 14 C, chlorine, such as 36 C1, fluorine, such as 18 F, iodine, such as 123 I and 125 I, nitrogen, such as 13 N and 15 N, oxygen, such as 15 0, 17 0 and 18 0, phosphorus, such as 32 P, and sulphur, such as 35 S.
• Substitution with heavier isotopes such as deuterium, i.e. 2 H, may afford certain therapeutic advantages resulting from greater metabolic stability, for example, increased in vivo half-life or reduced dosage requirements, and hence may be preferred in some circumstances.
• Isotopically-labeled compounds of the present invention can generally be prepared by conventional techniques known to those skilled in the art or by processes analogous to those described in the accompanying Examples and Preparations Sections using an appropriate isotopically-labeled reagent in place of the non-labeled reagent previously employed.
• solvates are considered pharmaceutical compositions, e.g., a compound of the present invention in combination with an excipient, wherein the excipient is a solvent.
• the present invention also relates to a pharmaceutical composition
• a pharmaceutical composition comprising a compound of the present invention and a pharmaceutically acceptable excipient.
• Suitable excipients generally include corn starch, potato starch, tapioca starch, starch paste, pre-gelatinized starch, sugars, gelatin, natural gums, synthetic gums, sodium alginate, alginic acid, tragacanth, guar gum, cellulose, ethyl cellulose, cellulose acetate, carboxymethyl cellulose calcium, sodium carboxymethylcellulose, methyl cellulose, hydroxypropyl methylcellulose, microcrystalline cellulose, magnesium aluminum silicate, polyvinyl pyrrolidone, talc, calcium carbonate, powdered cellulose, dextrates, kaolin, mannitol, silicic acid, sorbitol, agar-agar, sodium carbonate, croscarmellose sodium, crospovidone, polacrilin potassium, sodium starch glycolate, clays, sodium stearate, calcium stearate, magnesium stearate, stearic acid, mineral oil, light mineral oil, g
• a typical formulation is prepared by mixing a compound of the present invention and a carrier, diluent or excipient.
• Suitable carriers, diluents and excipients are well known to those skilled in the art and include materials such as carbohydrates, waxes, water soluble and/or swellable polymers, hydrophilic or hydrophobic materials, gelatin, oils, solvents, water, and the like.
• the particular carrier, diluent or excipient used will depend upon the means and purpose for which the compound of the present invention is being applied. Solvents are generally selected based on solvents recognized by persons skilled in the art as safe (GRAS) to be administered to a mammal.
• GRAS solvents recognized by persons skilled in the art as safe
• safe solvents are non-toxic aqueous solvents such as water and other non-toxic solvents that are soluble or miscible in water.
• Suitable aqueous solvents include water, ethanol, propylene glycol, polyethylene glycols (e.g., PEG400, PEG300), etc. and mixtures thereof.
• the formulations may also include one or more buffers, stabilizing agents, surfactants, wetting agents, lubricating agents, emulsifiers, suspending agents, preservatives, antioxidants, opaquing agents, glidants, processing aids, colorants, sweeteners, perfuming agents, flavoring agents and other known additives to provide an elegant presentation of the drug (i.e., a compound of the present invention or
• composition thereof or aid in the manufacturing of the pharmaceutical product (i.e., medicament).
• the formulations may be prepared using conventional dissolution and mixing procedures.
• the bulk drug substance i.e., compound of the present invention or stabilized form of the compound (e.g., complex with a cyclodextrin derivative or other known complexation agent)
• a suitable solvent in the presence of one or more of the excipients.
• the compound of the present invention is typically formulated into pharmaceutical dosage forms to provide an easily controllable dosage of the drug and to give the patient an elegant and easily handleable product.
• composition is generally formulated into various dosage forms selected from a group comprising tablet, troches, lozenges, aqueous or oily suspensions, ointment, patch, gel, lotion, dentifrice, capsule, emulsion, creams, spray, drops, dispersible powders or granules, emulsion in hard or soft gel capsules, syrups and elixirs.
• the pharmaceutical composition (or formulation) for application may be packaged in a variety of ways depending upon the method used for administering the drug.
• an article for distribution includes a container having deposited therein the pharmaceutical formulation in an appropriate form.
• Suitable containers are well- known to those skilled in the art and include materials such as bottles (plastic and glass), sachets, ampoules, plastic bags, metal cylinders, and the like.
• the container may also include a tamper-proof assemblage to prevent indiscreet access to the contents of the package.
• the container has deposited thereon a label that describes the contents of the container. The label may also include appropriate warnings.
• the compounds of the present invention are useful as both prophylactic and therapeutic treatments for diseases or conditions related to the hyperactivity of MEK, as well as diseases or conditions modulated by the Raf/Ras/Mek pathway.
• the invention relates to a method for treating a disease or condition related to the hyperactivity of MEK, or a disease or condition modulated by the MEK cascade, comprising administration of an effective therapeutic amount of a compound of the present invention.
• the invention relates to a method for treating proliferative diseases, such as cancer, comprising administration of an effective amount of a compound of the present invention.
• cancers include but are not limited to: angiosarcoma, fibrosarcoma, rhabdomyosarcoma, liposarcoma, myxoma, rhabdomyoma, fibroma, lipoma, teratoma; bronchogenic carcinoma, squamous cell carcinoma, undifferentiated small cell carcinoma, undifferentiated large cell carcinoma, alveolar (bronchiolar) carcinoma, bronchial adenoma, lymphoma, chondromatous hanlartoma, inesothelioma, esophageal squamous cell carcinoma, leiomyosarcoma, leiomyosarcoma, ductal adenocarcinoma, insulinoma, glucagonoma, gastrinoma, vipoma, stomach and small bowel carcinoid tumors, adenocarcinoma, Karposi's sarcoma, leiomyoma, leiomyo
• osteoma chondromyxofibroma, osteoid osteoma and giant cell tumors, osteoma, granuloma, xanthoma, osteitis deformans, meningioma, meningiosarcoma, gliomatosis, astrocytoma, medulloblastoma, glioma, ependymoma, germinoma [pinealoma], glioblastoma multiform, oligodendroglioma, schwannoma, retinoblastoma, congenital tumors, spinal cord neurofibroma, meningioma, glioma, endometrial carcinoma, cervical carcinoma, pre-tumor cervical dysplasia, ovarian carcinoma, serous cystadenocarcinoma, mucinous cystadenocarcinoma, granulosa-thecal cell tumors, Sertoli-Leydig cell tumors, dysgermin
• rhabdomyosarcoma fallopian tube carcinoma, acute and chronic myeloid leukemia, acute lymphoblastic leukemia, chronic lymphocytic leukemia, myeloproliferative diseases, multiple myeloma, myelodysplastic syndrome, Hodgkin's disease, non- Hodgkin's lymphoma, malignant lymphoma, malignant melanoma, basal cell carcinoma, moles, dysplastic nevi, angioma, dermatofibroma, keloids, psoriasis, and neuroblastoma.
• the present invention includes a method of treating cancer selected from non-small cell lung carcinoma, pancreatic carcinoma, bladder carcinoma, colon carcinoma, myeloid disorders, breast cancer, prostate cancer, thyroid cancer, melanoma, adenomas and carcinomas of the ovary, eye, liver, biliary tract, and nervous system and advanced solid tumors with KRAS, NRAS and BRAF mutations, comprising
• the compounds of the present invention may also be useful in the treatment of other diseases or conditions related to the hyperactivity of MEK.
• the invention relates to a method of treatment of a disorder selected from: xenograft (cellos), skin, limb, organ or bone marrow transplant) rejection; osteoarthritis; rheumatoid arthritis; cystic fibrosis; complications of diabetes (including diabetic retinopathy and diabetic nephropathy); hepatomegaly; cardiomegaly; stroke (such as acute focal ischemic stroke and global cerebral ischemia); heart failure; septic shock; asthma; chronic obstructive pulmonary disorder; Alzheimer's disease; and chronic or neuropathic pain.
• a disorder selected from: xenograft (cellos), skin, limb, organ or bone marrow transplant) rejection; osteoarthritis; rheumatoid arthritis; cystic fibrosis; complications of diabetes (including diabetic retinopathy and diabetic nephropathy);
• chronic pain for purposes of the present invention includes, but is not limited to, idiopathic pain, and pain associated with chronic alcoholism, vitamin deficiency, uremia, or hypothyroidism. Chronic pain is associated with numerous conditions including, but not limited to, inflammation, and post-operative pain.
• neurodegeneration pain is associated with numerous conditions which include, but are not limited to, inflammation, postoperative pain, phantom limb pain, burn pain, gout, trigeminal neuralgia, acute herpetic and postherpetic pain, causalgia, diabetic neuropathy, plexus avulsion, neuroma, vasculitis, viral infection, crush injury, constriction injury, tissue injury, limb amputation, and nerve injury between the peripheral nervous system and the central nervous system.
• Compounds of the present invention may also be useful as antiviral agents for treating viral infections such as HIV, hepatitis (B) virus (HBV) human papilloma virus (HPV), cytomegalovirus (CMV], and Epstein-Barr virus (EBV).
• HIV hepatitis
• HBV hepatitis virus
• HPV human papilloma virus
• CMV cytomegalovirus
• EBV Epstein-Barr virus
• Compounds of the present invention may also be useful in the treatment of restenosis, psoriasis, allergic contact dermatitis, autoimmune disease, atherosclerosis and inflammatory bowel diseases, e.g. Crohn's disease and ulcerative colitis.
• a MEK inhibitor of the present invention may be usefully combined with another pharmacologically active compound (additional therapeutic agent), or with two or more other pharmacologically active compounds, particularly in the treatment of cancer.
• additional therapeutic agent pharmacologically active compound
• a compound of the present invention, as defined above may be administered simultaneously, sequentially or separately in combination with one or more additional therapeutic agents selected from an anticancer drug (or chemotherapy agent), a pain medication, an antiemetic, an antidepressant or an anti-inflammatory agent.
• Chemotherapy agents include, for example, mitotic inhibitors such as a taxane, a vinca alkaloid, paclitaxel, docetaxel, vincristine, vinblastine, vinorelbine or vinflunine, and other anticancer agents, e.g. cisplatin, 5-fluorouracil or 5-fluoro-2-4(l H,3H)- pyrimidinedione (5FU), flutamide or gemcitabine.
• mitotic inhibitors such as a taxane, a vinca alkaloid, paclitaxel, docetaxel, vincristine, vinblastine, vinorelbine or vinflunine
• other anticancer agents e.g. cisplatin, 5-fluorouracil or 5-fluoro-2-4(l H,3H)- pyrimidinedione (5FU), flutamide or gemcitabine.
• a compound of the present invention may also be used to advantage in combination with other antiproliferative compounds.
• antiproliferative compounds include, but are not limited to aromatase inhibitors; antiestrogens; topoisomerase I inhibitors; topoisomerase II inhibitors; microtubule active compounds; alkylating compounds; histone deacetylase inhibitors, such as LBH589; compounds which induce cell differentiation processes; cyclooxygenase inhibitors; MMP inhibitors; mTOR inhibitors, such as RAD001; antineoplastic antimetabolites; platin compounds;
• antiproliferative antibodies include heparanase inhibitors; inhibitors of Ras oncogenic isoforms; telomerase inhibitors; proteasome inhibitors; compounds used in the treatment of hematologic malignancies; compounds which target, decrease or inhibit the activity of Flt-3, such as PKC412; Hsp90 inhibitors such as 17-AAG (17-allylamino-gelda-namycin, NSC330507), 17-DMAG (17-dimethylaminoethylamino- 17-demethoxy-geldana-mycin, NSC707545), IPI-504, CNFIOIO, CNF2024, CNFIOIO from Conforma Therapeutics and AUY922; temozolomide (TEMODAL); kinesin spindle protein inhibitors, such as SB715992 or SB743921 from GlaxoSmithKline, or pentamidine/chlorpromazine from CombinatoRx; PI3K inhibitors, such as BEZ235; RAF inhibitors, such
• tumor treatment approaches including surgery, ionizing radiation, photodynamic therapy, implants, e.g. with corticosteroids, hormones, or they may be used as radiosensitizers.
• implants e.g. with corticosteroids, hormones, or they may be used as radiosensitizers.
• anti-inflammatory and/or antiproliferative treatment combination with anti-inflammatory drugs is included. Combination is also possible with antihistamine drug substances, bronchodilatatory drugs, NSAID or antagonists of chemokine receptors.
• aromatase inhibitor as used herein relates to a compound which inhibits the estrogen production, i.e. the conversion of the substrates
• the term includes, but is not limited to steroids, especially atame-stane, exemestane and formestane and, in part-icular, non-steroids, especially aminoglutethimide, roglethimide,
• Exemestane can be administered, e.g., in the form as it is marketed, e.g. under the trademark AROMASIN.
• Formestane can be administered, e.g., in the form as it is marketed, e.g. under the trademark LENTARON.
• Fadrozole can be administered, e.g., in the form as it is marketed, e.g. un-der the trademark AFEMA.
• Anastrozole can be administered, e.g., in the form as it is marketed, e.g. under the trademark ARIMIDEX.
• Letrozole can be administered, e.g., in the form as it is marketed, e.g. under the trademark FEMARA or FEMAR.
• Amino glutethimide can be
• anti-estrogen as used herein relates to a compound which antagonizes the effect of estrogens at the estrogen receptor level.
• the term includes, but is not limited to tamoxifen, fulvestrant, raloxifene and raloxifene hydrochloride.
• Tamoxifen can be administered, e.g., in the form as it is marketed, e.g. under the trademark NOLVADEX.
• Ralo-xifene hydrochloride can be administered, e.g., in the form as it is marketed, e.g. under the trademark EVISTA.
• Fulvestrant can be formulated as disclosed in US 4,659,516 or it can be administered, e.g., in the form as it is marketed, e.g. under the trademark FASLODEX.
• a combination of the invention comprising a chemotherapeutic agent which is an anti-estrogen is particularly useful for the treatment of estrogen receptor positive tumors, e.g. breast tumors.
• anti-androgen as used herein relates to any substance which is capable of in-hibiting the biological effects of androgenic hormones and includes, but is not limited to, bicalutamide (CASODEX), which can be formulated, e.g. as disclosed in US 4,636,505.
• CASODEX bicalutamide
• gonadorelin agonist includes, but is not limited to abarelix, goserelin and goserelin acetate. Goserelin is disclosed in US
• 4,100,274 and can be administered, e.g., in the form as it is marketed, e.g. under the trademark ZOLADEX.
• Abarelix can be formulated, e.g. as disclosed in US 5,843,901.
• topoisomerase I inhibitor includes, but is not limited to topotecan, gimatecan, irinotecan, camptothecin and its analogues, 9- nitrocamptothecin and the macromolecular camptothecin conjugate PNU-166148 (compound Al in W099/ 17804).
• Irinotecan can be administered, e.g. in the form as it is marketed, e.g. under the trademark CAMPTOSAR.
• Topotecan can be administered, e.g., in the form as it is marketed, e.g. under the trademark HYCAMTIN.
• topoisomerase II inhibitor includes, but is not limited to the an-thracyclines such as doxorubicin (including liposomal formulation, e.g. CAELYX), daunorubicin, epirubicin, idarubicin and nemorubicin, the anthraquinones mitoxantrone and losoxantrone, and the podophillotoxines etoposide and teniposide.
• Etoposide can be administered, e.g. in the form as it is marketed, e.g. under the trademark ETOPOPHOS.
• Teniposide can be administered, e.g. in the form as it is marketed, e.g.
• Doxorubicin can be administered, e.g. in the form as it is marketed, e.g. under the trademark ADRIBLASTIN or ADRIAMYCIN.
• Epirubicin can be administered, e.g. in the form as it is marketed, e.g. under the trademark FARMORUBICIN.
• Idarubicin can be administered, e.g. in the form as it is marketed, e.g. under the trademark ZAVEDOS.
• Mitoxantrone can be administered, e.g. in the form as it is marketed, e.g. under the trademark NOVANTRON.
• microtubule active compound relates to microtubule stabilizing, microtubule destabilizing compounds and microtublin polymerization inhibitors including, but not limited to taxanes, e.g. paclitaxel and docetaxel, vinca alkaloids, e.g., vinblastine, especially vinblastine sulfate, vincristine especially vincristine sulfate, and vinorelbine, discodermolides, cochicine and epothilones and derivatives thereof, e.g. epothilone B or D or derivatives thereof.
• Paclitaxel may be administered e.g. in the form as it is marketed, e.g. TAXOL.
• Docetaxel can be administered, e.g., in the form as it is marketed, e.g. under the trademark TAXOTERE.
• Vinblastine sulfate can be administered, e.g., in the form as it is marketed, e.g. under the trademark VINBLASTIN R.P.
• Vincristine sulfate can be administered, e.g., in the form as it is marketed, e.g. under the trademark FARMISTIN.
• Discodermolide can be obtained, e.g., as disclosed in US 5,010,099. Also included are Epothilone derivatives which are disclosed in WO
• Epothilone A and/or B are Especially preferred.
• alkylating compound includes, but is not limited to, cyclophosphamide, ifosfamide, melphalan or nitrosourea (BCNU or Gliadel).
• Cyclophosphamide can be administered, e.g., in the form as it is marketed, e.g. under the trademark CYCLOSTIN.
• Ifosfamide can be administered, e.g., in the form as it is marketed, e.g., under the trademark HOLOXAN.
• histone deacetylase inhibitors or "HDAC inhibitors” relates to compounds which inhibit the histone deacetylase and which possess antiproliferative activity. This includes compounds such as sodium butyrate, LDH589 disclosed in WO 02/22577, especially N-hydroxy-3-[4-[[(2-hydroxyethyl)[2-(lH-indol-3-yl)ethyl]- amino]methyl] phenyl]-2E-2-propenamide, N-hydroxy-3-[4-[[[2-(2 -methyl- lH-indol-3- yl)-ethyl] -amino] methyl]phenyl]-2E-2-propenamide and pharmaceutically acceptable salts thereof, especially the lactate salt.
• SAHA suberoylanilide hydroxamic acid
• MS275 MS275
• FK228 formerly FR9012228
• trichostatin A compounds disclosed in US 6,552,065, in particular, N-hydroxy-3-[4-[[[2-(2-methyl-lH- indol-3-yl)-ethyl]-amino]-methyl]phenyl]-2E-2-propenamide, or a pharmaceutically acceptable salt thereof.
• antiproliferative antimetabolite includes, but is not limited to,
• 5-Fluorouracil or 5-FU capecitabine, gemcitabine, DNA demethylating compounds, such as 5-azacy-ti-dine and decitabine, methotrexate and edatrexate, and folic acid antagonists such as pemetrexed.
• Capecitabine can be administe-red, e.g., in the form as it is marketed, e.g. under the trademark XELODA.
• Gemcitabine can be administered, e.g., in the form as it is marketed, e.g. under the trademark GEMZAR.
• platinum compound as used herein includes, but is not limited to, carboplatin, cis-platin, cisplatinum and oxaliplatin.
• Carboplatin can be administered, e.g., in the form as it is marketed, e.g. under the trademark CARBOPLAT.
• Oxaliplatin can be administered, e.g., in the form as it is marketed, e.g. under the trademark
• the term "compounds targeting/decreasing a protein or lipid kinase activity”; or a “protein or lipid phosphatase activity”; or “further anti-angiogenic compounds” as used herein includes, but is not limited to, protein tyrosine kinase and/or serine and/or threonine kinase inhibitors or lipid kinase inhibitors, e.g.,
• PDGFR platelet-derived growth factor-receptors
• FGFR fibroblast growth factor-receptors
• IGF-IR insulin-like growth factor receptor I
• compounds targeting, decreasing or inhibiting the activity of the insulin-like growth factor receptor I such as compounds which target, decrease or inhibit the activity of IGF-IR, especially compounds which inhibit the kinase activity of IGF-I receptor, such as those compounds disclosed in WO 02/092599, or antibodies that target the extracellular domain of IGF-I receptor or its growth factors;
• Trk receptor tyrosine kinase family Trk receptor tyrosine kinase family, or ephrin B4 inhibitors; [0055] e) compounds targeting, decreasing or inhibiting the activity of the
• Kit/SCFR receptor tyrosine kinase i.e C-kit receptor tyrosine kinases - (part of the PDGFR family), such as compounds which target, decrease or inhibit the activity of the c- Kit receptor tyrosine kinase family, especially compounds which inhibit the c-Kit receptor, e.g. imatinib;
• a N-phenyl-2-pyrimidine-amine derivative e.g. imatinib or nilotinib (AMN107)
• PD180970 AG957; NSC 680410; PD173955 from ParkeDavis; or dasatinib (BMS-354825)
• LY333531/LY379196 isochinoline compounds such as those disclosed in WO 00/09495; FTIs; BEZ235 (a P13K inhibitor) or AT7519 (CDK inhibitor);
• [0060] j) compounds targeting, decreasing or inhibiting the activity of protein- tyrosine kinase inhibitors such as compounds which target, decrease or inhibit the activity of protein-tyrosine kinase inhibitors include imatinib mesylate (GLEEVEC) or tyrphostin.
• a tyrphostin is preferably a low molecular weight (mw ⁇ 1500) compound, or a pharmaceutically acceptable salt thereof, especially a compound selected from the benzylidenemalonitrile class or the S-arylbenzenemalonirile or bisubstrate quinoline class of compounds, more especially any compound selected from the group consisting of Tyrphostin A23/RG-50810; AG 99; Tyrphostin AG 213; Tyrphostin AG 1748; Tyrphostin AG 490; Tyrphostin B44; Tyrphostin B44 (+) enantiomer; Tyrphostin AG 555; AG 494; Tyrphostin AG 556, AG957 and adaphostin (4- ⁇ [(2,5- dihydroxyphenyl)methyl] amino ⁇ -benzoic acid adamantyl ester; NSC 680410, adaphostin);
• k) compounds targeting, decreasing or inhibiting the activity of the epidermal growth factor family of receptor tyrosine kinases (EGFR, ErbB2, ErbB3, ErbB4 as homo- or heterodimers) and their mutants, such as compounds which target, decrease or inhibit the activity of the epidermal growth factor receptor family are especially compounds, proteins or antibodies which inhibit members of the EGF receptor tyrosine kinase family, e.g. EGF receptor, ErbB2, ErbB3 and ErbB4 or bind to EGF or EGF related ligands, and are in particular those compounds, proteins or monoclonal antibodies generically and specifically disclosed in WO 97/02266, e.g. the compound of ex.
• trastuzumab Herceptin
• cetuximab Erbitux
• Iressa Tarceva
• OSI-774 OSI-774
• CI-1033 EKB-569, GW- 2016, El. l, E2.4, E2.5, E6.2, E6.4, E2. l l, E6.3 or E7.6.3, and 7H-pyrrolo-[2,3- d]pyrimidine derivatives which are disclosed in WO 03/013541; and
• Met receptor such as compounds which target, decrease or inhibit the activity of c-Met, especially compounds which inhibit the kinase activity of c-Met receptor, or antibodies that target the extracellular domain of c-Met or bind to HGF.
• Further anti-angiogenic compounds include compounds having another mechanism for their activity, e.g. unrelated to protein or lipid kinase inhibition e.g.
• TAALOMID thalidomide
• Compounds which target, decrease or inhibit the activity of a protein or lipid phosphatase are e.g., inhibitors of phosphatase 1, phosphatase 2A, or CDC25, e.g. okadaic acid or a derivative thereof.
• cyclooxygenase inhibitor as used herein includes, but is not limited to, e.g. Cox-2 inhibitors, 5-alkyl substituted 2-arylaminophenylacetic acid and derivatives, such as celecoxib (CELEBREX), rofecoxib (VIOXX), etoricoxib, valdecoxib or a 5-alkyl-2-arylaminophenylacetic acid, e.g. 5-methyl-2-(2'-chloro-6'- fluoroanilino)phenyl acetic acid, lumiracoxib.
• Cox-2 inhibitors 5-alkyl substituted 2-arylaminophenylacetic acid and derivatives, such as celecoxib (CELEBREX), rofecoxib (VIOXX), etoricoxib, valdecoxib or a 5-alkyl-2-arylaminophenylacetic acid, e.g. 5-methyl-2-(2'-chloro-6'- fluor
• bisphosphonates as used herein includes, but is not limited to, etridonic, clodronic, tiludronic, pamidronic, alendronic, ibandronic, risedronic and zoledronic acid.
• Etridonic acid can be administered, e.g., in the form as it is marketed, e.g. under the trademark DIDRONEL.
• Clodronic acid can be administered, e.g., in the form as it is marketed, e.g. under the trademark BONEFOS.
• titaniumudronic acid can be administered, e.g., in the form as it is marketed, e.g. under the trademark SKELID.
• “Pamidronic acid” can be administered, e.g. in the form as it is marketed, e.g. under the trademark AREDIA.
• “Alendronic acid” can be administered, e.g., in the form as it is marketed, e.g. under the trademark FOSAMAX.
• “Ibandronic acid” can be administered, e.g., in the form as it is marketed, e.g. under the trademark BONDRANAT.
• “Risedronic acid” can be administered, e.g., in the form as it is marketed, e.g. under the trademark ACTONEL.
• "Zoledronic acid” can be administered, e.g. in the form as it is marketed, e.g. under the trademark ZOMETA.
• mTOR inhibitors relates to compounds which inhibit the mammalian target of rapamycin (mTOR) and which possess antiproliferative activity such as sirolimus (Rapamune), everolimus (CerticanO), CCI-779 and ABT578.
• heparanase inhibitor refers to compounds which target, decrease or inhibit heparin sulfate degradation.
• the term includes, but is not limited to, PI-88.
• biological response modifier refers to a lymphokine or interferons, e.g. interferon.
• inhibitor of Ras oncogenic isoforms e.g. H-Ras, K-Ras, or
• N-Ras refers to compounds which target, decrease or inhibit the oncogenic activity of Ras e.g. a "farnesyl transferase inhibitor” e.g. L-744832, DK8G557 or Rl 15777 (Zarnestra).
• a "farnesyl transferase inhibitor” e.g. L-744832, DK8G557 or Rl 15777 (Zarnestra).
• telomerase inhibitor refers to compounds which target, decrease or inhibit the activity of telomerase.
• Compounds which target, decrease or inhibit the activity of telomerase are especially compounds which inhibit the telomerase receptor, e.g. telomestatin.
• methionine aminopeptidase inhibitor refers to compounds which target, decrease or inhibit the activity of methionine aminopeptidase.
• Compounds which target, decrease or inhibit the activity of methionine aminopeptidase are e.g. bengamide or a derivative thereof.
• proteasome inhibitor refers to compounds which target, decrease or inhibit the activity of the proteasome.
• Compounds which target, decrease or inhibit the activity of the proteasome include e.g. Bortezomid (Velcade) and MLN 341.
• matrix metalloproteinase inhibitor or (“MMP” inhibitor) as used herein includes, but is not limited to, collagen peptidomimetic and
• nonpeptidomimetic inhibitors tetracycline derivatives, e.g. hydroxamate peptidomimetic inhibitor batimastat and its orally bioavailable analogue marimastat (BB-2516), prinomastat (AG3340), metastat (NSC 683551) BMS-279251, BAY 12-9566, TAA211, MMI270B or AAJ996.
• FMS-like tyrosine kinase inhibitors e.g. compounds targeting, decreasing or inhibiting the activity of FMS-like tyrosine kinase receptors (Flt-3R); interferon, 1 -b-D-arabinofuransylcytosine (ara-c) and bisulfan; and ALK inhibitors e.g. compounds which target, decrease or inhibit anaplastic lymphoma kinase.
• FMS-like tyrosine kinase receptors are especially compounds, proteins or antibodies which inhibit members of the Flt-3R receptor kinase family, e.g. PKC412, TKI258, midostaurin, a staurosporine derivative, SU11248 and MLN518.
• HSP90 inhibitors includes, but is not limited to, compounds targeting, decreasing or inhibiting the intrinsic ATPase activity of HSP90; degrading, targeting, decreasing or inhibiting the HSP90 client proteins via the ubiquitin proteosome pathway.
• Compounds targeting, decreasing or inhibiting the intrinsic ATPase activity of HSP90 are especially compounds, proteins or antibodies which inhibit the ATPase activity of HSP90 e.g., 17-allylamino,17-demethoxygeldanamycin (17AAG), a geldanamycin derivative; other geldanamycin related compounds, and radicicol.
• antiproliferative antibodies includes, but is not limited to, trastuzumab (Herceptin), Trastuzumab-DMl,erbitux, bevacizumab (Avastin), rituximab (Rituxan), PR064553 (anti-CD40) and 2C4 Antibody.
• trastuzumab Herceptin
• Trastuzumab-DMl bevacizumab
• bevacizumab Avastin
• rituximab Rauximab
• PR064553 anti-CD40
• 2C4 Antibody 2C4 Antibody.
• antibodies is meant e.g. intact monoclonal antibodies, polyclonal antibodies,
• multispe-cific antibodies formed from at least 2 intact antibodies, and antibodies fragments so long as they exhibit the desired biological activity.
• compounds of formula (I) can be used in combination with standard leukemia therapies, especially in combination with therapies used for the treatment of AML.
• compounds of formula (I) can be administered in combination with, e.g., farnesyl transferase inhibitors and/or other drugs useful for the treatment of AML, such as Daunorubicin, Adriamycin, Ara-C, VP- 16, Teniposide, Mitoxantrone, Idarubicin, Carboplatinum and PKC412.
• antigenemic compounds includes, for example, Ara-C, a pyrimidine analog, which is the 2-alpha-hydroxy ribose (arabinoside) derivative of deoxycytidine. Also included is the purine analog of hypoxanthine, 6-mercaptopurine (6- MP) and fludarabine phosphate.
• Somatostatin receptor antagonists refers to compounds which target, treat or inhibit the somatostatin receptor such as octreotide, and SOM230 (pasireotide).
• Tumor cell damaging approaches refer to approaches such as ionizing radiation.
• ionizing radiation means ionizing radiation that occurs as either electromagnetic rays (such as X-rays and gamma rays) or particles (such as alpha and beta particles). Ionizing radiation is provided in, but not limited to, radiation therapy and is known in the art. See Hellman, Principles of
• EDG binders refers a class of
• immunosuppressants that modulates lymphocyte recirculation, such as FTY720.
• ribonucleotide reductase inhibitors refers to pyrimidine or purine nucleoside analogs including, but not limited to, fludarabine and/or cytosine arabinoside (ara-C), 6-thioguanine, 5-fluorouracil, cladribine, 6-mercaptopurine
• Ribonucleotide reductase inhibitors are especially hydroxyurea or 2-hydroxy-lH-isoindole-l,3-dione derivatives, such as PL-1, PL-2, PL-3, PL-4, PL-5, PL-6, PL-7 or PL-8 mentioned in Nandy et al., Acta Oncologica, Vol. 33, No. 8, pp. 953-961 (1994).
• S-adenosylmethionine decarboxylase inhibitors includes, but is not limited to the compounds disclosed in US 5,461,076.
• VEGF vascular endothelial growth factor
• WO 98/35958 e.g. l-(4-chloroanilino)-4- (4-pyridylmethyl)phthalazine or a pharmaceutically acceptable salt thereof, e.g. the succinate, or in WO 00/09495, WO 00/27820, WO 00/59509, WO 98/11223, WO 00/27819 and EP 0 769 947; those as described by Prewett et al, Cancer Res, Vol. 59, pp. 5209-5218 (1999); Yuan et al., Proc Natl Acad Sci U S A, Vol. 93, pp.
• anthranilic acid amides ZD4190; ZD6474; SU5416; SU6668; bevacizumab; or anti- VEGF antibodies or anti-VEGF receptor antibodies, e.g. rhuMAb and RHUFab, VEGF aptamer e.g. Macugon; FLT-4 inhibitors, FLT-3 inhibitors, VEGFR-2 IgGl antibody, Angiozyme (RPI 4610) and Bevacizumab (Avastin).
• Photodynamic therapy refers to therapy which uses certain chemicals known as photosensitizing compounds to treat or prevent cancers.
• Examples of photodynamic therapy includes treatment with compounds, such as e.g. VISUDYNE and porfimer sodium.
• Angiostatic steroids refers to compounds which block or inhibit angiogenesis, such as, e.g., anecortave, triamcinolone, hydrocortisone, 11— epihydrocotisol, cortexolone, 17-hydroxyprogesterone, corticosterone,
• Implants containing corticosteroids refers to compounds, such as e.g. fluocinolone, dexamethasone.
• “Other chemotherapeutic compounds” include, but are not limited to, plant alkaloids, hormonal compounds and antagonists; biological response modifiers, preferably lymphokines or interferons; antisense oligonucleotides or oligonucleotide derivatives; shRNA or siRNA; or miscellaneous compounds or compounds with other or unknown mechanism of action.
• the compounds of the present invention may also be administered simultaneously, separately or sequentially in combination with one or more other suitable active agents selected from the following classes of agents: Anti IL-1 agents, e.g:
• Anakinra anti cytokine and anti-cytokine receptor agents, e.g. anti IL-6 R Ab, anti IL-15 Ab, anti IL-17 Ab, anti IL-12 Ab; B-cell and T-cell modulating drugs, e.g. anti CD20 Ab; CTL4-Ig, disease-modifying anti-rheumatic agents (DMARDs), e.g. methotrexate, leflunamide, sulfasalazine; gold salts, penicillamine, hydroxychloroquine and
• DMARDs disease-modifying anti-rheumatic agents
• chloroquine chloroquine, azathioprine, glucocorticoids and non-steroidal anti-inflammatories
• NSAIDs e.g. cyclooxygenase inhibitors, selective COX-2 inhibitors, agents which modulate migration of immune cells, e.g. chemokine receptor antagonists, modulators of adhesion molecules, e.g. inhibitors of LFA-1, VLA-4.
• the pharmaceutical composition or combination of the present invention can be in unit dosage of about 1-1000 mg of active ingredient(s) for a subject of about 50-70 kg, or about 1-500 mg or about 1-250 mg or about 1-150 mg or about 0.5- 100 mg, or about 1-50 mg of active ingredients.
• suitable daily dosages for oral administration are from about 0.1 to about 10 mg/kg.
• the therapeutically effective dosage of a compound, the pharmaceutical composition, or the combinations thereof is dependent on the species of the subject, the body weight, age and individual condition, the disorder or disease or the severity thereof being treated.
• a physician, clinician or veterinarian of ordinary skill can readily determine the effective amount of each of the active ingredients necessary to prevent, treat or inhibit the progress of the disorder or disease.
• the above-cited dosage properties are demonstrable in vitro and in vivo tests using advantageously mammals, e.g., mice, rats, dogs, monkeys or isolated organs, tissues and preparations thereof.
• the compounds of the present invention can be applied in vitro in the form of solutions, e.g., aqueous solutions, and in vivo either enterally, parenterally, advantageously intravenously, e.g., as a suspension or in aqueous solution.
• the dosage in vitro may range between about 10 ⁇ 3 molar and 10 ⁇ 9 molar concentrations.
• a therapeutically effective amount in vivo may range depending on the route of administration, between about 0.1-500 mg/kg, or between about 1-100 mg/kg.
• a therapeutically effective amount of a compound of the present invention is administered to a patient in need of treatment.
• a therapeutically effective amount of a compound of the present invention refers to an amount of the compound of the present invention that will elicit the biological or medical response of a subject, for example, reduction or inhibition of an enzyme or a protein activity, or ameliorate symptoms, alleviate conditions, slow or delay disease progression, or prevent a disease, etc.
• a method for treating cancer in a subject comprises administering to a mammal in need of such treatment an effective amount of a compound of the present invention.
• the term "subject" refers to an animal. Typically the animal is a mammal. A subject also refers to for example, primates ⁇ e.g., humans, male or female), cows, sheep, goats, horses, dogs, cats, rabbits, rats, mice, fish, birds and the like. In certain embodiments, the subject is a primate. Preferably, the subject is a human.
• the term “inhibit”, “inhibition” or “inhibiting” refers to the reduction or suppression of a given condition, symptom, or disorder, or disease, or a significant decrease in the baseline activity of a biological activity or process.
• the term “treat”, “treating” or “treatment” of any disease or disorder refers (i) to ameliorating the disease or disorder (i.e., slowing or arresting or reducing the development of the disease or at least one of the clinical symptoms thereof); (ii) to alleviating or ameliorating at least one physical parameter including those which may not be discernible by the patient; or (iii) to preventing or delaying the onset or development or progression of the disease or disorder.
• treating or “treatment” describes the management and care of a patient for the purpose of combating the disease, condition, or disorder and includes the administration of a compound of the present invention to prevent the onset of the symptoms or complications, alleviating the symptoms or complications, or eliminating the disease, condition or disorder.
• a subject is "in need of a treatment if such subject would benefit biologically, medically or in quality of life from such treatment.
• Another aspect of the invention is a product comprising a compound of the present invention and at least one other therapeutic agent (or pharmaceutical agent) as a combined preparation for simultaneous, separate or sequential use in therapy to enhance apoptosis.
• the compound of the present invention and the other therapeutic agent may be manufactured and/or formulated by the same or different manufacturers. Moreover, the compound of the present invention and the other therapeutic (or pharmaceutical agent) may be brought together into a combination therapy: (i) prior to release of the combination product to physicians (e.g. in the case of a kit comprising the compound of the invention and the other therapeutic agent); (ii) by the physician themselves (or under the guidance of the physician) shortly before administration; (iii) in the patient themselves, e.g. during sequential administration of the compound of the invention and the other therapeutic agent.
• the invention provides the use of a compound of the present invention for treating a disease or condition by inhibiting the MAP kinase pathway, wherein the medicament is prepared for administration with another therapeutic agent.
• the invention also provides for the use of another therapeutic agent, wherein the medicament is administered as a combination of a compound of the present invention with the other therapeutic agent.
• the present invention also includes processes for the preparation of compounds of the invention.
• it can be necessary to protect reactive functional groups, where these are desired in the final product, to avoid their unwanted participation in the reactions.
• Conventional protecting groups can be used in accordance with standard practice, for example, see T.W. Greene and P. G. M. Wuts in "Protective Groups in Organic Chemistry", John Wiley and Sons, 1991.
• a compound of formula 4a can be prepared by reacting a compound of formula 2 with a compound of formula 3 a in the presence of a suitable amine (for example, triethylamine, N,N-Diisopropy3ethy3amme, Triisopropylamine, or the like), a suitable catalyst (for example, N,N-dimethylpyridin-4-amine, or the like) and a suitable solvent (for example, DCM, 1,2-dichloromethane, toluene, tetrahydrofuranyl, or the like). The reaction proceeds at a temperature of about 25°C and can take up to about 12 hours to complete.
• a suitable amine for example, triethylamine, N,N-Diisopropy3ethy3amme, Triisopropylamine, or the like
• a suitable catalyst for example, N,N-dimethylpyridin-4-amine, or the like
• a suitable solvent for example, DCM, 1,2-d
• a compound of formula 5a can be prepared from a compound of formula 4a in the presence of a suitable base (for example, potassium trimethylsilanoate, or the like). The reaction proceeds at a temperature of about 25°C and can take up to about 12 hours to complete.
• a suitable base for example, potassium trimethylsilanoate, or the like.
• the S-enantiomer can be prepared from a compound of formula 5a in the presence of a suitable Lewis acid (for example, trichlorborane, boron trifluoride, boron tribromide, or the like) and a suitable solvent (for example, dichloromethane, 1,2- dichloromethane, or the like).
• a suitable Lewis acid for example, trichlorborane, boron trifluoride, boron tribromide, or the like
• a suitable solvent for example, dichloromethane, 1,2- dichloromethane, or the like.
• a compound of formula 4b can be prepared by reacting a compound of formula 2 with a compound of formula 3b in the presence of a suitable amine (for example, triethylamine, , ⁇ -Diisopropylethylamine, Triisopropylamine or the like), a suitable catalyst (for example, N,N-dimethylpyridin-4-amine or the like) and a suitable solvent (for example, DCM, 1,2-dichloromethane, toluene, tetrahydrofuran, or the like). The reaction proceeds at a temperature of about 25°C and can take up to about 12 hours to complete.
• a suitable amine for example, triethylamine, , ⁇ -Diisopropylethylamine, Triisopropylamine or the like
• a suitable catalyst for example, N,N-dimethylpyridin-4-amine or the like
• a suitable solvent for example, DCM, 1,2-dichloromethane, toluen
• a compound of formula 5b can be prepared from a compound of formula 4b in the presence of a suitable base (for example, potassium trimethylsilanoate, or the like). The reaction proceeds at a temperature of about 25°C and can take up to about 12 hours to complete.
• a suitable base for example, potassium trimethylsilanoate, or the like.
• the R-enantiomer can be prepared from a compound of formula 5b in the presence of a suitable Lewis acid(for example, trichlorborane boron trifluoride, boron tribromide, or the like) and a suitable solvent (for example, dichloromethane, 1,2- dichloromethane, or the like).
• a suitable Lewis acid for example, trichlorborane boron trifluoride, boron tribromide, or the like
• a suitable solvent for example, dichloromethane, 1,2- dichloromethane, or the like.
• TEA Triethylamine
• EA Ethyl acetate
• MCC Microcrystalline cellulose
• DMAP 4-Dimethylaminopyridine
• DCM Dichloromethane
• THF Tetrahydrofuran
• DMF Dimethylformamide
• LHMDS lithium bis(trimethylsilyl)amide
• CDI 1,1- Carbonyldiimidazole
• PTSA p-toluene sulfonic acid
• RT room temperature
• TLC thin layer chromatography
• NMR nuclear magnetic resonance
• LC-MS liquid chromatography- mass spectrometry
• HPLC high pressure liquid chromatography or high performance liquid chromatography
• l,2,3,5-tetrafluoro-4-nitrobenzene was prepared by precooling a solution of HN0 3 (990 g) in H 2 S0 4 (973 mL) and adding it into a cold solution of 1,2,3, 5-tetrafluorobenzene (973.1 g) in H 2 S0 4 (2920 mL) at a temperature between 0°C and 10°C over a 1.5 hour period. After the addition, the yellow solution was stirred at a temperature between 0°C and 10°C for 1 hour until analysis showed the reaction had gone to completion.
• LiHMDS (1732 mL) was added to a solution of aniline in 2.5 L of THF at between -72°C and -65°C over 30 minutes. The resultant suspension was stirred at - 70°C for 30 minutes. A solution of nitrobenzene in 300 mL of THF was then added dropwise into the above suspension at between -75°C and -70°C over 1 hour and stirred at this temperature for a further 15 minutes. An additional lOOmL of LiHMDS was added at -70°C and stirred at this temperature for 15 minutes. HPLC analysis revealed 23.4% of aniline, 6.6% nitrobenzene and 68.3%) product. The solution was concentrated at 35°C.
• 6-yl)-(2-fluoro-4-iodo-phenyl)-amine (165g) and NH 4 Cl in 2.5 L of THF and 2.5 L of water was cooled to between 0°C and 5°C.
• Zinc dust was added (portionwise) at a temperature of less than 25°C (exothermic during first half of addition) over 30 minutes. The addition of zinc dust resulted in a color change from yellow to dark purple and to pale yellow. Complete conversion was achieved with the addition of NH 4 C1 (50 g) and zinc (60 g), followed by vigorous stirring for 20 minutes and a further 1.5 hours. Purity was 89.5%). The suspension was filtered and rinsed with 1 L of water and 1.5 L of THF.
• CDI (0.144g, 0.891mmol) was added to a solution of 4,5-difluoro-N6-
• IPA (1 1 L) was added to pyridine (1850 g; 1890 mL) at 20°C with stirring for 1 hour. The solution was cooled to between - 10°C and 2°C and cyclopropane sulfonic chloride (1 100 g) was added dropwise followed by stirring at this temperature for 70 hours. NMR analysis showed 85%) of sulfonic chloride was consumed in the reaction. The reaction was cooled to 0°C and NaOH (312 g) in water (600 mL) was added dropwise. The resultant product was concentrated to dryness and 45°C and diluted with EtOAc (2750 mL), MTBE (1375 mL). The solution was stirred for 10 minutes and filtered.
• MBTE (5L) was added with stirring for 30 minutes.
• the organic and aqueous layers were separated and two additions of MBTE (5 L; 3.5 L) were used to extract the aqueous layer.
• the combined organic layer was washed with brine (1.8 L (x2)) and dried over Na 2 S0 4 for 1 hour.
• the organic layer was concentrated to dryness, distilled under vacuum (5mm Hg, 85°C) to give the product as a yellow liquid (760 g).
• N-methyl morpholine oxide (0.035g, 0.3041mmol) was added to a solution of 1-allyl-cyclopropane sulfonic acid [4,5-difluoro-6-(2-fluoro-4-iodo- phenylamino)-benzofuran-7-yl]-amide (0.167g, 0.3041mmol) in THF (5mL). This was followed by the addition of osmium tetroxide (0.0077g, 0.03041mmol) in water (lmL). The reaction mass was stirred for 16 hours at 30-40 °C. The reaction was monitored by TLC (10%) methanol in chloroform).
• racemic l-(2,3-dihydroxy-propyl)-cyclopropanesulfonic acid [4,5-difluoro-6-(2-fluoro-4-iodo-phenylamino)-benzofuran-7-yl]amide was applied to a Chiralpak® AD-HTM/03 semi prep column at 25°C.
• the mobile phase was composed of 80%) hexane/20%) IPA with a 1 mL/minute flow rate.
• the diluent was ethanol.
• the first eluted enantiomer, Example 1A had a retention time of 10.465 minutes (49%>) and the second eluted enantiomer, Example IB, had a retention time 13.535 minutes (46%>).
• the enantiomers were further purified by preparative HPLC to give
• a BRAF-MEK-ERK cascade assay is used to evaluate the effects of these compounds as inhibitors of the MAP kinase pathway.
• An enzymatic cascade assay is set up using recombinant human activated BRAF (V599E) kinase (Cat No. 14-557), human full length MEK1 kinase (Cat No. 14-706) and human full length active MAP Kinase 2/ERK2 (Cat No. 14-536) enzymes procured from Upstate.
• TR-FRET Time resolved fluorescence resonance energy transfer
• the assay buffer solution contains 50 mM Tris pH 7.5, 10 mM MgC12 , 1 mM DTT, 0.01 % Tween 20, 0.1 nM activated BRAF, 2 nM inactive ⁇ , ⁇ nM inactive ERK2, 100 ⁇ ATP and 500 nM long chain biotin-peptide substrate (LCB- FFKNIVTPRTPPP) in a 384 well format.
• the kinase reaction is stopped after 90 minutes with 10 mM EDTA and Lance detection mix (2 nM Eu-labeled phospho-serine/threonine antibody (Cat. No.ADOl 76-Perkin Elmer), 20 nM SA-APC (Cat No.
• the TR-FRET signal (Excitation at 340 nm, Emission at 615 nm and 665 nm) is read with 50 ⁇ 8 delay time on a Victor3 V fluorimeter. The data is calculated using the ratio of readings at 665nm to 615 nm.
• the final concentration of DMSO is 2.5 % in the assay. Compounds are screened at 10 ⁇ concentration with pre-incubation of the enzymes in the presence of test compound for 45 minutes.
• Each individual IC50 is determined using a 10 point dose response curve generated by GraphPad Prism software Version 4 (San Diego, California, USA) using non linear regression curve fit for sigmoidal dose response (variable slope).
• the assay buffer solution contains 50 mM Tris pH 7.5, 10 mM MgCl 2 ,
• nM DTT 1 mM DTT, 0.01 % Tween 20, 1 nM activated ERK2, 100 ⁇ ATP and 500 nM long chain biotin-peptide substrate (LCB- FFKNIVTPRTPPP) in a 384 well format.
• the kinase reaction is stopped after 90 minutes with 10 mM EDTA and Lance detection mix (2 nM Eu-labeled phospho-serine/threonine antibody (Cat.No. ADO 176-Perkin Elmer), 20 nM SA-APC (Cat. No. CR130-100-Perkin Elmer) is added.
• the TR-FRET signal (excitation at 340 nm, emission at 615 nm and 665 nm) is read with 50 ⁇ 8 delay time on Victor3 V fluorimeter. The data is calculated using the ratio of readings at 665nm to 615 nm. The final concentration of DMSO is 2.5 % in the assay. Compounds are screened at 10 ⁇ concentration with pre-incubation of the enzymes in the presence of test compound for 45 minutes.
• the radioactive filter binding assay is standardized using recombinant human activated BRAF (V599E) kinase (Cat No. 14-557) and kinase dead MEKl (K97R) ( Cat No. 14-737) procured from Upstate.
• the incorporation of 32P into MEKl (K97R) by BRAF (V599E) is measured with final assay buffer conditions of 50 mM Tris pH 7.5, 10 mM MgC12 , 1 mM DTT, 100 mM sucrose, 100 ⁇ sodium orthovanadate,5 ⁇ ATP and 2 ⁇ [ ⁇ 32P] ATP and 500 mg MEKl Kinase dead substrate.
• the enzymatic reaction is stopped after 120 minutes with 8N HC1 (hydrochloric acid) and 1 mM ATP.
• the solution is spotted on P81 filter paper and washed 4 times with 0.75 %
• the cell viability assay in A375 cells is set up in a 96-well plate format using XTT.
• XTT is a yellow tetrazolium salt that is cleaved to an orange formazan dye by the mitochondria of metabolically active cells. The procedure allows for rapid determination in a microtitre plate, to give reproducible and sensitive results.
• A375 cells are grown in DMEM media containing 10% FBS and ImM sodium pyruvate. Cells are trypsinized and seeded at 1000 cells/well. After allowing the cells to adhere overnight, compound is added to the wells at the following final concentrations: 10, 3, 1, 0.3, 0.1, 0.03, 0.01, 0.001, and 0.0001 ⁇ . The assay is set up in triplicates for each concentration. DMSO concentrations are kept at 0.5% /well. Three days after compound addition, the XTT assay is performed. Wells are washed once with PBS. 100 ⁇ of DMEM media without phenol red or FBS is added to each well. A working solution of XTT containing lmg/ml XTT and 100 ⁇ ⁇ of PMS (stock
• Percentage viability is calculated considering the blank subtracted value from wells treated with DMSO alone as 100% viable.
• GI5 0 values are calculated using Graphpad Prism, using non-linear regression curve fit for sigmoidal dose response (variable slope). Compounds of the invention were evaluated in this cell viability assay.
• Example 1A has a GI5 0 of 13.4 nM and
• Example IB has a GI 50 of 29.6 nM, while Example 1 has a GI5 0 of 17.6 nM.
• test compounds were diluted in DMSO to generate a concentration curve. A 5 mM stock was used for the highest concentration at 500 times; to yield a final concentration of 10 ⁇ with 3-fold dilutions down to 0.000 ⁇ . 1 ⁇ of diluted compound was added to 500 ⁇ cell culture media and mixed well. Media was removed from cells and 200 ⁇ of media containing compound was added. Cells were treated for 3 hrs with compound at 37°C, 5% C0 2 .
• EC 50 values were calculated using DMSO diluent values as 0% inhibition and counts of the highest tested concentration of the reference inhibitor as 100% inhibition. All the concentrations along with DMSO were done in triplicates.
• the EC50 of Example 1A is 6.4 nM compared with 15.6nM for Example IB and 12.2nM for Example 1.
• Examples 1A and IB show improved PK-parameters compared to
• Example 1 (one-way analysis of variance).
• A375 B-RafV 600E human melanoma model in rat - PK-PD experiment [00163] A375 cells were thawed using a 37°C water bath. Cells were transferred to a tube containing lOmls of warm DMEM medium. The tube was centrifuged for 5 minutes at 1200 rpm and the supernatant was discarded. The cell pellet was re-suspended and transferred to a 75 cm 2 tissue culture flask containing 15mls of medium and cultured at 37°C in a 5% CO 2 incubator.
• Rats were treated orally with a single dose of 10, 30 and 60 mg/kg p.o. of Example 1A and 30 mg/kg of Example IB. Plasma and tumor samples were taken at 4, 12, 24 and 36 hours, post dosing. The mRNA expression levels of two direct target genes of MEK substrate P-Erk (DUSP6 and SPRY4) and an indirect target (BMF) can be measured. Upon treatment with MEK inhibitors, these genes have been shown to be regulated in a dose-dependent manner in tumor cell lines grown in vitro and in vivo. The tumor samples were pulverized, extracted and studied for expression of the transcription factor DUSP6 using a real time quantitative PCR.
• DUSP6 and SPRY4 direct target genes of MEK substrate P-Erk
• BMF indirect target

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  • 2012-06-06 JP JP2014514204A patent/JP2014517004A/ja active Pending
  • 2012-06-06 WO PCT/IB2012/052860 patent/WO2012168884A1/en active Application Filing

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